US20130064846A1 - Neisserial antigens - Google Patents

Abstract

The invention provides proteins from Neisseria meningitidis (strains A & B) and from Neisseria gonorrhoeae, including amino acid sequences, the corresponding nucleotide sequences, expression data, and serological data. The proteins are useful antigens for vaccines, immunogenic compositions, and/or diagnostics.

Description

CROSS REFERENCE TO RELATED APPLICATIONS
• This application is a Divisional application of U.S. patent application Ser. No. 12/653,954, filed, Dec. 18, 2009, which is a Divisional of U.S. patent application Ser. No. 10/864,684, filed Jun. 8, 2004, now U.S. Pat. No. 7,655,245, which is a continuation application of U.S. patent application Ser. No. 09/303,518, filed Apr. 30, 1999, now U.S. Pat. No. 6,914,131, which is a continuation-in-part of International Patent Application PCT/IB1998/001665, filed Oct. 9, 1998, from which applications priority is claimed pursuant to 35 U.S.C. §120. PCT/IB1998/001665 claims priority to Great Britain Patent Applications No. 9723516.2, filed Nov. 6, 1997; No. 9724190.5, filed Nov. 14, 1997; No. 9724386.9, filed Nov. 18, 1997; No. 9725158.1, filed Nov. 27, 1997; No. 9726147.3, filed Dec. 10, 1997; No. 9800759.4, filed Jan. 14, 1998; No. 9819016.8, filed Sep. 1, 1998. All of the above applications are incorporated herein by reference in their entirety.
SUBMISSION OF SEQUENCE LISTING ON ASCII TEXT FILE
• The content of the following submission on ASCII text file is incorporated herein by reference in its entirety: a computer readable form (CRF) of the Sequence Listing (file name: 223002099611SUBSEQLIST.txt, date recorded: Oct. 1, 2012, size: 2,099 KB).
• This invention relates to antigens from Neisseria bacteria.
BACKGROUND ART
• Neisseria meningitidis and Neisseria gonorrhoeae are non-motile, gram negative diplococci that are pathogenic in humans. N. meningitidis colonises the pharynx and causes meningitis (and, occasionally, septicaemia in the absence of meningitis); N. gonorrhoeae colonises the genital tract and causes gonorrhea. Although colonising different areas of the body and causing completely different diseases, the two pathogens are closely related, although one feature that clearly differentiates meningococcus from gonococcus is the presence of a polysaccharide capsule that is present in all pathogenic meningococci.
• N. gonorrhoeae caused approximately 800,000 cases per year during the period 1983-1990 in the United States alone (chapter by Meitzner & Cohen, “Vaccines Against Gonococcal Infection”, In: New Generation Vaccines, 2nd edition, ed. Levine, Woodrow, Kaper, & Cobon, Marcel Dekker, New York, 1997, pp. 817-842). The disease causes significant morbidity but limited mortality. Vaccination against N. gonorrhoeae would be highly desirable, but repeated attempts have failed. The main candidate antigens for this vaccine are surface-exposed proteins such as pili, porins, opacity-associated proteins (Opas) and other surface-exposed proteins such as the Lip, Laz, IgA1 protease and transferrin-binding proteins. The lipooligosaccharide (LOS) has also been suggested as vaccine (Meitzner & Cohen, supra).
• N. meningitidis causes both endemic and epidemic disease. In the United States the attack rate is 0.6-1 per 100,000 persons per year, and it can be much greater during outbreaks (see Lieberman et al. (1996) Safety and Immunogenicity of a Serogroups A/C Neisseria meningitidis Oligosaccharide-Protein Conjugate Vaccine in Young Children. JAMA 275(19):1499-1503; Schuchat et al (1997) Bacterial Meningitis in the United States in 1995. N Engl J Med 337(14):970-976). In developing countries, endemic disease rates are much higher and during epidemics incidence rates can reach 500 cases per 100,000 persons per year. Mortality is extremely high, at 10-20% in the United States, and much higher in developing countries. Following the introduction of the conjugate vaccine against Haemophilus influenzae, N. meningitidis is the major cause of bacterial meningitis at all ages in the United States (Schuchat et al (1997) supra).
• Based on the organism's capsular polysaccharide, 12 serogroups of N. meningitidis have been identified. Group A is the pathogen most often implicated in epidemic disease in sub-Saharan Africa. Serogroups B and C are responsible for the vast majority of cases in the United States and in most developed countries. Serogroups W135 and Y are responsible for the rest of the cases in the United States and developed countries. The meningococcal vaccine currently in use is a tetravalent polysaccharide vaccine composed of serogroups A, C, Y and W135. Although efficacious in adolescents and adults, it induces a poor immune response and short duration of protection, and cannot be, used in infants [eg. Morbidity and Mortality weekly report, Vol. 46, No. RR-5 (1997)]. This is because polysaccharides are T-cell independent antigens that induce a weak immune response that cannot be boosted by repeated immunization. Following the success of the vaccination against H. influenzae, conjugate vaccines against serogroups A and C have been developed and are at the final stage of clinical testing (Zollinger W D “New and Improved Vaccines Against Meningococcal Disease” in: New Generation Vaccines, supra, pp. 469-488; Lieberman et al (1996) supra; Costantino et al (1992) Development and phase I clinical testing of a conjugate vaccine against meningococcus A and C. Vaccine 10:691-698).
• Meningococcus B remains a problem, however. This serotype currently is responsible for approximately 50% of total meningitis in the United States, Europe, and South America. The polysaccharide approach cannot be used because the menB capsular polysaccharide is a polymer of α(2-8)-linked N-acetyl neuraminic acid that is also present in mammalian tissue. This results in tolerance to the antigen; indeed, if an immune response were elicited, it would be anti-self, and therefore undesirable. In order to avoid induction of autoimmunity and to induce a protective immune response, the capsular polysaccharide has, for instance, been chemically modified substituting the N-acetyl groups with N-propionyl groups, leaving the specific antigenicity unaltered (Romero & Outschoorn (1994) Current status of Meningococcal group B vaccine candidates: capsular or non-capsular? Clin Microbiol Rev 7(4):559-575).
• Alternative approaches to menB vaccines have used complex mixtures of outer membrane proteins (OMPs), containing either the OMPs alone, or OMPs enriched in porins, or deleted of the class 4 OMPs that are believed to induce antibodies that block bactericidal activity. This approach produces vaccines that are not well characterized. They are able to protect against the homologous strain, but are not effective at large where there are many antigenic variants of the outer membrane proteins. To overcome the antigenic variability, multivalent vaccines containing up to nine different porins have been constructed (eg. Poolman J T (1992) Development of a meningococcal vaccine. Infect. Agents Dis. 4:13-28). Additional proteins to be used in outer membrane vaccines have been the opa and opc proteins, but none of these approaches have been able to overcome the antigenic variability (eg. Ala'Aldeen & Borriello (1996) The meningococcal transferrin-binding proteins 1 and 2 are both surface exposed and generate bactericidal antibodies capable of killing homologous and heterologous strains. Vaccine 14(1):49-53).
• A certain amount of sequence data is available for meningococcal and gonoccocal genes and proteins (eg. EP-A-0467714, WO96/29412), but this is by no means complete. The provision of further sequences could provide an opportunity to identify secreted or surface-exposed proteins that are presumed targets for the immune system and which are not antigenically variable. For instance, some of the identified proteins could be components of efficacious vaccines against meningococcus B, some could be components of vaccines against all meningococcal serotypes, and others could be components of vaccines against all pathogenic Neisseriae.
THE INVENTION
• The invention provides proteins comprising the Neisserial amino acid sequences disclosed in the examples. These sequences relate to N. meningitidis or N. gonorrhoeae.
• It also provides proteins comprising sequences homologous (ie. having sequence identity) to the Neisserial amino acid sequences disclosed in the examples. Depending on the particular sequence, the degree of identity is preferably greater than 50% (eg. 65%, 80%, 90%, or more). These homologous proteins include mutants and allelic variants of the sequences disclosed in the examples. Typically, 50% identity or more between two proteins is considered to be an indication of functional equivalence. Identity between the proteins is preferably determined by the Smith-Waterman homology search algorithm as implemented in the MPSRCH program (Oxford Molecular), using an affine gap search with parameters gap open penalty=12 and gap extension penalty=1.
• The invention further provides proteins comprising fragments of the Neisserial amino acid sequences disclosed in the examples. The fragments should comprise at least n consecutive amino acids from the sequences and, depending on the particular sequence, n is 7 or more (eg. 8, 10, 12, 14, 16, 18, 20 or more). Preferably the fragments comprise an epitope from the sequence.
• The proteins of the invention can, of course, be prepared by various means (eg. recombinant expression, purification from cell culture, chemical synthesis etc.) and in various forms (eg. native, fusions etc.). They are preferably prepared in substantially pure or isolated form (ie. substantially free from other Neisserial or host cell proteins)
• According to a further aspect, the invention provides antibodies which bind to these proteins. These may be polyclonal or monoclonal and may be produced by any suitable means.
• According to a further aspect, the invention provides nucleic acid comprising the Neisserial nucleotide sequences disclosed in the examples. In addition, the invention provides nucleic acid comprising sequences homologous (ie. having sequence identity) to the Neisserial nucleotide sequences disclosed in the examples.
• Furthermore, the invention provides nucleic acid which can hybridise to the Neisserial nucleic acid disclosed in the examples, preferably under “high stringency” conditions (eg. 65° C. in a 0.1×SSC, 0.5% SDS solution).
• Nucleic acid comprising fragments of these sequences are also provided. These should comprise at least n consecutive nucleotides from the Neisserial sequences and, depending on the particular sequence, n is 10 or more ( eg 12, 14, 15, 18, 20, 25, 30, 35, 40 or more).
• According to a further aspect, the invention provides nucleic acid encoding the proteins and protein fragments of the invention.
• It should also be appreciated that the invention provides nucleic acid comprising sequences complementary to those described above (eg. for antisense or probing purposes).
• Nucleic acid according to the invention can, of course, be prepared in many ways (eg. by chemical synthesis, from genomic or cDNA libraries, from the organism itself etc.) and can take various forms (eg. single stranded, double stranded, vectors, probes etc.).
• In addition, the term “nucleic acid” includes DNA and RNA, and also their analogues, such as those containing modified backbones, and also peptide nucleic acids (PNA) etc.
• According to a further aspect, the invention provides vectors comprising nucleotide sequences of the invention (eg. expression vectors) and host cells transformed with such vectors.
• According to a further aspect, the invention provides compositions comprising protein, antibody, and/or nucleic acid according to the invention. These compositions may be suitable as vaccines, for instance, or as diagnostic reagents, or as immunogenic compositions.
• The invention also provides nucleic acid, protein, or antibody according to the invention for use as medicaments (eg. as vaccines) or as diagnostic reagents. It also provides the use of nucleic acid, protein, or antibody according to the invention in the manufacture of: (i) a medicament for treating or preventing infection due to Neisserial bacteria; (ii) a diagnostic reagent for detecting the presence of Neisserial bacteria or of antibodies raised against Neisserial bacteria; and/or (iii) a reagent which can raise antibodies against Neisserial bacteria. Said Neisserial bacteria may be any species or strain (such as N. gonorrhoeae, or any strain of N. meningitidis, such as strain A, strain B or strain C).
• The invention also provides a method of treating a patient, comprising administering to the patient a therapeutically effective amount of nucleic acid, protein, and/or antibody according to the invention.
• According to further aspects, the invention provides various processes.
• A process for producing proteins of the invention is provided, comprising the step of culturing a host cell according to the invention under conditions which induce protein expression.
• A process for producing protein or nucleic acid of the invention is provided, wherein the protein or nucleic acid is synthesised in part or in whole using chemical means.
• A process for detecting polynucleotides of the invention is provided, comprising the steps of: (a) contacting a nucleic probe according to the invention with a biological sample under hybridizing conditions to form duplexes; and (b) detecting said duplexes.
• A process for detecting proteins of the invention is provided, comprising the steps of: (a) contacting an antibody according to the invention with a biological sample under conditions suitable for the formation of an antibody-antigen complexes; and (b) detecting said complexes.
• A summary of standard techniques and procedures which may be employed in order to perform the invention (eg. to utilise the disclosed sequences for vaccination or diagnostic purposes) follows. This summary is not a limitation on the invention but, rather, gives examples that may be used, but are not required.
General
• The practice of the present invention will employ, unless otherwise indicated, conventional techniques of molecular biology, microbiology, recombinant DNA, and immunology, which are within the skill of the art. Such techniques are explained fully in the literature eg. Sambrook Molecular Cloning; A Laboratory Manual, Second Edition (1989); DNA Cloning, Volumes I and ii (D. N Glover ed. 1985); Oligonucleotide Synthesis (M. J. Gait ed, 1984); Nucleic Acid Hybridization (B. D. Hames & S. J. Higgins eds. 1984); Transcription and Translation (B. D. Hames & S. J. Higgins eds. 1984); Animal Cell Culture (R. I. Freshney ed. 1986); Immobilized Cells and Enzymes (IRL Press, 1986); B. Perbal, A Practical Guide to Molecular Cloning (1984); the Methods in Enzymology series (Academic Press, Inc.), especially volumes 154 & 155; Gene Transfer Vectors for Mammalian Cells (J. H. Miller and M. P. Calm eds. 1987, Cold Spring Harbor Laboratory); Mayer and Walker, eds. (1987), Immunochemical Methods in Cell and Molecular Biology (Academic Press, London); Scopes, (1987) Protein Purification: Principles and Practice, Second Edition (Springer-Verlag, N.Y.), and Handbook of Experimental Immunology, Volumes I-IV (D. M. Weir and C. C. Blackwell eds 1986).
• Standard abbreviations for nucleotides and amino acids are used in this specification.
• All publications, patents, and patent applications cited herein are incorporated in full by reference. In particular, the contents of UK patent applications 9723516.2, 9724190.5, 9724386.9, 9725158.1, 9726147.3, 9800759.4, and 9819016.8 are incorporated herein.
DEFINITIONS
• A composition containing X is “substantially free of” Y when at least 85% by weight of the total X+Y in the composition is X. Preferably, X comprises at least about 90% by weight of the total of X+Y in the composition, more preferably at least about 95% or even 99% by weight.
• The term “comprising” means “including” as well as “consisting” eg. a composition “comprising” X may consist exclusively of X or may include something additional to X, such as X+Y.
• The term “heterologous” refers to two biological components that are not found together in nature. The components may be host cells, genes, or regulatory regions, such as promoters. Although the heterologous components are not found together in nature, they can function together, as when a promoter heterologous to a gene is operably linked to the gene. Another example is where a Neisserial sequence is heterologous to a mouse host cell. A further examples would be two epitopes from the same or different proteins which have been assembled in a single protein in an arrangement not found in nature.
• An “origin of replication” is a polynucleotide sequence that initiates and regulates replication of polynucleotides, such as an expression vector. The origin of replication behaves as an autonomous unit of polynucleotide replication within a cell, capable of replication under its own control. An origin of replication may be needed for a vector to replicate in a particular host cell. With certain origins of replication, an expression vector can be reproduced at a high copy number in the presence of the appropriate proteins within the cell. Examples of origins are the autonomously replicating sequences, which are effective in yeast; and the viral T-antigen, effective in COS-7 cells.
• A “mutant” sequence is defined as DNA, RNA or amino acid sequence differing from but having sequence identity with the native or disclosed sequence. Depending on the particular sequence, the degree of sequence identity between the native or disclosed sequence and the mutant sequence is preferably greater than 50% (eg. 60%, 70%, 80%, 90%, 95%, 99% or more, calculated using the Smith-Waterman algorithm as described above). As used herein, an “allelic variant” of a nucleic acid molecule, or region, for which nucleic acid sequence is provided herein is a nucleic acid molecule, or region, that occurs essentially at the same locus in the genome of another or second isolate, and that, due to natural variation caused by, for example, mutation or recombination, has a similar but not identical nucleic acid sequence. A coding region allelic variant typically encodes a protein having similar activity to that of the protein encoded by the gene to which it is being compared. An allelic variant can also comprise an alteration in the 5′ or 3′ untranslated regions of the gene, such as in regulatory control regions (eg. see U.S. Pat. No. 5,753,235).
Expression Systems
• The Neisserial nucleotide sequences can be expressed in a variety of different expression systems; for example those used with mammalian cells, baculoviruses, plants, bacteria, and yeast.
• i. Mammalian Systems
• Mammalian expression systems are known in the art. A mammalian promoter is any DNA sequence capable of binding mammalian RNA polymerase and initiating the downstream (3′) transcription of a coding sequence (eg. structural gene) into mRNA. A promoter will have a transcription initiating region, which is usually placed proximal to the 5′ end of the coding sequence, and a TATA box, usually located 25-30 base pairs (bp) upstream of the transcription initiation site. The TATA box is thought to direct RNA polymerase II to begin RNA synthesis at the correct site. A mammalian promoter will also contain an upstream promoter element, usually located within 100 to 200 bp upstream of the TATA box. An upstream promoter element determines the rate at which transcription is initiated and can act in either orientation [Sambrook et al. (1989) “Expression of Cloned Genes in Mammalian Cells.” In Molecular Cloning: A Laboratory Manual, 2nd ed.].
• Mammalian viral genes are often highly expressed and have a broad host range; therefore sequences encoding mammalian viral genes provide particularly useful promoter sequences. Examples include the SV40 early promoter, mouse mammary tumor virus LTR promoter, adenovirus major late promoter (Ad MLP), and herpes simplex virus promoter. In addition, sequences derived from non-viral genes, such as the murine metallotheionein gene, also provide useful promoter sequences. Expression may be either constitutive or regulated (inducible), depending on the promoter can be induced with glucocorticoid in hormone-responsive cells.
• The presence of an enhancer element (enhancer), combined with the promoter elements described above, will usually increase expression levels. An enhancer is a regulatory DNA sequence that can stimulate transcription up to 1000-fold when linked to homologous or heterologous promoters, with synthesis beginning at the normal RNA start site. Enhancers are also active when they are placed upstream or downstream from the transcription initiation site, in either normal or flipped orientation, or at a distance of more than 1000 nucleotides from the promoter [Maniatis et al. (1987) Science 236:1237; Alberts et al. (1989) Molecular Biology of the Cell, 2nd ed.]. Enhancer elements derived from viruses may be particularly useful, because they usually have a broader host range. Examples include the SV40 early gene enhancer [Dijkema et al (1985) EMBO J. 4:761] and the enhancer/promoters derived from the long terminal repeat (LTR) of the Rous Sarcoma Virus [Gorman et al. (1982b) Proc. Natl. Acad. Sci. 79:6777] and from human cytomegalovirus [Boshart et al. (1985) Cell 41:521]. Additionally, some enhancers are regulatable and become active only in the presence of an inducer, such as a hormone or metal ion [Sassone-Corsi and Borelli (1986) Trends Genet. 2:215; Maniatis et al. (1987) Science 236:1237].
• A DNA molecule may be expressed intracellularly in mammalian cells. A promoter sequence may be directly linked with the DNA molecule, in which case the first amino acid at the N-terminus of the recombinant protein will always be a methionine, which is encoded by the ATG start codon. If desired, the N-terminus may be cleaved from the protein by in vitro incubation with cyanogen bromide.
• Alternatively, foreign proteins can also be secreted from the cell into the growth media by creating chimeric DNA molecules that encode a fusion protein comprised of a leader sequence fragment that provides for secretion of the foreign protein in mammalian cells. Preferably, there are processing sites encoded between the leader fragment and the foreign gene that can be cleaved either in vivo or in vitro. The leader sequence fragment usually encodes a signal peptide comprised of hydrophobic amino acids which direct the secretion of the protein from the cell. The adenovirus triparite leader is an example of a leader sequence that provides for secretion of a foreign protein in mammalian cells.
• Usually, transcription termination and polyadenylation sequences recognized by mammalian cells are regulatory regions located 3′ to the translation stop codon and thus, together with the promoter elements, flank the coding sequence. The 3′ terminus of the mature mRNA is formed by site-specific post-transcriptional cleavage and polyadenylation [Birnstiel et al. (1985) Cell 41:349; Proudfoot and Whitelaw (1988) “Termination and 3′ end processing of eukaryotic RNA. In Transcription and splicing (ed. B. D. Hames and D. M. Glover); Proudfoot (1989) Trends Biochem. Sci. 14:105]. These sequences direct the transcription of an mRNA which can be translated into the polypeptide encoded by the DNA. Examples of transcription terminater/polyadenylation signals include those derived from SV40 [Sambrook et al (1989) “Expression of cloned genes in cultured mammalian cells.” In Molecular Cloning: A Laboratory Manual].
• Usually, the above described components, comprising a promoter, polyadenylation signal, and transcription termination sequence are put together into expression constructs. Enhancers, introns with functional splice donor and acceptor sites, and leader sequences may also be included in an expression construct, if desired. Expression constructs are often maintained in a replicon, such as an extrachromosomal element (eg. plasmids) capable of stable maintenance in a host, such as mammalian cells or bacteria. Mammalian replication systems include those derived from animal viruses, which require trans-acting factors to replicate. For example, plasmids containing the replication systems of papovaviruses, such as SV40 [Gluzman (1981) Cell 23:175] or polyomavirus, replicate to extremely high copy number in the presence of the appropriate viral T antigen. Additional examples of mammalian replicons include those derived from bovine papillomavirus and Epstein-Barr virus. Additionally, the replicon may have two replicaton systems, thus allowing it to be maintained, for example, in mammalian cells for expression and in a prokaryotic host for cloning and amplification. Examples of such mammalian-bacteria shuttle vectors include pMT2 [Kaufman et al. (1989) Mol. Cell. Biol. 9:946] and pHEBO [Shimizu et al. (1986) Mol. Cell. Biol. 6:1074].
• The transformation procedure used depends upon the host to be transformed. Methods for introduction of heterologous polynucleotides into mammalian cells are known in the art and include dextran-mediated transfection, calcium phosphate precipitation, polybrene mediated transfection, protoplast fusion, electroporation, encapsulation of the polynucleotide(s) in liposomes, and direct microinjection of the DNA into nuclei.
• Mammalian cell lines available as hosts for expression are known in the art and include many immortalized cell lines available from the American Type Culture Collection (ATCC), including but not limited to, Chinese hamster ovary (CHO) cells, HeLa cells, baby hamster kidney (BHK) cells, monkey kidney cells (COS), human hepatocellular carcinoma cells (eg. Hep G2), and a number of other cell lines.
• ii. Baculovirus Systems
• The polynucleotide encoding the protein can also be inserted into a suitable insect expression vector, and is operably linked to the control elements within that vector. Vector construction employs techniques which are known in the art. Generally, the components of the expression system include a transfer vector, usually a bacterial plasmid, which contains both a fragment of the baculovirus genome, and a convenient restriction site for insertion of the heterologous gene or genes to be expressed; a wild type baculovirus with a sequence homologous to the baculovirus-specific fragment in the transfer vector (this allows for the homologous recombination of the heterologous gene in to the baculovirus genome); and appropriate insect host cells and growth media.
• After inserting the DNA sequence encoding the protein into the transfer vector, the vector and the wild type viral genome are transfected into an insect host cell where the vector and viral genome are allowed to recombine. The packaged recombinant virus is expressed and recombinant plaques are identified and purified. Materials and methods for baculovirus/insect cell expression systems are commercially available in kit form from, inter alia, Invitrogen, San Diego Calif. (“MaxBac” kit). These techniques are generally known to those skilled in the art and fully described in Summers and Smith, Texas Agricultural Experiment Station Bulletin No. 1555 (1987) (hereinafter “Summers and Smith”).
• Prior to inserting the DNA sequence encoding the protein into the baculovirus genome, the above described components, comprising a promoter, leader (if desired), coding sequence of interest, and transcription termination sequence, are usually assembled into an intermediate transplacement construct (transfer vector). This construct may contain a single gene and operably linked regulatory elements; multiple genes, each with its owned set of operably linked regulatory elements; or multiple genes, regulated by the same set of regulatory elements. Intermediate transplacement constructs are often maintained in a replicon, such as an extrachromosomal element (eg. plasmids) capable of stable maintenance in a host, such as a bacterium. The replicon will have a replication system, thus allowing it to be maintained in a suitable host for cloning and amplification.
• Currently, the most commonly used transfer vector for introducing foreign genes into AcNPV is pAc373. Many other vectors, known to those of skill in the art, have also been designed. These include, for example, pVL985 (which alters the polyhedrin start codon from ATG to ATT, and which introduces a BamHI cloning site 32 basepairs downstream from the ATT; see Luckow and Summers, Virology (1989) 17:31.
• The plasmid usually also contains the polyhedrin polyadenylation signal (Miller et al. (1988) Ann. Rev. Microbiol., 42:177) and a prokaryotic ampicillin-resistance (amp) gene and origin of replication for selection and propagation in E. coli.
• Baculovirus transfer vectors usually contain a baculovirus promoter. A baculovirus promoter is any DNA sequence capable of binding a baculovirus RNA polymerase and initiating the downstream (5′ to 3′) transcription of a coding sequence (eg. structural gene) into mRNA. A promoter will have a transcription initiation region which is usually placed proximal to the 5′ end of the coding sequence. This transcription initiation region usually includes an RNA polymerase binding site and a transcription initiation site. A baculovirus transfer vector may also have a second domain called an enhancer, which, if present, is usually distal to the structural gene. Expression may be either regulated or constitutive.
• Structural genes, abundantly transcribed at late times in a viral infection cycle, provide particularly useful promoter sequences. Examples include sequences derived from the gene encoding the viral polyhedron protein, Friesen et al., (1986) “The Regulation of Baculovirus Gene Expression,” in: The Molecular Biology of Baculoviruses (ed. Walter Doerfler); EPO Publ. Nos. 127 839 and 155 476; and the gene encoding the p10 protein, Vlak et al., (1988), J. Gen. Virol. 69:765.
• DNA encoding suitable signal sequences can be derived from genes for secreted insect or baculovirus proteins, such as the baculovirus polyhedrin gene (Carbonell et al. (1988) Gene, 73:409). Alternatively, since the signals for mammalian cell posttranslational modifications (such as signal peptide cleavage, proteolytic cleavage, and phosphorylation) appear to be recognized by insect cells, and the signals required for secretion and nuclear accumulation also appear to be conserved between the invertebrate cells and vertebrate cells, leaders of non-insect origin, such as those derived from genes encoding human α-interferon, Maeda et al., (1985), Nature 315:592; human gastrin-releasing peptide, Lebacq-Verheyden et al., (1988), Molec. Cell. Biol. 8:3129; human IL-2, Smith et al., (1985) Proc. Nat'l Acad. Sci. USA, 82:8404; mouse IL-3, (Miyajima et al., (1987) Gene 58:273; and human glucocerebrosidase, Martin et al. (1988) DNA, 7:99, can also be used to provide for secretion in insects.
• A recombinant polypeptide or polyprotein may be expressed intracellularly or, if it is expressed with the proper regulatory sequences, it can be secreted. Good intracellular expression of nonfused foreign proteins usually requires heterologous genes that ideally have a short leader sequence containing suitable translation initiation signals preceding an ATG start signal. If desired, methionine at the N-terminus may be cleaved from the mature protein by in vitro incubation with cyanogen bromide.
• Alternatively, recombinant polyproteins or proteins which are not naturally secreted can be secreted from the insect cell by creating chimeric DNA molecules that encode a fusion protein comprised of a leader sequence fragment that provides for secretion of the foreign protein in insects. The leader sequence fragment usually encodes a signal peptide comprised of hydrophobic amino acids which direct the translocation of the protein into the endoplasmic reticulum.
• After insertion of the DNA sequence and/or the gene encoding the expression product precursor of the protein, an insect cell host is co-transformed with the heterologous DNA of the transfer vector and the genomic DNA of wild type baculovirus—usually by co-transfection. The promoter and transcription termination sequence of the construct will usually comprise a 2-5 kb section of the baculovirus genome. Methods for introducing heterologous DNA into the desired site in the baculovirus virus are known in the art. (See Summers and Smith supra; Ju et al. (1987); Smith et al., Mol. Cell. Biol. (1983) 3:2156; and Luckow and Summers (1989)). For example, the insertion can be into a gene such as the polyhedrin gene, by homologous double crossover recombination; insertion can also be into a restriction enzyme site engineered into the desired baculovirus gene. Miller et al., (1989), Bioessays 4:91. The DNA sequence, when cloned in place of the polyhedrin gene in the expression vector, is flanked both 5′ and 3′ by polyhedrin-specific sequences and is positioned downstream of the polyhedrin promoter.
• The newly formed baculovirus expression vector is subsequently packaged into an infectious recombinant baculovirus. Homologous recombination occurs at low frequency (between about 1% and about 5%); thus, the majority of the virus produced after cotransfection is still wild-type virus. Therefore, a method is necessary to identify recombinant viruses. An advantage of the expression system is a visual screen allowing recombinant viruses to be distinguished. The polyhedrin protein, which is produced by the native virus, is produced at very high levels in the nuclei of infected cells at late times after viral infection. Accumulated polyhedrin protein forms occlusion bodies that also contain embedded particles. These occlusion bodies, up to 15 μm in size, are highly refractile, giving them a bright shiny appearance that is readily visualized under the light microscope. Cells infected with recombinant viruses lack occlusion bodies. To distinguish recombinant virus from wild-type virus, the transfection supernatant is plagued onto a monolayer of insect cells by techniques known to those skilled in the art. Namely, the plaques are screened under the light microscope for the presence (indicative of wild-type virus) or absence (indicative of recombinant virus) of occlusion bodies. “Current Protocols in Microbiology” Vol. 2 (Ausubel et al. eds) at 16.8 (Supp. 10, 1990); Summers and Smith, supra; Miller et al. (1989).
• Recombinant baculovirus expression vectors have been developed for infection into several insect cells. For example, recombinant baculoviruses have been developed for, inter alis: Aedes aegypti, Autographa californica, Bombyx mori, Drosophila melanogaster, Spodoptera frugiperda, and Trichoplusia ni (WO 89/046699; Carbonell et al., (1985) J. Virol. 56:153; Wright (1986) Nature 321:718; Smith et al., (1983) Mol. Cell. Biol. 3:2156; and see generally, Fraser, et al. (1989) In Vitro Cell. Dev. Biol. 25:225).
• Cells and cell culture media are commercially available for both direct and fusion expression of heterologous polypeptides in a baculovirus/expression system; cell culture technology is generally known to those skilled in the art. See, eg. Summers and Smith supra.
• The modified insect cells may then be grown in an appropriate nutrient medium, which allows for stable maintenance of the plasmid(s) present in the modified insect host. Where the expression product gene is under inducible control, the host may be grown to high density, and expression induced. Alternatively, where expression is constitutive, the product will be continuously expressed into the medium and the nutrient medium must be continuously circulated, while removing the product of interest and augmenting depleted nutrients. The product may be purified by such techniques as chromatography, eg. HPLC, affinity chromatography, ion exchange chromatography, etc.; electrophoresis; density gradient centrifugation; solvent extraction, or the like. As appropriate, the product may be further purified, as required, so as to remove substantially any insect proteins which are also secreted in the medium or result from lysis of insect cells, so as to provide a product which is at least substantially free of host debris, eg. proteins, lipids and polysaccharides.
• In order to obtain protein expression, recombinant host cells derived from the transformants are incubated under conditions which allow expression of the recombinant protein encoding sequence. These conditions will vary, dependent upon the host cell selected. However, the conditions are readily ascertainable to those of ordinary skill in the art, based upon what is known in the art.
• iii. Plant Systems
• There are many plant cell culture and whole plant genetic expression systems known in the art. Exemplary plant cellular genetic expression systems include those described in patents, such as: U.S. Pat. No. 5,693,506; U.S. Pat. No. 5,659,122; and U.S. Pat. No. 5,608,143. Additional examples of genetic expression in plant cell culture has been described by Zenk, Phytochemistry 30:3861-3863 (1991). Descriptions of plant protein signal peptides may be found in addition to the references described above in Vaulcombe et al., Mol. Gen. Genet. 209:33-40 (1987); Chandler et al., Plant Molecular Biology 3:407-418 (1984); Rogers, J. Biol. Chem. 260:3731-3738 (1985); Rothstein et al., Gene 55:353-356 (1987); Whittier et al., Nucleic Acids Research 15:2515-2535 (1987); Wirsel et al., Molecular Microbiology 3:3-14 (1989); Yu et al., Gene 122:247-253 (1992). A description of the regulation of plant gene expression by the phytohormone, gibberellic acid and secreted enzymes induced by gibberellic acid can be found in R. L. Jones and J. MacMillin, Gibberellins: in: Advanced Plant Physiology, Malcolm B. Wilkins, ed., 1984 Pitman Publishing Limited, London, pp. 21-52. References that describe other metabolically-regulated genes: Sheen, Plant Cell, 2:1027-1038 (1990); Maas et al., EMBO J. 9:3447-3452 (1990); Benkel and Hickey, Proc. Natl. Acad. Sci. 84:1337-1339 (1987)
• Typically, using techniques known in the art, a desired polynucleotide sequence is inserted into an expression cassette comprising genetic regulatory elements designed for operation in plants. The expression cassette is inserted into a desired expression vector with companion sequences upstream and downstream from the expression cassette suitable for expression in a plant host. The companion sequences will be of plasmid or viral origin and provide necessary characteristics to the vector to permit the vectors to move DNA from an original cloning host, such as bacteria, to the desired plant host. The basic bacterial/plant vector construct will preferably provide a broad host range prokaryote replication origin; a prokaryote selectable marker; and, for Agrobacterium transformations, T DNA sequences for Agrobacterium-mediated transfer to plant chromosomes. Where the heterologous gene is not readily amenable to detection, the construct will preferably also have a selectable marker gene suitable for determining if a plant cell has been transformed. A general review of suitable markers, for example for the members of the grass family, is found in Wilmink and Dons, 1993, Plant Mol. Biol. Reptr, 11(2):165-185.
• Sequences suitable for permitting integration of the heterologous sequence into the plant genome are also recommended. These might include transposon sequences and the like for homologous recombination as well as Ti sequences which permit random insertion of a heterologous expression cassette into a plant genome. Suitable prokaryote selectable markers include resistance toward antibiotics such as ampicillin or tetracycline. Other DNA sequences encoding additional functions may also be present in the vector, as is known in the art.
• The nucleic acid molecules of the subject invention may be included into an expression cassette for expression of the protein(s) of interest. Usually, there will be only one expression cassette, although two or more are feasible. The recombinant expression cassette will contain in addition to the heterologous protein encoding sequence the following elements, a promoter region, plant 5′ untranslated sequences, initiation codon depending upon whether or not the structural gene comes equipped with one, and a transcription and translation termination sequence. Unique restriction enzyme sites at the 5′ and 3′ ends of the cassette allow for easy insertion into a pre-existing vector.
• A heterologous coding sequence may be for any protein relating to the present invention. The sequence encoding the protein of interest will encode a signal peptide which allows processing and translocation of the protein, as appropriate, and will usually lack any sequence which might result in the binding of the desired protein of the invention to a membrane. Since, for the most part, the transcriptional initiation region will be for a gene which is expressed and translocated during germination, by employing the signal peptide which provides for translocation, one may also provide for translocation of the protein of interest. In this way, the protein(s) of interest will be translocated from the cells in which they are expressed and may be efficiently harvested. Typically secretion in seeds are across the aleurone or scutellar epithelium layer into the endosperm of the seed. While it is not required that the protein be secreted from the cells in which the protein is produced, this facilitates the isolation and purification of the recombinant protein.
• Since the ultimate expression of the desired gene product will be in a eucaryotic cell it is desirable to determine whether any portion of the cloned gene contains sequences which will be processed out as introns by the host's splicosome machinery. If so, site-directed mutagenesis of the “intron” region may be conducted to prevent losing a portion of the genetic message as a false intron code, Reed and Maniatis, Cell 41:95-105, 1985.
• The vector can be microinjected directly into plant cells by use of micropipettes to mechanically transfer the recombinant DNA. Crossway, Mol. Gen. Genet, 202:179-185, 1985. The genetic material may also be transferred into the plant cell by using polyethylene glycol, Krens, et al., Nature, 296, 72-74, 1982. Another method of introduction of nucleic acid segments is high velocity ballistic penetration by small particles with the nucleic acid either within the matrix of small beads or particles, or on the surface, Klein, et al., Nature, 327, 70-73, 1987 and Knudsen and Muller, 1991, Planta, 185:330-336 teaching particle bombardment of barley endosperm to create transgenic barley. Yet another method of introduction would be fusion of protoplasts with other entities, either minicells, cells, lysosomes or other fusible lipid-surfaced bodies, Fraley, et al., Proc. Natl. Acad. Sci. USA, 79, 1859-1863, 1982.
• The vector may also be introduced into the plant cells by electroporation. (Fromm et al., Proc. Natl. Acad. Sci. USA 82:5824, 1985). In this technique, plant protoplasts are electroporated in the presence of plasmids containing the gene construct. Electrical impulses of high field strength reversibly permeabilize biomembranes allowing the introduction of the plasmids. Electroporated plant protoplasts reform the cell wall, divide, and form plant callus.
• All plants from which protoplasts can be isolated and cultured to give whole regenerated plants can be transformed by the present invention so that whole plants are recovered which contain the transferred gene. It is known that practically all plants can be regenerated from cultured cells or tissues, including but not limited to all major species of sugarcane, sugar beet, cotton, fruit and other trees, legumes and vegetables. Some suitable plants include, for example, species from the genera Fragaria, Lotus, Medicago, Onobrychis, Trifolium, Trigonella, Vigna, Citrus, Linum, Geranium, Manihot, Daucus, Arabidopsis, Brassica, Raphanus, Sinapis, Atropa, Capsicum, Datura, Hyoscyamus, Lycopersion, Nicotiana, Solanum, Petunia, Digitalis, Majorana, Cichorium, Helianthus, Lactuca, Bromus, Asparagus, Antirrhinum, Hererocallis, Nemesia, Pelargonium, Panicum, Pennisetum, Ranunculus, Senecio, Salpiglossis, Cucumis, Browaalia, Glycine, Lolium, Zea, Triticum, Sorghum, and Datura.
• Means for regeneration vary from species to species of plants, but generally a suspension of transformed protoplasts containing copies of the heterologous gene is first provided. Callus tissue is formed and shoots may be induced from callus and subsequently rooted. Alternatively, embryo formation can be induced from the protoplast suspension. These embryos germinate as natural embryos to form plants. The culture media will generally contain various amino acids and hormones, such as auxin and cytokinins. It is also advantageous to add glutamic acid and proline to the medium, especially for such species as corn and alfalfa. Shoots and roots normally develop simultaneously. Efficient regeneration will depend on the medium, on the genotype, and on the history of the culture. If these three variables are controlled, then regeneration is fully reproducible and repeatable.
• In some plant cell culture systems, the desired protein of the invention may be excreted or alternatively, the protein may be extracted from the whole plant. Where the desired protein of the invention is secreted into the medium, it may be collected. Alternatively, the embryos and embryoless-half seeds or other plant tissue may be mechanically disrupted to release any secreted protein between cells and tissues. The mixture may be suspended in a buffer solution to retrieve soluble proteins. Conventional protein isolation and purification methods will be then used to purify the recombinant protein. Parameters of time, temperature pH, oxygen, and volumes will be adjusted through routine methods to optimize expression and recovery of heterologous protein.
• iv. Bacterial Systems
• Bacterial expression techniques are known in the art. A bacterial promoter is any DNA sequence capable of binding bacterial RNA polymerase and initiating the downstream (3′) transcription of a coding sequence (eg. structural gene) into mRNA. A promoter will have a transcription initiation region which is usually placed proximal to the 5′ end of the coding sequence. This transcription initiation region usually includes an RNA polymerase binding site and a transcription initiation site. A bacterial promoter may also have a second domain called an operator, that may overlap an adjacent RNA polymerase binding site at which RNA synthesis begins. The operator permits negative regulated (inducible) transcription, as a gene repressor protein may bind the operator and thereby inhibit transcription of a specific gene. Constitutive expression may occur in the absence of negative regulatory elements, such as the operator. In addition, positive regulation may be achieved by a gene activator protein binding sequence, which, if present is usually proximal (5′) to the RNA polymerase binding sequence. An example of a gene activator protein is the catabolite activator protein (CAP), which helps initiate transcription of the lac operon in Escherichia coli (E. coli) [Raibaud et al. (1984) Annu. Rev. Genet. 18:173]. Regulated expression may therefore be either positive or negative, thereby either enhancing or reducing transcription.
• Sequences encoding metabolic pathway enzymes provide particularly useful promoter sequences. Examples include promoter sequences derived from sugar metabolizing enzymes, such as galactose, lactose (lac) [Chang et al. (1977) Nature 198:1056], and maltose. Additional examples include promoter sequences derived from biosynthetic enzymes such as tryptophan (trp) [Goeddel et al. (1980) Nuc. Acids Res. 8:4057; Yelverton et al. (1981) Nucl. Acids Res. 9:731; U.S. Pat. No. 4,738,921; EP-A-0036776 and EP-A-0121775]. The g-laotamase (bla) promoter system [Weissmann (1981) “The cloning of interferon and other mistakes.” In Interferon 3 (ed. I. Gresser)], bacteriophage lambda PL [Shimatake et al. (1981) Nature 292:128] and T5 [U.S. Pat. No. 4,689,406] promoter systems also provide useful promoter sequences.
• In addition, synthetic promoters which do not occur in nature also function as bacterial promoters. For example, transcription activation sequences of one bacterial or bacteriophage promoter may be joined with the operon sequences of another bacterial or bacteriophage promoter, creating a synthetic hybrid promoter [U.S. Pat. No. 4,551,433]. For example, the tac promoter is a hybrid trp-lac promoter comprised of both trp promoter and lac operon sequences that is regulated by the lac repressor [Amann et al. (1983) Gene 25:167; de Boer et al. (1983) Proc. Natl. Acad. Sci. 80:21].
• Furthermore, a bacterial promoter can include naturally occurring promoters of non-bacterial origin that have the ability to bind bacterial RNA polymerase and initiate transcription. A naturally occurring promoter of non-bacterial origin can also be coupled with a compatible RNA polymerase to produce high levels of expression of some genes in prokaryotes. The bacteriophage T7 RNA polymerase/promoter system is an example of a coupled promoter system [Studier et al. (1986) J. Mol. Biol. 189:113; Tabor et al., (1985) Proc Natl. Acad. Sci. 82:1074]. In addition, a hybrid promoter can also be comprised of a bacteriophage promoter and an E. coli operator region (EPO-A-0 267 851).
• In addition to a functioning promoter sequence, an efficient ribosome binding site is also useful for the expression of foreign genes in prokaryotes. In E. coli, the ribosome binding site is called the Shine-Dalgarno (SD) sequence and includes an initiation codon (ATG) and a sequence 3-9 nucleotides in length located 3-11 nucleotides upstream of the initiation codon [Shine et al. (1975) Nature 254:34]. The SD sequence is thought to promote binding of mRNA to the ribosome by the pairing of bases between the SD sequence and the 3′ and of E. coli 16S rRNA [Steitz et al: (1979) “Genetic signals and nucleotide sequences in messenger RNA.” In Biological Regulation and Development: Gene Expression (ed. R. F. Goldberger)]. To express eukaryotic genes and prokaryotic genes with weak ribosome-binding site [Sambrook et al. (1989) “Expression of cloned genes in Escherichia coli.” In Molecular Cloning: A Laboratory Manual].
• A DNA molecule may be expressed intracellularly. A promoter sequence may be directly linked with the DNA molecule, in which case the first amino acid at the N-terminus will always be a methionine, which is encoded by the ATG start codon. If desired, methionine at the N-terminus may be cleaved from the protein by in vitro incubation with cyanogen bromide or by either in vivo on in vitro incubation with a bacterial methionine N-terminal peptidase (EPO-A-0 219 237).
• Fusion proteins provide an alternative to direct expression. Usually, a DNA sequence encoding the N-terminal portion of an endogenous bacterial protein, or other stable protein, is fused to the 5′ end of heterologous coding sequences. Upon expression, this construct will provide a fusion of the two amino acid sequences. For example, the bacteriophage lambda cell gene can be linked at the 5′ terminus of a foreign gene and expressed in bacteria. The resulting fusion protein preferably retains a site for a processing enzyme (factor Xa) to cleave the bacteriophage protein from the foreign gene [Nagai et al. (1984) Nature 309:810]. Fusion proteins can also be made with sequences from the lacZ [Jia et al. (1987) Gene 60:197], trpE [Allen et al. (1987) J. Biotechnol. 5:93; Makoff et al. (1989) J. Gen. Microbiol. 135:11], and Chey [EP-A-0 324 647] genes. The DNA sequence at the junction of the two amino acid sequences may or may not encode a cleavable site. Another example is a ubiquitin fusion protein. Such a fusion protein is made with the ubiquitin region that preferably retains a site for a processing enzyme (eg. ubiquitin specific processing-protease) to cleave the ubiquitin from the foreign protein. Through this method, native foreign protein can be isolated [Miller et al. (1989) Bio/Technology 7:698].
• Alternatively, foreign proteins can also be secreted from the cell by creating chimeric DNA molecules that encode a fusion protein comprised of a signal peptide sequence fragment that provides for secretion of the foreign protein in bacteria [U.S. Pat. No. 4,336,336]. The signal sequence fragment usually encodes a signal peptide comprised of hydrophobic amino acids which direct the secretion of the protein from the cell. The protein is either secreted into the growth media (gram-positive bacteria) or into the periplasmic space, located between the inner and outer membrane of the cell (gram-negative bacteria). Preferably there are processing sites, which can be cleaved either in vivo or in vitro encoded between the signal peptide fragment and the foreign gene.
• DNA encoding suitable signal sequences can be derived from genes for secreted bacterial proteins, such as the E. coli outer membrane protein gene (ompA) [Masui et al. (1983), in: Experimental Manipulation of Gene Expression; Ghrayeb et al. (1984) EMBO J. 3:2437] and the E. coli alkaline phosphatase signal sequence (phoA) [Oka et al. (1985) Proc. Natl. Acad. Sci. 82:7212]. As an additional example, the signal sequence of the alpha-amylase gene from various Bacillus strains can be used to secrete heterologous proteins from B. subtilis [Palva et al. (1982) Proc. Natl. Acad. Sci. USA 79:5582; EP-A-0 244 042].
• Usually, transcription termination sequences recognized by bacteria are regulatory regions located 3′ to the translation stop codon, and thus together with the promoter flank the coding sequence. These sequences direct the transcription of an mRNA which can be translated into the polypeptide encoded by the DNA. Transcription termination sequences frequently include DNA sequences of about 50 nucleotides capable of forming stem loop structures that aid in terminating transcription. Examples include transcription termination sequences derived from genes with strong promoters, such as the irp gene in E. coli as well as other biosynthetic genes.
• Usually, the above described components, comprising a promoter, signal sequence (if desired), coding sequence of interest, and transcription termination sequence, are put together into expression constructs. Expression constructs are often maintained in a replicon, such as an extrachromosomal element (eg. plasmids) capable of stable maintenance in a host, such as bacteria. The replicon will have a replication system, thus allowing it to be maintained in a prokaryotic host either for expression or for cloning and amplification. In addition, a replicon may be either a high or low copy number plasmid. A high copy number plasmid will generally have a copy number ranging from about 5 to about 200, and usually about 10 to about 150. A host containing a high copy number plasmid will preferably contain at least about 10, and more preferably at least about 20 plasmids. Either a high or low copy number vector may be selected, depending upon the effect of the vector and the foreign protein on the host.
• Alternatively, the expression constructs can be integrated into the bacterial genome with an integrating vector. Integrating vectors usually contain at least one sequence homologous to the bacterial chromosome that allows the vector to integrate. Integrations appear to result from recombinations between homologous DNA in the vector and the bacterial chromosome. For example, integrating vectors constructed with DNA from various Bacillus strains integrate into the Bacillus chromosome (EP-A-0 127 328). Integrating vectors may also be comprised of bacteriophage of transposon sequences.
• Usually, extrachromosomal and integrating expression constructs may contain selectable markers to allow for the selection of bacterial strains that have been transformed. Selectable markers can be expressed in the bacterial host and may include genes which render bacteria resistant to drugs such as ampicillin, chloramphenicol, erythromycin, kanamycin (neomycin), and tetracycline [Davies et al. (1978) Annu. Rev. Microbiol. 32:469]. Selectable markers may also include biosynthetic genes, such as those in the histidine, tryptophan, and leucine biosynthetic pathways.
• Alternatively, some of the above described components can be put together in transformation vectors. Transformation vectors are usually comprised of a selectable market that is either maintained in a replicon or developed into an integrating vector, as described above.
• Expression and transformation vectors, either extra-chromosomal replicons or integrating vectors, have been developed for transformation into many bacteria. For example, expression vectors have been developed for, inter alfa, the following bacteria: Bacillus subtilis [Palva et al. (1982) Proc. Natl. Acad. Sci. USA 79:5582; EP-A-0 036 259 and EP-A-0 063 953; WO 84/04541], Escherichia coli [Shimatake et al. (1981) Nature 292:128; Amann et al. (1985) Gene 40:183; Studier et al. (1986) J. Mol. Biol. 189:113; EP-A-0 036 776, EP-A-0 136 829 and EP-A-0 136 907], Streptococcus cremoris [Powell et al. (1988) Appl. Environ. Microbiol. 54:655]; Streptococcus lividans [Powell et al. (1988) Appl. Environ. Microbiol. 54:655], Streptomyces lividans [U.S. Pat. No. 4,745,056].
• Methods of introducing exogenous DNA into bacterial hosts are well-known in the art, and usually include either the transformation of bacteria treated with CaCl₂ or other agents, such as divalent cations and DMSO. DNA can also be introduced into bacterial cells by electroporation. Transformation procedures usually vary with the bacterial specie to be transformed. See eg. [Masson et al. (1989) FEMS Microbiol. Lett. 60:273; Palva et al. (1982) Proc. Natl. Acad. Sci. USA 79:5582; EP-A-0 036 259 and EP-A-0 063 953; WO 84/04541, Bacillus], [Miller et al. (1988) Proc. Natl. Acad. Sci. 85:856; Wang et al. (1990) J. Bacteriol. 172:949, Campylobacter], [Cohen et al. (1973) Proc. Natl. Acad. Sci. 69:2110; Dower et al. (1988) Nucleic Acids Res. 16:6127; Kushner (1978) “An improved method for transformation of Escherichia coli with ColE1-derived plasmids. In Genetic Engineering: Proceedings of the International Symposium on Genetic Engineering (eds. H. W. Boyer and S. Nicosia); Mandel et al. (1970) J. Mol. Biol. 53:159; Taketo (1988) Biochim. Biophys. Acta 949:318; Escherichia], [Chassy et al. (1987) FEMS Microbiol. Lett. 44:173 Lactobacillus]; [Fiedler et al. (1988) Anal. Biochem 170:38, Pseudomonas]; [Augustin et al. (1990) FEMS Microbiol. Lett. 66:203, Staphylococcus], [Barany et al. (1980) J. Bacteriol. 144:698; Harlander (1987) “Transformation of Streptococcus lactis by electroporation, in: Streptococcal Genetics (ed. J. Ferretti and R. Curtiss III); Perry et al. (1981) Infect. Immun. 32:1295; Powell et al. (1988) Appl. Environ. Microbiol. 54:655; Somkuti et al. (1987) Proc. 4th Evr. Cong. Biotechnology 1:412, Streptococcus].
• v. Yeast Expression
• Yeast expression systems are also known to one of ordinary skill in the art. A yeast promoter is any DNA sequence capable of binding yeast RNA polymerase and initiating the downstream (3′) transcription of a coding sequence (eg. structural gene) into mRNA. A promoter will have a transcription initiation region which is usually placed proximal to the 5′ end of the coding sequence. This transcription initiation region usually includes an RNA polymerase binding site (the “TATA Box”) and a transcription initiation site. A yeast promoter may also have a second domain called an upstream activator sequence (UAS), which, if present, is usually distal to the structural gene. The UAS permits regulated (inducible) expression. Constitutive expression occurs in the absence of a UAS. Regulated expression may be either positive or negative, thereby either enhancing or reducing transcription.
• Yeast is a fermenting organism with an active metabolic pathway, therefore sequences encoding enzymes in the metabolic pathway provide particularly useful promoter sequences. Examples include alcohol dehydrogenase (ADH) (EP-A-0 284 044), enolase, glucokinase, glucose-6-phosphate isomerase, glyceraldehyde-3-phosphate-dehydrogenase (GAP or GAPDH), hexokinase, phosphofructokinase, 3-phosphoglycerate mutase, and pyruvate kinase (PyK) (EPO-A-0 329 203). The yeast PHO5 gene, encoding acid phosphatase, also provides useful promoter sequences [Myanohara et al. (1983) Proc. Natl. Acad. Sci. USA 80:1).
• In addition, synthetic promoters which do not occur in nature also function as yeast promoters. For example, UAS sequences of one yeast promoter may be joined with the transcription activation region of another yeast promoter, creating a synthetic hybrid promoter. Examples of such hybrid promoters include the ADH regulatory sequence linked to the GAP transcription activation region (U.S. Pat. Nos. 4,876,197 and 4,880,734). Other examples of hybrid promoters include promoters which consist of the regulatory sequences of either the ADH2, GAL4, GAL10, OR PHO5 genes, combined with the transcriptional activation region of a glycolytic enzyme gene such as GAP or PyK (EP-A-0 164 556). Furthermore, a yeast promoter can include naturally occurring promoters of non-yeast origin that have the ability to bind yeast RNA polymerase and initiate transcription. Examples of such promoters include, inter alia, [Cohen et al. (1980) Proc. Natl. Acad. Sci. USA 77:1078; Henikoff et al. (1981) Nature 283:835; Hollenberg et al. (1981) Curr. Topics Microbiol. Immunol. 96:119; Hollenberg et al. (1979) “The Expression of Bacterial Antibiotic Resistance Genes in the Yeast Saccharomyces cerevisiae,” in: Plasmids of Medical, Environmental and Commercial Importance (eds. K. N. Timmis and A. Puhler); Mercerau-Puigalon et al. (1980) Gene 11:163; Panthier et al. (1980) Curr. Genet. 2:109;].
• A DNA molecule may be expressed intracellularly in yeast. A promoter sequence may be directly linked with the DNA molecule, in which case the first amino acid at the N-terminus of the recombinant protein will always be a methionine, which is encoded by the ATG start codon. If desired, methionine at the N-terminus may be cleaved from the protein by in vitro incubation with cyanogen bromide.
• Fusion proteins provide an alternative for yeast expression systems, as well as in mammalian, baculovirus, and bacterial expression systems. Usually, a DNA sequence encoding the N-terminal portion of an endogenous yeast protein, or other stable protein, is fused to the 5′ end of heterologous coding sequences. Upon expression, this construct will provide a fusion of the two amino acid sequences. For example, the yeast or human superoxide dismutase (SOD) gene, can be linked at the 5′ terminus of a foreign gene and expressed in yeast. The DNA sequence at the junction of the two amino acid sequences may or may not encode a cleavable site. See eg. EP-A-0 196 056. Another example is a ubiquitin fusion protein. Such a fusion protein is made with the ubiquitin region that preferably retains a site for a processing enzyme (eg, ubiquitin-specific processing protease) to cleave the ubiquitin from the foreign protein. Through this method, therefore, native foreign protein can be isolated (eg. WO88/024066).
• Alternatively, foreign proteins can also be secreted from the cell into the growth media by creating chimeric DNA molecules that encode a fusion protein comprised of a leader sequence fragment that provide for secretion in yeast of the foreign protein. Preferably, there are processing sites encoded between the leader fragment and the foreign gene that can be cleaved either in vivo or in vitro. The leader sequence fragment usually encodes a signal peptide comprised of hydrophobic amino acids which direct the secretion of the protein from the cell.
• DNA encoding suitable signal sequences can be derived from genes for secreted yeast proteins, such as the yeast invertase gene (EP-A-0 012 873; JPO. 62,096,086) and the A-factor gene (U.S. Pat. No. 4,588,684). Alternatively, leaders of non-yeast origin, such as an interferon leader, exist that also provide for secretion in yeast (EP-A-0 060 057).
• A preferred class of secretion leaders are those that employ a fragment of the yeast alpha-factor gene, which contains both a “pre” signal sequence, and a “pro” region. The types of alpha-factor fragments that can be employed include the full-length pre-pro alpha factor leader (about 83 amino acid residues) as well as truncated alpha-factor leaders (usually about 25 to about 50 amino acid residues) (U.S. Pat. Nos. 4,546,083 and 4,870,008; EP-A-0 324 274). Additional leaders employing an alpha-factor leader fragment that provides for secretion include hybrid alpha-factor leaders made with a presequence of a first yeast, but a pro-region from a second yeast alphafactor. (eg. see WO 89/02463.)
• Usually, transcription termination sequences recognized by yeast are regulatory regions located 3′ to the translation stop codon, and thus together with the promoter flank the coding sequence. These sequences direct the transcription of an mRNA which can be translated into the polypeptide encoded by the DNA. Examples of transcription terminator sequence and other yeast-recognized termination sequences, such as those coding for glycolytic enzymes.
• Usually, the above described components, comprising a promoter, leader (if desired), coding sequence of interest, and transcription termination sequence, are put together into expression constructs. Expression constructs are often maintained in a replicon, such as an extrachromosomal element (eg. plasmids) capable of stable maintenance in a host, such as yeast or bacteria. The repl icon may have two replication systems, thus allowing it to be maintained, for example, in yeast for expression and in a prokaryotic host for cloning and amplification. Examples of such yeast-bacteria shuttle vectors include YEp24 [Botstein et al. (1979) Gene 8:17-24], pCl/1 [Brake et al. (1984) Proc. Natl. Acad. Sci. USA 81:4642-4646], and YRp17 [Stinchcomb et al. (1982) J. Mol. Biol. 158:157]. In addition, a replicon may be either a high or low copy number plasmid. A high copy number plasmid will generally have a copy number ranging from about 5 to about 200, and usually about 10 to about 150. A host containing a high copy number plasmid will preferably have at least about 10, and more preferably at least about 20. Enter a high or low copy number vector may be selected, depending upon the effect of the vector and the foreign protein on the host. See eg. Brake et al., supra.
• Alternatively, the expression constructs can be integrated into the yeast genome with an integrating vector. Integrating vectors usually contain at least one sequence homologous to a yeast chromosome that allows the vector to integrate, and preferably contain two homologous sequences flanking the expression construct. Integrations appear to result from recombinations between homologous DNA in the vector and the yeast chromosome [Orr-Weaver et al. (1983) Methods in Enzymol. 101:228-245]. An integrating vector may be directed to a specific locus in yeast by selecting the appropriate homologous sequence for inclusion in the vector. See Orr-Weaver et al., supra. One or more expression construct may integrate, possibly affecting levels of recombinant protein produced [Rine et al. (1983) Proc. Natl. Acad. Sci. USA 80:6750]. The chromosomal sequences included in the vector can occur either as a single segment in the vector, which results in the integration of the entire vector, or two segments homologous to adjacent segments in the chromosome and flanking the expression construct in the vector, which can result in the stable integration of only the expression construct.
• Usually, extrachromosomal and integrating expression constructs may contain selectable markers to allow for the selection of yeast strains that have been transformed. Selectable markers may include biosynthetic genes that can be expressed in the yeast host, such as ADE2, HIS4, LEU2, TRP1, and ALG7, and the G418 resistance gene, which confer resistance in yeast cells to tunicamycin and G418, respectively. In addition, a suitable selectable marker may also provide yeast with the ability to grow in the presence of toxic compounds, such as metal. For example, the presence of CUP1 allows yeast to grow in the presence of copper ions [Butt et al. (1987) Microbiol, Rev. 51:351].
• Alternatively, some of the above described components can be put together into transformation vectors. Transformation vectors are usually comprised of a selectable marker that is either maintained in a replicon or developed into an integrating vector, as described above.
• Expression and transformation vectors, either extrachromosomal replicons or integrating vectors, have been developed for transformation into many yeasts. For example, expression vectors have been developed for, inter alia, the following yeasts: Candida albicans [Kurtz, et al. (1986) Mol. Cell. Biol. 6:142], Candida maltosa [Kunze, et al. (1985) J. Basic Microbiol. 25:141]. Hansenula polymorpha [Gleeson, et al. (1986) J. Gen. Microbiol. 132:3459; Roggenkamp et al. (1986) Mol. Gen. Genet. 202:302], Kluyveromyces fragilis [Das, et al. (1984) J. Bacteriol. 158:1165], Kluyveromyces lactis [De Louvencourt et al. (1983) J. Bacteriol. 154:737; Van den Berg et al. (1990) Bio/Technology 8:135], Pichia guillerimondii [Kunze et al. (1985) J. Basic Microbiol. 25:141], Pichia pastoris [Cregg, et al. (1985) Mol. Cell. Biol. 5:3376; U.S. Pat. Nos. 4,837,148 and 4,929,555], Saccharomyces cerevisiae [Hinnen et al. (1978) Proc. Natl. Acad. Sci. USA 75:1929; Ito et al. (1983) J. Bacteriol. 153:163], Schizosaccharomyces pombe [Beach and Nurse (1981) Nature 300:706], and Yarrowia lipolytica [Davidow, et al. (1985) Curr. Genet. 10:380471 Gaillardin, et al. (1985) Curr. Genet. 10:49].
• Methods of introducing exogenous DNA into yeast hosts are well-known in the art, and usually include either the transformation of spheroplasts or of intact yeast cells treated with alkali cations. Transformation procedures usually vary with the yeast species to be transformed. See eg. [Kurtz et al. (1986) Mol. Cell. Biol. 6:142; Kunze et al. (1985) J. Basic Microbiol. 25:141; Candida]; [Gleeson et al. (1986) J. Gen. Microbiol. 132:3459; Roggenkamp et al. (1986) Mol. Gen. Genet. 202:302; Hansenula]; [Das et al. (1984) J. Bacteriol. 158:1165; De Louvencourt et al. (1983) J. Bacteriol. 154:1165; Van den Berg et al. (1990) Bio/Technology 8:135; Kluyveromyces]; [Cregg et al. (1985) Mol. Cell. Biol. 5:3376; Kunze et al. (1985) J. Basic Microbiol. 25:141; U.S. Pat. Nos. 4,837,148 and 4,929,555; Pichia]; [Hinnen et al. (1978) Proc. Natl. Acad. Sci. USA 75; 1929; Ito et al. (1983) J. Bacteriol. 153:163 Saccharomyces]; [Beach and Nurse (1981) Nature 300:706; Schizosaccharomyces]; [Davidow et al. (1985) Curr. Genet. 10:39; Gaillardin et al. (1985) Curr. Genet. 10:49; Yarrowia].
Antibodies
• As used herein, the term “antibody” refers to a polypeptide or group of polypeptides composed of at least one antibody combining site. An “antibody combining site” is the three-dimensional binding space with an internal surface shape and charge distribution complementary to the features of an epitope of an antigen, which allows a binding of the antibody with the antigen. “Antibody” includes, for example, vertebrate antibodies, hybrid antibodies, chimeric antibodies, humanised antibodies, altered antibodies, univalent antibodies, Fab proteins, and single domain antibodies.
• Antibodies against the proteins of the invention are useful for affinity chromatography, immunoassays, and distinguishing/identifying Neisserial proteins.
• Antibodies to the proteins of the invention, both polyclonal and monoclonal, may be prepared by conventional methods. In general, the protein is first used to immunize a suitable animal, preferably a mouse, rat, rabbit or goat. Rabbits and goats are preferred for the preparation of polyclonal sera due to the volume of serum obtainable, and the availability of labeled anti-rabbit and anti-goat antibodies. Immunization is generally performed by mixing or emulsifying the protein in saline, preferably in an adjuvant such as Freund's complete adjuvant, and injecting the mixture or emulsion parenterally (generally subcutaneously or intramuscularly): A dose of 50-200 μg/injection is typically sufficient. Immunization is generally boosted 2-6 weeks later with one or more injections of the protein in saline, preferably using Freund's incomplete adjuvant. One may alternatively generate antibodies by in vitro immunization using methods known in the art, which for the purposes of this invention is considered equivalent to in vivo immunization. Polyclonal antisera is obtained by bleeding the immunized animal into a glass or plastic container, incubating the blood at 25° C. for one hour, followed by incubating at 4° C. for 2-18 hours. The serum is recovered by centrifugation (eg. 1,000 g for 10 minutes). About 20-50 ml per bleed may be obtained from rabbits.
• Monoclonal antibodies are prepared using the standard method of Kohler & Milstein [Nature (1975) 256:495-96], or a modification thereof. Typically, a mouse or rat is immunized as described above. However, rather than bleeding the animal to extract serum, the spleen (and optionally several large lymph nodes) is removed and dissociated into single cells. If desired, the spleen cells may be screened (after removal of nonspecifically adherent cells) by applying a cell suspension to a plate or well coated with the protein antigen. B-cells expressing membrane-bound immunoglobulin specific for the antigen bind to the plate, and are not rinsed away with the rest of the suspension. Resulting B-cells, or all dissociated spleen cells, are then induced to fuse with myeloma cells to form hybridomas, and are cultured in a selective medium (eg. hypoxanthine, aminopterin, thymidine medium, “HAT”). The resulting hybridomas are plated by limiting dilution, and are assayed for the production of antibodies which bind specifically to the immunizing antigen (and which do not bind to unrelated antigens). The selected MAb-secreting hybridomas are then cultured either in vitro (eg. in tissue culture bottles or hollow fiber reactors), or in vivo (as ascites in mice).
• If desired, the antibodies (whether polyclonal or monoclonal) may be labeled using conventional techniques. Suitable labels include fluorophores, chromophores, radioactive atoms (particularly ³²P and ¹²⁵I), electron-dense reagents, enzymes, and ligands having specific binding partners. Enzymes are typically detected by their activity. For example, horseradish peroxidase is usually detected by its ability to convert 3,3′,5,5′-tetramethylbenzidine (TMB) to a blue pigment, quantifiable with a spectrophotometer. “Specific binding partner” refers to a protein capable of binding a ligand molecule with high specificity, as for example in the case of an antigen and a monoclonal antibody specific therefor. Other specific binding partners include biotin and avidin or streptavidin, IgG and protein A, and the numerous receptor-ligand couples known in the art. It should be understood that the above description is not meant to categorize the various labels into distinct classes, as the same label may serve in several different modes. For example, ¹²⁵I may serve as a radioactive label or as an electron-dense reagent. HRP may serve as enzyme or as antigen for a MAb. Further, one may combine various labels for desired effect. For example, MAbs and avidin also require labels in the practice of this invention: thus, one might label a MAb with biotin, and detect its presence with avidin labeled with ¹²⁵I, or with an anti-biotin MAb labeled with HRP. Other permutations and possibilities will be readily apparent to those of ordinary skill in the art, and are considered as equivalents within the scope of the instant invention.
Pharmaceutical Compositions
• Pharmaceutical compositions can comprise either polypeptides, antibodies, or nucleic acid of the invention. The pharmaceutical compositions will comprise a therapeutically effective amount of either polypeptides, antibodies, or polynucleotides of the claimed invention.
• The term “therapeutically effective amount” as used herein refers to an amount of a therapeutic agent to treat, ameliorate, or prevent a desired disease or condition, or to exhibit a detectable therapeutic or preventative effect. The effect can be detected by, for example, chemical markers or antigen levels. Therapeutic effects also include reduction in physical symptoms, such as decreased body temperature. The precise effective amount for a subject will depend upon the subject's size and health, the nature and extent of the condition, and the therapeutics or combination of therapeutics selected for administration. Thus, it is not useful to specify an exact effective amount in advance. However, the effective amount for a given situation can be determined by routine experimentation and is within the judgement of the clinician.
• For purposes of the present invention, an effective dose will be from about 0.01 mg/kg to 50 mg/kg or 0.05 mg/kg to about 10 mg/kg of the DNA constructs in the individual to which it is administered.
• A pharmaceutical composition can also contain a pharmaceutically acceptable carrier. The term “pharmaceutically acceptable carrier” refers to a carrier for administration of a therapeutic agent, such as antibodies or a polypeptide, genes, and other therapeutic agents. The term refers to any pharmaceutical carrier that does not itself induce the production of antibodies harmful to the individual receiving the composition, and which may be administered without undue toxicity. Suitable carriers may be large, slowly metabolized macromolecules such as proteins, polysaccharides, polylactic acids, polyglycolic acids, polymeric amino acids, amino acid copolymers, and inactive virus particles. Such carriers are well known to those of ordinary skill in the art.
• Pharmaceutically acceptable salts can be used therein, for example, mineral acid salts such as hydrochlorides, hydrobromides, phosphates, sulfates, and the like; and the salts of organic acids such as acetates, propionates, malonates, benzoates, and the like. A thorough discussion of pharmaceutically acceptable excipients is available in Remington's Pharmaceutical Sciences (Mack Pub. Co., N.J. 1991).
• Pharmaceutically acceptable carriers in therapeutic compositions may contain liquids such as water, saline, glycerol and ethanol. Additionally, auxiliary substances, such as wetting or emulsifying agents, pH buffering substances, and the like, may be present in such vehicles. Typically, the therapeutic compositions are prepared as injectables, either as liquid solutions or suspensions; solid forms suitable for solution in, or suspension in, liquid vehicles prior to injection may also be prepared. Liposomes are included within the definition of a pharmaceutically acceptable carrier.
Delivery Methods
• Once formulated, the compositions of the invention can be administered directly to the subject. The subjects to be treated can be animals; in particular, human subjects can be treated.
• Direct delivery of the compositions will generally be accomplished by injection, either subcutaneously, intraperitoneally, intravenously or intramuscularly or delivered to the interstitial space of a tissue. The compositions can also be administered into a lesion. Other modes of administration include oral and pulmonary administration, suppositories, and transdermal or transcutaneous applications (eg. see WO98/20734), needles, and gene guns or hyposprays. Dosage treatment may be a single dose schedule or a multiple dose schedule.
Vaccines
• Vaccines according to the invention may either be prophylactic (ie. to prevent infection) or therapeutic (ie. to treat disease after infection).
• Such vaccines comprise immunising antigen(s), immunogen(s), polypeptide(s), protein(s) or nucleic acid, usually in combination with “pharmaceutically acceptable carriers,” which include any carrier that does not itself induce the production of antibodies harmful to the individual receiving the composition. Suitable carriers are typically large, slowly metabolized macromolecules such as proteins, polysaccharides, polylactic acids, polyglycolic acids, polymeric amino acids, amino acid copolymers, lipid aggregates (such as oil droplets or liposomes), and inactive virus particles. Such carriers are well known to those of ordinary skill in the art. Additionally, these carriers may function as immunostimulating agents (“adjuvants”). Furthermore, the antigen or immunogen may be conjugated to a bacterial toxoid, such as a toxoid from diphtheria, tetanus, cholera, H. pylori, etc. pathogens.
• Preferred adjuvants to enhance effectiveness of the composition include, but are not limited to: (1) aluminum salts (alum), such as aluminum hydroxide, aluminum phosphate, aluminum sulfate, etc; (2) oil-in-water emulsion formulations (with or without other specific immunostimulating agents such as muramyl peptides (see below) or bacterial cell wall components), such as for example (a) MF59™ (WO 90/14837; Chapter 10 in Vaccine design: the subunit and adjuvant approach, eds. Powell & Newman, Plenum Press 1995), containing 5% Squalene, 0.5% Tween 80, and 0.5% Span 85 (optionally containing various amounts of MTP-PE (see below), although not required) formulated into submicron particles using a microfluidizer such as Model 110Y microfluidizer (Microfluidics, Newton, Mass.), (b) SAF, containing 10% Squalane, 0.4% Tween 80, 5% pluronic-blocked polymer L121, and thr-MDP (see below) either microfluidized into a submicron emulsion or vortexed to generate a larger particle size emulsion, and (c) Ribi™ adjuvant system (RAS), (Ribi Immunochem, Hamilton, Mont.) containing 2% Squalene, 0.2% Tween 80, and one or more bacterial cell wall components from the group consisting of monophosphorylipid A (MPL), trehalose dimycolate (TDM), and cell wall skeleton (CWS), preferably MPL+CWS (Detox™); (3) saponin adjuvants, such as Stimulon™ (Cambridge Bioscience, Worcester, Mass.) may be used or particles generated therefrom such as ISCOMs (immunostimulating complexes); (4) Complete Freund's Adjuvant (CFA) and Incomplete Freund's Adjuvant (IFA); (5) cytokines, such as interleukins (eg. IL-1, IL-2, IL-4, IL-5, IL-6, IL-7, IL-12, etc.), interferons (eg. gamma interferon), macrophage colony stimulating factor (M-CSF), tumor necrosis factor (TNF), etc; and (6) other substances that act as immunostimulating agents to enhance the effectiveness of the composition. Alum and MF59™ are preferred.
• As mentioned above, muramyl peptides include, but are not limited to, N-acetyl-muramyl-L-threonyl-D-isoglutamine (thr-MDP), N-acetyl-normuramyl-L-alanyl-D-isoglutamine (nor-MDP), N-acetylmuramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-(1′-2′-dipalmitoyl-sn-glycero-3-hydroxyphosphoryloxy)-ethylamine (MTP-PE), etc.
• The immunogenic compositions (eg. the immunising antigen/immunogen/polypeptide/protein/nucleic acid, pharmaceutically acceptable carrier, and adjuvant) typically will contain diluents, such as water, saline, glycerol, ethanol, etc. Additionally, auxiliary substances, such as wetting or emulsifying agents, pH buffering substances, and the like, may be present in such vehicles.
• Typically, the immunogenic compositions are prepared as injectables, either as liquid solutions or suspensions; solid forms suitable for solution in, or suspension in, liquid vehicles prior to injection may also be prepared. The preparation also may be emulsified or encapsulated in liposomes for enhanced adjuvant effect, as discussed above under pharmaceutically acceptable carriers.
• Immunogenic compositions used as vaccines comprise an immunologically effective amount of the antigenic or immunogenic polypeptides, as well as any other of the above-mentioned components, as needed. By “immunologically effective amount”, it is meant that the administration of that amount to an individual, either in a single dose or as part of a series, is effective for treatment or prevention. This amount varies depending upon the health and physical condition of the individual to be treated, the taxonomic group of individual to be treated (eg. nonhuman primate, primate, etc.), the capacity of the individual's immune system to synthesize antibodies, the degree of protection desired, the formulation of the vaccine, the treating doctor's assessment of the medical situation, and other relevant factors. It is expected that the amount will fall in a relatively broad range that can be determined through routine trials.
• The immunogenic compositions are conventionally administered parenterally, eg. by injection, either subcutaneously, intramuscularly, or transdermally/transcutaneously (eg. WO98/20734). Additional formulations suitable for other modes of administration include oral and pulmonary formulations, suppositories, and transdermal applications. Dosage treatment may be a single dose schedule or a multiple dose schedule. The vaccine may be administered in conjunction with other immunoregulatory agents.
• As an alternative to protein-based vaccines, DNA vaccination may be employed [eg. Robinson & Torres (1997) Seminars in Immunology 9:271-283; Donnelly et al. (1997) Annu Rev Immunol 15:617-648; see later herein].
Gene Delivery Vehicles
• Gene therapy vehicles for delivery of constructs including a coding sequence of a therapeutic of the invention, to be delivered to the mammal for expression in the mammal, can be administered either locally or systemically. These constructs can utilize viral or non-viral vector approaches in in vivo or ex vivo modality. Expression of such coding sequence can be induced using endogenous mammalian or heterologous promoters. Expression of the coding sequence in vivo can be either constitutive or regulated.
• The invention includes gene delivery vehicles capable of expressing the contemplated nucleic acid sequences. The gene delivery vehicle is preferably a viral vector and, more preferably, a retroviral, adenoviral, adeno-associated viral (AAV), herpes viral, or alphavirus vector. The viral vector can also be an astrovirus, coronavirus, orthomyxovirus, papovavirus, paramyxovirus, parvovirus, picornavirus, poxvirus, or togavirus viral vector. See generally, Jolly (1994) Cancer Gene Therapy 1:51-64; Kimura (1994) Human Gene Therapy 5:845-852; Connelly (1995) Human Gene Therapy 6:185-193; and Kaplitt (1994) Nature Genetics 6:148-153.
• Retroviral vectors are well known in the art and we contemplate that any retroviral gene therapy vector is employable in the invention, including B, C and D type retroviruses, xenotropic retroviruses (for example, NZB-X1, NZB-X2 and NZB9-1 (see O'Neill (1985) J. Virol. 53:160) polytropic retroviruses eg. MCF and MCF-M LV (see Kelly (1983) J. Virol. 45:291), spumaviruses and lentiviruses. See RNA Tumor Viruses, Second Edition, Cold Spring Harbor Laboratory, 1985.
• Portions of the retroviral gene therapy vector may be derived from different retroviruses. For example, retrovector LTRs may be derived from a Murine Sarcoma Virus, a tRNA binding site from a Rous Sarcoma Virus, a packaging signal from a Murine Leukemia Virus, and an origin of second strand synthesis from an Avian Leukosis Virus.
• These recombinant retroviral vectors may be used to generate transduction competent retroviral vector particles by introducing them into appropriate packaging cell lines (see U.S. Pat. No. 5,591,624). Retrovirus vectors can be constructed for site-specific integration into host cell DNA by incorporation of a chimeric integrase enzyme into the retroviral particle (see WO96/37626). It is preferable that the recombinant viral vector is a replication defective recombinant virus.
• Packaging cell lines suitable for use with the above-described retrovirus vectors are well known in the art, are readily prepared (see WO95/30763 and WO92/05266), and can be used to create producer cell lines (also termed vector cell lines or “VCLs”) for the production of recombinant vector particles. Preferably, the packaging cell lines are made from human parent cells (eg. HT1080 cells) or mink parent cell lines, which eliminates inactivation in human serum.
• Preferred retroviruses for the construction of retroviral gene therapy vectors include Avian Leukosis Virus, Bovine Leukemia, Virus, Murine Leukemia Virus, Mink-Cell Focus-Inducing Virus, Murine Sarcoma Virus, Reticuloendotheliosis Virus and Rous Sarcoma Virus. Particularly preferred Murine Leukemia Viruses include 4070A and 1504A (Hartley and Rowe (1976) J Virol 19:19-25), Abelson (ATCC No. VR-999), Friend (ATCC No. VR-245), Graffi, Gross (ATCC Nol VR-590), Kirsten, Harvey Sarcoma Virus and Rauscher (ATCC No. VR-998) and Moloney Murine Leukemia Virus (ATCC No. VR-190). Such retroviruses may be obtained from depositories or collections such as the American Type Culture Collection (“ATCC”) in Rockville, Md. or isolated from known sources using commonly available techniques.
• Exemplary known retroviral gene therapy vectors employable in this invention include those described in patent applications GB2200651, EP0415731, EP0345242, EP0334301, WO89/02468; WO89/05349, WO89/09271, WO90/02806, WO90/07936, WO94/03622, WO93/25698, WO93/25234, WO93/11230, WO93/10218, WO91/02805, WO91/02825, WO95/07994, U.S. Pat. No. 5,219,740, U.S. Pat. No. 4,405,712, U.S. Pat. No. 4,861,719, U.S. Pat. No. 4,980,289, U.S. Pat. No. 4,777,127, U.S. Pat. No. 5,591,624. See also Vile (1993) Cancer Res 53:3860-3864; Vile (1993) Cancer Res 53:962-967; Ram (1993) Cancer Res 53 (1993) 83-88; Takamiya (1992) J Neurosci Res 33:493-503; Baba (1993) J Neurosurg 79:729-735; Mann (1983) Cell 33:153; Cane (1984) Proc Natl Acad Sci 81:6349; and Miller (1990) Human Gene Therapy 1.
• Human adenoviral gene therapy vectors are also known in the art and employable in this invention. See, for example, Berkner (1988) Biotechniques 6:616 and Rosenfeld (1991) Science 252:431, and WO93/07283, WO93/06223, and WO93/07282. Exemplary known adenoviral gene therapy vectors employable in this invention include those described in the above referenced documents and in WO94/12649, WO93/03769, WO93/19191, WO94/28938, WO95/11984, WO95/00655, WO95/27071, WO95/29993, WO95/34671, WO96/05320, WO94/08026, WO94/11506, WO93/06223, WO94/24299, WO95/14102, WO95/24297, WO95/02697, WO94/28152, WO94/24299, WO95/09241, WO95/25807, WO95/05835, WO94/18922 and WO95/09654. Alternatively, administration of DNA linked to killed adenovirus as described in Curiel (1992) Hum. Gene Ther. 3:147-154 may be employed. The gene delivery vehicles of the invention also include adenovirus associated virus (AAV) vectors. Leading and preferred examples of such vectors for use in this invention are the AAV-2 based vectors disclosed in Srivastava, WO93/09239. Most preferred AAV vectors comprise the two AAV inverted terminal repeats in which the native D-sequences are modified by substitution of nucleotides, such that at least 5 native nucleotides and up to 18 native nucleotides, preferably at least 10 native nucleotides up to 18 native nucleotides, most preferably 10 native nucleotides are retained and the remaining nucleotides of the D-sequence are deleted or replaced with non-native nucleotides. The native D-sequences of the AAV inverted terminal repeats are sequences of 20 consecutive nucleotides in each AAV inverted terminal repeat (ie. there is one sequence at each end) which are not involved in HP formation. The non-native replacement nucleotide may be any nucleotide other than the nucleotide found in the native D-sequence in the same position. Other employable exemplary AAV vectors are pWP-19, pWN-1, both of which are disclosed in Nahreini (1993) Gene 124:257-262. Another example of such an AAV vector is psub201 (see Samulski (1987) J. Virol. 61:3096). Another exemplary AAV vector is the Double-D ITR vector. Construction of the Double-D ITR vector is disclosed in U.S. Pat. No. 5,478,745. Still other vectors are those disclosed in Carter U.S. Pat. No. 4,797,368 and Muzyczka U.S. Pat. No. 5,139,941, Chartejee U.S. Pat. No. 5,474,935, and Kotin WO94/288157. Yet a further example of an AAV vector employable in this invention is SSV9AFABTKneo, which contains the AFP enhancer and albumin promoter and directs expression predominantly in the liver. Its structure and construction are disclosed in Su (1996) Human Gene Therapy 7:463-470. Additional AAV gene therapy vectors are described in U.S. Pat. No. 5,354,678, U.S. Pat. No. 5,173,414, U.S. Pat. No. 5,139,941, and U.S. Pat. No. 5,252,479.
• The gene therapy vectors of the invention also include herpes vectors. Leading and preferred examples are herpes simplex virus vectors containing a sequence encoding a thymidine kinase polypeptide such as those disclosed in U.S. Pat. No. 5,288,641 and EP0176170 (Roizman). Additional exemplary herpes simplex virus vectors include HFEWICP6-LacZ disclosed in WO95/04139 (Wistar Institute), pHSVlac described in Geller (1988) Science 241:1667-1669 and in WO90/09441 and WO92/07945, HSV Us3::pgC-lacZ described in Fink (1992) Human Gene Therapy 3:11-19 and HSV 7134, 2 RH 105 and GAL4 described in EP 0453242 (Breakefield), and those deposited with the ATCC as accession numbers ATCC VR-977 and ATCC VR-260.
• Also contemplated are alpha virus gene therapy vectors that can be employed in this invention. Preferred alpha virus vectors are Sindbis viruses vectors. Togaviruses, Semliki Forest virus (ATCC VR-67; ATCC VR-1247), Middleberg virus (ATCC VR-370), Ross River virus (ATCC VR-373; ATCC VR-1246), Venezuelan equine encephalitis virus (ATCC VR923; ATCC VR-1250; ATCC VR-1249; ATCC VR-532), and those described in U.S. Pat. Nos. 5,091,309, 5,217,879, and WO92/10578. More particularly, those alpha virus vectors described in U.S. Ser. No. 08/405,627, filed Mar. 15, 1995, WO94/21792, WO92/10578, WO95/07994, U.S. Pat. No. 5,091,309 and U.S. Pat. No. 5,217,879 are employable. Such alpha viruses may be obtained from depositories or collections such as the ATCC in Rockville, Md. or isolated from known sources using commonly available techniques. Preferably, alphavirus vectors with reduced cytotoxicity are used (see U.S. Ser. No. 08/679,640).
• DNA vector systems such as eukarytic layered expression systems are also useful for expressing the nucleic acids of the invention. See WO95/07994 for a detailed description of eukaryotic layered expression systems. Preferably, the eukaryotic layered expression systems of the invention are derived from alphavirus vectors and most preferably from Sindbis viral vectors.
• Other viral vectors suitable for use in the present invention include those derived from poliovirus, for example ATCC VR-58 and those described in Evans, Nature 339 (1989)385 and Sabin (1973) J. Biol. Standardization 1:115; rhinovirus, for example ATCC VR-1110 and those described in Arnold (1990) J Cell Biochem L401; pox viruses such as canary pox virus or vaccinia virus, for example ATCC VR-111 and ATCC VR-2010 and those described in Fisher-Hoch (1989) Proc Nan Acad Sci 86:317; Flexner (1989) Ann NY Acad Sci 569:86, Flexner (1990) Vaccine 8:17; in U.S. Pat. No. 4,603,112 and U.S. Pat. No. 4,769,330 and WO89/01973; SV40 virus, for example ATCC VR-305 and those described in Mulligan (1979) Nature 277:108 and Madzak (1992) J Gen Virol 73:1533; influenza virus, for example ATCC VR-797 and recombinant influenza viruses made employing reverse genetics techniques as described in U.S. Pat. No. 5,166,057 and in Enami (1990) Proc Nall Acad Sci 87:3802-3805; Enami & Palese (1991) J Virol 65:2711-2713 and Luytjes (1989) Cell 59:110, (see also McMichael (1983) NEJ Med 309:13, and Yap (1978) Nature 273:238 and Nature (1979) 277:108); human immunodeficiency virus as described in EP-0386882 and in Buchschacher (1992) J. Virol. 66:2731; measles virus, for example ATCC VR-67 and VR-1247 and those described in EP-0440219; Aura virus, for example ATCC VR-368; Bebaru virus, for example ATCC VR-600 and ATCC VR-1240; Cabassou virus, for example ATCC VR-922; Chikungunya virus, for example ATCC VR-64 and ATCC VR-1241; Fort Morgan Virus, for example ATCC VR-924; Getah virus, for example ATCC VR-369 and ATCC VR-1243; Kyzylagach virus, for example ATCC VR-927; Mayaro virus, for example ATCC VR-66; Mucambo virus, for example ATCC VR-580 and ATCC VR-1244; Ndumu virus, for example ATCC VR-371; Pixuna virus, for example ATCC VR-372 and ATCC VR-1245; Tonate virus, for example ATCC VR-925; Triniti virus, for example ATCC VR-469; Una virus, for example ATCC VR-374; Whataroa virus, for example ATCC VR-926; Y-62-33 virus, for example ATCC VR-375; O'Nyong virus, Eastern encephalitis virus, for example ATCC VR-65 and ATCC VR-1242; Western encephalitis virus, for example ATCC VR-70, ATCC VR-1251, ATCC VR-622 and ATCC VR-1252; and coronavirus, for example ATCC VR-740 and those described in Hamre (1966) Proc Soc Exp Biol Med 121:190.
• Delivery of the compositions of this invention into cells is not limited to the above mentioned viral vectors. Other delivery methods and media may be employed such as, for example, nucleic acid expression vectors, polycationic condensed DNA linked or unlinked to killed adenovirus alone, for example see U.S. Ser. No. 08/366,787, filed Dec. 30, 1994 and Curie] (1992) Hum Gene Titer 3:147-154 ligand linked DNA, for example see Wu (1989) J Biol Chem 264:16985-16987, eucaryotic cell delivery vehicles cells, for example see U.S. Ser. No. 08/240,030, filed May 9, 1994, and U.S. Ser. No. 08/404,796, deposition of photopolymerized hydrogel materials, hand-held gene transfer particle gun, as described in U.S. Pat. No. 5,149,655, ionizing radiation as described in U.S. Pat. No. 5,206,152 and in WO92/11033, nucleic charge neutralization or fusion with cell membranes. Additional approaches are described in Philip (1994) Mol Cell Biol 14:2411-2418 and in Woffendin (1994) Proc Natl Acad Sci 91:1581-1585.
• Particle mediated gene transfer may be employed, for example see U.S. Ser. No. 60/023,867. Briefly, the sequence can be inserted into conventional vectors that contain conventional control sequences for high level expression, and then incubated with synthetic gene transfer molecules such as polymeric DNA-binding cations like polylysine, protamine, and albumin, linked to cell targeting ligands such as asialoorosomucoid, as described in Wu & Wu (1987) J. Biol. Chem. 262:4429-4432, insulin as described in Hucked (1990) Biochem Pharmacol 40:253-263, galactose as described in Plank (1992) Bioconjugate Chem 3:533-539, lactose or transferrin.
• Naked DNA may also be employed. Exemplary naked DNA introduction methods are described in WO 90/11092 and U.S. Pat. No. 5,580,859. Uptake efficiency may be improved using biodegradable latex beads. DNA coated latex beads are efficiently transported into cells after endocytosis initiation by the beads. The method may be improved further by treatment of the beads to increase hydrophobicity and thereby facilitate disruption of the endosome and release of the DNA into the cytoplasm.
• Liposomes that can act as gene delivery vehicles are described in U.S. Pat. No. 5,422,120, WO95/13796, WO94/23697, WO91/14445 and EP-524,968. As described in U.S. Ser. No. 60/023,867, on non-viral delivery, the nucleic acid sequences encoding a polypeptide can be inserted into conventional vectors that contain conventional control sequences for high level expression, and then be incubated with synthetic gene transfer molecules such as polymeric DNA-binding cations like polylysine, protamine, and albumin, linked to cell targeting ligands such as asialoorosomucoid, insulin, galactose, lactose, or transferrin. Other delivery systems include the use of liposomes to encapsulate DNA comprising the gene under the control of a variety of tissue-specific or ubiquitously-active promoters. Further non-viral delivery suitable for use includes mechanical delivery systems such as the approach described in Woffendin et al (1994) Proc. Natl. Acad. Sci. USA 91(24):11581-11585. Moreover, the coding sequence and the product of expression of such can be delivered through deposition of photopolymerized hydrogel materials. Other conventional methods for gene delivery that can be used for delivery of the coding sequence include, for example, use of hand-held gene transfer particle gun, as described in U.S. Pat. No. 5,149,655; use of ionizing radiation for activating transferred gene, as described in U.S. Pat. No. 5,206,152 and WO92/11033
• Exemplary liposome and polycationic gene delivery vehicles are those described in U.S. Pat. Nos. 5,422,120 and 4,762,915; in WO 95/13796; WO94/23697; and WO91/14445; in EP-0524968; and in Stryer, Biochemistry, pages 236-240 (1975) W.H. Freeman, San Francisco; Szoka (1980) Biochem Biophys Acta 600:1; Bayer (1979) Biochem Biophys Acta 550:464; Rivnay (1987) Meth Enzymol 149:119; Wang (1987) Proc Natl Acad Sci 84:7851; Plant (1989) Anal Biochem 176:420.
• A polynucleotide composition can comprises therapeutically effective amount of a gene therapy vehicle, as the term is defined above. For purposes of the present invention, an effective dose will be from about 0.01 mg/kg to 50 mg/kg or 0.05 mg/kg to about 10 mg/kg of the DNA constructs in the individual to which it is administered.
Delivery Methods
• Once formulated, the polynucleotide compositions of the invention can be administered (1) directly to the subject; (2) delivered ex vivo, to cells derived from the subject; or (3) in vitro for expression of recombinant proteins. The subjects to be treated can be mammals or birds. Also, human subjects can be treated.
• Direct delivery of the compositions will generally be accomplished by injection, either subcutaneously, intraperitoneally, intravenously or intramuscularly or delivered to the interstitial space of a tissue. The compositions can also be administered into a lesion. Other modes of administration include oral and pulmonary administration, suppositories, and transdermal or transcutaneous applications (eg. see WO98/20734), needles, and gene guns or hyposprays. Dosage treatment may be a single dose schedule or a multiple dose schedule.
• Methods for the ex vivo delivery and reimplantation of transformed cells into a subject are known in the art and described in eg. WO93/14778. Examples of cells useful in ex vivo applications include, for example, stem cells, particularly hematopoetic, lymph cells, macrophages, dendritic cells, or tumor cells.
• Generally, delivery of nucleic acids for both ex vivo and in vitro applications can be accomplished by the following procedures, for example, dextran-mediated transfection, calcium phosphate precipitation, polybrene mediated transfection, protoplast fusion, electroporation, encapsulation of the polynucleotide(s) in liposomes, and direct microinjection of the DNA into nuclei, all well known in the art.
Polynucleotide and Polypeptide Pharmaceutical Compositions
• In addition to the pharmaceutically acceptable carriers and salts described above, the following additional agents can be used with polynucleotide and/or polypeptide compositions.
A. Polypeptides
• One example are polypeptides which include, without limitation: asioloorosomucoid (ASOR); transferrin; asialoglycoproteins; antibodies; antibody fragments; ferritin; interleukins; interferons, granulocyte, macrophage colony stimulating factor (GM-CSF), granulocyte colony stimulating factor (G-CSF), macrophage colony stimulating factor (M-CSF), stem cell factor and erythropoietin. Viral antigens, such as envelope proteins, can also be used. Also, proteins from other invasive organisms, such as the 17 amino acid peptide from the circumsporozoite protein of plasmodium falciparum known as RH.
B. Hormones, Vitamins, etc.
• Other groups that can be included are, for example: hormones, steroids, androgens, estrogens, thyroid hormone, or vitamins, folic acid.
C. Polyalkylenes, Polysaccharides, etc.
• Also, polyalkylene glycol can be included with the desired polynucleotides/polypeptides. In a preferred embodiment, the polyalkylene glycol is polyethlylene glycol. Iri addition, mono-, di-, or polysaccarides can be included. In a preferred embodiment of this aspect, the polysaccharide is dextran or DEAE-dextran. Also, chitosan and poly(lactide-co-glycolide)
D. Lipids, and Liposomes
• The desired polynucleotide/polypeptide can also be encapsulated in lipids or packaged in liposomes prior to delivery to the subject or to cells derived therefrom.
• Lipid encapsulation is generally accomplished using liposomes which are able to stably bind or entrap and retain nucleic acid. The ratio of condensed polynucleotide to lipid preparation can vary but will generally be around 1:1 (mg DNA:micromoles lipid), or more of lipid. For a review of the use of liposomes as carriers for delivery of nucleic acids, see, Hug and Sleight (1991) Biochim. Biophys. Acta. 1097:1-17; Straubinger (1983) Meth. Enzymol. 101:512-527.
• Liposomal preparations for use in the present invention include cationic (positively charged), anionic (negatively charged) and neutral preparations. Cationic liposomes have been shown to mediate intracellular delivery of plasmid DNA (Feigner (1987) Proc. Natl. Acad. Sci. USA 84:7413-7416); mRNA (Malone (1989) Proc. Natl. Acad. Sci. USA 86:6077-6081); and purified transcription factors (Debs (1990) J. Biol. Chem. 265:10189-10192), in functional form.
• Cationic liposomes are readily available. For example, N[1-2,3-dioleyloxy)propyl]-N,N,N-triethylammonium (DOTMA) liposomes are available under the trademark Lipofectin, from GIBCO BRL, Grand Island, N.Y. (See, also, Feigner supra). Other commercially available liposomes include transfectace (DDAB/DOPE) and DOTAP/DOPE (Boerhinger). Other cationic liposomes can be prepared from readily available materials using techniques well known in the art. See, eg. Szoka (1978) Proc. Natl. Acad. Sci. USA 75:4194-4198; WO90/11092 fora description of the synthesis of DOTAP (1,2-bis(oleoyloxy)-3-(trimethylammonio)propane) liposomes.
• Similarly, anionic and neutral liposomes are readily available, such as from Avanti Polar Lipids (Birmingham, Ala.), or can be easily prepared using readily available materials. Such materials include phosphatidyl choline, cholesterol, phosphatidyl ethanolamine, dioleoylphosphatidyl choline (DOPC), dioleoylphosphatidyl glycerol (DOPG), dioleoylphoshatidyl ethanolamine (DOPE), among others. These materials can also be mixed with the DOTMA and DOTAP starting materials in appropriate ratios. Methods for making liposomes using these materials are well known in the art.
• The liposomes can comprise multilammelar vesicles (MLVs), small unilamellar vesicles (SUVs), or large unilamellar vesicles (LUVs). The various liposome-nucleic acid complexes are prepared using methods known in the art. See eg. Straubinger (1983) Meth. Immunol. 101:512-527; Szoka (1978) Proc. Natl. Acad. Sci. USA 75:4194-4198; Papahadjopoulos (1975) Biochim. Biophys. Acta 394:483; Wilson (1979) Cell 17:77); Deamer & Bangham (1976) Biochim. Biophys. Ada 443:629; Ostro (1977) Biochem. Biophys. Res. Commun. 76:836; Fraley (1979) Proc. Natl. Acad. Sci. USA 76:3348); Enoch & Strittmatter (1979) Proc. Natl. Acad. Sci. USA 76:145; Fraley (1980) J. Biol. Chem. (1980) 255:10431; Szoka & Papahadjopoulos (1978) Proc. Natl. Acad. Sci. USA 75:145; and Schaefer-Ridder (1982) Science 215:166.
E. Lipoproteins
• In addition, lipoproteins can be included with the polynucleotide/polypeptide to be delivered. Examples of lipoproteins to be utilized include: chylomicrons, HDL, IDL, LDL, and VLDL. Mutants, fragments, or fusions of these proteins can also be used. Also, modifications of naturally occurring lipoproteins can be used, such as acetylated LDL. These lipoproteins can target the delivery of polynucleotides to cells expressing lipoprotein receptors. Preferably, if lipoproteins are including with the polynucleotide to be delivered, no other targeting ligand is included in the composition.
• Naturally occurring lipoproteins comprise a lipid and a protein portion. The protein portion are known as apoproteins. At the present, apoproteins A, B, C, D, and E have been isolated and identified. At least two of these contain several proteins, designated by Roman numerals, AI, AII, AIV; CI, CII, CIII.
• A lipoprotein can comprise more than one apoprotein. For example, naturally occurring chylomicrons comprises of A, B, C, and E, over time these lipoproteins lose A and acquire C and E apoproteins. VLDL comprises A, B, C, and E apoproteins, LDL comprises apoprotein B; and HDL comprises apoproteins A, C, and E.
• The amino acid of these apoproteins are known and are described in, for example, Breslow (1985) Annu Rev. Biochem 54:699; Law (1986) Adv. Exp Med. Biol. 151:162; Chen (1986) J Biol Chem 261:12918; Kane (1980) Proc Natl Acad Sci USA 77:2465; and Utermann (1984) Hum Genet 65:232.
• Lipoproteins contain a variety of lipids including, triglycerides, cholesterol (free and esters), and phopholipids. The composition of the lipids varies in naturally occurring lipoproteins. For example, chylomicrons comprise mainly triglycerides. A more detailed description of the lipid content of naturally occurring lipoproteins can be found, for example, in Meth. Enzymol. 128 (1986). The composition of the lipids are chosen to aid in conformation of the apoprotein for receptor binding activity. The composition of lipids can also be chosen to facilitate hydrophobic interaction and association with the polynucleotide binding molecule.
• Naturally occurring lipoproteins can be isolated from serum by ultracentrifugation, for instance. Such methods are described in Meth. Enzymol. (supra); Pitas (1980) J. Biochem. 255:5454-5460 and Mahey (1979) J. Clin. Invest 64:743-750. Lipoproteins can also be produced by in vitro or recombinant methods by expression of the apoprotein genes in a desired host cell. See, for example, Atkinson (1986) Annu Rev Biophys Chem 15:403 and Radding (1958) Biochim Biophys Acta 30: 443. Lipoproteins can also be purchased from commercial suppliers, such as Biomedical Techniologies, Inc., Stoughton, Mass., USA. Further description of lipoproteins can be found in Zuckermann et al. PCT/US97/14465.
F. Polycationic Agents
• Polycationic agents can be included, with or without lipoprotein, in a composition with the desired polynucleotide/polypeptide to be delivered.
• Polycationic agents, typically, exhibit a net positive charge at physiological relevant pH and are capable of neutralizing the electrical charge of nucleic acids to facilitate delivery to a desired location. These agents have both in vitro, ex vivo, and in vivo applications. Polycationic agents can be used to deliver nucleic acids to a living subject either intramuscularly, subcutaneously, etc.
• The following are examples of useful polypeptides as polycationic agents: polylysine, polyarginine, polyornithine, and protamine. Other examples include histones, protamines, human serum albumin, DNA binding proteins, non-histone chromosomal proteins, coat proteins from DNA viruses, such as (X174, transcriptional factors also contain domains that bind DNA and therefore may be useful as nucleic aid condensing agents. Briefly, transcriptional factors such as C/CEBP, c-jun, c-fos, AP-1, AP-2, AP-3, CPF, Prot-1, Sp-1, Oct-1, Oct-2, CREP, and TFIID contain basic domains that bind DNA sequences.
• Organic polycationic agents include: spermine, spermidine, and purtrescine.
• The dimensions and of the physical properties of a polycationic agent can be extrapolated from the list above, to construct other polypeptide polycationic agents or to produce synthetic polycationic agents.
• Synthetic polycationic agents which are useful include, for example, DEAE-dextran, polybrene. Lipofectin™, and lipofectAMINE™ are monomers that form polycationic complexes when combined with polynucleotides/polypeptides.
Immunodiagnostic Assays
• Neisserial antigens of the invention can be used in immunoassays to detect antibody levels (or, conversely, anti-Neisserial antibodies can be used to detect antigen levels). Immunoassays based on well defined, recombinant antigens can be developed to replace invasive diagnostics methods. Antibodies to Neisserial proteins within biological samples, including for example, blood or serum samples, can be detected. Design of the immunoassays is subject to a great deal of variation, and a variety of these are known in the art. Protocols for the immunoassay may be based, for example, upon competition, or direct reaction, or sandwich type assays. Protocols may also, for example, use solid supports, or may be by immunoprecipitation. Most assays involve the use of labeled antibody or polypeptide; the labels may be, for example, fluorescent, chemiluminescent, radioactive, or dye molecules. Assays which amplify the signals from the probe are also known; examples of which are assays which utilize biotin and avidin, and enzyme-labeled and mediated immunoassays, such as ELISA assays.
• Kits suitable for immunodiagnosis and containing the appropriate labeled reagents are constructed by packaging the appropriate materials, including the compositions of the invention, in suitable containers, along with the remaining reagents and materials (for example, suitable buffers, salt solutions, etc.) required for the conduct of the assay, as well as suitable set of assay instructions.
Nucleic Acid Hybridisation
• “Hybridization” refers to the association of two nucleic acid sequences to one another by hydrogen bonding. Typically, one sequence will be fixed to a solid support and the other will be free in solution. Then, the two sequences will be placed in contact with one another under conditions that favor hydrogen bonding. Factors that affect this bonding include: the type and volume of solvent; reaction temperature; time of hybridization; agitation; agents to block the non-specific attachment of the liquid phase sequence to the solid support (Denhardt's reagent or BLOTTO); concentration of the sequences; use of compounds to increase the rate of association of sequences (dextran sulfate or polyethylene glycol); and the stringency of the washing conditions following hybridization. See Sambrook et al. [supra] Volume 2, chapter 9, pages 9.47 to 9.57.
• “Stringency” refers to conditions in a hybridization reaction that favor association of very similar sequences over sequences that differ. For example, the combination of temperature and salt concentration should be chosen that is approximately 120 to 200° C. below the calculated Tm of the hybrid under study. The temperature and salt conditions can often be determined empirically in preliminary experiments in which samples of genomic DNA immobilized on filters are hybridized to the sequence of interest and then washed under conditions of different stringencies. See Sambrook et al. at page 9.50.
• Variables to consider when performing, for example, a Southern blot are (1) the complexity of the DNA being blotted and (2) the homology between the probe and the sequences being detected. The total amount of the fragment(s) to be studied can vary a magnitude of 10, from 0.1 to 1 μg for a plasmid or phage digest to 10⁻⁹ to 10⁻⁸ g for a single copy gene in a highly complex eukaryotic genome. For lower complexity polynucleotides, substantially shorter blotting, hybridization, and exposure times, a smaller amount of starting polynucleotides, and lower specific activity of probes can be used. For example, a single-copy yeast gene can be detected with an exposure time of only 1 hour starting with 1 μg of yeast DNA, blotting for two hours, and hybridizing for 4-8 hours with a probe of 10⁸ cpm/μg. For a single-copy mammalian gene a conservative approach would start with 10 μg of DNA, blot overnight, and hybridize overnight in the presence of 10% dextran sulfate using a probe of greater than 10⁸ cpm/μg, resulting in an exposure time of ˜24 hours.
• Several factors can affect the melting temperature (Tm) of a DNA-DNA hybrid between the probe and the fragment of interest, and consequently, the appropriate conditions for hybridization and washing. In many cases the probe is not 100% homologous to the fragment. Other commonly encountered variables include the length and total G+C content of the hybridizing sequences and the ionic strength and formamide content of the hybridization buffer. The effects of all of these factors can be approximated by a single equation:
• 
  Tm=81+16.6(log₁₀Ci)+0.4[%(G+C)]−0.6(% formamide)−600/n−1.5(% mismatch).
• where Ci is the salt concentration (monovalent ions) and n is the length of the hybrid in base pairs (slightly modified from Meinkoth & Wahl (1984) Anal. Biochem. 138: 267-284).
• In designing a hybridization experiment, some factors affecting nucleic acid hybridization can be conveniently altered. The temperature of the hybridization and washes and the salt concentration during the washes are the simplest to adjust. As the temperature of the hybridization increases (ie. stringency), it becomes less likely for hybridization to occur between strands that are nonhomologous, and as a result, background decreases. If the radiolabeled probe is not completely homologous with the immobilized fragment (as is frequently the case in gene family and interspecies hybridization experiments), the hybridization temperature must be reduced, and background will increase. The temperature of the washes affects the intensity of the hybridizing band and the degree of background in a similar manner. The stringency of the washes is also increased with decreasing salt concentrations.
• In general, convenient hybridization temperatures in the presence of 50% formamide are 42° C. for a probe with is 95% to 100% homologous to the target fragment, 37° C. for 90% to 95% homology, and 32° C. for 85% to 90% homology. For lower homologies, formamide content should be lowered and temperature adjusted accordingly, using the equation above. If the homology between the probe and the target fragment are not known, the simplest approach is to start with both hybridization and wash conditions which are nonstringent. If non-specific bands or high background are observed after autoradiography, the filter can be washed at high stringency and reexposed. If the time required for exposure makes this approach impractical, several hybridization and/or washing stringencies should be tested in parallel.
Nucleic Acid Probe Assays
• Methods such as PCR, branched DNA probe assays, or blotting techniques utilizing nucleic acid probes according to the invention can determine the presence of cDNA or mRNA. A probe is said to “hybridize” with a sequence of the invention if it can form a duplex or double stranded complex, which is stable enough to be detected.
• The nucleic acid probes will hybridize to the Neisserial nucleotide sequences of the invention (including both sense and antisense strands). Though many different nucleotide sequences will encode the amino acid sequence, the native Neisserial sequence is preferred because it is the actual sequence present in cells. mRNA represents a coding sequence and so a probe should be complementary to the coding sequence; single-stranded cDNA is complementary to mRNA, and so a cDNA probe should be complementary to the non-coding sequence.
• The probe sequence need not be identical to the Neisserial sequence (or its complement)—some variation in the sequence and length can lead to increased assay sensitivity if the nucleic acid probe can form a duplex with target nucleotides, which can be detected. Also, the nucleic acid probe can include additional nucleotides to stabilize the formed duplex. Additional Neisserial sequence may also be helpful as a label to detect the formed duplex. For example, a non-complementary nucleotide sequence may be attached to the 5′ end of the probe, with the remainder of the probe sequence being complementary to a Neisserial sequence. Alternatively, non-complementary bases or longer sequences can be interspersed into the probe, provided that the probe sequence has sufficient complementarity with the a Neisserial sequence in order to hybridize therewith and thereby form a duplex which can be detected.
• The exact length and sequence of the probe will depend on the hybridization conditions, such as temperature, salt condition and the like. For example, for diagnostic applications, depending on the complexity of the analyte sequence, the nucleic acid probe typically contains at least 10-20 nucleotides, preferably 15-25, and more preferably at least 30 nucleotides, although it may be shorter than this. Short primers generally require cooler temperatures to form sufficiently stable hybrid complexes with the template.
• Probes may be produced by synthetic procedures, such as the triester method of Matteucci et al. [J. Am. Chem. Soc. (1981) 103:3185], or according to Urdea et al. [Proc. Natl. Acad. Sci. USA (1983) 80: 7461], or using commercially available automated oligonucleotide synthesizers.
• The chemical nature of the probe can be selected according to preference. For certain applications, DNA or RNA are appropriate. For other applications, modifications may be incorporated eg. backbone modifications, such as phosphorothioates or methylphosphonates, can be used to increase in vivo half-life, alter RNA affinity, increase nuclease resistance etc. [eg. see Agrawal & Iyer (1995) Curr Opin Biotechnol 6:12-19; Agrawal (1996) TIBTECH 14:376-387]; analogues such as peptide nucleic acids may also be used [eg. see Corey (1997) TIBTECH 15:224-229; Buchardt et al. (1993) TIBTECH 11:384-386].
• Alternatively, the polymerase chain reaction (PCR) is another well-known means for detecting small amounts of target nucleic acids. The assay is described in: Mullis et al. [Meth. Enzymol. (1987) 155: 335-350]; U.S. Pat. Nos. 4,683,195 and 4,683,202. Two “primer” nucleotides hybridize with the target nucleic acids and are used to prime the reaction. The primers can comprise sequence that does not hybridize to the sequence of the amplification target (or its complement) to aid with duplex stability or, for example, to incorporate a convenient restriction site. Typically, such sequence will flank the desired Neisserial sequence.
• A thermostable polymerase creates copies of target nucleic acids from the primers using the original target nucleic acids as a template. After a threshold amount of target nucleic acids are generated by the polymerase, they can be detected by more traditional methods, such as Southern blots. When using the Southern blot method, the labelled probe will hybridize to the Neisserial sequence (or its complement).
• Also, mRNA or cDNA can be detected by traditional blotting techniques described in Sambrook et al [supra]. mRNA, or cDNA generated from mRNA using a polymerase enzyme, can be purified and separated using gel electrophoresis. The nucleic acids on the gel are then blotted onto a solid support, such as nitrocellulose. The solid support is exposed to a labelled probe and then washed to remove any unhybridized probe. Next, the duplexes containing the labeled probe are detected. Typically, the probe is labelled with a radioactive moiety.
BRIEF DESCRIPTION OF THE DRAWINGS
• FIG. 1A-E: For ORF37-1, (A) shows the results of affinity purification of the GST-fusion protein, (B) shows the results of expression of the His-fusion in E. coli. Purified GST-fusion protein was used to immunise mice, whose sera were used for ELISA (positive result), (C) shows FACS analysis, and (D) shows a bactericidal assay (FIG. 1D), and (E) shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF37-1.
• FIG. 2A-B: For ORF5-1, (A) shows the results of affinity purification of the GST-fusion protein, and (B) shows the Western blot analysis of sera from mice immunized with purified GST-fusion protein.
• FIG. 3A-D: For ORF2-1, (A) shows the results of affinity purification of the GST-fusion protein, (B) shows the results of expression of the His-fusion in E. coli, (C) shows the Western blot analysis of sera from mice immunized with turified GST-fusion protein, (D) shows the ELISA (positive result), and (D) shows the FACS analysis.
• FIG. 4A-C: For ORF15-1, (A) shows the results of affinity purification of the GST-fusion protein, (B) shows the results of expression of the His-fusion in E. coli, and (C) shows the Western blot analysis of sera from mice immunized with purified GST-fusion protein.
• FIG. 5A-C: For ORF22-1, (A) shows the results of affinity purification of the GST-fusion protein, (B) shows the results of expression of the His-fusion in E. coli, and (C) shows the FACS analysis using sera from mice immunized with the purified GST-fusion protein.
• FIG. 6A-B: For ORF28-1, (A) shows the results of affinity purification of the GST-fusion protein, and (B) shows the results of expression of the His-fusion in E. coli.
• FIG. 7A-B: For ORF32-1, (A) shows the results of affinity purification of the His-fusion protein, and (B) shows the results of expression of the GST-fusion in E. coli.
• FIG. 8A-F: For ORF4-1, (A) shows the results of affinity purification of the His-fusion, (B) shows the results of affinity purification of the GST-fusion proteins, (C) shows the Western blot analysis of sera from mice immunized with the His-fusion protein, (D) shows the FACS analysis, (E) shows a bactericidal assay, and (F) shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF37-1.
• FIG. 9 shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF61-1.
• FIG. 10A-C: For ORF76-1, (A) shows the results of affinity purification of the His-fusion protein, (B) shows the Western blot analysis of sera from mice immunized with the purified His-fusion protein, and (C) shows the FACS analysis.
• FIG. 11 shows the results of affinity purification of the GST-ORF89-1 fusion protein.
• FIG. 12A-E: For ORF97-1, (A) show the results of affinity purification of the GST-fusion protein, (B) shows the results of affinity purification of the His-fusion protein, (C) shows the Western blot analysis of sera from mice immunized with purified GST-fusion protein, (D) shows the FACS analysis, and (E) shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF97-1.
• FIG. 13A-C: For ORF106-1, (A) shows the results of affinity purification of the His-fusion protein, (B) shows the results of expression of the GST-fusion in E. coli, (C) shows the FACS analysis of sera from mice immunized with the purified His-fusion protein.
• FIG. 14A-B: For ORF138-1, (A) shows the results of affinity purification of the GST-fusion protein, and (B) shows the FACS analysis of sera from mice immunized with the purified GST-fusion protein.
• FIG. 15A-C: For ORF23-1, (A) shows the results of affinity purification of the His-fusion protein, (B) shows the results of expression of the GST-fusion in E. coli, (C) shows the Western blot analysis of sera from mice immunized with the purified His-fusion protein.
• FIG. 16A-E: For ORF25-1, (A) shows the results of affinity purification of the GST-fusion protein, (B) shows the results of expression of the His-fusion in E. coli, (C) shows the Western blot analysis of sera from mice immunized with purified His-fusion protein, (D) shows the FACS analysis, and (E) shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF25-1.
• FIG. 17A-B: For ORF27-1, (A) shows the results of affinity purification of the GST-fusion protein, and (B) shows the results of expression of the His-fusion in E. coli.
• FIG. 18A-B: For ORF79-1, (A) shows the results of affinity purification of the His-fusion protein, and (B) shows the FACS analysis of sera from mice immunized with purified His-fusion protein.
• FIG. 19A-D: For ORF85a, (A) shows the results of affinity purification of the GST-fusion protein, (B) shows Western blot analysis of sera from mice immunized with purified GST-fusion protein, (C) shows FACS analysis, and (D) shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF85a.
• FIG. 20A-C: For ORF132-1, (A) shows the results of affinity purification of the His-fusion protein, (B) shows the results of expression of the GST-fusion in E. coli, (C) shows the FACS analysis of sera from mice immunized with purified His-fusion protein.
EXAMPLES
• The examples describe nucleic acid sequences which have been identified in N. meningitidis, along with their putative translation products, and also those of N. gonorrhoeae. Not all of the nucleic acid sequences are complete i.e. they encode less than the full-length wild-type protein.
• The examples are generally in the following format:
• • a nucleotide sequence which has been identified in N. meningitidis (strain B)
  • the putative translation product of this sequence
  • a computer analysis of the translation product based on database comparisons
  • corresponding gene and protein sequences identified in N. meningitidis (strain A) and in N. gonorrhoeae
  • a description of the characteristics of the proteins which indicates that they might be suitably antigenic
  • results of biochemical analysis (expression, purification, ELISA, FACS etc.)
  • The examples typically include details of sequence identity between species and strains. Proteins that are similar in sequence are generally similar in both structure and function, and the sequence identity often indicates a common evolutionary origin. Comparison with sequences of proteins of known function is widely used as a guide for the assignment of putative protein function to a new sequence and has proved particularly useful in whole-genome analyses.
• Sequence comparisons were performed at NCBI (http://www.ncbi.nlm.nih.gov) using the algorithms BLAST, BLAST2, BLASTn, BLASTp, tBLASTn, BLASTx, & tBLASTx [eg. see also Altschul et al. (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Research 25:2289-3402]. Searches were performed against the following databases: non-redundant GenBank+EMBL+DDBJ+PDB sequences and non-redundant GenBank CDS translations+PDB+SwissProt+SPupdate+PIR sequences.
• To compare Meningococcal and Gonococcal sequences, the tBLASTx algorithm was used, as implemented at http://www.genome.ou.edu/gono_blast.html. The FASTA algorithm was also used to compare the ORFs (from GCG Wisconsin Package, version 9.0).
• Dots within nucleotide sequences (eg. position 495 in SEQ ID 11) represent nucleotides which have been arbitrarily introduced in order to maintain a reading frame. In the same way, double-underlined nucleotides were removed. Lower case letters (eg. position 496 in SEQ ID 11) represent ambiguities which arose during alignment of independent sequencing reactions (some of the nucleotide sequences in the examples are derived from combining the results of two or more experiments).
• Nucleotide sequences were scanned in all six reading frames to predict the presence of hydrophobic domains using an algorithm based on the statistical studies of Esposti et al. [Critical evaluation of the hydropathy of membrane proteins (1990) Eur J Biochem 190:207-219]. These domains represent potential transmembrane regions or hydrophobic leader sequences.
• Open reading frames were predicted from fragmented nucleotide sequences using the program ORFFINDER (NCBI).
• Underlined amino acid sequences indicate possible transmembrane domains or leader sequences in the ORFs, as predicted by the PSORT algorithm (http://www.psort.nibb.ac.jp). Functional domains were also predicted using the MOTIFS program (GCG Wisconsin & PROSITE).
• Various tests can be used to assess the in vivo immunogencity of the proteins identified in the examples. For example, the proteins can be expressed recombinantly and used to screen patient sera by immunoblot. A positive reaction between the protein and patient serum indicates that the patient has previously mounted an immune response to the protein in question ie. the protein is an immunogen. This method can also be used to identify immunodominant proteins.
• The recombinant protein can also be conveniently used to prepare antibodies eg. in a mouse. These can be used for direct confirmation that a protein is located on the cell-surface. Labelled antibody (eg. fluorescent labelling for FACS) can be incubated with intact bacteria and the presence of label on the bacterial surface confirms the location of the protein.
• In particular, the following methods (A) to (S) were used to express, purify and biochemically characterise the proteins of the invention:
A) Chromosomal DNA Preparation
• N. meningitidis strain 2996 was grown to exponential phase in 100 ml of GC medium, harvested by centrifugation, and resuspended in 5 ml buffer (20% Sucrose, 50 mM Tris-HCl, 50 mM EDTA, pH8). After 10 minutes incubation on ice, the bacteria were lysed by adding 10 ml lysis solution (50 mM NaCl, 1% Na-Sarkosyl, 50 μg/ml Proteinase K), and the suspension was incubated at 37° C. for 2 hours. Two phenol extractions (equilibrated to pH 8) and one ChCl₃/isoamylalcohol (24:1) extraction were performed. DNA was precipitated by addition of 0.3M sodium acetate and 2 volumes ethanol, and was collected by centrifugation. The pellet was washed once with 70% ethanol and redissolved in 4 ml buffer (10 mM Tris-HCl, 1 mM EDTA, pH 8). The DNA concentration was measured by reading the OD at 260 nm.
B) Oligonucleotide Design
• Synthetic oligonucleotide primers were designed on the basis of the coding sequence of each ORF, using (a) the meningococcus B sequence when available, or (b) the gonococcus/meningococcus A sequence, adapted to the codon preference usage of meningococcus as necessary. Any predicted signal peptides were omitted, by deducing the 5′-end amplification primer sequence immediately downstream from the predicted leader sequence.
• For most ORFs, the 5′ primers included two restriction enzyme recognition sites (BamHI-NdeI, BamHI-NheI, or EcoRI-NheI, depending on the gene's own restriction pattern); the 3′ primers included a XhoI restriction site. This procedure was established in order to direct the cloning of each amplification product (corresponding to each ORF) into two different expression systems: pGEX-KG (using either BamHI-XhoI or EcoRI-XhoI), and pET21b+ (using either NdeI-XhoI or NheI-XhoI).

• 5′-end primer tail:	CGCGGATCCCATATG	(BamHI-NdeI)
  	CGCGGATCCGCTAGC	(BamHI-NheI)
  	CCGGAATTCTAGCTAGC	(EcoRI-NheI)
  3′-end primer tail:	CCCGCTCGAG	(XhoI)

• For ORFs 5, 15, 17, 19, 20, 22, 27, 28, 65 & 89, two different amplifications were performed to clone each ORF in the two expression systems. Two different 5′ primers were used for each ORF; the same 3′ XhoI primer was used as before:

• 5′-end primer tail:	GGAATTCCATATGGCCATGG	(NdeI)
  5′-end primer tail:	CGGGATCC	(BamHI)

• ORF 76 was cloned in the pTRC expression vector and expressed as an amino-terminus His-tag fusion. In this particular case, the predicted signal peptide was included in the final product. NheI-BamHI restriction sites were incorporated using primers:

• 5′-end primer tail:	GATCAGCTAGCCATATG	(NheI)
  3′-end primer tail:	CGGGATCC	(BamHI)

• As well as containing the restriction enzyme recognition sequences, the primers included nucleotides which hybridized to the sequence to be amplified. The number of hybridizing nucleotides depended on the melting temperature of the whole primer, and was determined for each primer using the formulae:
• 
  T _m=4(G+C)+2(A+T) (tail excluded)
• 
  T _m=64.9+0.41(% GC)−600/N (whole primer)
• The average melting temperature of the selected oligos were 65-70° C. for the whole oligo and 50-55° C. for the hybridising region alone.
• Table I (page 487) shows the forward and reverse primers used for each amplification. In certain cases, it will be noted that the sequence of the primer does not exactly match the sequence in the ORF. When initial amplifications were performed, the complete 5′ and/or 3′ sequence was not known for some meningococcal ORFs, although the corresponding sequences had been identified in gonococcus. For amplification, the gonococcal sequences could thus be used as the basis for primer design, altered to take account of codon preference. In particular, the following codons were changed: ATA→ATT; TCG→TCT; CAG→CAA; AAG→AAA; GAG→GAA; CGA→CGC; CGG→CGC; GGG→GGC. Italicised nucleotides in Table I indicate such a change. It will be appreciated that, once the complete sequence has been identified, this approach is generally no longer necessary.
• Oligos were synthesized by a Perkin Elmer 394 DNA/RNA Synthesizer, eluted from the columns in 2 ml NH₄OH, and deprotected by 5 hours incubation at 56° C. The oligos were precipitated by addition of 0.3M Na-Acetate and 2 volumes ethanol. The samples were then centrifuged and the pellets resuspended in either 100 μl or 1 ml of water. OD₂₆₀ was determined using a Perkin Elmer Lambda Bio spectophotometer and the concentration was determined and adjusted to 2-10 pmol/μl.
C) Amplification
• The standard PCR protocol was as follows: 50-200 ng of genomic DNA were used as a template in the presence of 20-40 μM of each oligo, 400-800 μM dNTPs solution, 1×PCR buffer (including 1.5 mM MgCl₂), 2.5 units TaqI DNA polymerase (using Perkin-Elmer AmpliTaQ, GIBCO Platinum, Pwo DNA polymerase, or Tahara Shuzo Taq polymerase).
• In some cases, PCR was optimsed by the addition of 10 μl DMSO or 50 μl 2M betaine.
• After a hot start (adding the polymerase during a preliminary 3 minute incubation of the whole mix at 95° C.), each sample underwent a double-step amplification: the first 5 cycles were performed using as the hybridization temperature the one of the oligos excluding the restriction enzymes tail, followed by 30 cycles performed according to the hybridization temperature of the whole length oligos. The cycles were followed by a final 10 minute extension step at 72° C.
• The standard cycles were as follows:

• 	Denaturation	Hybridisation	Elongation
  First 5 cycles	30 seconds	30 seconds	30-60 seconds
  	95° C.	50-55° C.	72° C.
  Last 30 cycles	30 seconds	30 seconds	30-60 seconds
  	95° C.	65-70° C.	72° C.

• The elongation time varied according to the length of the ORF to be amplified.
• The amplifications were performed using either a 9600 or a 2400 Perkin Elmer GeneAmp PCR System. To check the results, 1/10 of the amplification volume was loaded onto a 1-1.5% agarose gel and the size of each amplified fragment compared with a DNA molecular weight marker.
• The amplified DNA was either loaded directly on a 1% agarose gel or first precipitated with ethanol and resuspended in a suitable volume to be loaded on a 1% agarose gel. The DNA fragment corresponding to the right size band was then eluted and purified from gel, using the Qiagen Gel Extraction Kit, following the instructions of the manufacturer. The final volume of the DNA fragment was 30 μl or 50 μl of either water or 10 mM Tris, pH 8.5.
D) Digestion of PCR Fragments
• The purified DNA corresponding to the amplified fragment was split into 2 aliquots and double-digested with:
• • NdeI/XhoI or NheI/XhoI for cloning into pET-21b+ and further expression of the protein as a C-terminus His-tag fusion
  • BamHI/XhoI or EcoRI/XhoI for cloning into pGEX-KG and further expression of the protein as N-terminus GST fusion.
  • For ORF 76, NheI/BamHI for cloning into pTRC-H isA vector and further expression of the protein as N-terminus His-tag fusion.
  • EcoRI/PstI, EcoRI/SalI, SalI/PstI for cloning into pGex-His and further expression of the protein as N-terminus His-tag fusion
• Each purified DNA fragment was incubated (37° C. for 3 hours to overnight) with 20 units of each restriction enzyme (New England Biolabs) in a either 30 or 40 μl final volume in the presence of the appropriate buffer. The digestion product was then purified using the QIAquick PCR purification kit, following the manufacturer's instructions, and eluted in a final volume of 30 or 50 μl of either water or 10 mM Tris-HCl, pH 8.5. The final DNA concentration was determined by 1% agarose gel electrophoresis in the presence of titrated molecular weight marker.
• E) Digestion of the Cloning Vectors (pET22B, pGEX-KG, pTRC-His A, and pGex-His)
• 10 μg plasmid was double-digested with 50 units of each restriction enzyme in 200 μl reaction volume in the presence of appropriate buffer by overnight incubation at 37° C. After loading the whole digestion on a 1% agarose gel, the band corresponding to the digested vector was purified from the gel using the Qiagen QIAquick Gel Extraction Kit and the DNA was eluted in 50 μl of 10 mM Tris-HCl, pH 8.5. The DNA concentration was evaluated by measuring OD₂₆₀ of the sample, and adjusted to 50 μg/μl. 1 μl of plasmid was used for each cloning procedure.
• The vector pGEX-His is a modified pGEX-2T vector carrying a region encoding six histidine residues upstream to the thrombin cleavage site and containing the multiple cloning site of the vector pTRC99 (Pharmacia).
F) Cloning
• The fragments corresponding to each ORF, previously digested and purified, were ligated in both pET22b and pGEX-KG. In a final volume of 20 μl, a molar ratio of 3:1 fragment/vector was ligated using 0.5 μl of NEB T4 DNA ligase (400 units/μl), in the presence of the buffer supplied by the manufacturer. The reaction was incubated at room temperature for 3 hours. In some experiments, ligation was performed using the Boheringer “Rapid Ligation Kit”, following the manufacturer's instructions.
• In order to introduce the recombinant plasmid in a suitable strain, 100 μl E. coli DH5 competent cells were incubated with the ligase reaction solution for 40 minutes on ice, then at 37° C. for 3 minutes, then, after adding 800 μl LB broth, again at 37° C. for 20 minutes. The cells were then centrifuged at maximum speed in an Eppendorf microfuge and resuspended in approximately 200 μl of the supernatant. The suspension was then plated on LB ampicillin (100 mg/ml).
• The screening of the recombinant clones was performed by growing 5 randomly-chosen colonies overnight at 37° C. in either 2 ml (pGEX or pTC clones) or 5 ml (pET clones) LB broth+100 μg/ml ampicillin. The cells were then pelletted and the DNA extracted using the Qiagen QIAprep Spin Miniprep Kit, following the manufacturer's instructions, to a final volume of 30 μl. 5 μl of each individual miniprep (approximately 1 g) were digested with either NdeI/XhoI or BamHI/XhoI and the whole digestion loaded onto a 1-1.5% agarose gel (depending on the expected insert size), in parallel with the molecular weight marker (1 Kb DNA Ladder, GIBCO). The screening of the positive clones was made on the base of the correct insert size.
• For the cloning of ORFs 110, 111, 113, 115, 119, 122, 125 & 130, the double-digested PCR product was ligated into double-digested vector using EcoRI-PstI cloning sites or, for ORFs 115 & 127, EcoRI-SalI or, for ORF 122, SalI-PstI. After cloning, the recombinant plasmids were introduced in the E. coli host W3110. Individual clones were grown overnight at 37° C. in L-broth with 50 μl/ml ampicillin.
G) Expression
• Each ORF cloned into the expression vector was transformed into the strain suitable for expression of the recombinant protein product. 1 μl of each construct was used to transform 30 μl of E. coli BL21 (pGEX vector), E. coli TOP 10 (pTRC vector) or E. coli BL21-DE3 (pET vector), as described above. In the case of the pGEX-His vector, the same E. coli strain (W3110) was used for initial cloning and expression. Single recombinant colonies were inoculated into 2 ml LB+Amp (100 μg/ml), incubated at 37° C. overnight, then diluted 1:30 in 20 ml of LB+Amp (100 μg/ml) in 100 ml flasks, making sure that the OD₆₀₀ ranged between 0.1 and 0.15. The flasks were incubated at 30° C. into gyratory water bath shakers until OD indicated exponential growth suitable for induction of expression (0.4-0.8 OD for pET and pTRC vectors; 0.8-1 OD for pGEX and pGEX-His vectors). For the pET, pTRC and pGEX-His vectors, the protein expression was induced by addition of 1 mM IPTG, whereas in the case of pGEX system the final concentration of IPTG was 0.2 mM. After 3 hours incubation at 30° C., the final concentration of the sample was checked by OD. In order to check expression, 1 ml of each sample was removed, centrifuged in a microfuge, the pellet resuspended in PBS, and analysed by 12% SDS-PAGE with Coomassie Blue staining. The whole sample was centrifuged at 6000 g and the pellet resuspended in PBS for further use.
H) GST-Fusion Proteins Large-Scale Purification.
• A single colony was grown overnight at 37° C. on LB+Amp agar plate. The bacteria were inoculated into 20 ml of LB+Amp liquid colture in a water bath shaker and grown overnight. Bacteria were diluted 1:30 into 600 ml of fresh medium and allowed to grow at the optimal temperature (20-37° C.) to OD₅₅₀ 0.8-1. Protein expression was induced with 0.2 mM IPTG followed by three hours incubation. The culture was centrifuged at 8000 rpm at 4° C. The supernatant was discarded and the bacterial pellet was resuspended in 7.5 ml cold PBS. The cells were disrupted by sonication on ice for 30 sec at 40 W using a Branson sonifier B-15, frozen and thawed twice and centrifuged again. The supernatant was collected and mixed with 150 μl Glutatione-Sepharose 4B resin (Pharmacia) (previously washed with PBS) and incubated at room temperature for 30 minutes. The sample was centrifuged at 700 g for 5 minutes at 4° C. The resin was washed twice with 10 ml cold PBS for 10 minutes, resuspended in 1 ml cold PBS, and loaded on a disposable column. The resin was washed twice with 2 ml cold PBS until the flow-through reached OD₂₈₀ of 0.02-0.06. The GST-fusion protein was eluted by addition of 700 μl cold Glutathione elution buffer (10 mM reduced glutathione, 50 mM Tris-HCl) and fractions collected until the OD₂₈₀ was 0.1.21 μl of each fraction were loaded on a 12% SDS gel using either Biorad SDS-PAGE Molecular weight standard broad range (M1) (200, 116.25, 97.4, 66.2, 45, 31, 21.5, 14.4, 6.5 kDa) or Amersham Rainbow Marker (M2) (220, 66, 46, 30, 21.5, 14.3 kDa) as standards. As the MW of GST is 26 kDa, this value must be added to the MW of each GST-fusion protein.
I) His-Fusion Solubility Analysis (ORFs 111-129)
• To analyse the solubility of the His-fusion expression products, pellets of 3 ml cultures were resuspended in buffer M1 [500 μl PBS pH 7.2]. 25 μl lysozyme (10 mg/ml) was added and the bacteria were incubated for 15 min at 4° C. The pellets were sonicated for 30 sec at 40 W using a Branson sonifier B-15, frozen and thawed twice and then separated again into pellet and supernatant by a centrifugation step. The supernatant was collected and the pellet was resuspended in buffer M2 [8M urea, 0.5M NaCl, 20 mM imidazole and 0.1 M NaH₂ PO₄] and incubated for 3 to 4 hours at 4° C. After centrifugation, the supernatant was collected and the pellet was resuspended in buffer M3 [6M guanidinium-HCl, 0.5M NaCl, 20 mM imidazole and 0.1M NaH₂PO₄] overnight at 4° C. The supernatants from all steps were analysed by SDS-PAGE.
• The proteins expressed from ORFs 113, 119 and 120 were found to be soluble in PBS, whereas ORFs 111, 122, 126 and 129 need urea and ORFs 125 and 127 need guanidium-HCl for their solubilization.
J) His-Fusion Large-Scale Purification.
• A single colony was grown overnight at 37° C. on a LB+Amp agar plate. The bacteria were inoculated into 20 ml of LB+Amp liquid culture and incubated overnight in a water bath shaker. Bacteria were diluted 1:30 into 600 ml fresh medium and allowed to grow at the optimal temperature (20-37° C.) to OD₅₅₀ 0.6-0.8. Protein expression was induced by addition of 1 mM IPTG and the culture further incubated for three hours. The culture was centrifuged at 8000 rpm at 4° C., the supernatant was discarded and the bacterial pellet was resuspended in 7.5 ml of either (i) cold buffer A (300 mM NaCl, 50 mM phosphate buffer, 10 mM imidazole, pH 8) for soluble proteins or (ii) buffer B (urea 8M, 10 mM Tris-HCl, 100 mM phosphate buffer, pH 8.8) for insoluble proteins. The cells were disrupted by sonication on ice for 30 sec at 40 W using a Branson sonifier B-15, frozen and thawed two times and centrifuged again.
• For insoluble proteins, the supernatant was stored at −20° C., while the pellets were resuspended in 2 ml buffer C (6M guanidine hydrochloride, 100 mM phosphate buffer, 10 mM Tris-HCl, pH 7.5) and treated in a homogenizer for 10 cycles. The product was centrifuged at 13000 rpm for 40 minutes.
• Supernatants were collected and mixed with 150 μl Ni²⁺-resin (Pharmacia) (previously washed with either buffer A or buffer B, as appropriate) and incubated at room temperature with gentle agitation for 30 minutes. The sample was centrifuged at 700 g for 5 minutes at 4° C. The resin was washed twice with 10 ml buffer A or B for 10 minutes, resuspended in 1 ml buffer A or B and loaded on a disposable column. The resin was washed at either (i) 4° C. with 2 ml cold buffer A or (ii) room temperature with 2 ml buffer B, until the flow-through reached OD₂₈₀ of 0.02-0.06.
• The resin was washed with either (i) 2 ml cold 20 mM imidazole buffer (300 mM NaCl, 50 mM phosphate buffer, 20 mM imidazole, pH 8) or (ii) buffer D (urea 8M, 10 mM Tris-HCl, 100 mM phosphate buffer, pH 6.3) until the flow-through reached the OD₂₈₀ of 0.02-0.06. The His-fusion protein was eluted by addition of 700 μl of either (i) cold elution buffer A (300 mM NaCl, 50 mM phosphate buffer, 250 mM imidazole, pH 8) or (ii) elution buffer B (urea 8M, 10 mM Tris-HCl, 100 mM phosphate buffer, pH 4.5) and fractions collected until the OD₂₈₀ was 0.1. 21 μl of each fraction were loaded on a 12% SDS gel.
K) His-Fusion Proteins Renaturation
• 10% glycerol was added to the denatured proteins. The proteins were then diluted to 20 μg/ml using dialysis buffer I (10% glycerol, 0.5M arginine, 50 mM phosphate buffer, 5 mM reduced glutathione, 0.5 mM oxidised glutathione, 2M urea, pH 8.8) and dialysed against the same buffer at 4° C. for 12-14 hours. The protein was further dialysed against dialysis buffer II (10% glycerol, 0.5M arginine, 50 mM phosphate buffer, 5 mM reduced glutathione, 0.5 mM oxidised glutathione, pH 8.8) for 12-14 hours at 4° C. Protein concentration was evaluated using the formula:
• 
  Protein (mg/ml)=(1.55×OD₂₈₀)−(0.76×OD₂₆₀)
L) His-Fusion Large-Scale Purification (ORFs 111-129)
• 500 ml of bacterial cultures were induced and the fusion proteins were obtained soluble in buffer M1, M2 or M3 using the procedure described above. The crude extract of the bacteria was loaded onto a Ni-NTA superflow column (Quiagen) equilibrated with buffer M1, M2 or M3 depending on the solubilization buffer of the fusion proteins. Unbound material was eluted by washing the column with the same buffer. The specific protein was eluted with the corresponding buffer containing 500 mM imidazole and dialysed against the corresponding buffer without imidazole. After each run the columns were sanitized by washing with at least two column volumes of 0.5 M sodium hydroxide and reequilibrated before the next use.
M) Mice Immunisations
• 20 μg of each purified protein were used to immunise mice intraperitoneally. In the case of ORFs 2, 4, 15, 22, 27, 28, 37, 76, 89 and 97, Balb-C mice were immunised with Al(OH)₃ as adjuvant on days 1, 21 and 42, and immune response was monitored in samples taken on day 56. For ORFs 44, 106 and 132, CD1 mice were immunised using the same protocol. For ORFs 25 and 40, CD1 mice were immunised using Freund's adjuvant, rather than AL(OH)₃, and the same immunisation protocol was used, except that the immune response was measured on day 42, rather than 56. Similarly, for ORFs 23, 32, 38 and 79, CD1 mice were immunised with Freund's adjuvant, but the immune response was measured on day 49.
N) ELISA Assay (Sera Analysis)
• The acapsulated MenB M7 strain was plated on chocolate agar plates and incubated overnight at 37° C. Bacterial colonies were collected from the agar plates using a sterile dracon swab and inoculated into 7 ml of Mueller-Hinton Broth (Difco) containing 0.25% Glucose. Bacterial growth was monitored every 30 minutes by following OD₆₂₀. The bacteria were let to grow until the OD reached the value of 0.3-0.4. The culture was centrifuged for 10 minutes at 10000 rpm. The supernatant was discarded and bacteria were washed once with PBS, resuspended in PBS containing 0.025% formaldehyde, and incubated for 2 hours at room temperature and then overnight at 4° C. with stirring. 100 μl bacterial cells were added to each well of a 96 well Greiner plate and incubated overnight at 4° C. The wells were then washed three times with PBT washing buffer (0.1% Tween-20 in PBS). 200 μl of saturation buffer (2.7% Polyvinylpyrrolidone 10 in water) was added to each well and the plates incubated for 2 hours at 37° C. Wells were washed three times with PBT. 200 μl of diluted sera (Dilution buffer: 1% BSA, 0.1% Tween-20, 0.1% NaN₃ in PBS) were added to each well and the plates incubated for 90 minutes at 37° C. Wells were washed three times with PBT. 100 μl of HRP-conjugated rabbit anti-mouse (Dako) serum diluted 1:2000 in dilution buffer were added to each well and the plates were incubated for 90 minutes at 37° C. Wells were washed three times with PBT buffer. 100 μl of substrate buffer for HRP (25 ml of citrate buffer pH5, 10 mg of O-phenildiamine and 10 μl of H₂O) were added to each well and the plates were left at room temperature for 20 minutes. 100 μl H₂SO₄ was added to each well and OD₄₉₀ was followed. The ELISA was considered positive when OD₄₉₀ was 2.5 times the respective pre-immune sera.
O) FACScan Bacteria Binding Assay Procedure.
• The acapsulated MenB M7 strain was plated on chocolate agar plates and incubated overnight at 37° C. Bacterial colonies were collected from the agar plates using a sterile dracon swab and inoculated into 4 tubes containing 8 ml each Mueller-Hinton Broth (Difco) containing 0.25% glucose. Bacterial growth was monitored every 30 minutes by following OD₆₂₀. The bacteria were let to grow until the OD reached the value of 0.35-0.5. The culture was centrifuged for 10 minutes at 4000 rpm. The supernatant was discarded and the pellet was resuspended in blocking buffer (1% BSA, 0.4% NaN₃) and centrifuged for 5 minutes at 4000 rpm. Cells were resuspended in blocking buffer to reach OD₆₂₀ of 0.07. 100 μl bacterial cells were added to each well of a Costar 96 well plate. 100 μl of diluted (1:200) sera (in blocking buffer) were added to each well and plates incubated for 2 hours at 4° C. Cells were centrifuged for 5 minutes at 4000 rpm, the supernatant aspirated and cells washed by addition of 200 μl/well of blocking buffer in each well. 100 μl of R-Phicoerytrin conjugated F(ab)₂ goat anti-mouse, diluted 1:100, was added to each well and plates incubated for 1 hour at 4° C. Cells were spun down by centrifugation at 4000 rpm for 5 minutes and washed by addition of 200 μl/well of blocking buffer. The supernatant was aspirated and cells resuspended in 200 μl/well of PBS, 0.25% formaldehyde. Samples were transferred to FACScan tubes and read. The condition for FACScan setting were: FL1 on, FL2 and FL3 off; FSC-H threshold:92; FSC PMT Voltage: E 02; SSC PMT: 474; Amp. Gains 7.1; FL-2 PMT: 539; compensation values: 0.
P) OMV Preparations
• Bacteria were grown overnight on 5 GC plates, harvested with a loop and resuspended in 10 ml 20 mM Tris-HCl. Heat inactivation was performed at 56° C. for 30 minutes and the bacteria disrupted by sonication for 10 minutes on ice (50% duty cycle, 50% output). Unbroken cells were removed by centrifugation at 5000 g for 10 minutes and the total cell envelope fraction recovered by centrifugation at 50000 g at 4° C. for 75 minutes. To extract cytoplasmic membrane proteins from the crude outer membranes, the whole fraction was resuspended in 2% sarkosyl (Sigma) and incubated at room temperature for 20 minutes. The suspension was centrifuged at 10000 g for 10 minutes to remove aggregates, and the supernatant further ultracentrifuged at 50000 g for 75 minutes to pellet the outer membranes. The outer membranes were resuspended in 10 mM Tris-HCl, pH8 and the protein concentration measured by the Bio-Rad Protein assay, using BSA as a standard.
Q) Whole Extracts Preparation
• Bacteria were grown overnight on a GC plate, harvested with a loop and resuspended in 1 ml of 20 mM Tris-HCl. Heat inactivation was performed at 56° C. for 30 minutes.
R) Western Blotting
• Purified proteins (500 ng/lane), outer membrane vesicles (5 μg) and total cell extracts (25 μg) derived from MenB strain 2996 were loaded on 15% SDS-PAGE and transferred to a nitrocellulose membrane. The transfer was performed for 2 hours at 150 mA at 4° C., in transferring buffer (0.3% Tris base, 1.44% glycine, 20% methanol). The membrane was saturated by overnight incubation at 4° C. in saturation buffer (10% skimmed milk, 0.1% Triton X100 in PBS). The membrane was washed twice with washing buffer (3% skimmed milk, 0.1% Triton X100 in PBS) and incubated for 2 hours at 37° C. with mice sera diluted 1:200 in washing buffer. The membrane was washed twice and incubated for 90 minutes with a 1:2000 dilution of horseradish peroxidase labelled anti-mouse Ig. The membrane was washed twice with 0.1% Triton X100 in PBS and developed with the Opti-4CN Substrate Kit (Bio-Rad). The reaction was stopped by adding water.
S) Bactericidal Assay
• MC58 strain was grown overnight at 37° C. on chocolate agar plates. 5-7 colonies were collected and used to inoculate 7 ml Mueller-Hinton broth. The suspension was incubated at 37° C. on a nutator and let to grow until OD₆₂₀ was 0.5-0.8. The culture was aliquoted into sterile 1.5 ml Eppendorf tubes and centrifuged for 20 minutes at maximum speed in a microfuge. The pellet was washed once in Gey's buffer (Gibco) and resuspended in the same buffer to an OD₆₂₀ of 0.5, diluted 1:20000 in Gey's buffer and stored at 25° C.
• 50 μl of Gey's buffer/]% BSA was added to each well of a 96-well tissue culture plate. 25 μl of diluted mice sera (1:100 in Gey's buffer/0.2% BSA) were added to each well and the plate incubated at 4° C. 25 μl of the previously described bacterial suspension were added to each well. 25 μl of either heat-inactivated (56° C. waterbath for 30 minutes) or normal baby rabbit complement were added to each well. immediately after the addition of the baby rabbit complement, 22 μl of each sample/well were plated on Mueller-Hinton agar plates (time 0). The 96-well plate was incubated for 1 hour at 37° C. with rotation and then 22 μl of each sample/well were plated on Mueller-Hinton agar plates (time 1). After overnight incubation the colonies corresponding to time 0 and time 1 hour were counted.
• Table II (page 493) gives a summary of the cloning, expression and prurification results.
Example 1
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 1>:

• 1	ATGAAACAGA CAGTCAA.AT GCTTGCCGCC GCCCTGATTG
  	CCTTGGGCTT
  51	GAACCGACCG GTGTGGNCGG ATGACGTATC GGATTTTCGG
  	GAAAACTTGC
  101	A.GCGGCAGC ACAGGGAAAT GCAGCAGCCC AATACAATTT
  	GGGCGCAATG
  151	TAT.TACAAA GGACGCGCGT GCGCCGGGAT GATGCTGAAG
  	CGGTCAGATG
  201	GTATCGGCAG CCGGCGGAAC AGGGGTTAGC CCAAGCCCAA
  	TACAATTTGG
  251	GCTGGATGTA TGCCAACGGG CGCGC.GTGC GCCAAGATGA
  	TACCGAAGCG
  301	GTCAGATGGT ATCGGCAGGC GGCAGCGCAG GGGGTTGTCC
  	AAGCCCAATA
  351	CAATTTGGGC GTGATATATG CCGAAGGACG TGGAGTGCGC
  	CAAGACGATG
  401	TCGAAGCGGT CAGATGGTTT CGGCAGGCGG CAGCGCAGGG
  	GGTAGCCCAA
  451	GCCCAAAACA ATTTGGGCGT GATGTATGCC GAAAGANCGC
  	GCGTGCGCCA
  501	AGACCG...

• This corresponds to the amino acid sequence <SEQ ID 2; ORF37>:

• 1	MKQTVXMLAA ALIALGLNRP VWXDDVSDFR ENLXAAAQGN
  	AAAQYNLGAM
  51	YXQRTRVRRD DAEAVRWYRQ PAEQGLAQAQ YNLGWMYANG
  	RXVRQDDTEA
  101	VRWYRQAAAQ GVVQAQYNLG VIYAEGRGVR QDDVEAVRWF
  	RQAAAQGVAQ
  151	AQNNLGVMYA ERXRVRQD...

• Further work revealed the complete nucleotide sequence <SEQ ID 3>:

• 1	ATGAAACAGA CAGTCAAATG GCTTGCCGCC GCCCTGATTG
  	CCTTGGGCTT
  51	GAACCGAGCG GTGTGGGCGG ATGACGTATC GGATTTTCGG
  	GAAAACTTGC
  101	AGGCGGCAGC ACAGGGAAAT GCAGCAGCCC AATACAATTT
  	GGGCGCAATG
  151	TATTACAAAG GACGCGGCGT GCGCCGGGAT GATGCTGAAG
  	CGGTCAGATG
  201	GTATCGGCAG GCGGCGGAAC AGGGGTTAGC CCAAGCCCAA
  	TACAATTTGG
  251	GCTGGATGTA TGCCAACGGG CGCGGCGTGC GCCAAGATGA
  	TACCGAAGCG
  301	GTCAGATGGT ATCGGCAGGC GGCAGCGCAG GGGGTTGTCC
  	AAGCCCAATA
  351	CAATTTGGGC GTGATATATG CCGAAGGACG TGGAGTGCGC
  	CAAGACGATG
  401	TCGAAGCGGT CAGATGGTTT CGGCAGGCGG CAGCGCAGGG
  	GGTAGCCCAA
  451	GCCCAAAACA ATTTGGGCGT GATGTATGCC GAAAGACGCG
  	GCGTGCGCCA
  501	AGACCGCGCC CTTGCACAAG AATGGTTTGG CAAGGCTTGT
  	CAAAACGGAG
  551	ACCAAGACGG CTGCGACAAT GACCAACGCC TGAAGGCGGG
  	TTATTGA

• This corresponds to the amino acid sequence <SEQ ID 4; ORF37-1>:

• 1	MKQTVKWLAA ALIALGLNRA VWADDVSDFR ENLQAAAQGN
  	AAAQYNLGAM
  51	YYKGRGVRRD DAEAVRWYRQ AAEQGLAQAQ YNLGWMYANG
  	RGVRQDDTEA
  101	VRWYRQAAAQ GVVQAQYNLG VIYAEGRGVR QDDVEAVRWF
  	RQAAAQGVAQ
  151	AQNNLGVMYA ERRGVRQDRA LAQEWFGKAC QNGDQDGCDN
  	DQRLKAGY*

• Further work identified the corresponding gene in strain A of N. meningitidis <SEQ ID 5>:

• 1	ATGAAACAGA CAGTCAAATG GCTTGCCGCC GCCCTGATTG
  	CCTTGGGCTT
  51	GAACCAAGCG GTGTGGGCGG ATGACGTATC GGATTTTCGG
  	GAAAACTTGC
  101	AGGCGGCAGC ACAGGGAAAT GCAGCAGCCC AAAACAATTT
  	GGGCGTGATG
  151	TATGCCGAAA GACGCGGCGT GCGCCAAGAC CGCGCCCTTG
  	CACAAGAATG
  201	GCTTGGCAAG GCTTGTCAAA ACGGATACCA AGACAGCTGC
  	GACAATGACC
  251	AACGCCTGAA AGCGGGTTAT TGA

• This encodes a protein having amino acid sequence <SEQ ID 6; ORF37a>:

• 1	MKQTVKWLAA ALIALGLNQA VWADDVSDFR ENLQAAAQGN
  	AAAQNNLGVM
  51	YAERRGVRQD RALAQEWLGK ACQNGYQDSC DNDQRLKAGY
  	*

• The originally-identified partial strain B sequence (ORF37) shows 68.0% identity over a 75aa overlap with ORF37a:
• 
• Further work identified the corresponding gene in N. gonorrhoeae <SEQ ID 7>:

• 1	ATGAAACAGA CAGTCAAATG GCTTGCCGCC GCCCTGATTG
  	CCTTGGGCTT
  51	GAACCAAGCG GTGTGGGCGG GTGACGTATC GGATTTTCGG
  	GAAAACTTGC
  101	AGgcggcaGA ACaggGAAAT GCAGCAGCCC AATTCAATTT
  	GGGCGTGATG
  151	TATGAAAATG GACAAGGAGT TCGTCAAGAT TATGTACAGG
  	CAGTGCAGTG
  201	GTATCGCAAG GCTTCAGAAC AAGGGGATGC CCAAGCCCAA
  	TACAATTTGG
  251	GCTTGATGTA TTACGATGGA CGCGGCGTGC GCCAAGACCT
  	TGCGCTCGCT
  301	CAACAATGGC TTGGCAAGGC TTGTCAAAAC GGAGACCAAA
  	ACAGCTGCGA
  351	CAATGACCAA CGCCTGAAGG CGGGTTATTA A

• This encodes a protein having amino acid sequence <SEQ ID 8; ORF37ng>:

• 1	MKQTVKWLAA ALIALGLNQA VWAGDVSDFR ENLQAAEQGN
  	AAAQFNLGVM
  51	YENGQGVRQD YVQAVQWYRK ASEQGDAQAQ YNLGLMYYDG
  	RGVRQDLALA
  101	QQWLGKACQN GDQNSCDNDQ RLKAGY*

• The originally-identified partial strain B sequence (ORF37) shows 64.9% identity over a 111aa overlap with ORF37ng:
• 
• The complete strain B sequence (ORF37-1) and ORF37ng show 51.5% identity in 198 aa overlap:
• 
• Computer analysis of these amino acid sequences indicates a putative leader sequence, and it was predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF37-1 (11 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 1A shows the results of affinity purification of the GST-fusion protein, and FIG. 1B shows the results of expression of the His-fusion in E. coli. Purified GST-fusion protein was used to immunise mice, whose sera were used for ELISA (positive result), FACS analysis (FIG. 1C), and a bactericidal assay (FIG. 1D). These experiments confirm that ORF37-1 is a surface-exposed protein, and that it is a useful immunogen.
• FIG. 1E shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF37-1.
Example 2
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 9>:

• 	TTCGGCGA CATCGGCGGT TTGAAGGTCA ATGCCCCCGT
  	CAAATCCGCA GGCGTATTGG TCGGGCGCGT CGGCGCTATC
  	GGACTTGACC CGAAATCCTA TCAGGCGAGG GTGCGCCTCG
  	ATTTGGACGG CAAGTATCAG TTCAGCAGCG ACGTTTCCGC
  	GCAAATCCTG ACTTCsGGAC TTTTGGGCGA GCAGTACATC
  	GGGCTGCAGC AGGGCGGCGA CACGGAAAAC CTTGCTGCCG
  	GCGACACCAT CTCCGTAACC AGTTCTGCAA TGGTTCTGGA
  	AAACCTTATC GGCAAATTCA TGACGAGTTT TGCCGAGAAA
  	AATGCCGACG GCGGCAATGC GGAAAAAGCC GCCGAATAA

• This corresponds to the amino acid sequence <SEQ ID 10>:

• 1 FGDIGGLKVN APVKSAGVLV GRVGAIGLDP KSYQARVRLD LDGKYQFSSD
  51 VSAQILTSGL LGEQYIGLQQ GGDTENLAAG DTISVTSSAM VLENLIGKFM
  101 TSFAEKNADG GNAEKAAE*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Hypothetical H. influenzae Protein (ybrd.haein; Accession Number p45029)
• SEQ ID 9 and ybrd.haein show 48.4% aa identity in 122 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• SEQ ID 9 shows 99.2% identity over a 118aa overlap with a predicted ORF from N. gonorrhoeae:
• 
• The complete yrbd H. influenzae sequence has a leader sequence and it is expected that the full-length homologous N. meningitidis protein will also have one. This suggests that it is either a membrane protein, a secreted protein, or a surface protein and that the protein, or one of its epitopes, could be a useful antigen for vaccines or diagnostics.
Example 3
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 11>:

• 1	..ATTTTGATAT ACCTCATCCG CAAGAATCTA GGTTCGCCCG
  	TCTTCTTCTT
  51	TCAGGAACGC CCCGGAAAGG ACGGAAAACC TTTTAAAATG
  	GTCAAATTCC
  101	GTTCCATGCG CGACGGCTTG TATTCAGACG GCATTCCGCT
  	GCCCGACGGA
  151	GAACGCCTGA CACCGTTCGG CAAAAAACTG CGTGCCGcCA
  	GTwTGGACGA
  201	ACTGCCTGAA TTATGGAATA TCTTAAAAGG CGAGATGAGC
  	CTGGTCGGCC
  251	CCCGCCCGCT GCTGATGCAA TATCTGCCGC TGTACGACAA
  	CTTCCAAAAC
  301	CGCCGCCACG AAATGAAACC CGGCATTACC GGCTGGGCGC
  	AGGTCAACGG
  351	GCGCAACGCg CTTTCGTGGG ACGAAAAATT CGCCTGCGAT
  	GTTTGGTATA
  401	TCGACCACTT CAGCCTGTGC CTCGACATCA AAATCCTACT
  	GCTGACGGTT
  451	AAAAAAGTAT TAATCAAGGA AGGGATTTCC GCACAGGGCG
  	AACA.aCCAT
  501	GCCCCCTTTC ACAGGAAAAC GCAAACTCGC CGTCGTCGGT
  	GCGGGCGGAC
  551	ACGGAAAAGT CGTTGCCGAC CTTGCCGCCG CACTCGGCCG
  	GTACAGGGAA
  601	ATCGTTTTTC TGGACGACCG CGCACAAGGC AGCGTCAACG
  	GCTTTTCCGT
  651	CATCGGCACG ACGCTGCTGC TTGAAAACAG TTTATCGCCC
  	GAACAATACG
  701	ACGTCGCCGT CGCCGTCGGC AACAACCGCA TCCGCCGCCA
  	AATCGCCGAA
  751	AAAGCCGCCG CGCTCGGCTT CGCCCTGCCC GTACTGGTTC
  	ATCCGGACGC
  801	GACCGTCTCG CCTTCTGCAA CAGTCGGACA AGGCAGCGTC
  	GTTATGGCGA
  851	AAGCGGTCG..

• This corresponds to the amino acid sequence <SEQ ID 12; ORF3>:

• 1	. . . ILIYLIRKNL GSPVFFFQER PGKDGKPFKM VKFRSMRDGL YSDGIPLPDG
  51	ERLTPFGKKL RAASXDELPE LWNILKGEMS LVGPRPLLMQ YLPLYDNFQN
  101	RRHEMKPGIT GWAQVNGRNA LSWDEKFACD VWYIDHFSLC LDIKILLLTV
  151	KKVLIKEGIS AQGEXTMPPF TGKRKLAVVG AGGHGKVVAD LAAALGRYRE
  201	IVFLDDRAQG SVNGFSVIGT TLLLENSLSP EQYDVAVAVG NNRIRRQIAE
  251	KAAALGFALP VLVHPDATVS PSATVGQGSV VMAKAV . . .

• Further sequence analysis revealed the complete nucleotide sequence <SEQ ID 13>:

• 1	ATGAGTAAAT TCTTCAAACG CCTGTTTGAC ATTGTTGCCT
  	CCGCCTCGGG
  51	ACTGATTTTC CTCTCGCCAG TATTTTTGAT TTTGATATAC
  	CTCATCCGCA
  101	AGAATCTAGG TTCGCCCGTC TTCTTCTTTC AGGAACGCCC
  	CGGAAAGGAC
  151	GGAAAACCTT TTAAAATGGT CAAATTCCGT TCCATGCGCG
  	ACGCGCTTGA
  201	TTCAGACGGC ATTCCGCTGC CCGACGGAGA ACGCCTGACA
  	CCGTTCGGCA
  251	AAAAACTGCG TGCCGCCAGT TTGGACGAAC TGCCTGAATT
  	ATGGAATATC
  301	TTAAAAGGCG AGATGAGCCT GGTCGGCCCC CGCCCGCTGC
  	TGATGCAATA
  351	TCTGCCGCTG TACGACAACT TCCAAAACCG CCGCCACGAA
  	ATGAAACCCG
  401	GCATTACCGG CTGGGCGCAG GTCAACGGGC GCAACGCGCT
  	TTCGTGGGAC
  451	GAAAAATTCG CCTGCGATGT TTGGTATATC GACCACTTCA
  	GCCTGTGCCT
  501	CGACATCAAA ATCCTACTGC TGACGGTTAA AAAAGTATTA
  	ATCAAGGAAG
  551	GGATTTCCGC ACAGGGCGAA GCCACCATGC CCCCTTTCAC
  	AGGAAAACGC
  601	AAACTCGCCG TCGTCGGTGC GGGCGGACAC GGAAAAGTCG
  	TTGCCGACCT
  651	TGCCGCCGCA CTCGGCCGGT ACAGGGAAAT CGTTTTTCTG
  	GACGACCGCG
  701	CACAAGGCAG CGTCAACGGC TTTTCCGTCA TCGGCACGAC
  	GCTGCTGCTT
  751	GAAAACAGTT TATCGCCCGA ACAATACGAC GTCGCCGTCG
  	CCGTCGGCAA
  801	CAACCGCATC CGCCGCCAAA TCGCCGAAAA AGCCGCCGCG
  	CTCGGCTTCG
  851	CCCTGCCCGT TCTGGTTCAT CCGGACGCGA CCGTCTCGCC
  	TTCTGCAACA
  901	GTCGGACAAG GCAGCGTCGT TATGGCGAAA GCCGTCGTAC
  	AGGCAGGCAG
  951	CGTATTGAAA GACGGCGTGA TTGTGAACAC TGCCGCCACC
  	GTCGATCACG
  1001	ACTGCCTGCT TAACGCTTTC GTCCACATCA GCCCAGGCGC
  	GCACCTGTCG
  1051	GGCAACACGC ATATCGGCGA AGAAAGCTGG ATAGGCACGG
  	GCGCGTGCAG
  1101	CCGCCAGCAG ATCCGTATCG GCAGCCGCGC AACCATTGGA
  	GCGGGCGCAG
  1151	TCGTCGTACG CGACGTTTCA GACGGCATGA CCGTCGCGGG
  	CAATCCGGCA
  1201	AAGCCGCTGC CGCGCAAAAA CCCCGAGACC TCGACAGCAT
  	AA

• This corresponds to the amino acid sequence <SEQ ID 14; ORF3-1>:

• 1	MSKFFKRLFD IVASASGLIF LSPVFLILIY LIRKNLGSPV
  	FFFQERPGKD
  51	GKPFKMVKFR SMRDALDSDG IPLPDGERLT PFGKKLRAAS
  	LDELPELWNI
  101	LKGEMSLVGP RPLLMQYLPL YDNFQNRRHE MKPGITGWAQ
  	VNGRNALSWD
  151	EKFACDVWYI DHFSLCLDIK ILLLTVKKVL IKEGISAQGE
  	ATMPPFTGKR
  201	KLAVVGAGGH GKVVADLAAA LGRYREIVFL DDRAQGSVNG
  	FSVIGTTLLL
  251	ENSLSPEQYD VAVAVGNNRI RRQIAEKAAA LGFALPVLVH
  	PDATVSPSAT
  301	VGQGSVVMAK AVVQAGSVLK DGVIVNTAAT VDHDCLLNAF
  	VHISPGAHLS
  351	GNTHIGEESW IGTGACSRQQ IRIGSRATIG AGAVVVRDVS
  	DGMTVAGNPA
  401	KPLPRKNPET STA*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF3 shows 93.0% identity over a 286aa overlap with an ORF (ORF3a) from strain A of N. meningitidis:
• 
• The complete length ORF3a nucleotide sequence <SEQ ID 15> is:

• 1	ATGAGTAAAT TCTTCAAACG CCTGTTTGAC ATTGTTGCCT
  	CCGCCTCGGG
  51	ACTGATTTTC CTCTCGCCAG TATTTTTGAT TTTGATATAC
  	CTCATCCGCA
  101	AGAATCTGGG TTCGCCCGTC TTCTTCTTTC AGGAACGCCC
  	CGGAAAGGAC
  151	GGAAAACCTT TTAAAATGGT CAAATTCCGT TCCATGCACG
  	ACGCGCTTGA
  201	TTCAGACGGC ATTCTGCTGC CCGACGGAGA ACGCCTGACA
  	CCGTTCGGCA
  251	AAAAACTGCG TGCCGCCAGT TTGGACGAAC TGCCCGAACT
  	GTGGAACGTC
  301	CTCAAAGGCG ACATGAGCCT GGTCGGCCCC CGCCCGCTGC
  	TGATGCAATA
  351	TCTGCCGCTG TACGACAACT TCCAAAACCG CCGCCACGAA
  	ATGAAACCGG
  401	GCATTACCGG CTGGGCGCAG GTCAACGGGC GCAACGCGCT
  	TTCGTGGGAC
  451	GAACGCTTCG CATGCGACAT CTGGTATATC GACCACTTCA
  	GCCTGTGCCT
  501	CGAGATCAAA ATCCTACTGC TGACGGTTAA AAAAGTATTA
  	ATCAAAGAAG
  551	GGATTTCCGC ACAGGGCGAA GCCACCATGC CCCCTTTCAC
  	AGGAAAACGC
  601	AAACTTGCCG TCGTCGGTGC GGGCGGACAC GGCAAAGTCG
  	TTGCCGAGCT
  651	TGCCGCCGCA CTCGGCACAT ACGGCGAAAT CGTTTTTCTG
  	GACGACCGCG
  701	TCCAAGGCAG CGTCAACGGC TTCCCCGTCA TCGGCACGAC
  	GCTGCTGCTT
  751	GAAAACAGTT TATCGCCCGA ACAATTCGAC ATCGCCGTCG
  	CCGTCGGCAA
  801	CAACCGCATC CGCCGCCAAA TCGCCGAAAA AGCCGCCGCG
  	CTCGGCTTCG
  851	CCCTGCCCGT CCTGATTCAT CCGGACTCGA CCGTCTCGCC
  	TTCTGCAACA
  901	GTCGGACAAG GCGGCGTCGT TATGGCGAAA GCCGTCGTAC
  	AGGCTGACAG
  951	CGTATTGAAA GACGGCGTAA TTGTGAACAC TGCCGCCACC
  	GTCGATCACG
  1001	ATTGCCTGCT TGATGCTTTC GTCCACATCA GCCCGGGCGC
  	GCACCTGTCG
  1051	GGCAACACGC GTATCGGCGA AGAAAGCTGG ATAGGCACAG
  	GCGCGTGCAG
  1101	CCGCCAGCAG ATCCGTATCG GCAGCCGCGC AACCATTGGA
  	GCGGGCGCAG
  1151	TCGTCGTGCG CGACGTTTCA GACGGCATGA CCGTCGCGGG
  	CAACCCGGCA
  1201	AAACCATTGG CAGGCAAAAA TACCGAGACC CTGCGGTCGT
  	AA

• This is predicted to encode a protein having amino acid sequence <SEQ ID 16>:

• 1	MSKFFKRLFD IVASASGLIF LSPVFLILIY LIRKNLGSPV
  	FFFQERPGKD
  51	GKPFKMVKFR SMHDALDSDG ILLPDGERLT PFGKKLRAAS
  	LDELPELWNV
  101	LKGDMSLVGP RPLLMQYLPL YDNFQNRRHE MKPGITGWAQ
  	VNGRNALSWD
  151	ERFACDIWYI DHFSLCLDIK ILLLTVKKVL IKEGISAQGE
  	ATMPPFTGKR
  201	KLAVVGAGGH GKVVAELAAA LGTYGEIVFL DDRVQGSVNG
  	FPVIGTTLLL
  251	ENSLSPEQFD IAVAVGNNRI RRQIAEKAAA LGFALPVLIH
  	PDSTVSPSAT
  301	VGQGGVVMAK AVVQADSVLK DGVIVNTAAT VDHDCLLDAF
  	VHISPGAHLS
  351	GNTRIGEESW IGTGACSRQQ IRIGSRATIG AGAVVVRDVS
  	DGMTVAGNPA
  401	KPLAGKNTET LRS*

• Two transmembrane domains are underlined.
• ORF3-1 shows 94.6% identity in 410 aa overlap with ORF3a:
• 
• Homology with Hypothetical Protein Encoded by yvfc Gene (Accession Z71928) of B. subtilis
• ORF3 and YVFC proteins show 55% aa identity in 170 aa overlap (BLASTp):

• ORF3	3	IYLIRKNLGSPVFFFQERPGKDGKPFKMVKFRSMRDGLYSDGIPLPDGERLTPFGKKLRA	62
  		I ++R  +GSPVFF Q RPG  GKPF + KFR+M D   S G  LPD  RLT  G+ +R
  yvfc	27	IAVVRLKIGSPVFFKQVRPGLHGKPFTLYKFRTMTDERDSKGNLLPDEVRLTKTGRLIRK	86
  ORF3	63	ASXDELPELWNILKGEMSLVGPRPLLMQYLPLYDNFQNRRHEMKPGITGWAQVNGRNALS	122
  		S DELP+L N+LKG++SLVGPRPLLM YLPLY   Q RRHE+KPGITGWAQ+NGRNA+S
  yvfc	87	LSIDELPQLLNVLKGDLSLVGPRPLLMDYLPLYTEKQARRHEVKPGITGWAQINGRNAIS	146
  ORF3	123	WDEKFACDVWYIDHFSLCLDXXXXXXXXXXXXXXEGISAQGEXTMPPFTG	172
  		W++KF  DVWY+D++S  LD              EGI      T   FTG
  yvfc	147	WEKKFELDVWYVDNWSFFLDLKILCLTVRKVLVSEGIQQTNHVTAERFTG	196

  
  Homology with a Predicted ORF from N. gonorrhoeae
• ORF3 shows 86.3% identity over a 286aa overlap with a predicted ORF (ORF3.ng) from N. gonorrhoeae:
• 
• The complete length ORF3ng nucleotide sequence <SEQ ID 17> is:

• 1	ATGAGTAAAG CCGTCAAACG CCTGTTCGAC ATCATCGCAT
  	CCGCATCGGG
  51	GCTGATTGTC CTGTCGCCCG TGTTTTTGGT TTTAATATAC
  	CTCATCCGCA
  101	AAAACTTAGG TTCGCCCGTC TTCTTCattC GGGAACGCCc
  	cgGAAAGGAc
  151	ggaaaacCTT TTAAAATGGT CAAATTCCGT TCCAtgcgcg
  	acgcgcttGA
  201	TTCAGACGGC ATTCCGCTGC CCGATAGCGA ACGCCTGACC
  	GATTTCGGCA
  251	AAAAATTACG CGCCACCAGT TTGGACGAAC TTCCTGAATT
  	ATGGAATGTC
  301	CTCAAAGGCG AGATGAGCCT GGTCGGCCCC CGCCCGCTTT
  	TGATGCAGTA
  351	TCTGCCGCTT TACAACAAAT TTCAAAACCG CCGCCACGAA
  	ATGAAACCGG
  401	GCATTACCGG CTGGGCGCAG GTCAACGGGC GCAACGCGCT
  	TTCGTGGGAC
  451	GAAAAGTTCT CCTGCGATGT TTGGTACACC GACAATTTCA
  	GCTTTTGGCT
  501	GGATATGAAA ATCCTGTTTC TGACAGTCAA AAAAGTCTTG
  	ATTAAAGAAG
  551	GCATTTCGGC GCAAGGGGAA GCCACCATGC CCCCTTTCGC
  	GGGGAATCGC
  601	AAACTCGCCG TTATCGGCGC GGGCGGACAC GGCAAAGTCG
  	TTGCCGAGCT
  651	TGCCGCCGCA CTCGGCACAT ACGGCGAAAT CGTTTTTCTG
  	GACGACCGCA
  701	CCCAAGGCAG CGTCAACGGC TTCCCCGTCA TCGGCACGAC
  	GCTGCTGCTT
  751	GAAAACAGTT TATCGCCCGA ACAATTCGAC ATCACCGTCG
  	CCGTCGGCAA
  801	CAACCGCATC CGCCGCCAAA TCACCGAAAA CGCCGCCGCG
  	CTCGGCTTCA
  851	AACTGCCCGT TCTGATTCAT CCCGACGCGA CCGTCTCGCC
  	TTCTGCAATA
  901	ATCGGACAAG GCAGCGTCGT AATGGCGAAA GCCGTCGTAC
  	AGGCCGGCAG
  951	CGTATTGAAA GACGGCGTGA TTGTGAACAC TGCCGCCACC
  	GTCGATCACG
  1001	ACTGCCTGCT TGACGCTTTC GtccaCATCA GCCCGGGCGC
  	GCACCTGTCG
  1051	GGCAACACGC GTATCGGCGA AGAAAGCCGG ATAGGCACGG
  	GCGCGTGCAG
  1101	CCGCCAGCAG ACAACCGTCG GCAGCGGGGT TACCgccgGT
  	GCAGGGgcGG
  1151	TTATCGTATG CGACATCCCG GACGGCATGA CCGTCGCGGG
  	CAACCCGGCA
  1201	AAGCCCCTTA CGGGCAAAAA CCCCAAGACC GGGACGGCAT
  	AA

• This encodes a protein having amino acid sequence <SEQ ID 18>:

• 1	MSKAVKRLFD IIASASGLIV LSPVFLVLIY LIRKNLGSPV
  	FFIRERPGKD
  51	GKPFKMVKFR SMRDALDSDG IPLPDSERLT DFGKKLRATS
  	LDELPELWNV
  101	LKGEMSLVGP RPLLMQYLPL YNKFQNRRHE MKPGITGWAQ
  	VNGRNALSWD
  151	EKFSCDVWYT DNFSFWLDMK ILFLTVKKVL IKEGISAQGE
  	ATMPPFAGNR
  201	KLAVIGAGGH GKVVAELAAA LGTYGEIVFL DDRTQGSVNG
  	FPVIGTTLLL
  251	ENSLSPEQFD ITVAVGNNRI RRQITENAAA LGFKLPVLIH
  	PDATVSPSAI
  301	IGQGSVVMAK AVVQAGSVLK DGVIVNTAAT VDHDCLLDAF
  	VHISPGAHLS
  351	GNTRIGEESR IGTGACSRQQ TTVGSGVTAG AGAVIVCDIP
  	DGMTVAGNPA
  401	KPLTGKNPKT GTA*

• This protein shows 86.9% identity in 413 aa overlap with ORF3-1:
• 
• In addition, ORF3ng shows significant homology with a hypothetical protein from B. subtilis:

• gnl|PID|e238668 (Z71928) hypothetical protein [Bacillus subtilis]
  >gi|1945702|gnl|PID|e313004 (Z94043) hypothetical protein
  [Bacillus subtilis]
  >gi|2635938|gnl|PID|e1186113 (Z99121) similar to capsular polysaccharide
  biosynthesis [Bacillus subtilis] Length = 202
  Score = 235 bits (594), Expect = 3e−61
  Identities = 114/195 (58%), Positives = 142/195 (72%)

  Query:	5	VKRLFDIIASASGLIVLSPVFLVLIYLIRKNLGSPVFFIRERPGKDGKPFKMVKFRSMRD	64
  		+KRLFD+ A+   L  S + L  I ++R  +GSPVFF + RPG  GKPF + KFR+M D
  Sbjct:	3	LKRLFDLTAAIFLLCCTSVIILFTIAVVRLKIGSPVFFKQVRPGLHGKPFTLYKFRTMTD	62
  Query:	65	ALDSDGIPLPDSERLTDFGKKLRATSLDELPELWNVLKGEMSLVGPRPLLMQYLPLYNKF	124
  		DS G  LPD  RLT  G+ +R  S+DELP+L NVLKG++SLVGPRPLLM YLPLY +
  Sbjct:	63	ERDSKGNLLPDEVRLTKTGRLIRKLSIDELPQLLNVLKGDLSLVGPRPLLMDYLPLYTEK	122
  Query:	125	QNRRHEMKPGITGWAQVNGRNALSWDEKFSCDVWYTDNFSFWLDMKILFLTVKKVLIKEG	184
  		Q RRHE+KPGITGWAQ+NGRNA+SW++KF  DVWY DN+SF+LD+KIL LTV+KVL+ EG
  Sbjct:	123	QARRHEVKPGITGWAQINGRNAISWEKKFELDVWYVDNWSFFLDLKILCLTVRKVLVSEG	182
  Query:	185	ISAQGEATMPPFAGN	199
  		I      T   F G+
  Sbjct:	183	IQQTNHVTAERFTGS	197

• The hypothetical product of yvfc gene shows similarity to EXOY of R. meliloti, an exopolysaccharide production protein. Based on this and on the two predicted transmembrane regions in the homologous N. gonorrhoeae sequence, it is predicted that these proteins, or their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 4
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 19>:

• 1	..AACCATATGG CGATTGTCAT CGACGAATAC GGCGGCACAT
  	CCGGCTTGGT
  51	CACCTTTGAA GACATCATCG AGCAAATCGT CGGCGAAATC
  	GAAGACGAGT
  101	TTGACGAAGA CGATAGCGCC GACAATATCC ATGCCGTTTC
  	TTCAGACACG
  151	TGGCGCATCC ATGCAGCTAC CGAAATCGAA GACATCAACA
  	CCTTCTTCGG
  201	CACGGAATAC AGCATCGAAG AAGCCGACAC CATT.GGCGG
  	CCTGGTCATT
  251	CAAGAGTTGG GACATCTGCC CGTGCGCGGC GAAAAAGTCC
  	TTATCGGCGG
  301	TTTGCAGTTC ACCGTCGCAC GCGCCGACAA CCGCCGCCTG
  	CATACGCTGA
  351	TGGCGACCCG CGTGAAGTAA GC........ .....ACCGC
  	CGTTTCTGCA
  401	CAGTTTAG

• This corresponds to amino acid sequence <SEQ ID 20; ORF5>:

• 1	..NHMAIVIDEY GGTSGLVTFE DIIEQIVGEI EDEFDEDDSA
  	DNIHAVSSDT
  51	WRIHAATEIE DINTFFGTEY SIEEADTIXR PGHSRVGTSA
  	RARRKSPYRR
  101	FAVHRRTRRQ PPPAYADGDP REVS....XR RFCTV*

• Further sequence analysis revealed the complete DNA sequence to be <SEQ ID 21>:

• 1	ATGGACGGCG CACAACCGAA AACGAATTTT TTTGAACGCC
  	TGATTGCCCG
  51	ACTCGCCCGC GAACCCGATT CCGCCGAAGA CGTATTAAAC
  	CTGCTTCGGC
  101	AGGCGCACGA GCAGGAAGTT TTTGATGCGG ATACGCTTTT
  	AAGATTGGAA
  151	AAAGTCCTCG ATTTTTCCGA TTTGGAAGTG CGCGACGCGA
  	TGATTACGCG
  201	CAGCCGTATG AACGTTTTAA AAGAAAACGA CAGCATCGAG
  	CGCATCACCG
  251	CCTACGTTAT CGATACCGCC CATTCGCGCT TCCCCGTCAT
  	CGGCGAAGAC
  301	AAAGACGAAG TTTTGGGCAT TTTGCACGCC AAAGACCTGC
  	TCAAATATAT
  351	GTTTAACCCC GAGCAGTTCC ACCTCAAATC CATTCTCCGC
  	CCCGCCGTCT
  401	TCGTCCCCGA AGGCAAATCG CTGACCGCCC TTTTAAAAGA
  	GTTCCGCGAA
  451	CAGCGCAACC ATATGGCGAT TGTCATCGAC GAATACGGCG
  	GCACATCCGG
  501	CTTGGTCACC TTTGAAGACA TCATCGAGCA AATCGTCGGC
  	GAAATCGAAG
  551	ACGAGTTTGA CGAAGACGAT AGCGCCGACA ATATCCATGC
  	CGTTTCTTCC
  601	GAACGCTGGC GCATCCATGC AGCTACCGAA ATCGAAGACA
  	TCAACACCTT
  651	CTTCGGCACG GAATACAGCA GCGAAGAAGC CGACACCATT
  	CGGCCTGGTC
  701	ATTCAAGAGT TGGGACATCT GCCCGTGCGC GGCGAAAAAG
  	TCCTTATCGG
  751	CGGTTTGCAG TTCACCGTCG CACGCGCCGA CAACCGCCGC
  	CTGCATACGC
  801	TGATGGCGAC CCGCGTGAAG TAAGCACCGC CGTTTCTGCA
  	CAGTTTAGGA
  851	TGACGGTACG GGCGTTTTCT GTTTCAATCC GCCCCATCCG
  	CCAAACATAA

• This corresponds to amino acid sequence <SEQ ID 22; ORF5-1>:

• 1	MDGAQPKTNF FERLIARLAR EPDSAEDVLN LLRQAHEQEV
  	FDADTLLRLE
  51	KVLDFSDLEV RDAMITRSRM NVLKENDSIE RITAYVIDTA
  	HSRFPVIGED
  101	KDEVLGILHA KDLLKYMFNP EQFHLKSILR PAVFVPEGKS
  	LTALLKEFRE
  151	QRNHMAIVID EYGGTSGLVT FEDIIEQIVG EIEDEFDEDD
  	SADNIHAVSS
  201	ERWRIHAATE IEDINTFFGT EYSSEEADTI RPGHSRVGTS
  	ARARRKSPYR
  251	RFAVHRRTRR QPPPAYADGD PREVSTAVSA QFRMTVRAFS
  	VSIRPIRQT*

• Further work identified the corresponding gene in strain A of N. meningitidis <SEQ ID 23>:

• 1	ATGGACGGCG CACAACCGAA AACAAATTTT TTNNAACGCC
  	TGATTGCCCG
  51	ACTCGCCCGC GAACCCGATT CCGCCGAAGA CGTATTGACC
  	CTGTTGCGCC
  101	AAGCGCACGA ACAGGAAGTA TTTGATGCGG ATACGCTTTT
  	AAGATTGGAA
  151	AAAGTCCTCG ATTTTTCTGA TTTGGAAGTG CGCGACGCGA
  	TGATTACGCG
  201	CAGCCGTATG AACGTTTTAA AAGAAAACGA CAGCATCGAA
  	CGCATCACCG
  251	CCTACGTTAT CGATACCGCC CATTCGCGCT TCCCCGTCAT
  	CGGTGAAGAC
  301	AAAGACGAAG TTTTGGGTAT TTTGCACGCC AAAGACCTGC
  	TCAAATATAT
  351	GTTCAACCCC GAGCAGTTCC ACCTCAAATC GATATTGCGC
  	CCTGCCGTCT
  401	TCGTCCCCGA AGGCAAATCG CTGACCGCCC TTTTAAAAGA
  	GTTCCGCGAA
  451	CAGCGCAACC ATATGGCAAT CGTCATCGAC GAATACGGCG
  	GCACGTCGGG
  501	TTTGGTAACT TTTGAAGACA TCATCGAGCA AATCGTCGGC
  	GACATCGAAG
  551	ATGAGTTTGA CGAAGACGAA AGCGCGGACA ACATCCACGC
  	CGTTTCCGCC
  601	GAACGCTGGC GCATCCACGC GGCTACCGAA ATCGAAGACA
  	TCAACGCCTT
  651	TTTCGGCACG GAATACAGCA GCGAAGAAGC CGACACCATC
  	GGCGGCCNTG
  701	GTCATTCAGG AATTGGNACA CCTGCCCGTG CGCGGCGAAA
  	AAGTCNTTAT
  751	CGGCGNNTTG CANTTCACNG TCGCCNGCGC NGACAACCGC
  	CGCCTGCATA
  801	CGCTGATGGC GACCCGCGTG AAGTAAGCTC CGCCGTTTCT
  	GTACAGTTTA
  851	GGATGACGGT ACGGGCGTTT TCTGTTTCAA TCCGCCCCAT
  	CCGCCANACA
  901	TAA

• This encodes a protein having amino acid sequence <SEQ ID 24; ORF5a>:

• 1	MDGAQPKTNF XXRLIARLAR EPDSAEDVLT LLRQAHEQEV
  	FDADTLLRLE
  51	KVLDFSDLEV RDAMITRSRM NVLKENDSIE RITAYVIDTA
  	HSRFPVIGED
  101	KDEVLGILHA KDLLKYMFNP EQFHLKSILR PAVFVPEGKS
  	LTALLKEFRE
  151	QRNHMAIVID EYGGTSGLVT FEDIIEQIVG DIEDEFDEDE
  	SADNIHAVSA
  201	ERWRIHAATE IEDINAFFGT EYSSEEADTI GGXGHSGIGT
  	PARARRKSXY
  251	RRXAXHXRXR XQPPPAYADG DPREVSSAVS VQFRMTVRAF
  	SVSIRPIRXT
  301	*

• The originally-identified partial strain B sequence (ORF5) shows 54.7% identity over a 124aa overlap with ORF5a:
• 
• The complete strain B sequence (ORF5-1) and ORF5a show 92.7% identity in 300 aa overlap:
• 
• Further work identified the a partial DNA sequence in N. gonorrhoeae <SEQ ID 25> which encodes a protein having amino acid sequence <SEQ ID 26; ORF5ng>:

• 1	MDGAQPKTNF FERLIARLAR EPDSAEDVLN LLRQAHEQEV
  	FDADTLTRLE
  51	KVLDFAELEV RDAMITRSRM NVLKENDSIE RITAYVIDTA
  	HSRFPVIGED
  101	KDEVLGILHA KDLLKYMFNP EQFHLKSVLR PAVFVPEGKS
  	LTALLKEFRE
  151	QRNHMAIVID EYGGTSGLVT FEDIIEQIVG DIEDEFDEDE
  	SADDIHSVSA
  201	ERWRIHAATE IEDINAFFGT EYGSEEADTI RRLGHSGIGT
  	PARARRKSPY
  251	RRFAVHRRPR RQPPPAHADG DPREVSRACP HRRFCTV*

• Further analysis revealed the complete gonococcal nucleotide sequence <SEQ ID 27> to be:

• 1	ATGGACGGCG CACAACCGAA AACAAATTTT TTTGAACGCC
  	TGATTGCCCG
  51	ACTCGCCCGC GAACCCGATT CCGCCGAAGA CGTATTAAAC
  	CTGCTTCGGC
  101	AGGCGCACGA ACAGGAAGTT TTTGATGCCG ACACACTGAC
  	CCGGCTGGAA
  151	AAAGTATTGG ACTTTGCCGA GCTGGAAGTG CGCGATGCGA
  	TGATTACGCG
  201	CAGCCGCATG AACGTATTGA AAGAAAACGA CAGCATCGAA
  	CGCATCACCG
  251	CCTACGTCAT CGATACCGCC CATTCGCGCT TCCCCGTCAT
  	CGGCGAAGAC
  301	AAAGACGAAG TTTTGGGCAT TTTGCACGCC AAAGACCTGC
  	TCAAATATAT
  351	GTTCAACCCC GAGCAGTTCC ACCTGAAATC CGTCTTGCGC
  	CCTGCCGTTT
  401	TCGTGCCCGA AGGCAAATCT TTGACCGCCC TTTTAAAAGA
  	GTTCCGCGAA
  451	CAGCGCAACC ATATGGCAAT CGTCATCGAC GAATACGGCG
  	GCACGTCGGG
  501	TTTGGTCACC TTTGAAGACA TCATCGAGCA AATCGTCGGT
  	GACATCGAAG
  551	ACGAGTTTGA CGAAGACGAA AGCGccgacg acatCCACTC
  	cgTTTccgCC
  601	GAACGCTGGC GCATCCacgc ggctaCCGAA ATCGAAGaca
  	TCAACGCCTT
  651	TTTCGGTACG GAatacggca gcgaagaagc cgacaccatc
  	cggcggctTG
  701	GTCATTCAGG AATTGGGACA CCTGCCCGTG CGCGGCGAAA
  	AAGTCCTTAt
  751	cggcgGTTTG Cagttcaccg tCGCCCGCGC CGACAACCGC
  	CGCCTGCACA
  801	CGCTGATGGC GACCCGCGTG AAGTAAGCAG AGCCTGCCcg
  	AccgccgttT
  851	CTGCacAGTT TAGGatgACG gtaCGGTCGT TTTCTGTTTC
  	AATCCGCCCC
  901	ATCCGCCAAA CATAA

• This encodes a protein having amino acid sequence <SEQ ID 28; ORF5ng-1>:

• 1	MDGAQPKTNF FERLIARLAR EPDSAEDVLN LLRQAHEQEV
  	FDADTLTRLE
  51	KVLDFAELEV RDAMITRSRM NVLKENDSIE RITAYVIDTA
  	HSRFPVIGED
  101	KDEVLGILHA KDLLKYMFNP EQFHLKSVLR PAVFVPEGKS
  	LTALLKEFRE
  151	QRNHMAIVID EYGGTSGLVT FEDIIEQIVG DIEDEFDEDE
  	SADDIHSVSA
  201	ERWRIHAATE IEDINAFFGT EYGSEEADTI RRLGHSGIGT
  	PARARRKSPY
  251	RRFAVHRRPR RQPPPAHADG DPREVSRACP TAVSAQFRMT
  	VRSFSVSIRP
  301	IRQT*

• The originally-identified partial strain B sequence (ORFS) shows 83.1% identity over a 135aa overlap with the partial gonococcal sequence (ORF5ng):
• 
• The complete strain B and gonococcal sequences (ORFS-1 & ORF5ng-1) show 92.4% identity in 304 aa overlap:
• 
• Computer analysis of these amino acid sequences indicates a putative leader sequence, and identified the following homologies:
• Homology with Hemolysin Homolog TlyC (Accession U32716) of H. influenzae
• ORF5 and TlyC proteins show 58% aa identity in 77 aa overlap (BLASTp).

• ORF5	2	HMAIVIDEYGGTSGLVTFEDIIEQIVGEIEDEFDEDDSADNIHAVSSDTWRIHAATEIED	61
  		HMAIV+DE+G  SGLVT EDI+EQIVG+IEDEFDE++ AD I  +S  T+ + A T+I+D
  TlyC	166	HMAIVVDEFGAVSGLVTIEDILEQIVGDIEDEFDEEEIAD-IRQLSRHTYAVRALTDIDD	224
  ORF5	62	INTFFGTEYSIEEADTI	78
  		N  F T++  EE DTI
  TlyC
  	225	FNAQFNTDFDDEEVDTI	241

• ORF5ng-1 also shows significant homology with TlyC:
• • SCORES Init1: 301 Initn: 419 Opt: 668
  • Smith-Waterman score: 668; 45.9% identity in 242 aa overlap
• 
• Homology with a Hypothetical Secreted Protein from E. coli:
• ORF5a shows homology to a hypothetical secreted protein from E. coli:

• sp|P77392|YBEX_ECOLI HYPOTHETICAL 33.3 KD PROTEIN IN CUTE-ASNB INTERGENIC
  REGION
  >gi|1778577 (U82598) similar to H. influenzae [Escherichia coli]
  >gi|1786879 (AE000170) f292; This 292 aa ORF is 23% identical (9 gaps)
  to 272 residues of an approx. 440 aa protein YTFL_HAEIN SW: P44717
  [Escherichia coli] Length = 292
  Score = 212 bits (533), Expect = 3e−54
  Identities = 112/230 (48%), Positives = 149/230 (64%), Gaps = 3/230 (1%)

  Query:	2	DGAQPKTNFXXRLIARLAR-EPDSAEDVLTLLRQAHEQEVFDADTLLRLEKVLDFSDLEV	60
  		D    K  F   L+++L   EP + +++L L+R + + ++ D DT   LE V+D +D  V
  Sbjct:	10	DTISNKKGFFSLLLSQLFHGEPKNRDELLALIRDSGQNDLIDEDTRDMLEGVMDIADQRV	69
  Query:	61	RDAMITRSRMNVLKENDSIERITAYVIDTAHSRFPVIGEDKDEVLGILHAKDLLKYM-FN	119
  		RD MI RS+M  LK N +++     +I++AHSRFPVI EDKD + GIL AKDLL +M  +
  Sbjct:	70	RDIMIPRSQMITLKRNQTLDECLDVIIESAHSRFPVISEDKDHIEGILMAKDLLPFMRSD	129
  Query:	120	PEQFHLKSILRPAVFVPEGKSLTALLKEFREQRNHMAIVIDEYGGTSGLVTFEDIIEQIV	179
  		E F +  +LR AV VPE K +  +LKEFR QR HMAIVIDE+GG SGLVT EDI+E IV
  Sbjct:	130	AEAFSMDKVLRQAVVVPESKRVDRMLKEFRSQRYHMAIVIDEFGGVSGLVTIEDILELIV	189
  Query:	180	GDIEDEFDEDESADNIHAVSAERWRIHAATEIEDINAFFGTEYSSEEADT	229
  		G+IEDE+DE++  D    +S   W + A   IED N  FGT +S EE DT
  Sbjct:	190	GEIEDEYDEEDDID-FRQLSRHTWTVRALASIEDFNEAFGTHFSDEEVDT	238

• Based on this analysis, including the amino acid homology to the TlyC hemolysin-homologue from H. influenzae (hemolysins are secreted proteins), it was predicted that the proteins from N. meningitidis and N. gonorrhoeae are secreted and could thus be useful antigens for vaccines or diagnostics.
• ORF5-1 (30.7 kDa) was cloned in the pGex vector and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 2A shows the results of affinity purification of the GST-fusion protein. Purified GST-fusion protein was used to immunise mice, whose sera were used for Western blot analysis (FIG. 1B). These experiments confirm that ORFS-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 5
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 29>:

• 1	ATGCGCGGCG GCAGGCCGGA TTCCGTTACC GTGCAGATTA
  	TCGAAGGTTC
  51	GCGTTTTTCG CATATGAGGA AAGTCATCGA CGCAACGCCC
  	GACATCGGAC
  101	ACGACACCAA AGGCTGGAGC AATGAAAAAC TGATGGCGGA
  	AGTTGCGCCC
  151	GATGCCTTCA GCGGCAATCC TGAAgGGCAG TTTTTCCCCG
  	ACAGCTACGA
  201	AATCGATGCG GGCGGCAGTG ATTTGCAGAT TTACCAAACC
  	GCCTACAAgG
  251	GCGATGCAAC GCCGCCTGAA TGAgGGCATG GGAAAGCAGG
  	CAGGACGGGC
  301	TGCCTTATAA AAACCCTTAT GAAATGCTGA TTATGGCGAr
  	CCTGGTCGAA
  351	AAGGAAACAG GGCATGAAGC CGAsCsCGAC CATGTcGCTT
  	CCGTCTTCGT
  401	CAACCGCCTG AAAATCGGTA TGCGCCTGCA AACCgAssCG
  	TCCGTGATTT
  451	ACGGCATGGG TGCGGCATAC AAGGGCAAAA TCCGTAAAGC
  	CGACCTGCGC
  501	CGCGACACGC CGTACAACAC CTACACGCGC GGCGGTCTGC
  	CGCCAACCCC
  551	GATTGCGCTG CCC..

• This corresponds to the amino acid sequence <SEQ ID 30; ORF7>:

• 1	MRGGRPDSVT VQIIEGSRFS HMRKVIDATP DIGHDTKGWS
  	NEKLMAEVAP
  51	DAFSGNPEGQ FFPDSYEIDA GGSDLQIYQT AYKAMQRRLN
  	EAWESRQDGL
  101	PYKNPYEMLI MAXLVEKETG HEAXXDHVAS VFVNRLKIGM
  	RLQTXXSVIY
  151	GMGAAYKGKI RKADLRRDTP YNTYTRGGLP PTPIALP..

• Further sequence analysis revealed the complete DNA sequence <SEQ ID 31>:

• 1	ATGTTGAGAA AATTGTTGAA ATGGTCTGCC GTTTTTTTGA
  	CCGTGTCGGC
  51	AGCCGTTTTC GCCGCGCTGC TTTTTGTTCC TAAGGATAAC
  	GGCAGGGCAT
  101	ACCGAATCAA AATTGCCAAA AACCAGGGTA TTTCGTCGGT
  	CGGCAGGAAA
  151	CTTGCCGAAG ACCGCATCGT GTTCAGCAGG CATGTTTTGA
  	CGGCGGCGGC
  201	CTACGTTTTG GGTGTGCACA ACAGGCTGCA TACGGGGACG
  	TACAGATTGC
  251	CTTCGGAAGT GTCTGCTTGG GATATCTTGC AGAAAATGCG
  	CGGCGGCAGG
  301	CCGGATTCCG TTACCGTGCA GATTATCGAA GGTTCGCGTT
  	TTTCGCATAT
  351	GAGGAAAGTC ATCGACGCAA CGCCCGACAT CGGACACGAC
  	ACCAAAGGCT
  401	GGAGCAATGA AAAACTGATG GCGGAAGTTG CGCCCGATGC
  	CTTCAGCGGC
  451	AATCCTGAAG GGCAGTTTTT CCCCGACAGC TACGAAATCG
  	ATGCGGGCGG
  501	CAGTGATTTG CAGATTTACC AAACCGCCTA CAAGGCGATG
  	CAACGCCGCC
  551	TGAATGAGGC ATGGGAAAGC AGGCAGGACG GGCTGCCTTA
  	TAAAAACCCT
  601	TATGAAATGC TGATTATGGC GAGCCTGGTC GAAAAGGAAA
  	CAGGGCATGA
  651	AGCCGACCGC GACCATGTCG CTTCCGTCTT CGTCAACCGC
  	CTGAAAATCG
  701	GTATGCGCCT GCAAACCGAC CCGTCCGTGA TTTACGGCAT
  	GGGTGCGGCA
  751	TACAAGGGCA AAATCCGTAA AGCCGACCTG CGCCGCGACA
  	CGCCGTACAA
  801	CACCTACACG CGCGGCGGTC TGCCGCCAAC CCCGATTGCG
  	CTGCCCGGCA
  851	AGGCGGCACT CGATGCCGCC GCCCATCCGT CCGGCGAAAA
  	ATACCTGTAT
  901	TTCGTGTCCA AAATGGACGG CACGGGCTTG AGCCAGTTCA
  	GCCATGATTT
  951	GACCGAACAC AATGCCGCCG TCCGCAAATA TATTTTGAAA
  	AAATAA

• This corresponds to the amino acid sequence <SEQ ID 32; ORF7-1>:

• 1	MLRKLLKWSA VFLTVSAAVF AALLFVPKDN GRAYRIKIAK
  	NQGISSVGRK
  51	LAEDRIVFSR HVLTAAAYVL GVHNRLHTGT YRLPSEVSAW
  	DILQKMRGGR
  101	PDSVTVQIIE GSRFSHMRKV IDATPDIGHD TKGWSNEKLM
  	AEVAPDAFSG
  151	NPEGQFFPDS YEIDAGGSDL QIYQTAYKAM QRRLNEAWES
  	RQDGLPYKNP
  201	YEMLIMASLV EKETGHEADR DHVASVFVNR LKIGMRLQTD
  	PSVIYGMGAA
  251	YKGKIRKADL RRDTPYNTYT RGGLPPTPIA LPGKAALDAA
  	AHPSGEKYLY
  301	FVSKMDGTGL SQFSHDLTEH NAAVRKYILK K*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with Hypothetical Protein Encoded by yceg Gene (Accession P44270) of H. influenzae
• ORF7 and yceg proteins show 44% aa identity in 192 aa overlap:

• ORF7   1 MRGGRPDSVTVQIIEGSRFSHMRKVIDATPDIGHDTKGWSNEKLMA-----EVAPDAFSG  55
  +  G+     V+ IEG  F   RK ++  P +    K  SNE++ A      ++  +
  yceg 102 LNSGKEVQFNVKWIEGKTFKDWRKDLENAPHLVQTLKDKSNEEIFALLDLPDIGQNLELK 161
  ORF7  56 NPEGQFFPDSYEIDAGGSDLQIYQTAYKAMQRRLNEAWESRQDGLPYKNPYEMLIMAXLV 115
  N EG  +PD+Y      +DL++ + + + M++ LN+AW  R + LP   NPYEMLI+A +V
  yceg 162 NVEGWLYPDTYNYTPKSTDLELLKRSAERMKKALNKAWNERDEDLPLANPYEMLILASIV 221
  ORF7 116 EKETGHEAXXDHVASVFVNRLKIGMRLQTXXSVIYGMGAAYKGKIRKADLRRDTPYNTYT 175
  EKETG       VASVF+NRLK  M+LQT  +VIYGMG  Y G IRK DL   TPYNTY
  yceg 222 EKETGIANERAKVASVFINRLKAKMKLQTDPTVIYGMGENYNGNIRKKDLETKTPYNTYV 281
  ORF7 176 RGGLPPTPIALP                                                 187
  GLPPTPIA+P
  yceg 282 IDGLPPTPIAMP                                                 293

• The complete length YCEG protein has sequence:

• 1	MKKFLIAILL LILILAGVAS FSYYKMTEFV KTPVNVQADE
  	LLTIERGTTS
  51	SKLATLFEQE KLIADGKLLP YLLKLKPELN KIKAGTYSLE
  	NVKTVQDLLD
  101	LLNSGKEVQF NVKWIEGKTF KDWRKDLENA PHLVQTLKDK
  	SNEEIFALLD
  151	LPDIGQNLEL KNVEGWLYPD TYNYTPKSTD LELLKRSAER
  	MKKALNKAWN
  201	ERDEDLPLAN PYEMLILASI VEKETGIANE RAKVASVFIN
  	RLKAKMKLQT
  251	DPTVIYGMGE NYNGNIRKKD LETKTPYNTY VIDGLPPTPI
  	AMPSESSLQA
  301	VANPEKTDFY YFVADGSGGH KFTRNLNEHN KAVQEYLRWY
  	RSQKNAK

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF7 shows 95.2% identity over a 187aa overlap with an ORF (ORF7a) from strain A of N. meningitidis:
• 
• The complete length ORF7a nucleotide sequence <SEQ ID 33> is:

• 1	ATGTTGAGAA AATTGTTGAA ATGGTCTGCC GTTTTTTTGA
  	CCGTATCGGC
  51	AGCCGTTTTC GCCGCGCTGC TTTTCGTCCC TAAAGACAAC
  	GGCAGGGCAT
  101	ACAGGATTAA AATTGCCAAA AACCAGGGTA TTTCGTCGGT
  	CGGCAGGAAA
  151	CTTGCCGAAG ACCGCATCGT GTTCAGCAGG CATGTTTTGA
  	CGGCGGCGGC
  201	CTACGTTTTG GGTGTGCACA ACAGGCTGCA TACGGGGACG
  	TACAGACTGC
  251	CTTCGGAAGT GTCTGCTTGG GATATCTTGC AGAAAATGCG
  	CGGCGGCAGG
  301	CCGGATTCCG TTACCGTGCA GATTATCGAA GGTTCGCGTT
  	TTTCGCATAT
  351	GAGGAAAGTC ATCGACGCAA CGCCCGACAT CGAACACGAC
  	ACCAAAGGCT
  401	GGAGCAATGA AAAACTGATG GCGGAAGTTG CCCCTGATGC
  	CTTCAGCGGC
  451	AATCCTGAAG GGCAGTTTTT CCCCGACAGC TACGAAATCG
  	ATGCGGGCGG
  501	CAGCGATTTA CGGATTTACC AAATCGCCTA CAAGGCGATG
  	CAACGCCGAC
  551	TGAATGAGGC ATGGGAAAGC AGGCAGGACG GGCTGCCTTA
  	TAAAAACCCT
  601	TATGAAATGC TGATTATGGC GAGCCTGATC GAAAAGGAAA
  	CAGGGCATGA
  651	AGCCGACCGC GACCATGTCG CTTCCGTCTT CGTCAACCGC
  	CTGAAAATCG
  701	GTATGCGCCT GCAAACCGAC CCGTCCGTGA TTTACGGCAT
  	GGGTGCGGCA
  751	TACAAGGGCA AAATCCGTAA AGCCGACCTG CGCCGCGACA
  	CGCCGTACAA
  801	CACCTACACG CGCGGCGGTC TGCCGCCAAC CCCGATCGCG
  	CTGCCCGGCA
  851	AGGCGGCACT CGATGCCGCC GCCCATCCGT CCGGTGAAAA
  	ATACCTGTAT
  901	TTCGTGTCCA AAATGGACGG TACGGGCTTG AGCCAGTTCA
  	GCCATGATTT
  951	GACCGAACAC AACGCCGCCG TTCGCAAATA TATTTTGAAA
  	AAATAA

• This is predicted to encode a protein having amino acid sequence <SEQ ID 34>:

• 1	MLRKLLKWSA VFLTVSAAVF AALLFVPKDN GRAYRIKIAK
  	NQGISSVGRK
  51	LAEDRIVFSR HVLTAAAYVL GVHNRLHTGT YRLPSEVSAW
  	DILQKMRGGR
  101	PDSVTVQIIE GSRFSHMRKV IDATPDIEHD TKGWSNEKLM
  	AEVAPDAFSG
  151	NPEGQFFPDS YEIDAGGSDL RIYQIAYKAM QRRLNEAWES
  	RQDGLPYKNP
  201	YEMLIMASLI EKETGHEADR DHVASVFVNR LKIGMRLQTD
  	PSVIYGMGAA
  251	YKGKIRKADL RRDTPYNTYT RGGLPPTPIA LPGKAALDAA
  	AHPSGEKYLY
  301	FVSKMDGTGL SQFSHDLTEH NAAVRKYILK K*

• A leader peptide is underlined.
• ORF7a and ORF7-1 show 98.8% identity in 331 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF7 shows 94.7% identity over a 187aa overlap with a predicted ORF (ORF7.ng) from N. gonorrhoeae:
• 
• An ORF7ng nucleotide sequence <SEQ ID 35> is predicted to encode a protein having amino acid sequence <SEQ ID 36>:

• 1	MRGGRPDSVT VQIIEGSRFS HMRKVIDATP DIGHDTKGWS
  	NEKLMAEVAP
  51	DAFSGNPEGQ FFPDSYEIDA GGSDLQIYQT AYKAMQRRLN
  	EAWAGRQDGL
  101	PYKNPYEMLI MASLIEKETG HEADRDHVAS VFVNRLKIGM
  	RLQTDPSVIY
  151	GMGAAYKGKI RKADLRRDTP YNTYTGGGLP PTRIALPGKA
  	AMDAAAHPSG
  201	EKYLYFVSKM DGTGLSQFSH DLTEHNAAVR KYILKK*

• Further sequence analysis revealed a partial DNA sequence of ORF7ng <SEQ ID 37>:

• 1	..taccgaatca AGATTGCCAA AAATCAGGGT ATTTCGTCGG
  	TCGGCAGGAA
  51	ACTTGCcgaA GACCGCATCG TGTTCAGCAG GCATGTTTTG
  	ACAGCGGCGG
  101	CCTACGTTTT GGGTGTGCAC AACAGGCTGC ATACGGGGAC
  	gTACAGATTG
  151	CCTTCGGAAG TGTCTGCTTG GGATATCTTG CAGAAAATGC
  	GCGGCGGCAG
  201	GCCGGATTCC GTTACCGTGC AGATTATCGA AGGTTCGCGT
  	TTTTCGCATA
  251	TGAGGAAAGT CATCGACGCA ACGCCCGACA TCGGACACGA
  	CACCAAAGGC
  301	TGGAGCAATG AAAAACTGAT GGCGGAAGTT GCGCCCGATG
  	CCTTCAGCGG
  351	CAATCCTGAA GGGCAGTTTT TTCCCGACAG CTACGAAATC
  	GATGCGGGCG
  401	GCAGCGATTT GCAGATTTAC CAAACCGCCT ACAAGGCGAT
  	GCAACGCCGC
  451	CTGAACGAGG CATGGGCAGG CAGGCAGGAC GGGCTGCCTT
  	ATAAAAACCC
  501	TTATGAAATG CTGATTATGG CGAGCCTGAT CGAAAAGGAA
  	ACGGGGCATG
  551	AGGCCGACCG CGACCATGTC GCTTCCGTCT TCGTCAACCG
  	CCTGAAAATC
  601	GGTATGCGCC TGCAAACCGA CCCGTCCGTG ATTTACGGCA
  	TGGGTGCGGC
  651	ATACAAGGGC AAAATCCGTA AAGCCGACCT GCGCCGCGAC
  	ACGCCGTACA
  701	aCAccTAtac gggcgggggc ttgccgccaa cccggattgc
  	gctgcccggC
  751	Aaggcggcaa tggatgccgc cgcccacccg tccggcgaAa
  	aatacctgTa
  801	tttcgtgtcC AAAATGGACG GCACGGGCTT GAGCCAGTTC
  	AGCCATGATT
  851	TGACCGAACA CAACGCCGCc gTcCGCAAAT ATATTTTGAA
  	AAAATAA

• This corresponds to the amino acid sequence <SEQ ID 38; ORF7ng-1>:

• 1	..YRIKIAKNQG ISSVGRKLAE DRIVFSRHVL TAAAYVLGVH
  	NRLHTGTYRL
  51	PSEVSAWDIL QKMRGGRPDS VTVQIIEGSR FSHMRKVIDA
  	TPDIGHDTKG
  101	WSNEKLMAEV APDAFSGNPE GQFFPDSYEI DAGGSDLQIY
  	QTAYKAMQRR
  151	LNEAWAGRQD GLPYKNPYEM LIMASLIEKE TGHEADRDHV
  	ASVFVNALKI
  201	GMRLQTDPSV IYGMGAAYKG KIRKADLRRD TPYNTYTGGG
  	LPPTRIALPG
  251	KAAMDAAAHP SGEKYLYFVS KMDGTGLSQF SHDLTEHNAA
  	VRKYILKK*

• ORF7ng-1 and ORF7-1 show 98.0% identity in 298 aa overlap:
• 
• In addition, ORF7ng-1 shows significant homology with a hypothetical E. coli protein:

• sp|P28306|YCEG_ECOLI HYPOTHETICAL 38.2 KD PROTEIN IN PABC-HOLB
  INTERGENIC REGION gi|1787339 (AE000210) o340; 100% identical to fragment
  YCEG_ECOLI SW: P28306 but has 97 additional C-terminal residues
  [Escherichia coli] Length = 340
  Score = 79 (36.2 bits), Expect = 5.0e−57, Sum P(2) = 5.0e−57
  Identities = 20/87 (22%), Positives = 40/87 (45%)

  Query:	10	GISSVGRKLAEDRIVFSRHVLTAAAYVLGVHNRLHTGTYRLPSEVSAWDILQKMRGGRPD	69
  		G  ++G +L  D+I+    V      +    +    GTYR   +++  ++L+ +  G+
  Sbjct:	49	GRLALGEQLYADKIINRPRVFQWLLRIEPDLSHFKAGTYRFTPQMTVREMLKLLESGKEA	108
  Query:	70	SVTVQIIEGSRFSHMRKVIDATPDIGH	96
  		++++EG R S   K +   P I H
  Sbjct:	109	QFPLRLVEGMRLSDYLKQLREAPYIKH	135

  Score = 438 (200.7 bits), Expect = 5.0e−57, Sum P(2) = 5.0e−57
  Identities = 84/155 (54%), Positives = 111/155 (71%)

  Query:	120	EGQFFPDSYEIDAGGSDLQIYQTAYKAMQRRLNEAWAGRQDGLPYKNPYEMLIMASLIEK	179
  		EG F+PD++   A  +D+ + + A+K M + ++ AW GR DGLPYK+  +++ MAS+IEK
  Sbjct:	158	EGWFWPDTWMYTANTTDVALLKRAHKKMVKAVDSAWEGRADGLPYKDKNQLVTMASIIEK	217
  Query:	180	ETGHEADRDHVASVFVNRLKIGMRLQTDPSVIYGMGAAYKGKIRKADLRRDTPYNTYTGG	239
  		ET   ++RD VASVF+NRL+IGMRLQTDP+VIYGMG  Y GK+ +ADL   T YNTYT
  Sbjct:	218	ETAVASERDKVASVFINRLRIGMRLQTDPTVIYGMGERYNGKLSRADLETPTAYNTYTIT	277
  Query:	240	GLPPTRIALPGKAAMDAAAHPSGEKYLYFVSKMDG	274
  		GLPP  IA PG  ++ AAAHP+   YLYFV+   G
  Sbjct:	278	GLPPGAIATPGADSLKAAAHPAKTPYLYFVADGKG	312

• Based on this analysis, including the fact that the H. influenzae YCEG protein possesses a possible leader sequence, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 6
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 39>:

• 1	CGTTTCAAAA TGTTAACTGT GTTGACGGCA ACCTTGATTG
  	CCGGACAGGT
  51	ATCTGCCGCC GGAGGCGGTG CGGGGGATAT GAAACAGCCG
  	AAGGAAGTCG
  101	GAAAGGTTTT CAGAAAGCAG CAGCGTTACA GCGAGGAAGA
  	AATCAAAAAC
  151	GAACGCGCAC GGCTTGCGGC AGTGGGCGAG CGGGTTAATC
  	AGATATTTAC
  201	GTTGCTGGGA GGGGAAACCG CCTTGCAAAA GGGGCAGGCG
  	GGAACGGCTC
  251	TGGCAACCTA TATGCTGATG TTGGAACGCA CAAAATCCCC
  	CGAAGTCGCC
  301	GAACGCGCCT TGGAAATGGC CGTGTCGCTG AACGCGTTTG
  	AACAGGCGGA
  351	AATGATTTAT CAGAAATGGC GGCAGATTGA GCCTATACCG
  	GGTAAGGCGC
  401	AAAAACGGGC GGGGTGGCTG CGGAACGTGC TGAGGGAAAG
  	AGGAAATCAG
  451	CATCTGGACG GACGGGAAGA AGTGCTGGCT CAGGCGGACG
  	AAGGACAG

• This corresponds to the amino acid sequence <SEQ ID 40; ORF9>:

• 1	..RFKMLTVLTA TLIAGQVSAA GGGAGDMKQP KEVGKVFRKQ
  	QRYSEEEIKN
  51	ERARLAAVGE RVNQIFTLLG GETALQKGQA GTALATYMLM
  	LERTKSPEVA
  101	ERALEMAVSL NAFEQAEMIY QKWRQIEPIP GKAQKRAGWL
  	RNVLRERGNQ
  151	HLDGREEVLA QADEGQ

• Further sequence analysis revealed the complete DNA sequence <SEQ ID 41>:

• 1	ATGTTACCTA ACCGTTTCAA AATGTTAACT GTGTTGACGG
  	CAACCTTGAT
  51	TGCCGGACAG GTATCTGCCG CCGGAGGCGG TGCGGGGGAT
  	ATGAAACAGC
  101	CGAAGGAAGT CGGAAAGGTT TTCAGAAAGC AGCAGCGTTA
  	CAGCGAGGAA
  151	GAAATCAAAA ACGAACGCGC ACGGCTTGCG GCAGTGGGCG
  	AGCGGGTTAA
  201	TCAGATATTT ACGTTGCTGG GAGGGGAAAC CGCCTTGCAA
  	AAGGGGCAGG
  251	CGGGAACGGC TCTGGCAACC TATATGCTGA TGTTGGAACG
  	CACAAAATCC
  301	CCCGAAGTCG CCGAACGCGC CTTGGAAATG GCCGTGTCGC
  	TGAACGCGTT
  351	TGAACAGGCG GAAATGATTT ATCAGAAATG GCGGCAGATT
  	GAGCCTATAC
  101	CGGGTAAGGC GCAAAAACGG GCGGGGTGGC TGCGGAACGT
  	GCTGAGGGAA
  451	AGAGGAAATC AGCATCTGGA CGGACTGGAA GAAGTGCTGG
  	CTCAGGCGGA
  501	CGAAGGACAG AACCGCAGGG TGTTTTTATT GTTGGCACAA
  	GCCGCCGTGC
  551	AACAGGACGG GTTGGCGCAA AAAGCATCGA AAGCGGTTCG
  	CCGCGCGGCG
  601	TTGAAATATG AACATCTGCC CGAAGCGGCG GTTGCCGATG
  	TGGTGTTCAG
  651	CGTACAGGGA CGCGAAAAGG AAAAGGCAAT CGGAGCTTTG
  	CAGCGTTTGG
  701	CGAAGCTCGA TACGGAAATA TTGCCCCCCA CTTTAATGAC
  	GTTGCGTCTG
  751	ACTGCACGCA AATATCCCGA AATACTCGAC GGCTTTTTCG
  	AGCAGACAGA
  801	CACCCAAAAC CTTTCGGCCG TCTGGCAGGA AATGGAAATT
  	ATGAATCTGG
  851	TTTCCCTGCA CAGGCTGGAT GATGCCTATG CGCGTTTGAA
  	CGTGCTGTTG
  901	GAACGCAATC CGAATGCAGA CCTGTATATT CAGGCAGCGA
  	TATTGGCGGC
  951	AAACCGAAAA GAAGGTGCTT CCGTTATCGA CGGCTACGCC
  	GAAAAGGCAT
  1001	ACGGCAGGGG GACGGAGGAA CAGCGGAGCA GGGCGGCGCT
  	AACGGCGGCG
  1051	ATGATGTATG CCGACCGCAG GGATTACGCC AAAGTCAGGC
  	AGTGGCTGAA
  1101	AAAAGTATCC GCGCCGGAAT ACCTGTTCGA CAAAGGTGTG
  	CTGGCGGCTG
  1151	CGGCGGCTGT CGAGTTGGAC GGCGGCAGGG CGGCTTTGCG
  	GCAGATCGGC
  1201	AGGGTGCGGA AACTTCCCGA ACAGCAGGGG CGGTATTTTA
  	CGGCAGACAA
  1251	TTTGTCCAAA ATACAGATGC TCGCCCTGTC GAAGCTGCCC
  	GATAAACGGG
  1301	AGGCTTTGAG GGGGTTGGAC AAGATTATCG AAAAACCGCC
  	TGCCGGCAGT
  1351	AATACAGAGT TACAGGCAGA GGCATTGGTA CAGCGGTCAG
  	TTGTTTACGA
  1401	TCGGCTTGGC AAGCGGAAAA AAATGATTTC AGATCTTGAA
  	AGGGCGTTCA
  1451	GGCTTGCACC CGATAACGCT CAGATTATGA ATAATCTGGG
  	CTACAGCCTG
  1501	CTGACCGATT CCAAACGTTT GGACGAAGGT TTCGCCCTGC
  	TTCAGACGGC
  1551	ATACCAAATC AACCCGGACG ATACCGCTGT CAACGACAGC
  	ATAGGCTGGG
  1601	CGTATTACCT GAAAGGCGAC GCGGAAAGCG CGCTGCCGTA
  	TCTGCGGTAT
  1651	TCGTTTGAAA ACGACCCCGA GCCCGAAGTT GCCGCCCATT
  	TGGGCGAAGT
  1701	GTTGTGGGCA TTGGGCGAAC GCGATCAGGC GGTTGACGTA
  	TGGACGCAGG
  1751	CGGCACACCT TACGGGAGAC AAGAAAATAT GGCGGGAAAC
  	GCTCAAACGT
  1801	CACGGCATCG CATTGCCCCA ACCTTCCCGA AAACCTCGGA
  	AATAA

• This corresponds to the amino acid sequence <SEQ ID 42; ORF9-1>:

• 1	MLPNRFKMLT VLTATLIAGQ VSAAGGGAGD MKQPKEVGKV
  	FRKQQRYSEE
  51	EIKNERARLA AVGERVNQIF TLLGGETALQ KGQAGTALAT
  	YMLMLERTKS
  101	PEVAERALEM AVSLNAFEQA EMIYQKWRQI EPIPGKAQKR
  	AGWLRNVLRE
  151	RGNQHLDGLE EVLAQADEGQ NRRVFLLLAQ AAVQQDGLAQ
  	KASKAVRRAA
  201	LKYEHLPEAA VADVVFSVQG REKEKAIGAL QRLAKLDTEI
  	LPPTLMTLRL
  251	TARKYPEILD GFFEQTDTQN LSAVWQEMEI MNLVSLHRLD
  	DAYARLNVLL
  301	ERNPNADLYI QAAILAANRK EGASVIDGYA EKAYGRGTEE
  	QRSRAALTAA
  351	MMYADRRDYA KVRQWLKKVS APEYLFDKGV LAAAAAVELD
  	GGRAALRQIG
  401	RVRKLPEQQG RYFTADNLSK IQMLALSKLP DKREALRGLD
  	KIIEKPPAGS
  451	NTELQAEALV QRSVVYDRLG KRKKMISDLE RAFRLAPDNA
  	QIMNNLGYSL
  501	LTDSKRLDEG FALLQTAYQI NPDDTAVNDS IGWAYYLKGD
  	AESALPYLRY
  551	SFENDPEPEV AAHLGEVLWA LGERDQAVDV WTQAAHLTGD
  	KKIWRETLKR
  601	HGIALPQPSR KPRK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF9 shows 89.8% identity over a 166aa overlap with an ORF (ORF9a) from strain A of N. meningitidis:
• 
• The complete length ORF9a nucleotide sequence <SEQ ID 43> is:

• 1	ATGTTACCCG CCCGTTTCAC CATTTTATCT GTGCTCGCGG
  	CAGCCCTGCT
  51	TGCCGGGCAG GCGTATGCCG CCGGCGCGGC GGATGCGAAG
  	CCGCCGAAGG
  101	AAGTCGGAAA GGTTTTCAGA AAGCAGCAGC GTTACAGCGA
  	GGAAGAAATC
  151	AAAAACGAAC GCGCACGGCT TGCGGCAGTG GGCGAGCGGG
  	TTAATCAGAT
  201	ATTTACGTTG CTGGGANGGG AAACCGCCTT GCAAAAGGGG
  	CAGGCGGGAA
  251	CGGCTCTGGC AACCTATATG CTGATGTTGG AACGCACAAA
  	ATCCCCCGAA
  301	GTCGCCGAAC GCGCCTTGGA AATGGCCGTG TCNCTGAACG
  	CGTTTGAACA
  351	GGCGGAAATG ATTTATCAGA AATGGCGGCA GATTGAGCCT
  	ATACCGGGTA
  401	AGGCGCAAAA ACGGGCGGGG TGGCTGCGGA ACGTGCTGAG
  	GGAAAGAGGA
  451	AATCAGCATC TAGACGGACT GGAAGAANTG CTGGCTCAGG
  	CGGACGAANG
  501	ACAGAACCGC AGGGTGTTTT TATTGTTGGC ACAAGCCGCC
  	GTGCAACAGG
  551	ACGGGTTGGC GCAAAAAGCA TCGAAAGCGG TTCGCCGCGC
  	GGCGTTGAGA
  601	TATGAACATC TGCCCGAAGC GGCGGTTGCC GATGTGGTGT
  	TCAGCGTACA
  651	GGNACGCGAA AAGGAAAAGG CAATCGGAGC TTTGCAGCGT
  	TTGGCGAAGC
  701	TCGATACGGA AATATTGCCC CCCACTTTAA TGACGTTGCG
  	TCTGACTGCA
  751	CGCAAATATC CCGAAATACT CGACGGCTTT TTCGAGCAGA
  	CAGACACCCA
  801	AAACCTTTCG GCCGTCTGGC AGGAAATGGA AATTATGAAT
  	CTGGTTTCCC
  851	TGCACAGGCT GGATGATGCC TATGCGCGTT TGAACGTGCT
  	GTTGGAACGC
  901	AATCCGAATG CAGACCTGTA TATTCAGGCA GCGATATTGG
  	CGGCAAACCG
  951	AAAAGAANGT GCTTCCGTTA TCGACGGCTA CGCCGAAAAG
  	GCATACGGCA
  1001	GGGGGACGGG GGAACAGCGG GGCAGGGCGG CAATGACGGC
  	GGCGATGATA
  1051	TATGCCGACC GAAGGGATTA CACCAAAGTC AGGCAGTGGT
  	TGAAAAAAGT
  1101	GTCCGCGCCG GAATACCTGT TCGACAAAGG TGTGCTGGCG
  	GCTGCGGCGG
  1151	CTGTCGAGTT GGACNGCGGC AGGGCGGCTT TGCGGCAGAT
  	CGGCAGGGTG
  1201	CGGAAACTTC CCGAACAGCA GGGGCGGTAT TTTACGGCAG
  	ACAATTTGTC
  1251	CAAAATACAG ATGTTCGCCC TGTCGAAGCT GCCCGACAAA
  	CGGGAGGCTT
  1301	TGAGGGGGTT GGACAAGATT ATCGAAAAAC CGCCTGCCGG
  	CAGTAATACA
  1351	GAGTTACAGG CAGAGGCATT GGTACAGCGG TCAGTTGTTT
  	ACGATCGGCT
  1401	TGGCAAGCGG AAAAAAATGA TTTCAGATCT TGAAAGGGCG
  	TTCAGGCTTG
  1451	CACCCGATAA CGCTCAGATT ATGAATAATC TGGGCTACAG
  	CCTGCTTTCC
  1501	GATTCCAAAC GTTTGGACGA AGGCTTCGCC CTGCTTCAGA
  	CGGCATACCA
  1551	AATCAACCCG GACGATACCG CTGTCAACGA CAGCATAGGC
  	TGGGCGTATT
  1601	ACCTGAAANG CGACGCGGAA AGCGCGCTGC CGTATCTGCG
  	GTATTCGTTT
  1651	GAAAACGACC CCGAGCCCGA AGTTGCCGCC CATTTGGGCG
  	AAGTGTTGTG
  1701	GGCATTGGGC GAACGCGATC AGGCGGTTGA CGTATGGACG
  	CAGGCGGCAC
  1751	ACCTTACGGG AGACAAGAAA ATATGGCGGG AAACGCTCAA
  	ACGTCACGGC
  1801	ATCGCATTGC CCCAACCTTC CCGAAAACCT CGGAAATAA

• This encodes a protein having amino acid sequence <SEQ ID 44>:

• 1	MLPARFTILS VLAAALLAGQ AYAAGAADAK PPKEVGKVFR
  	KQQRYSEEEI
  51	KNERARLAAV GERVNQIFTL LGXETALQKG QAGTALATYM
  	LMLERTKSPE
  101	VAERALEMAV SLNAFEQAEM IYQKWRQIEP IPGKAQKRAG
  	WLRNVLRERG
  151	NQHLDGLEEX LAQADEXQNR RVFLLLAQAA VQQDGLAQKA
  	SKAVRRAALR
  201	YEHLPEAAVA DVVFSVQXRE KEKAIGALQR LAKLDTEILP
  	PTLMTLRLTA
  251	RKYPEILDGF FEQTDTQNLS AVWQEMEIMN LVSLHRLDDA
  	YARLNVLLER
  301	NPNADLYIQA AILAANRKEX ASVIDGYAEK AYGRGTGEQR
  	GRAAMTAAMI
  351	YADRRDYTKV RQWLKKVSAP EYLFDKGVLA AAAAVELDXG
  	RAALRQIGRV
  401	RKLPEQQGRY FTADNLSKIQ MFALSKLPDK REALRGLDKI
  	IEKPPAGSNT
  451	ELQAEALVQR SVVYDRLGKR KKMISDLERA FRLAPDNAQI
  	MNNLGYSLLS
  501	DSKRLDEGFA LLQTAYQINP DDTAVNDSIG WAYYLKXDAE
  	SALPYLRYSF
  551	ENDPEPEVAA HLGEVLWALG ERDQAVDVWT QAAHLTGDKK
  	IWRETLKRHG
  601	IALPQPSRKP RK*

• ORF9a and ORF9-1 show 95.3% identity in 614 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF9 shows 82.8% identity over a 163aa overlap with a predicted ORF (ORF9.ng) from N. gonorrhoeae:
• 
• The ORF9ng nucleotide sequence <SEQ ID 45> was predicted to encode a protein having including acid sequence <SEQ ID 46>:

• 1	MIMLPARFTI LSVLAAALLA GQAYAAGAAD VELPKEVGKV
  	LRKHRRYSEE
  51	EIKNERARLA AVGERVNRVF TLLGGETALQ KGQAGTALAT
  	YMLMLERTKS
  101	PEVAERALEM AVSLNAFEQA EMIYQKWRQI EPIPGEAQKP
  	AGWLRNVLKE
  151	GGNPHLDRLE EVPAQSDYVH QPMIFLLLVQ AAVQHGGVAQ
  	KPSKAVRPAA
  201	YNYEVLPETA GADAVFCVQG PQYEKAIQSF PPCGRNPQTE
  	NIAPPFNELF
  251	RPTARPISPK LLQRFFRTEP NLAKPFRPPG PEMETYQTGF
  	PRPLTRNNPT

• Amino acids 1-28 are a putative leader sequence, and 173-189 are predicted to be a transmembrane domain.
• Further sequence analysis revealed the complete length ORF9ng DNA sequence <SEQ ID 47>:

• 1	ATGTTACCCG CCCGTTTCAC TATTTTATCT GTCCTCGCAG
  	CAGCCCTGCT
  51	TGCCGGACAG GCGTATGCTG CCGGCGCGGC GGATGTGGAG
  	CTGCCGAAGG
  101	AAGTCGGAAA GGTTTTAAGG AAACATCGGC GTTACAGCGA
  	GGAAGAAATC
  151	AAAAACGAAC GCGCACGGCT TGCGGCAGTG GGCGAACGGG
  	TCAACAGGGT
  201	GTTTACGCTG TTGGGCGGTG AAACGGCTTT GCAGAAAGGG
  	CAGGCGGGAA
  251	CGGCTCTGGC AACCTATATG CTGATGTTGG AACGCACAAA
  	ATCCCCCGAA
  301	GTCGCCGAAC GCGCCTTGGA AATGGCCGTG TCGCTGAACG
  	CGTTTGAACA
  351	GGCGGAAATG ATTTATCAGA AATGgcggca gatcgagcct
  	ataCcgggtg
  401	aggcgcaaaa accgGcgggG tggctgcgga acgtattgaa
  	ggaagggGGa
  451	aaTCAGCATC TGGAcgggtt gaaagaggTG CtggcgcaAT
  	cggacgatGT
  501	GCAAAAAcgc aggaTATTTT TGCTGCTGGT GCAAGCCGCC
  	GTGCagcagg
  551	gTGGGGTGGC TCAAAAAGCA TCGAAAGCGG TTCGCcgtgc
  	GGcgttgaAG
  601	TATGAACATC TGCCcgaagc ggcggTTGCC GATGcggTGT
  	TCGGCGTACA
  651	GGGACGCGAA AAGGAAAagg caaTCGAAGC TTTGCAGCGT
  	TTGGCGAAGC
  701	TCGATACGGA AATATTGCCC CCCACTTTAA TGACGTTGCG
  	TCTGACTGCA
  751	CGCAAATATC CCGAAATACT CGACGGCTTT TTCGAGCAGA
  	CAGACACCCA
  801	AAACCTTTCG GCCGTCTGGC AGGAAATGGA AATTATGAAT
  	CTGGTTTCCC
  851	TGCGTAAGCC GGATGATGCC TATGCGCGTT TGAACGTGCT
  	GTTGGAACAC
  901	AACCCGAATG CAAACCTGTA TATTCAGGCG GCGATATTGG
  	CGGCAAACCG
  951	AAAAGAAGGT GCGTCCGTTA TCGACGGCTA CGCCGAAAAG
  	GCATACGGCA
  1001	GGGGGACGGG GGAACAGCGG GGCagggcgg cAATgacggc
  	GGCGATGATA
  1051	TATGCCGACC GCAGGGATTA CGCCAAAGTC AGGCAGTGGT
  	TGAAAAAAGT
  1101	GTCCGCGCCG GAATACCTGT TCGACAAAGG CGTGCTGGCG
  	GCTGCGGCGG
  1151	CTGCCGAATT GGACGGAGGC CGGGCGGCTT TGCGGCAGAT
  	CGGCAGGGTG
  1201	CGGAAACTTC CCGAACAGCA GGGGCGGTAT TTTACGGCAG
  	ACAATTTGTC
  1251	CAAAATACAG ATGCTCGCCC TGTCGAAGCT GCCCGACAAA
  	CGGGAAGCCC
  1301	TGATCGGGCT GAACAACATC ATCGCCAAAC TTTCGGCGGC
  	GGGAAGCACG
  1351	GAACCTTTGG CGGAAGCATT GGCACAGCGT TCCATTATTT
  	ACGaacAGTT
  1401	cggCAAACGG GGAAAAATGA TTGCCGACCT tgaAACcgcg
  	CTCAAACTTA
  1451	CGCCCGATAA TGCACAAATT ATGAATAATC TGGGCTACAG
  	CCTGCTTTCC
  1501	GATTCCAAAC GTTTGGACGA GGGTTTCGCC CTGCTTCAGA
  	CGGCATACCA
  1551	AATCAACCCG GACGATACCG CCGTTAACGA CAGCATAGGC
  	TGGGCGTATT
  1601	ACCTGAAAGG CGACgcggaA AGCGCGCTGC CGTATCTGcg
  	gtattcgttt
  1651	gAAAACGACC CCGAGCCCGA AGTTGCCGCC CATTTGGGCG
  	AAGTGTTGTG
  1701	GGCATTGGGC GAACGCGATC AGGCGGTTGA CGTATGGACG
  	CAGGCGGCAC
  1751	ACCTTAGGGG AGACAAGAAA ATATGGCGGG AGACGCTCAA
  	ACGCTACGGA
  1801	ATCGCCTTGC CCGAGCCTTC CCGAAAACCC CGGAAATAA

• This encodes a protein having amino acid sequence <SEQ ID 48>:

• 1	MLPARFTILS VLAAALLAGQ AYAAGAADVE LPKEVGKVLR
  	KHRRYSEEEI
  51	KNERARLAAV GERVNRVFTL LGGETALQKG QAGTALATYM
  	LMLERTKSPE
  101	VAERALEMAV SLNAFEQAEM IYQKWRQIEP IPGEAQKPAG
  	WLRNVLKEGG
  151	NQHLDGLKEV LAQSDDVQKR RIFLLLVQAA VQQGGVAQKA
  	SKAVRRAALK
  201	YEHLPEAAVA DAVFGVQGRE KEKAIEALQR LAKLDTEILP
  	PTLMTLRLTA
  251	RKYPEILDGF FEQTDTQNLS AVWQEMEIMN LVSLRKPDDA
  	YARLNVLLEH
  301	NPNANLYIQA AILAANRKEG ASVIDGYAEK AYGRGTGEQR
  	GRAAMTAAMI
  351	YADRRDYAKV RQWLKKVSAP EYLFDKGVLA AAAAAELDGG
  	RAALRQIGRV
  401	RKLPEQQGRY FTADNLSKIQ MLALSKLPDK REALIGLNNI
  	IAKLSAAGST
  451	EPLAEALAQR SIIYEQFGKR GKMIADLETA LKLTPDNAQI
  	MNNLGYSLLS
  501	DSKRLDEGFA LLQTAYQINP DDTAVNDSIG WAYYLKGDAE
  	SALPYLRYSF
  551	ENDPEPEVAA HLGEVLWALG ERDQAVDVWT QAAHLRGDKK
  	IWRETLKRYG
  601	IALPEPSRKP RK*

• ORF9ng and ORF9-1 show 88.1% identity in 614 aa overlap:
• 

  
• In addition, ORF9ng shows significant homology with a hypothetical protein from P. aeruginosa:

• sp|P42810|YHE3_PSEAE HYPOTHETICAL 64.8 KD PROTEIN IN HEMM-HEMA INTERGENIC
  REGION (ORF3)
  >gi|1072999|pir||S49376 hypothetical protein 3 - Pseudomonas aeruginosa
  >gi|557259 (X82071) orf3 [Pseudomonas aeruginosa] Length = 576
  Score = 128 bits (318), Expect = 1e−28
  Identities = 138/587 (23%), Positives = 228/587 (38%), Gaps = 125/587 (21%)

  Query:	67	VFTLLGGETALQKGQAGTALATYMLMLERTKSPEVAERALEMAVSLNAFEQAEMIYQKWR	126
  		+++LL  E A Q+ +   AL+ Y++  ++T+ P V+ERA  +A  L A ++A      W
  Sbjct:	53	LYSLLVAELAGQRNRFDIALSNYVVQAQKTRDPGVSERAFRIAEYLGADQEALDTSLLWA	112
  Query:	127	QIEPIPGEAQKPAG--------------WLRNVLKEGGNQHLDGLKEVLAQSDDVQKRRI	172
  		+  P   +AQ+ A               ++  VL   G+ H D L    A++D   +  +
  Sbjct:	113	RSAPDNLDAQRAAAIQLARAGRYEESMVYMEKVLNGQGDTHFDFLALSAAETDPDTRAGL	172
  Query:	173	FXXXXXXXXXXXXXXXKASKAVRRAALKYEHLPEAAVADAVFGVQGREKEKAIEALQRLA	232
  		++      KY +  +     A+   Q    ++A+  L+  +
  Sbjct:	173	L------------------QSFDHLLKKYPNNGQLLFGKALLLQQDGRPDEALTLLEDNS	214
  Query:	233	KLDTEILPPTLMTLRLTARK-----YPEILDGFFEQTDTQNLSAVWQEMEIMNLVSLRKP	287
  		E+ P  L +  L + K      P +  G  E  D + +   +  +    LV   +
  Sbjct:	215	ASRHEVAPLLLRSRLLQSMKRSDEALPLLKAGIKEHPDDKRVRLAYARL----LVEQNRL	270
  Query:	288	DDAYARLNVLLEHNPN---------------------ANLYIQAAI--------------	312
  		DDA A    L++  P+                     A +Y++  +
  Sbjct:	271	DDAKAEFAGLVQQFPDDDDDLRFSLALVCLEAQAWDEARIYLEELVERDSHVDAAHFNLG	330
  Query:	313	-LAANRKEGASVIDGYAEKAYGRGTGEQRGRAAMTAAMIYADRRDYAKVRQWLKKVSAPE	371
  		LA  +K+ A  +D YA+   G G      +   T  ++ A R D A  R    +   P+
  Sbjct:	331	RLAEEQKDTARALDEYAQ--VGPGNDFLPAQLRQTDVLLKAGRVDEAAQRLDKARSEQPD	388
  Query:	372	YLFDKXXXXXXXXXXXXXXXXXXRQIGRVRKLPEQQGRYFTADNLSKIQMLALSKLPDKR	431
  		Y                                        A  L  I+  ALS    +
  Sbjct:	389	Y----------------------------------------AIQLYLIEAEALSNNDQQE	408
  Query:	432	EALIGLNNIIAKLSAAGSTEPLAEALAQRSIIYEQFGKRGKMIADLETALKLTPDNAQIM	491
  		+A   +   + +       E L   L  RS++ E+     +M  DL   +   PDNA  +
  Sbjct:	409	KAWQAIQEGLKQYP-----EDL-NLLYTRSMLAEKRNDLAQMEKDLRFVIAREPDNAMAL	462
  Query:	492	NNLGYSLLSDSKRLDEGFALLQTAYQINPDDTAVNDSIGWAYYLKGDAESALPYLRYSFE	551
  		N LGY+L   + R  E   L+  A+++NPDD A+ DS+GW  Y +G    A  YLR + +
  Sbjct:	463	NALGYTLADRTTRYGEARELILKAHKLNPDDPAILDSMGWINYRQGKLADAERYLRQALQ	522
  Query:	552	NDPEPEVAAHLGEVLWALGERDQAVDVWTQAAHLRGDKKIWRETLKR	598
  		P+ EVAAHLGEVLWA G +  A  +W +    + D  + R T+KR
  Sbjct:	523	RYPDHEVAAHLGEVLWAQGRQGDARAIWREYLDKQPDSDVLRRTIKR	569

  gi|2983399 (AE000710) hypothetical protein [Aquifex aeolicus] Length = 545
  Score = 81.5 bits (198), Expect = 1e−14
  Identities = 61/198 (30%), Positives = 98/198 (48%), Gaps = 19/198 (9%)

  Query:	408	GRYFTADNL-SKIQMLALSKLPDKREALIGLNNIIAKLSAAGSTEPLAEALAQ-------	459
  		G Y  A  L  K ++LA    PDK+E L    +  +K         + + L +
  Sbjct:	335	GNYEDAKRLIEKAKVLA----PDKKEILFLEADYYSKTKQYDKALEILKKLEKDYPNDSR	390
  Query:	460	----RSIIYEQFGKRGKMIADLETALKLTPDNAQIMNNLGYSLLS--DSKRLDEGFALLQ	513
  		+I+Y+  G        L  A++L P+N    N LGYSLL     +R++E   L++
  Sbjct:	391	VYFMEAIVYDNLGDIKNAEKALRKAIELDPENPDYYNYLGYSLLLWYGKERVEEAEELIK	450
  Query:	514	TAYQINPDDTAVNDSIGWAYYLKGDAESALPYLRYSF-ENDPEPEVAAHLGEVLWALGER	572
  		A + +P++ A  DS+GW YYLKGD E A+ YL  +  E   +P V  H+G+VL  +G +
  Sbjct:	451	KALEKDPENPAYIDSMGWVYYLKGDYERAMQYLLKALREAYDDPVVNEHVGDVLLKMGYK	510
  Query:	573	DQAVDVWTQAAHLRGDKK	590
  		++A + + +A  L  + K
  Sbjct:	511	EEARNYYERALKLLEEGK	528

• Based on this analysis, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 7
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 49>:

• 1	AACCTCTACG CCGGCCCGCA GACCACATCC GTCATCGCAA
  	ACATCGCCGA
  51	CAACCTGCAA CTGGCCAAAG ACTACGGCAA AGTACACTGG
  	TTCGCCTCCC
  101	CGCTCTTCTG GCTCCTGAAC CAACTGCACA ACATCATCGG
  	CAACTGGGGC
  151	TGGGCGATTA TCGTTTTAAC CATCATCGTC AAAGCCGTAC
  	TGTATCCATT
  201	GACCAACGCC TCTTACCGCT CTATGGCGAA AATGCGTGCC
  	GCCGCACCCA
  251	AACTGCAAGC CATCAAAGAG AAATACGGCG ACGACCGTAT
  	GGCGCAACAA
  301	CAGGCGATGA TGCAGCTTTA CACAGACGAG AAAATCAACC
  	CGaCTGGGCG
  351	GCTGCCTGCC TATGCTGTTG CAAATCCCCG TCTTCATCGG
  	ATTGTATTGG
  401	GCATTGTTCG CCTCCGTAGA ATTGCGCCAG GCACCTTGGC
  	TGGGTTGGAT
  451	TACCGACCTC AGCCGCGCCG ACCCCTACTA CATCCTGCCC
  	ATCATTATGG
  501	CGGCAACGAT GTTCGCCCAA ACTTATCTGA ACCCGCCGCC
  	GAcCGACCCG
  551	ATGCagGCGA AAATGATGAA AATCATGCCG TTGGTTTTCT
  	CsGwCrTGTT
  601	CTTCTTCTTC CCTGCCGGks TGGTATTGTA CTGGGTAGTC
  	AACAACCTCC
  651	TGACCATCGC CCAGCAATGG CACATCAACC GCAGCATCGA
  	AAAACAACGC
  701	GCCCAAGGCG AAGTCGTTTC CTAA

• This corresponds to the amino acid sequence <SEQ ID 50; ORF11>:

• 1	..NLYAGPQTTS VIANIADNLQ LAKDYGKVHW FASPLFWLLN
  	QLHNIIGNWG
  51	WAIIVLTIIV KAVLYPLTNA SYRSMAKMRA AAPKLQAIKE
  	KYGDDRMAQQ
  101	QAMMQLYTDE KINPLGGCLP MLLQIPVFIG LYWALFASVE
  	LRQAPWLGWI
  151	TDLSRADPYY ILPIIMAATM FAQTYLNPPP TDPMQAKMMK
  	IMPLVFSXXF
  201	FFFPAGXVLY WVVNNLLTIA QQWHINRSIE KQRAQGEVVS *

• Further sequence analysis revealed the complete DNA sequence <SEQ ID 51>:

• 1	ATGGATTTTA AAAGACTCAC GGCGTTTTTC GCCATCGCGC
  	TGGTGATTAT
  51	GATCGGCTGG GAAAAGATGT TCCCCACTCC GAAGCCAGTC
  	CCCGCGCCCC
  101	AACAGGCAGC ACAACAACAG GCCGTAACCG CTTCCGCCGA
  	AGCCGCGCTC
  151	GCGCCCGCAA CGCCGATTAC CGTAACGACC GACACGGTTC
  	AAGCCGTCAT
  201	TGATGAAAAA AGCGGCGACC TGCGCCGGCT GACCCTGCTC
  	AAATACAAAG
  251	CAACCGGCGA CGAAAATAAA CCGTTCATCC TGTTTGGCGA
  	CGGCAAAGAA
  301	TACACCTACG TCGCCCAATC CGAACTTTTG GACGCGCAGG
  	GCAACAACAT
  351	TCTAAAAGGC ATCGGCTTTA GCGCACCGAA AAAACAGTAC
  	AGCTTGGAAG
  401	GCGACAAAGT TGAAGTCCGC CTGAGCGCGC CTGAAACACG
  	CGGTCTGAAA
  451	ATCGACAAAG TTTATACTTT CACCAAAGGC AGCTATCTGG
  	TCAACGTCCG
  501	CTTCGACATC GCCAACGGCA GCGGTCAAAC CGCCAACCTG
  	AGCGCGGACT
  551	ACCGCATCGT CCGCGACCAC AGCGAACCCG AGGGTCAAGG
  	TTACTTTACC
  601	CACTCTTACG TCGGCCCTGT TGTTTATACC CCTGAAGGCA
  	ACTTCCAAAA
  651	AGTCAGCTTT TCCGACTTGG ACGACGATGC CAAATCCGGC
  	AAATCCGAGG
  701	CCGAATACAT CCGCAAAACC CCGACCGGCT GGCTCGGCAT
  	GATTGAACAC
  751	CACTTCATGT CCACCTGGAT TCTCCAACCT AAAGGCAGAC
  	AAAGCGTTTG
  801	CGCCGCAGGC GAGTGCAACA TCGACATCAA ACGCCGCAAC
  	GACAAGCTGT
  851	ACAGCACCAG CGTCAGCGTG CCTTTAGCCG CCATCCAAAA
  	CGGCGCGAAA
  901	GCCGAAGCCT CCATCAACCT CTACGCCGGC CCGCAGACCA
  	CATCCGTCAT
  951	CGCAAACATC GCCGACAACC TGCAACTGGC CAAAGACTAC
  	GGCAAAGTAC
  1001	ACTGGTTCGC CTCCCCGCTC TTCTGGCTCC TGAACCAACT
  	GCACAACATC
  1051	ATCGGCAACT GGGGCTGGGC GATTATCGTT TTAACCATCA
  	TCGTCAAAGC
  1101	CGTACTGTAT CCATTGACCA ACGCCTCTTA CCGCTCTATG
  	GCGAAAATGC
  1151	GTGCCGCCGC ACCCAAACTG CAAGCCATCA AAGAGAAATA
  	CGGCGACGAC
  1201	CGTATGGCGC AACAACAGGC GATGATGCAG CTTTACACAG
  	ACGAGAAAAT
  1251	CAACCCGCTG GGCGGCTGCC TGCCTATGCT GTTGCAAATC
  	CCCGTCTTCA
  1301	TCGGATTGTA TTGGGCATTG TTCGCCTCCG TAGAATTGCG
  	CCAGGCACCT
  1351	TGGCTGGGTT GGATTACCGA CCTCAGCCGC GCCGACCCCT
  	ACTACATCCT
  1401	GCCCATCATT ATGGCGGCAA CGATGTTCGC CCAAACTTAT
  	CTGAACCCGC
  1451	CGCCGACCGA CCCGATGCAG GCGAAAATGA TGAAAATCAT
  	GCCGTTGGTT
  1501	TTCTCCGTCA TGTTCTTCTT CTTCCCTGCC GGTCTGGTAT
  	TGTACTGGGT
  1551	AGTCAACAAC CTCCTGACCA TCGCCCAGCA ATGGCACATC
  	AACCGCAGCA
  1601	TCGAAAAACA ACGCGCCCAA GGCGAAGTCG TTTCCTAA

• This corresponds to the amino acid sequence <SEQ ID 52; ORF11-1>:

• 1	MDFKRLTAFF AIALVIMIGW EKMFPTPKPV PAPQQAAQQQ
  	AVTASAEAAL
  51	APATPITVTT DTVQAVIDEK SGDLRRLTLL KYKATGDENK
  	PFILFGDGKE
  101	YTYVAQSELL DAQGNNILKG IGFSAPKKQY SLEGDKVEVR
  	LSAPETRGLK
  151	IDKVYTFTKG SYLVNVRFDI ANGSGQTANL SADYRIVRDH
  	SEPEGQGYFT
  201	HSYVGPVVYT PEGNFQKVSF SDLDDDAKSG KSEAEYIRKT
  	PTGWLGMIEH
  251	HFMSTWILQP KGRQSVCAAG ECNIDIKRRN DKLYSTSVSV
  	PLAAIQNGAK
  301	AEASINLYAG PQTTSVIANI ADNLQLAKDY GKVHWFASPL
  	FWLLNQLHNI
  351	IGNWGWAIIV LTIIVKAVLY PLTNASYRSM AKMRAAAPKL
  	QAIKEKYGDD
  401	RMAQQQAMMQ LYTDEKINPL GGCLPMLLQI PVFIGLYWAL
  	FASVELRQAP
  451	WLGWITDLSR ADPYYILPII MAATMFAQTY LNPPPTDPMQ
  	AKMMKIMPLV
  501	FSVMFFFFPA GLVLYWVVNN LLTIAQQWHI NRSIEKQRAQ
  	GEVVS*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a 60 kDa Inner-Membrane Protein (Accession P25754) of Pseudomonas putida
• ORF11 and the 60 kDa protein show 58% aa identity in 229 aa overlap (BLASTp).

• ORF11	2	LYAGPQTTSVIANIADNLQLAKDYGKVHWFASPLFWLLNQLHNIIGNWGWAIIVLTIIVK	61
  		LYAGP+  S +  ++  L+L  DYG + + A P+FWLL  +H+++GNWGW+IIVLT+++K
  60K	324	LYAGPKIQSKLKELSPGLELTVDYGFLWFIAQPIFWLLQHIHSLLGNWGWSIIVLTMLIK	383
  ORF11	62	AVLYPLTNASYRSMAKMRAAAPKLQAIKEKYGDDRXXXXXXXXXLYTDEKINPLGGCLPM	121
  		+ +PL+ ASYRSMA+MRA APKL A+KE++GDDR         LY  EKINPLGGCLP+
  60K	384	GLFFPLSAASYRSMARMRAVAPKLAALKERFGDDRQKMSQAMMELYKKEKINPLGGCLPI	443
  ORF11	122	LLQIPVFIGLYWALFASVELRQAPWLGWITDLSRADPYYILPIIMAATMFAQTYLNPPPT	181
  		L+Q+PVF+ LYW L  SVE+RQAPW+ WITDLS  DP++ILPIIM ATMF Q  LNP P
  60K	444	LVQMPVFLALYWVLLESVEMRQAPWILWITDLSIKDPFFILPIIMGATMFIQQRLNPTPP	503
  ORF11	182	DPMQAKMMKIMPLVXXXXXXXXPAGXVLYWVVNNLLTIAQQWHINRSIE	230
  		DPMQAK+MK+MP++        PAG VLYWVVNN L+I+QQW+I R IE
  60K	504	DPMQAKVMKMMPIIFTFFFLWFPAGLVLYWVVNNCLSISQQWYITRRIE	552

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF 11 shows 97.9% identity over a 240aa overlap with an ORF (ORF11a) from strain A of N. meningitidis:
• 
• The complete length ORF11a nucleotide sequence <SEQ ID 53> is:

• 	ANGGATTTTA AAAGACTCAC NGNGTTTTTC GCCATCGCAC
  	TGGTGATTAT
  51	GATCGGATNG NAAANGATGT TCCCCACTCC GAAGCCCGTC
  	CCCGCGCCCC
  101	AACAGACGGC ACAACAACAG GCCGTAANCG CTTCCGCCGA
  	AGCCGCGCTC
  151	GCGCCCGNAN CGCCGATTAC CGTAACGACC GACACGGTTC
  	AAGCCGTCAT
  201	TGATGAAAAA AGCGGCGACC TGCGCCGGCT GACCCTGCTC
  	AAATACAAAG
  251	CAACCGGCGA CNAAAATAAA CCGTTCATCC TGTTTGGCGA
  	CGGCAAANAA
  301	TACACCTACN TCGCCCANTC CGAACTTTTG GACGCGCAGG
  	GCAACAACAT
  351	TCTAAAAGGC ATCGGCTTTA GCGCACCGAA AAAACAGTAC
  	AGCTTGGAAG
  401	GCGACAAAGT TGAAGTCCGC CTGAGCGCAC CTGAAACACG
  	CGGTCTGAAA
  451	ATCGACAAAG TTTATACTTT CACCAAAGGC AGCTATCTGG
  	TCAACGTCCG
  501	CTTCGACATC GCCAACGGCA GCGGTCAAAC CGCCAACCTG
  	AGCGCGGACT
  551	ACCGCATCGT CCGCGACCAC AGCGAACCCG AGGGTCAAGG
  	CTACTTTACC
  601	CACTCTTACG TCGGCCCTGT TGTTTATACC CCTGAAGGCA
  	ACTTCCAAAA
  651	AGTCAGCTTC TCCGACTTGG ACGACGATGC CAANTCCGGN
  	AAATCCGAGG
  701	CCGAATACAT CCGCAAAACC CNGACCGGCT GGCTCGGCAT
  	GATTGAACAC
  751	CACTTCATGT CCACCTGGAT CCTCCAACCC AAAGGCGGAC
  	AAAGCGTTTG
  801	CGCCGCTGGC GACTGCNGTA TNGACATCAA ACGCCGCAAC
  	GACAAGCTGT
  851	ACAGCACCAG CGTCAGCGTG CCTTTAGCCG CTATCCAAAA
  	CGGTGCGAAA
  901	TCCNAAGCCT CCATCAACCT CTACGCCGGC CCACAGACCA
  	CATCNGTTAT
  951	CGCAAACATC GCCGACAACC TGCAACTGGN CAAAGACTAC
  	GGCAAAGTAC
  1001	ACTGGTTCGC CTCCCCCCTC TTTTGGCTTT TGAACCAACT
  	GCACAACATC
  1051	ATCGGCAACT GGGGCTGGGC GATTATCGTT TTAACCATCA
  	TCGTCAAAGC
  1101	CGTACTGTAT CCATTGACCA ACGCCTCTTA CCGTTCGATG
  	GCGAAAATGC
  1151	GTGCCGCCGC GCCCAAACTG CAAGCCATCA AAGAGAAATA
  	CGGCGACGAC
  1201	CGTATGGCGC AGCAACAAGC CATGATGCAG CTTTACACAG
  	ACGAGAAAAT
  1251	CAACCCGCTG GGCGGCTGCC TGCCTATGCT GTTGCAAATC
  	CCCGTCTTCA
  1301	TCGGATTGTA TTGGGCATTG TTCGCCTCCG TAGAATTGCG
  	CCAGGCACCT
  1351	TGGCTGGGTT GGATTACCGA CCTCAGCCGC GCCGACCCNT
  	ACTACATCCT
  1401	GCCCATCATT ATGGCGGCAA CGATGTTCGC CCAAACCTAT
  	CTGAACCCGC
  1451	CGCCGACCGA CCCGATGCAG GCGAAAATGA TGAAAATCAT
  	GCCTTTGGTT
  1501	NTNTCNNNNA NGTTCTTCNN CTTCCCTGCC GGTCTGGTAT
  	TGTACTGGGT
  1551	GATCAACAAC CTCCTGACCA TCGCCCAGCA ATGGCACATC
  	AACCGCAGCA
  1601	TCGAAAAACA ACGCGCCCAA GGCGAAGTCG TTTCCTAA

• This encodes a protein having amino acid sequence <SEQ ID 54>:

• 1	XDFKRLTXFF AIALVIMIGX XXMFPTPKPV PAPQQTAQQQ
  	AVXASAEAAL
  51	APXXPITVTT DTVQAVIDEK SGDLRRLTLL KYKATGDXNK
  	PFILFGDGKX
  101	YTYXAXSELL DAQGNNILKG IGFSAPKKQY SLEGDKVEVR
  	LSAPETRGLK
  151	IDKVYTFTKG SYLVNVRFDI ANGSGQTANL SADYRIVRDH
  	SEPEGQGYFT
  201	HSYVGPVVYT PEGNFQKVSF SDLDDDAXSG KSEAEYIRKT
  	XTGWLGMIEH
  251	HFMSTWILQP KGGQSVCAAG DCXXDIKRRN DKLYSTSVSV
  	PLAAIQNGAK
  301	SXASINLYAG PQTTSVIANI ADNLQLXKDY GKVHWFASPL
  	FWLLNQLHNI
  351	IGNWGWAIIV LTIIVKAVLY PLTNASYRSM AKMRAAAPKL
  	QAIKEKYGDD
  401	RMAQQQAMMQ LYTDEKINPL GGCLPMLLQI PVFIGLYWAL
  	FASVELRQAP
  451	WLGWITDLSR ADPYYILPII MAATMFAQTY LNPPPTDPMQ
  	AKMMKIMPLV
  501	XSXXFFXFPA GLVLYWVINN LLTIAQQWHI NRSIEKQRAQ
  	GEVVS*

• ORF11a and ORF11-1 show 95.2% identity in 544 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF11 shows 96.3% identity over a 240aa overlap with a predicted ORF (ORF11.ng) from N. gonorrhoeae:
• 
• An ORF11ng nucleotide sequence <SEQ ID 55> was predicted to encode a protein having amino acid sequence <SEQ ID 56>:

• 1	MAVNLYAGPQ TTSVIANIAD NLQLAKDYGK VHWFASPLFW
  	LLNQLHNIIG
  51	NWGWAIVVLT IIVKAVLYPL T NASYRSMAK MRAAAPELQT
  	IKEKYGDDRM
  101	AQQQAMMQLF EDEEINPLGG CLPMLLQIPV FIGLYWALFA
  	SVELRQAPWL
  151	GWITDLSRAD PYYILPIIMA ATMFAQTYLN PPPTDPMQAK
  	MMKIMPLVFS
  201	VMFFFFPAGL VLYWVVNNLL TIAQQWHINR SIEKQRAQGE
  	VVS*

• Further sequence analysis revealed the complete gonococcal DNA sequence <SEQ ID 57> to be:

• 1	ATGGATTTTA AAAGACTCAC GGCGTTTTTC GCCATCGCGC
  	TGGTGATTAT
  51	GATCGGCTGG GAAAAAATGT TCCCCACCCC GAAACCCGTC
  	CCCGCGCCCC
  101	AACAGGCGGC ACAAAAACAG GCAGCAACCG CTTCCGCCGA
  	AGCCGCGCTC
  151	GCGCCCGCAA CGCCGATTAC CGTAACGACC GACACGGTTC
  	AAGCCGTTAT
  201	TGATGAAAAA AGTGGCGACC TGCGCCGGCT GACCCTGCTC
  	AAATACAAAG
  251	CAACCGGCGA CGAAAACAAA CCGTTCGTCC TGTTTGGCGA
  	CGGCAAAGAA
  301	TACACCTACG TCGCCCAATC CGAACTTTTG GACGCGCAGG
  	GCAACAACAT
  351	TCTGAAAGGC ATCGGCTTTA GCGCACCGAA AAAACAGTAC
  	ACCCTCAACG
  401	GCGACACAGT CGAAGTCCGC CTGAGCGCGC CCGAAACCAA
  	CGGACTGAAA
  451	ATCGACAAAG TCTATACCTT TACCAAAGAC AGCTATCTGG
  	TCAACGTCCG
  501	CTTCGACATC GCCAACGGCA GCGGTCAAAC CGCCAACCTG
  	AGCGCGGACT
  551	ACCGCATCGT CCGCGACCAC AGCGAACCCG AGGGTCAAGG
  	CTACTTTACC
  601	CACTCTTACG TCGGCCCTGT TGTTTATACC CCTGAAGGCA
  	ACTTCCAAAA
  651	AGTCAGCTTC TCCgacTTgg acgACGATGC gaaaTccggc
  	aaATccgagg
  701	ccgaatacaT CCGCAAAACC ccgaccggtt ggctcggcat
  	gattgaacac
  751	cacttcatgt ccacctggat cctccAAcct aaaggcggcc
  	aaaacgtttg
  801	cgcccaggga gactgccgta tcgacattaa aCgccgcaac
  	gacaagctgt
  851	acagcgcaag cgtcagcgtg cctttaaccg ctatcccaac
  	ccgggggcca
  901	aaaccgaaaa tggcggTCAA CCTGTATGCC GGTCCGCAAA
  	CCACATCCGT
  951	TATCGCAAAC ATCGCcgacA ACCTGCAACT GGCAAAAGAC
  	TACGGTAAAG
  1001	TACACTGGTT CGCATCGCCG CTCTTCTGGC TCCTGAACCA
  	ACTGCACAAC
  1051	ATTATCGGCA ACTGGGGCTG GGCAATCGTC GTTTTGACCA
  	TCATCGTCAA
  1101	AGCCGTACTG TATCCATTGA CCAACGcctc ctACCGTTCG
  	ATGGCGAAAA
  1151	TGCGTGccgc cgcacCcaaA CTGCAGACCA TCAAAGAAAA
  	ATAcgGCGAC
  1201	GACCGTATGG CGCAACAGCA AGCGATGATG CAGCTTTACA
  	AAgacgAGAA
  1251	AATCAACCCG CTGGGCGGCT GTctgcctat gctgttgCAA
  	ATCCCCGTCT
  1301	TCATCGGCTT GTACTGGGCA TTGTTCGCCT CCGTAGAATT
  	GCGCCAGGCA
  1351	CCTTGGCTGG GCTGGATTAC CGACCTCAGC CGCGCCGACC
  	CCTACTACAT
  1401	CCTGCCCATC ATTATGGCGG CAACGATGTT CGCCCAAACC
  	TATCTGAACC
  1451	CGCCGCCGAC CGACCCGATG CAGGCGAAAA TGATGAAAAT
  	CATGCCGTTG
  1501	GTTTTCTCCG TCATGTTCTT CTTCTTCCCT GCCGGTTTGG
  	TTCTCTACTG
  1551	GGTGGTCAAC AACCTCCTGA CCATCGCCCA GCAGTGGCAC
  	ATCAACCGCA
  1601	GCATCGAAAA ACAACGCGCC CAAGGCGAAG TCGTTTCCTA
  	A

• This encodes a protein having amino acid sequence <SEQ ID 58; ORF11ng-1>:

• 1	MDFKRLTAFF AIALVIMIGW EKMFPTPKPV PAPQQAAQKQ
  	AATASAEAAL
  51	APATPITVTT DTVQAVIDEK SGDLRRLTLL KYKATGDENK
  	PFVLFGDGKE
  101	YTYVAQSELL DAQGNNILKG IGFSAPKKQY TLNGDTVEVR
  	LSAPETNGLK
  151	IDKVYTFTKD SYLVNVRFDI ANGSGQTANL SADYRIVRDH
  	SEPEGQGYFT
  201	HSYVGPVVYT PEGNFQKVSF SDLDDDAKSG KSEAEYIRKT
  	PTGWLGMIEH
  251	HFMSTWILQP KGGQNVCAQG DCRIDIKRRN DKLYSASVSV
  	PLTAIPTRGP
  301	KPKMAVNLYA GPQTTSVIAN IADNLQLAKD YGKVHWFASP
  	LFWLLNQLHN
  351	IIGNWGWAIV VLTIIVKAVL YPLTNASYRS MAKMRAAAPK
  	LQTIKEKYGD
  401	DRMAQQQAMM QLYKDEKINP LGGCLPMLLQ IPVFIGLYWA
  	LFASVELRQA
  451	PWLGWITDLS RADPYYILPI IMAATMFAQT YLNPPPTDPM
  	QAKMMKIMPL
  501	VFSVMFFFFP AGLVLYWVVN NLLTIAQQWH INRSIEKQRA
  	QGEVVS*

• ORF11ng-1 and ORF11-1 shown 95.1% identity in 546 aa overlap:
• 
• In addition, ORF11ng-1 shows significant homology with an inner-membrane protein from the database (accession number p25754):
• 
• Based on this analysis, including the homology to an inner-membrane protein from P. putida and the predicted transmembrane domains (seen in both the meningococcal and gonoccal proteins), it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 8
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 59>:

• 1	..GCCGTCTTAA TCATCGAATT ATTGACGGGA ACGGTTTATC
  	TTTTGGTTGT
  51	NAGCGCGGCT TTGGCGGGTT CGGGCATTGC TTACGGGCTG
  	ACCGGCAGTA
  101	CGCCTGCCGC CGTCTTGACC GNCGCTCTGC TTTCCGCGCT
  	GGGTATTTNG
  151	TTCGTACACG CCAAAACCGC CGTTAGAAAA GTTGAAACGG
  	ATTCATATCA
  201	GGATTTGGAT GCCGGACAAT ATGTCGAAAT CCTCCGNCAC
  	ACAGGCGGCA
  251	ACCGTTACGA AGTT.TTTAT CGCGGTACG. ACTGGCAGGC
  	TCAAAATACG
  301	GGGCAAGAAG AGCTTGAACC AGGAACTCGC GCCCTCATTG
  	TCCGCAAGGA
  351	AGGCAACCTT CTTATTATCA CACACCCTTA A

• This corresponds to the amino acid sequence <SEQ ID 60; ORF13>:

• 1	..AVLIIELLTG TVYLLVVSAA LAGSGIAYGL TGSTPAAVLT
  	XALLSALGIX
  51	FVHAKTAVRK VETDSYQDLD AGQYVEILRH TGGNRYEVXY
  	RGTXWQAQNT
  101	GQEELEPGTR ALIVRKEGNL LIITHP*

• Further sequence analysis elaborated the DNA sequence slightly <SEQ ID 61>:

• 1	..GCCGTCTTAA TCATCGAATT ATTGACGGGA ACGGTTTATC
  	TTTTGGTTGT
  51	nAGCGCGGCT TTGGCGGGTT CGGGCATTGC TTACGGGCTG
  	ACCGGCAGTA
  101	CGCCTGCCGC CGTCTTGACC GnCGCTCTGC TTTCCGCGCT
  	GGGTATTTnG
  151	TTCGTACACG CCAAAACCGC CGTTAGAAAA GTTGAAACGG
  	ATTCATATCA
  201	GGATTTGGAT GCCGGACAAT ATGTCGAAAT CCTCCGACAC
  	ACAGGCGGCA
  251	ACCGTTACGA AGTTTTtTAT CGCGGTACGc ACTGGCAGGC
  	TCAAAATACG
  301	GGGCAAGAAG AGCTTGAACC AGGAACTCGC GCCCTCATTG
  	TCCGCAAGGA
  351	AGGCAACCTT CTTATTATCA CACACCCTTA A

• This corresponds to the amino acid sequence <SEQ ID 62; ORF13-1>:

• 1	..AVLIIELLTG TVYLLVVSAA LAGSGIAYGL TGSTPAAVLT
  	XALLSALGIX
  51	FVHAKTAVRK VETDSYQDLD AGQYVEILRH TGGNRYEVFY
  	RGTHWQAQNT
  101	GQEELEPGTR ALIVRKEGNL LIITHP*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF13 shows 92.9% identity over a 126aa overlap with an ORF (ORF13a) from strain A of N. meningitidis:
• 
• The complete length ORF13a nucleotide sequence <SEQ ID 63> is:

• 1	ATGACTGTAT GGTTTGTTGC CGCTGTTGCC GTCTTAATCA
  	TCGAATTATT
  51	GACGGGAACG GTTTATCTTT TGGTTGTCAG CGCGGCTTTG
  	GCGGGTTCGG
  101	GCATTGCTTA CGGGCTGACC GGCAGCACGC CTGCCGCCGT
  	CTTGACCGCC
  151	GCTCTGCTTT CCGCGCTGGG TATTTGGTTC GTACACGCCA
  	AAACCGCCGT
  201	GGGAAAAGTT GAAACGGATT CATATCAGGA TTTGGATGCC
  	GGGCAATATG
  251	CCGAAATCCT CCGGCACGCA GGCGGCAACC GTTACGAAGT
  	TTTTTATCGC
  301	GGTACGCACT GGCAGGCTCA AAATACGGGG CAAGAAGAGC
  	TTGAACCAGG
  351	AACGCGCGCC CTAATCGTCC GCAAGGAAGG CAACCTTCTT
  	ATCATCGCAA
  401	AACCTTAA

• This encodes a protein having amino acid sequence <SEQ ID 64>:

• 1	MTVWFVAAVA VLIIELLTGT VYLLVVSAAL AGSGIAYGLT
  	GSTPAAVLTA
  51	ALLSALGIWF VHAKTAVGKV ETDSYQDLDA GQYAEILRHA
  	GGNRYEVFYR
  101	GTHWQAQNTG QEELEPGTRA LIVRKEGNLL IIAKP*

• ORF13a and ORF13-1 show 94.4% identity in 126 aa overlap
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF13 shows 89.7% identity over a 126aa overlap with a predicted ORF (ORF13.ng) from N. gonorrhoeae:
• 
• The complete length ORF13ng nucleotide sequence <SEQ ID 65> is:

• 1	ATGACTGTAT GGTTTGTTGC CGCTGTTGCC GTCTTAATCA
  	TCGAATTATT
  51	GACGGGAACG GTTTATCTTT TGGTTGTCAG CGCGGCTTTG
  	GCGGGTTCGG
  101	GCATTGCCTA CGGGCTGACT GGCAGCACGC CTGCCGCCGT
  	CTTGACCGCC
  151	GCACTGCTTT CCGCGCTGGG CATTTGGTTC GTACATGCCA
  	AAACCGCCGT
  201	GGGAAAAGTT GAAACGGATT CATATCAGGA TTTGGATACC
  	GGAAAATATG
  251	CCGAAATCCT CCGATACACA GGCGGCAACC GTTACGAAGT
  	TTTTTATCGC
  301	GGTACGCACT GGCAGGCGCA AAATACGGGG CAGGAAGTGT
  	TTGAACCGGG
  351	AACGCGCGCC CTCATCGTCC GCAAAGAAGG TAACCTTCTT
  	ATCATCGCAA
  401	ACCCTTAA

• This encodes a protein having amino acid sequence <SEQ ID 66>:

• 1	MTVWFVAAVA VLIIELLTGT VYLLVVSAAL AGSGIAYGLT
  	GSTPAAVLTA
  51	ALLSALGIWF VHAKTAVGKV ETDSYQDLDT GKYAEILRYT
  	GGNRYEVFYR
  101	GTHWQAQNTG QEVFEPGTRA LIVRKEGNLL IIANP*

• ORF13ng shows 91.3% identity in 126 aa overlap with ORF13-1:
• 
• Based on this analysis, including the extensive leader sequence in this protein, it is predicted that ORF13 and ORF13ng are likely to be outer membrane proteins. It is thus predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 9
• The following DNA sequence was identified in N. meningitidis <SEQ ID 67>:

• 1	ATGTwTGATT TCGGTTTrGG CGArCTGGTT TTTGTCGGCA
  	TTATCGCCCT
  51	GATwGtCCTC GGCCCCGAAC GCsTGCCCGA GGCCGCCCGC
  	AyCGCCGGAC
  101	GGcTCATCGG CAGGCTGCAA CGCTTTGTCG GcAGCGTCAA
  	ACAGGAATTT
  151	GACACTCAAA TCGAACTGGA AGAACTGAGG AAGGCAAAGC
  	AGGAATTTGA
  201	AGCTGCCGcC GCTCAGGTTC GAGACAGCCT CAAAGAAACC
  	GGTACGGATA
  251	TGGAAGGCAA TCTGCACGAC ATTTCCGACG GTCTGAAGCC
  	TTGGGAAAAA
  301	CTGCCCGAAC AGCGGACACC TGCCGATTTC GGTGTCGATG
  	AAAACGGCAA
  351	TCCGCT.TCC CGATGCGGCA AACACCCTAT CAGACGGCAT
  	TTCCGACGTT
  401	ATGCCGTC..

• This corresponds to the amino acid sequence <SEQ ID 68; ORF2>:

• 1	MXDFGLGELV FVGIIALIVL GPERXPEAAR XAGRLIGRLQ
  	RFVGSVKQEF
  51	DTQIELEELR KAKQEFEAAA AQVRDSLKET GTDMEGNLHD
  	ISDGLKPWEK
  101	LPEQRTPADF GVDENGNPXS RCGKHPIRRH FRRYAV..

• Further work revealed the complete nucleotide sequence <SEQ ID 69>:

• 1	ATGTTTGATT TCGGTTTGGG CGAGCTGGTT TTTGTCGGCA
  	TTATCGCCCT
  51	GATTGTCCTC GGCCCCGAAC GCCTGCCCGA GGCCGCCCGC
  	ACCGCCGGAC
  101	GGCTCATCGG CAGGCTGCAA CGCTTTGTCG GCAGCGTCAA
  	ACAGGAATTT
  151	GACACTCAAA TCGAACTGGA AGAACTGAGG AAGGCAAAGC
  	AGGAATTTGA
  201	AGCTGCCGCC GCTCAGGTTC GAGACAGCCT CAAAGAAACC
  	GGTACGGATA
  251	TGGAAGGCAA TCTGCACGAC ATTTCCGACG GTCTGAAGCC
  	TTGGGAAAAA
  301	CTGCCCGAAC AGCGGACACC TGCCGATTTC GGTGTCGATG
  	AAAACGGCAA
  351	TCCGCTTCCC GATGCGGCAA ACACCCTATC AGACGGCATT
  	TCCGACGTTA
  401	TGCCGTCCGA ACGTTCCTAC GCTTCCGCCG AAACCCTTGG
  	GGACAGCGGG
  451	CAAACCGGCA GTACAGCCGA ACCCGCGGAA ACCGACCAAG
  	ACCGCGCATG
  501	GCGGGAATAC CTGACTGCTT CTGCCGCCGC ACCCGTCGTA
  	CAGACCGTCG
  551	AAGTCAGCTA TATCGATACT GCTGTTGAAA CGCCTGTTCC
  	GCACACCACT
  601	TCCCTGCGCA AACAGGCAAT AAGCCGCAAA CGCGATTTTC
  	GTCCGAAACA
  651	CCGCGCCAAA CCTAAATTGC GCGTCCGTAA ATCATAA

• This corresponds to the amino acid sequence <SEQ ID 70; ORF2-1>:

• 1	MFDFGLGELV FVGIIALIVL GPERLPEAAR TAGRLIGRLQ
  	RFVGSVKQEF
  51	DTQIELEELR KAKQEFEAAA AQVRDSLKET GTDMEGNLHD
  	ISDGLKPWEK
  101	LPEQRTPADF GVDENGNPLP DAANTLSDGI SDVMPSERSY
  	ASAETLGDSG
  151	QTGSTAEPAE TDQDRAWREY LTASAAAPVV QTVEVSYIDT
  	AVETPVPHTT
  201	SLRKQAISRK RDFRPKHRAK PKLRVRKS*

• Further work identified the corresponding gene in strain A of N. meningitidis <SEQ ID 71>:

• 1	ATGTTTGATT TCGGTTTGGG CGAGCTGGTT TTTGTCGGCA
  	TTATCGCCCT
  51	GATTGTCCTC GGCCCCGAAC GCCTGCCCGA GGCCGCCCGC
  	ACCGCCGGAC
  101	GGCTCATCGG CAGGCTGCAA CGCTTTGTCG GCAGCGTCAA
  	ACAGGAATTT
  151	GACACGCAAA TCGAACTGGA AGAACTAAGG AAGGCAAAGC
  	AGGAATTTGA
  201	AGCTGCCGCT GCTCAGGTTC GAGACAGCCT CAAAGAAACC
  	GGTACGGATA
  251	TGGAGGGTAA TCTGCACGAC ATTTCCGACG GTCTGAAGCC
  	TTGGGAAAAA
  301	CTGCCCGAAC AGCGCACGCC TGCTGATTTC GGTGTCGATG
  	AAAACGGCAA
  351	TCCCTTTCCC GATGCGGCAA ACACCCTATT AGACGGCATT
  	TCCGACGTTA
  401	TGCCGTCCGA ACGTTCCTAC GCTTCCGCCG AAACCCTTGG
  	GGACAGCGGG
  451	CAAACCGGCA GTACAGCCGA ACCCGCGGAA ACCGACCAAG
  	ACCGTGCATG
  501	GCGGGAATAC CTGACTGCTT CTGCCGCCGC ACCCGTCGTA
  	CAGACCGTCG
  551	AAGTCAGCTA TATCGATACC GCTGTTGAAA CCCCTGTTCC
  	GCATACCACT
  601	TCGCTGCGTA AACAGGCAAT AAGCCGCAAA CGCGATTTGC
  	GTCCTAAATC
  651	CCGCGCCAAA CCTAAATTGC GCGTCCGTAA ATCATAA

• This encodes a protein having amino acid sequence <SEQ ID 72; ORF2a>:

• 1	MFDFGLGELV FVGIIALIVL GPERLPEAAR TAGRLIGRLQ
  	RFVGSVKQEF
  51	DTQIELEELR KAKQEFEAAA AQVRDSLKET GTDMEGNLHD
  	ISDGLKPWEK
  101	LPEQRTPADF GVDENGNPFP DAANTLLDGI SDVMPSERSY
  	ASAETLGDSG
  151	QTGSTAEPAE TDQDRAWREY LTASAAAPVV QTVEVSYIDT
  	AVETPVPHTT
  201	SLRKQAISRK RDLRPKSRAK PKLRVRKS*

• The originally-identified partial strain B sequence (ORF2) shows 97.5% identity over a 118aa overlap with ORF2a:
• 
• The complete strain B sequence (ORF2-1) and ORF2a show 98.2% identity in 228 aa overlap:
• 
• Further work identified a partial DNA sequence <SEQ ID 73> in N. gonorrhoeae encoding the following amino acid sequence <SEQ ID 74; ORF2ng>:

• 1	MFDFGLGELI FVGIIALIVL GPERLPEAAR TAGRLIGRLQ
  	RFVGSVKQEL
  51	DTQIELEELR KVKQAFEAAA AQVRDSLKET DTDMQNSLHD
  	ISDGLKPWEK
  101	LPEQRTPADF GVDEKGNSLS RYGKHRIRRH FRRYAV*

• Further work identified the complete gonococcal gene sequence <SEQ ID 75>:

• 1	ATGTTTGATT TCGGTTTGGG CGAGCTGATT TTTGTCGGCA
  	TTATCGCCCT
  51	GATTGTCCTT GGTCCAGAAC GCCTGCCCGA AGCCGCCCGC
  	ACTGCCGGAC
  101	GGCTTATCGG CAGGCTGCAA CGCTTTGTAG GAAGCGTCAA
  	ACAAGAACTT
  151	GACACTCAAA TCGAACTGGA AGAGCTGAGG AAGGTCAAGC
  	AGGCATTCGA
  201	AGCTGCCGCC GCTCAGGTTC GAGACAGCCT CAAAGAAACC
  	GATACGGATA
  251	TGCAGAACAG TCTGCACGAC ATTTCCGACG GTCTGAAGCC
  	TTGGGAAAAA
  301	CTGCCCGAAC AGCGCACGCc tgccgatttc gGTGTCGATg
  	AAAacggcaa
  351	tccccttccc gATACGGCAA ACACCGTATC AGACGGCATT
  	TCCGACGTTA
  401	TGCCGTCTGA ACGTTCCGAT ACTtccgcCG AAACCCTTGG
  	GGACGACAGG
  451	CAAACCGGCA GTACAGCCGA ACCTGCGGAA ACCGACAAAG
  	ACCGCGCATG
  501	GCGGGAATAC CTGactgctt ctgccgccgc acctgtcgta
  	Cagagggccg
  551	tcgaagtcag ctaTATCGAT ACTGCTGTTG AAacgcctgT
  	tccgcaCacc
  601	acttccctgc gcaAACAGGC AATAAACCGC AAACGCGATT
  	TttgtccgaA
  651	ACACCGCGCc aAACCGAAat tgcgcgtcCG TAAATCATAA

• This encodes a protein having the amino acid sequence <SEQ ID 76; ORF2ng-1>:

• 1	MFDFGLGELI FVGIIALIVL GPERLPEAAR TAGRLIGRLQ
  	RFVGSVKQEL
  51	DTQIELEELR KVKQAFEAAA AQVRDSLKET DTDMQNSLHD
  	ISDGLKPWEK
  101	LPEQRTPADF GVDENGNPLP DTANTVSDGI SDVMPSERSD
  	TSAETLGDDR
  151	QTGSTAEPAE TDKDRAWREY LTASAAAPVV QRAVEVSYID
  	TAVETPVPHT
  201	TSLRKQAINR KRDFCPKHRA KPKLRVRKS*

• The originally-identified partial strain B sequence (ORF2) shows 87.5% identity over a 136aa overlap with ORF2ng:
• 
• The complete strain B and gonococcal sequences (ORF2-1 & ORF2ng-1) show 91.7% identity in 229 aa overlap:
• 
• Computer analysis of these amino acid sequences indicates a transmembrane region (underlined), and also revealed homology (59% identity) between the gonococcal sequence and the TatB protein of E. coli:

• gnl|PID|e1292181 (AJ005830) TatB protein [Escherichia coli] Length = 171
  Score = 56.6 bits (134), Expect = 1e−07
  Identities = 30/88 (34%), Positives = 52/88 (59%), Gaps = 1/88 (1%)
  Query:  1 MFDFGLGELIFVGIIALIVLGPERLPEAARTAGRLIGRLQRFVGSVKQELDTQIELEELR 60
  MFD G  EL+ V II L+VLGP+RLP A +T    I  L+     +V+ EL +++L+E  +
  Sbjct:  1 MFDIGFSELLLVFIIGLVVLGPQRLPVAVKTVAGWIRALRSLATTVQNELTQELKLQEFQ 60
  Query: 61 -KVKQAFEAAAAQVRDSLKETDTDMQNS 87
  +K+  +A+   +   LK +  +++ +
  Sbjct: 61 DSLKKVEKASLTNLTPELKASMDELRQA 88

• Based on this analysis, it was predicted that ORF2, ORF2a and ORF2ng are likely to be membrane proteins and so the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF2-1 (16 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 3A shows the results of affinity purification of the GST-fusion protein, and FIG. 3B shows the results of expression of the His-fusion in E. coli. Purified GST-fusion protein was used to immunise mice, whose sera were used for Western blots (FIG. 3C), ELISA (positive result), and FACS analysis (FIG. 3D). These experiments confirm that ORF37-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 10
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 77>:

• 1	ATGCAAGCAC GGCTGCTGAT ACCTATTCTT TTTTCAGTTT
  	TTATTTTATC
  51	CGC.TGCGGG ACACTGACAG GTATTCCATC GCATGGCGgA
  	GkTAAACgCT
  101	TTgCGGTCGA ACAAGAACTT GTGGCCGCTT CTGCCAGAGC
  	TGCCGTTAAA
  151	GACATGGATT TACAGGCATT ACACGGACGA AAAGTTGCAT
  	TGTACATTGC
  201	CACTATGGGC GACCAAGGTT CAGGcAGTTT GACAGGGGGG
  	TCGCTACTCC
  251	ATTGATGCAC kGrTwCsTGG CGAATACATA AACAGCCCTG
  	CCGTCCGTAC
  301	CGATTACACC TATCCACGTT ACGAAACCAC CGCTGAAACA
  	ACATCAGGCG
  351	GTTTGACAGG TTTAACCACT TCTTTATCTA CACTTAATGC
  	CCCTGCACTC
  401	TCTCGCACCC AATCAGACGG TAGCGGAAGT AAAAGCAGTC
  	TGGGCTTAAA
  451	TATTGGCGGG ATGGGGGATT ATCGAAATGA AACCTTGACG
  	ACTAACCCGC
  501	GCGACACTGC CTTTCTTTCC CACTTGGTAC AGACCGTATT
  	TTTCCTGCGC
  551	GGCATAGACG TTGTTTCTCC TGCCAATGCC GATACAGATG
  	TGTTTATTAA
  601	CATCGACGTA TTCGGAACGA TACGCAACAG AACCGAAATG..

• This corresponds to the amino acid sequence <SEQ ID 78; ORF15>:

• 1	MQARLLIPIL FSVFILSACG TLTGIPSHGG XKRFAVEQEL
  	VAASARAAVK
  51	DMDLQALHGR KVALYIATMG DQGSGSLTGG RYSIDAXXXG
  	EYINSPAVRT
  101	DYTYPRYETT AETTSGGLTG LTTSLSTLNA PALSRTQSDG
  	SGSKSSLGLN
  151	IGGMGDYRNE TLTTNPRDTA FLSHLVQTVF FLRGIDVVSP
  	ANADTDVFIN
  201	IDVFGTIRNR TEM..

• Further work revealed the complete nucleotide sequence <SEQ ID 79>:

• 1	ATGCAAGCAC GGCTGCTGAT ACCTATTCTT TTTTCAGTTT
  	TTATTTTATC
  51	CGCCTGCGGG ACACTGACAG GTATTCCATC GCATGGCGGA
  	GGTAAACGCT
  101	TTGCGGTCGA ACAAGAACTT GTGGCCGCTT CTGCCAGAGC
  	TGCCGTTAAA
  151	GACATGGATT TACAGGCATT ACACGGACGA AAAGTTGCAT
  	TGTACATTGC
  201	CACTATGGGC GACCAAGGTT CAGGCAGTTT GACAGGGGGT
  	CGCTACTCCA
  251	TTGATGCACT GATTCGTGGC GAATACATAA ACAGCCCTGC
  	CGTCCGTACC
  301	GATTACACCT ATCCACGTTA CGAAACCACC GCTGAAACAA
  	CATCAGGCGG
  351	TTTGACAGGT TTAACCACTT CTTTATCTAC ACTTAATGCC
  	CCTGCACTCT
  401	CTCGCACCCA ATCAGACGGT AGCGGAAGTA AAAGCAGTCT
  	GGGCTTAAAT
  451	ATTGGCGGGA TGGGGGATTA TCGAAATGAA ACCTTGACGA
  	CTAACCCGCG
  501	CGACACTGCC TTTCTTTCCC ACTTGGTACA GACCGTATTT
  	TTCCTGCGCG
  551	GCATAGACGT TGTTTCTCCT GCCAATGCCG ATACAGATGT
  	GTTTATTAAC
  601	ATCGACGTAT TCGGAACGAT ACGCAACAGA ACCGAAATGC
  	ACCTATACAA
  651	TGCCGAAACA CTGAAAGCCC AAACAAAACT GGAATATTTC
  	GCAGTAGACA
  701	GAACCAATAA AAAATTGCTC ATCAAACCAA AAACCAATGC
  	GTTTGAAGCT
  751	GCCTATAAAG AAAATTACGC ATTGTGGATG GGGCCGTATA
  	AAGTAAGCAA
  801	AGGAATTAAA CCGACGGAAG GATTAATGGT CGATTTCTCC
  	GATATCCGAC
  851	CATACGGCAA TCATACGGGT AACTCCGCCC CATCCGTAGA
  	GGCTGATAAC
  901	AGTCATGAGG GGTATGGATA CAGCGATGAA GTAGTGCGAC
  	AACATAGACA
  951	AGGACAACCT TGA

• This corresponds to the amino acid sequence <SEQ ID 80; ORF15-1>:

• 1	MQARLLIPIL FSVFILSACG TLTGIPSHGG GKRFAVEQEL
  	VAASARAAVK
  51	DMDLQALHGR KVALYIATMG DQGSGSLTGG RYSIDALIRG
  	EYINSPAVRT
  101	DYTYPRYETT AETTSGGLTG LTTSLSTLNA PALSRTQSDG
  	SGSKSSLGLN
  151	IGGMGDYRNE TLTTNPRDTA FLSHLVQTVF FLRGIDVVSP
  	ANADTDVFIN
  201	IDVFGTIRNR TEMHLYNAET LKAQTKLEYF AVDRTNKKLL
  	IKPKTNAFEA
  251	AYKENYALWM GPYKVSKGIK PTEGLMVDFS DIRPYGNHTG
  	NSAPSVEADN
  301	SHEGYGYSDE VVRQHRQGQP *

• Further work identified the corresponding gene in strain A of N. meningitidis <SEQ ID 81>:

• 1	ATGCAAGCAC GGCTGCTGAT ACCTATTCTT TTTTCAGTTT
  	TTATTTTATC
  51	CGCCTGCGGG ACACTGACAG GTATTCCATC GCATGGCGGA
  	GGTAAACGCT
  101	TTGCGGTCGA ACAAGAACTT GTGGCCGCTT CTGCCAGAGC
  	TGCCGTTAAA
  151	GACATGGATT TACAGGCATT ACACGGACGA AAAGTTGCAT
  	TGTACATTGC
  201	AACTATGGGC GACCAAGGTT CAGGCAGTTT GACAGGGGGT
  	CGCTACTCCA
  251	TTGATGCACT GATTCGTGGC GAATACATAA ACAGCCCTGC
  	CGTCCGTACC
  301	GATTACACCT ATCCACGTTA CGAAACCACC GCTGAAACAA
  	CATCAGGCGG
  351	TTTGACAGGT TTAACCACTT CTTTATCTAC ACTTAATGCC
  	CCTGCACTCT
  401	CGCGCACCCA ATCAGACGGT AGCGGAAGTA AAAGCAGTCT
  	GGGCTTAAAT
  451	ATTGGCGGGA TGGGGGATTA TCGAAATGAA ACCTTGACGA
  	CTAACCCGCG
  501	CGACACTGCC TTTCTTTCCC ACTTGGTACA GACCGTATTT
  	TTCCTGCGCG
  551	GCATAGACGT TGTTTCTCCT GCCAATGCCG ATACGGATGT
  	GTTTATTAAC
  601	ATCGACGTAT TCGGAACGAT ACGCAACAGA ACCGAAATGC
  	ACCTATACAA
  651	TGCCGAAACA CTGAAAGCCC AAACAAAACT GGAATATTTC
  	GCAGTAGACA
  701	GAACCAATAA AAAATTGCTC ATCAAACCAA AAACCAATGC
  	GTTTGAAGCT
  751	GCCTATAAAG AAAATTACGC ATTGTGGATG GGACCGTATA
  	AAGTAAGCAA
  801	AGGAATTAAA CCGACAGAAG GATTAATGGT CGATTTCTCC
  	GATATCCAAC
  851	CATACGGCAA TCATATGGGT AACTCTGCCC CATCCGTAGA
  	GGCTGATAAC
  901	AGTCATGAGG GGTATGGATA CAGCGATGAA GCAGTGCGAC
  	GACATAGACA
  951	AGGGCAACCT TGA

• This encodes a protein having amino acid sequence <SEQ ID 82; ORF15a>:

• 1	MQARLLIPIL FSVFILSACG TLTGIPSHGG GKRFAVEQEL
  	VAASARAAVK
  51	DMDLQALHGR KVALYIATMG DQGSGSLTGG RYSIDALIRG
  	EYINSPAVRT
  101	DYTYPRYETT AETTSGGLTG LTTSLSTLNA PALSRTQSDG
  	SGSKSSLGLN
  151	IGGMGDYRNE TLTTNPRDTA FLSHLVQTVF FLRGIDVVSP
  	ANADTDVFIN
  201	IDVFGTIRNR TEMHLYNAET LKAQTKLEYF AVDRTNKKLL
  	IKPKTNAFEA
  251	AYKENYALWM GPYKVSKGIK PTEGLMVDFS DIQPYGNHMG
  	NSAPSVEADN
  301	SHEGYGYSDE AVRRHRQGQP *

• The originally-identified partial strain B sequence (ORF15) shows 98.1% identity over a 213aa overlap with ORF15a:
• 
• The complete strain B sequence (ORF15-1) and ORF15a show 98.8% identity in 320 aa overlap:
• 
• Further work identified the corresponding gene in N. gonorrhoeae <SEQ ID 83>:

• 1	ATGCGGGCAC GGCTGCTGAT ACCTATTCTT TTTTCAGTTT
  	TTATTTTATC
  51	CGCCTGCGGG ACACTGACAG GTATTCCATC GCATGGCGGA
  	GGCAAACGCT
  101	TCGCGGTCGA ACAAGAACTT GTGGCCGCTT CTGCCAGAGC
  	TGCCGTTAAA
  151	GACATGGATT TACAGGCATT ACACGGACGA AAAGTTGCAT
  	TGTACATTGC
  201	AACTATGGGC GACCAAGGTT CAGGCAGTTT GACAGGGGGT
  	CGCTACTCCA
  251	TTGATGCACT GATTCGCGGC GAATACATAA ACAGCCCTGC
  	CGTCCGCACC
  301	GATTACACCT ATCCGCGTTA CGAAACCACC GCTGAAACAA
  	CATCAGGCGG
  351	TTTGACGGGT TTAACCACTT CTTTATCTAC ACTTAATGCC
  	CCTGCACTCT
  401	CGCGCACCCA ATCAGACGGT AGCGGAAGTA GGAGCAGTCT
  	GGGCTTAAAT
  451	ATTGGCGGGA TGGGGGATTA TCGAAATGAA ACCTTGACGA
  	CCAACCCGCG
  501	CGACACTGCC TTTCTTTCCC ACTTGGTGCA GACCGTATTT
  	TTCCTGCGCG
  551	GCATAGACGT TGTTTCTCCT GCCAATGCCG ATACAGATGT
  	GTTTATTAAC
  601	ATCGACGTAT TCGGAACGAT ACGCAACAGA ACCGAAATGC
  	ACCTATACAA
  651	TGCCGAAACA CTGAAAGCCC AAACAAAACT GGAATATTTC
  	GCAGTAGACA
  701	GAACCAATAA AAAATTGCTC ATCAAACCCA AAACCAATGC
  	GTTTGAAGCT
  751	GCCTATAAAG AAAATTACGC ATTGTGGATG GGGCCGTATA
  	AAGTAAGCAA
  801	AGGAATCAAA CCGACGGAAG GATTGATGGT CGATTTCTCC
  	GATATCCAAC
  851	CATACGGCAA TCATACGGGT AACTCCGCCC CATCCGTAGA
  	GGCTGATAAC
  901	AGTCATGAGG GGTATGGATA CAGCGATGAA GCAGTGCGAC
  	AACATAGACA
  951	AGGGCAACCT TGA

• This encodes a protein having amino acid sequence <SEQ ID 84; ORF15ng>:

• 1	MRARLLIPIL FSVFILSACG TLTGIPSHGG GKRFAVEQEL
  	VAASARAAVK
  51	DMDLQALHGR KVALYIATMG DQGSGSLTGG RYSIDALIRG
  	EYINSPAVRT
  101	DYTYPRYETT AETTSGGLTG LTTSLSTLNA PALSRTQSDG
  	SGSRSSLGLN
  151	IGGMGDYRNE TLTTNPRDTA FLSHLVQTVF FLRGIDVVSP
  	ANADTDVFIN
  201	IDVFGTIRNR TEMHLYNAET LKAQTKLEYF AVDRTNKKLL
  	IKPKTNAFEA
  251	AYKENYALWM GPYKVSKGIK PTEGLMVDFS DIQPYGNHTG
  	NSAPSVEADN
  301	SHEGYGYSDE AVRQHRQGQP *

• The originally-identified partial strain B sequence (ORF15) shows 97.2% identity over a 213aa overlap with ORF15ng:
• 
• The complete strain B sequence (ORF15-1) and ORF15ng show 98.8% identity in 320 aa overlap:
• 
• Computer analysis of these amino acid sequences reveals an ILSAC motif (putative membrane lipoprotein lipid attachment site, as predicted by the MOTIFS program).
• Indicates a putative leader sequence, and it was predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF15-1 (31.7 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 4A shows the results of affinity purification of the GST-fusion protein, and FIG. 4B shows the results of expression of the His-fusion in E. coli. Purified GST-fusion protein was used to immunise mice, whose sera were used for Western blot (FIG. 4C) and ELISA (positive result). These experiments confirm that ORFX-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 11
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 85>:

• 1	..GG.CAGCACA AAAAACAGGC GGTTGAACGG AAAAACCGTA
  	TTTACGATGA
  51	TGCCGGGTAT GATATTCGGC GTATTCACGG GCGCATTCTC
  	CGCAAAATAT
  101	ATCCCCGCGT TCGGGCTTCA AATTTTCTTC ATCCTGTTTT
  	TAACCGCCGT
  151	CGCATTCAAA ACACTGCATA CCGACCCTCA GACGGCATCC
  	CGCCCGCTGC
  201	CCGGACTGCC CrGACTGACT GCGGTTTCCA CACTGTTCGG
  	CACAATGTCG
  251	AGCTGGGTCG GCATAGGCGG CGGTTCACTT TCCGTCCCCT
  	TCTTAATCCA
  301	CTGCGGCTTC CCCGCCCATA AAGCCATCGG CACATCATCC
  	GGCCTTGCCT
  351	GGCCGATTGC ACTCTCCGGC GCAATATCGT ATCTGCTCAA
  	CGGCCTGAAT
  401	ATTGCAGGAT TGCCCGAAGG GTCACTGGGC TTCCTTTACC
  	TGCCCGCCGT
  451	CGCCGTCCTC AGCGCGGCAA CCATTGCCTT TGCCCCGCTC
  	GGTGTCAAAA
  501	CCGCCCACAA ACTTTCTTCT GCCAAACTCA AAAAATC.TT
  	CGGCATTATG
  551	TTGCTTTTGA TTGCCGGAAA AATGCTGTAC AACCTGCTTT
  	AA

• This corresponds to the amino acid sequence <SEQ ID 86; ORF17>:

• 1	..GQHKKQAVNG KTVFTMMPGM IFGVFTGAFS AKYIPAFGLQ
  	IFFILFLTAV
  51	AFKTLHTDPQ TASRPLPGLP XLTAVSTLFG TMSSWVGIGG
  	GSLSVPFLIH
  101	CGFPAHKAIG TSSGLAWPIA LSGAISYLLN GLNIAGLPEG
  	SLGFLYLPAV
  151	AVLSAATIAF APLGVKTAHK LSSAKLKKSF GIMLLLIAGK
  	MLYNLL*

• Further work revealed the complete nucleotide sequence <SEQ ID 87>:

• 1	ATGTGGCATT GGGACATTAT CTTAATCCTG CTTGCCGTAG
  	GCAGTGCGGC
  51	AGGTTTTATT GCCGGCCTGT TCGGCGTAGG CGGCGGCACG
  	CTGATTGTCC
  101	CTGTCGTTTT ATGGGTGCTT GATTTGCAGG GTTTGGCACA
  	ACATCCTTAC
  151	GCGCAACACC TCGCCGTCGG CACATCCTTC GCCGTCATGG
  	TCTTCACCGC
  201	CTTTTCCAGT ATGCTGGGGC AGCACAAAAA ACAGGCGGTC
  	GACTGGAAAA
  251	CCGTATTTAC GATGATGCCG GGTATGATAT TCGGCGTATT
  	CACGGGCGCA
  301	CTCTCCGCAA AATATATCCC CGCGTTCGGG CTTCAAATTT
  	TCTTCATCCT
  351	GTTTTTAACC GCCGTCGCAT TCAAAACACT GCATACCGAC
  	CCTCAGACGG
  401	CATCCCGCCC GCTGCCCGGA CTGCCCGGAC TGACTGCGGT
  	TTCCACACTG
  451	TTCGGCACAA TGTCGAGCTG GGTCGGCATA GGCGGCGGTT
  	CACTTTCCGT
  501	CCCCTTCTTA ATCCACTGCG GCTTCCCCGC CCATAAAGCC
  	ATCGGCACAT
  551	CATCCGGCCT TGCCTGGCCG ATTGCACTCT CCGGCGCAAT
  	ATCGTATCTG
  601	CTCAACGGCC TGAATATTGC AGGATTGCCC GAAGGGTCAC
  	TGGGCTTCCT
  651	TTACCTGCCC GCCGTCGCCG TCCTCAGCGC GGCAACCATT
  	GCCTTTGCCC
  701	CGCTCGGTGT CAAAACCGCC CACAAACTTT CTTCTGCCAA
  	ACTCAAAAAA
  751	Tc.TTCGGCA TTATGTTGCT TTTGATTGCC GGAAAAATGC
  	TGTACAACCT
  801	GCTTTAA

• This corresponds to the amino acid sequence <SEQ ID 88; ORF17-1>:

• 1	MWHWDIILIL LAVGSAAGFI AGLFGVGGGT LIVPVVLWVL
  	DLQGLAQHPY
  51	AQHLAVGTSF AVMVFTAFSS ML GQHKKQAV DWKTVFTMMP
  	GMIFGVFTGA
  101	LSAKYIPAFG LQIFFILFLT AVAFKTLHTD PQTASRPLPG
  	LPGLTAVSTL
  151	FGTMSSWVGI GGGSLSVPFL IHCGFPAHKA IGTSSGLAWP
  	IALSGAISYL
  201	LNGLNIAGLP EGSLGFLYLP  AVAVLSAATI AFAPLGV KTA
  	HKLSSAKLKK
  251	XFGIMLLLIA GKMLYNLL*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with Hypothetical H. influenzae Transmembrane Protein H10902 (Accession number P44070)
• ORF17 and HI0902 proteins show 28% aa identity in 192 aa overlap:

• ORF17	3	HKKQAVNGKTVFTMMPGMIFGVFT-GAFSAKYIPAFGLQIF--FILFLTAVAFKTLHTDP	59
  		HK   +  + V  + P ++  VF  G F  +       +IF   +++L      ++  D
  HI0902	72	HKLGNIVWQAVRILAPVIMLSVFICGLFIGRLDREISAKIFACLVVYLATKMVLSIKKD-	130
  ORF17	60	QTASRPLPGLPXLTAVSTLFGTMSSWVGIGGGSLSVPFLIHCGFPAHKAIGTSSGLAWPI	119
  		Q  ++ L  L  +     L G  SS  GIGGG   VPFL   G    +AIG+S+     +
  HI0902	131	QVTTKSLTPLSSVIG-GILIGMASSAAGIGGGGFIVPFLTARGINIKQAIGSSAFCGMLL	189
  ORF17
  	120	ALSGAISYLLNGLNIAGLPEGSLGFLYLPAVAVLSAATIAFAPLGVXXXXXXXXXXXXXX	179
  		+SG  S++++G     +PE SLG++YLPAV  ++A +   + LG
  HI0902	190	GISGMFSFIVSGWGNPLMPEYSLGYIYLPAVLGITATSFFTSKLGASATAKLPVSTLKKG	249
  ORF17	180	FGIMLLLIAGKM	191
  		F + L+++A  M
  HI0902
  	250	FALFLIVVAINM	261

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF17 shows 96.9% identity over a 196aa overlap with an ORF (ORF17a) from strain A of N. meningitidis:
• 
• The complete length ORF17a nucleotide sequence <SEQ ID 89> is:

• 1	ATGTGGCATT GGGACATTAT CTTAATCCTG CTTGCCGTAG
  	GCAGTGCGGC
  51	AGGTTTTATT GCCGGCCTGT TCGGCGTAGG CGGCGGCACG
  	CTGATTGTCC
  101	CTGTCGTTTT ATGGGTGCTT GATTTGCAGG GTTTGGCACA
  	ACATCCTTAC
  151	GCGCAACACC TCGCCGTCGG CACATCCTTC GCCGTCATGG
  	TCTTCACCGC
  201	CTTTTCCAGT ATGCTGGGGC AGCACAAAAA ACAGGCGGTC
  	GACTGGAAAA
  251	CCGTATTTAC GATGATGCCG GGTATGGTAT TCGGCGTATT
  	CGCTGGCGCA
  301	CTCTCCGCAA AATATATCCC AGCGTTCGGG CTTCAAATTT
  	TCTTCATCCT
  351	GTTTTTAACC GCCGTCGCAT TCAAAACACT GCATACCGAC
  	CCTCAGACGG
  401	CATCCCGCCC GCTGCCCGGA CTGCCCGGAC TGACTGCGGT
  	TTCCACACTG
  451	TTCGGCACAA TGTCGAGCTG GGTCGGCATA GGCGGCGGTT
  	CACTTTCCGT
  501	CCCCTTCTTA ATCCACTGCG GCTTCCCCGC CCATAAAGCC
  	ATCGGCACAT
  551	CATCCGGCCT TGCCTGGCCG ATTGCACTCT CCGGCGCAAT
  	ATCGTATCTG
  601	CTCAACGGCC TGAATATTGC AGGATTGCCC GAAGGGTCAC
  	TGGGCTTCCT
  651	TTACCTGCCC GCCGTCGCCG TCCTCAGCGC GGCAACCATT
  	GCCTTTGCCC
  701	CGCTCGGTGT CAAAACCGCC CACAAACTTT CTTCTGCCAA
  	ACTCAAAAAA
  751	TCCTTCGGCA TTATGTTGCT TTTGATTGCC GGAAAAATGC
  	TGTACAACCT
  801	GCTTTAA

• This encodes a protein having amino acid sequence <SEQ ID 90>:

• 1	MWHWDIILIL LAVGSAAGFI AGLFGVGGGT LIVPVVLWVL
  	DLQGLAQHPY
  51	AQHLAVGTSF AVMVFTAFSS ML GQHKKQAV DWKTVFTMMP
  	GMVFGVFAGA
  101	LSAKYIPAFG LQIFFILFLT AVAFKTLHTD PQTASRPLPG
  	LPGLTAVSTL
  151	FGTMSSWVGI GGGSLSVPFL IHCGFPAHKA IGTSSGLAWP
  	IALSGAISYL
  201	LNGLNIAGLP EGSLGFLYLP  AVAVLSAATI AFAPLGV KTA
  	HKLSSAKLKK
  251	SFGIMLLLIA GKMLYNLL*

• ORF17a and ORF17-1 show 98.9% identity in 268 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF17 shows 93.9% identity over a 196aa overlap with a predicted ORF (ORF17.ng) from N. gonorrhoeae:
• 
• An ORF17ng nucleotide sequence <SEQ ID 91> is predicted to encode a protein having amino acid sequence <SEQ ID 92>:

• 1	MWHWDIILIL LAVGSAAGFI AGLFGVGGGT LIVPVVLWVL
  	DLQGLAQHPY
  51	AQHLAVGTSF AVMVFTAFSS MLGQHKKQAV DWKTIFAMMP
  	GMIFGVFAGA
  101	LSAKYIPAFG LQIFFILFLT AVAFKTLHTG RQTASRPLPG
  	LPGLTAVSTL
  151	FGAMSSWVGI GGGSLSVPFL IHCGFPAHKA IGTSSGLAWP
  	IALSGAISYL
  201	VNGLNIAGLP EGSLGFLYLP AVAVLSAATI AFAPLGVKTA
  	HKLSSAKLKE
  251	SFGIMLLLIA GKMLYNLL*

• Further work revealed the complete gonococcal DNA sequence <SEQ ID 93>:

• 1	ATGTGGCATT GGGACATTAT CTTAATCCTG CTTGCcgtag
  	gcAGTGCGGC
  51	AGGTTTTATT GCCGGCCTGT Tcggtgtagg cggcgGTACG
  	CTGATTGTCC
  101	CTGTCGTTTT ATGGGTGCTT GATTTGCAGG GTTTGGCACA
  	ACATCCTTAC
  151	GCGCAACACC TCGCCGTCGG CAcaTccttc gcCGTCATGG
  	TCTTCACCGC
  201	CTTTTCCAGT ATGTTGGGGC AGCACAAAAA ACAGGCGGTC
  	GACTGGAAAA
  251	CCATATTTGC GATGATGCCG GGTATGATAT TCGGCGTATT
  	CGCTGGCGCA
  301	CTCTCCGCAA AATATATCCC CGCGTTCGGG CTTCAAATTT
  	TCTTCATCCT
  351	GTTTTTAACC GCCGTCGCAT TCAAAACACT GCATACCGGT
  	CGTCAGACGG
  401	CATCCCGCCC GCTGCCCGGG CTGCCCGGAC TGACTGCGGT
  	TTCCACACTG
  451	TTCGGCGCAA TGTCGAGCTG GGTCGGCATA GGCGGCGGTT
  	CACTTTCCGT
  501	CCCCTTCTTA ATCCACTGCG GCTTCCCCGC CCATAAAGCC
  	ATCGGCACAT
  551	CATCCGGCCT TGCCTGGCCG ATTGCACTCT CCGGCGCAAT
  	ATCGTATCTG
  601	GTCAACGGTC TGAATATTGC AGGATTGCCC GAAGGGTCGC
  	TGGGCTTCCT
  651	TTACCTGCCC GCCGTCGCCG TCCTCAGCGC GGCAACCATT
  	GCCTTTGCCC
  701	CGCTCGGTGT CAAAACCGCC CACAAACTTT CTTCTGCCAA
  	ACTCAAAGAA
  751	TCCTTCGGCA TTATGTTGCT TTTGATTGCC GGAAAAATGC
  	TGTACAACCT
  801	GCTTTAA

• This corresponds to the amino acid sequence <SEQ ID 94; ORF17ng-1>:

• 1	MWHWDIILIL LAVGSAAGFI AGLFGVGGGT LIVPVVLWVL
  	DLQGLAQHPY
  51	AQHLAVGTSF AVMVFTAFSS ML GQHKKQAV DWKTIFAMMP
  	GMIFGVFAGA
  101	LSAKYIPAFG LQIFFILFLT AVAFKTLHTG RQTASRPLPG
  	LPGLTAVSTL
  151	FGAMSSWVGI GGGSLSVPFL IHCGFPAHKA IGTSSGLAWP
  	IALSGAISYL
  201	VNGLNIAGLP EGSLGFLYLP  AVAVLSAATI AFAPLGV KTA
  	HKLSSAKLKE
  251	SFGIMLLLIA GKMLYNLL*

• ORF17ng-1 and ORF17-1 show 96.6% identity in 268 aa overlap:
• 
• In addition, ORF17ng-1 shows significant homology with a hypothetical H. influenzae protein:

• sp|P44070|Y902_HAEIN HYPOTHETICAL PROTEIN HI0902 pir||G64015
  hypothetical protein HI0902 - Haemophilus influenzae (strain Rd KW20)
  gi|1573922 (U32772) H. influenzae
  predicted coding region HI0902 [Haemophilus influenzae]Length = 264
  Score = 74 (34.9 bits), Expect = 1.6e−23, Sum P(2) = 1.6e−23
  Identities = 15/43 (34%), Positives = 23/43 (53%)

  Query:	55	AVGTSFAVMVFTAFSSMLGQHKKQAVDWKTIFAMMPGMIFGVF	97
  		A+GTSFA +V T   S    HK   + W+ +  + P ++  VF
  Sbjct:	52	ALGTSFATIVITGIGSAQRHHKLGNIVWQAVRILAPVIMLSVF	94

  Score = 195 (91.9 bits), Expect = 1.6e−23, Sum P(2) = 1.6e−23
  Identities = 44/114 (38%), Positives = 65/114 (57%)

  Query:	150	LFGAMSSWVGIGGGSLSVPFLIHCGFPAHKAIGTSSGLAWPIALSGAISYLVNGLNIAGL	209
  		L G  SS  GIGGG   VPFL   G    +AIG+S+     + +SG  S++V+G     +
  Sbjct:	148	LIGMASSAAGIGGGGFIVPFLTARGINIKQAIGSSAFCGMLLGISGMFSFIVSGWGNPLM	207
  Query:	210	PEGSLGFLYLPAVAVLSAATIAFAPLGVKTAFIKLSSAKLKESFGIMLLLIAGKM	263
  		PE SLG++YLPAV  ++A +   + LG     KL  + LK+ F + L+++A  M
  Sbjct:	208	PEYSLGYIYLPAVLGITATSFFTSKLGASATAKLPVSTLKKGFALFLIVVAINM	261

• This analysis, including the homology with the hypothetical H. influenzae transmembrane protein, suggests that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 12
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 95>:

• 1	..GGAAACGGAT GGCAGGCAGA CCCCGAACAT CCGCTGCTCG
  	GGCTTTTTGC
  51	CGTCAGTAAT GTATCGATGA CGCTTGCTTT TGTCGGAATA
  	TGTGCGTTGG
  101	TGCATTATTG CTTTTCGGGA ACGGTTCAAG TGTTTGTGTT
  	TGCGGCACTG
  151	CTCAAACTTT ATGCGCTGAA GCCGGTTTAT TGGTTCGTGT
  	TGCAGTTTGT
  201	GCTGATGGCG GTTGCCTATG TCCACCGCTG CGGTATAGAC
  	CGGCAGCCGC
  251	CGTCAACGTT CGGCGGCTCG CAGCTGCGAC TCGGCGGGTT
  	GACGGCAGCG
  301	TTGATGCAGG TCTCGGTACT GGTGCTGCTG CTTTCAGAAA
  	TTGGAAGATA
  351	A

• This corresponds to the amino acid sequence <SEQ ID 96; ORF18>:

• 1	..GNGWQADPEH PLLGLFAVSN VSMTLAFVGI CALVHYCFSG
  	TVQVFVFAAL
  51	LKLYALKPVY WFVLQFVLMA VAYVHRCGID RQPPSTFGGS
  	QLRLGGLTAA
  101	LMQVSVLVLL LSEIGR*

• Further work revealed the complete nucleotide sequence <SEQ ID 97>:

• 1	ATGATTTTGC TGCATTTGGA TTTTTTGTCT GCCTTACTGT
  	ATGCGGCGGT
  51	TTTTCTGTTT CTGATATTCC GCGCAGGAAT GTTGCAATGG
  	TTTTGGGCGA
  101	GTATTATGCT GTGGCTGGGC ATATCGGTTT TGGGGGCAAA
  	GCTGATGCCC
  151	GGCATATGGG GAATGACCCG CGCCGCGCCC TTGTTCATCC
  	CCCATTTTTA
  201	CCTGACTTTG GGCAGCATAT TTTTTTTCAT CGGGCATTGG
  	AACCGGAAAA
  251	CAGATGGAAA CGGATGGCAG GCAGACCCCG AACATCCGCT
  	GCTCGGGCTT
  301	TTTGCCGTCA GTAATGTATC GATGACGCTT GCTTTTGTCG
  	GAATATGTGC
  351	GTTGGTGCAT TATTGCTTTT CGGGAACGGT TCAAGTGTTT
  	GTGTTTGCGG
  401	CACTGCTCAA ACTTTATGCG CTGAAGCCGG TTTATTGGTT
  	CGTGTTGCAG
  451	TTTGTGCTGA TGGCGGTTGC CTATGTCCAC CGCTGCGGTA
  	TAGACCGGCA
  501	GCCGCCGTCA ACGTTCGGCG GCTCGCAGCT GCGACTCGGC
  	GGGTTGACGG
  551	CAGCGTTGAT GCAGGTCTCG GTACTGGTGC TGCTGCTTTC
  	AGAAATTGGA
  601	AGATAA

• This corresponds to the amino acid sequence <SEQ ID 98; ORF18-1>:

• 1	MILLHLDFLS ALLYAAVFLF LIFRAGMLQW FWASIMLWLG
  	ISVLGAKLMP
  51	GIWGMTRAAP LFIPHFYLTL GSIFFFI GHW NRKTDGNGWQ
  	ADPEHPLLGL
  101	FAVSNVSMTL AFVGICALVH YCFSGTVQVF VFAALLKLYA
  	LKPVYWFVLQ
  151	FVLMAVAYVH RCGIDRQPPS TFGGSQLRLG GLTAALMQVS
  	VLVLLLSEIG
  201	R*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF18 shows 98.3% identity over a 116aa overlap with an ORF (ORF18a) from strain A of N. meningitidis:
• 
• The complete length ORF18a nucleotide sequence <SEQ ID 99> is:

• 1	ATGATTTTGC TGCATTTGGA TTTTTTGTCT GCCTTACTGT
  	ATGCGGCGGT
  51	TTTTCTGTTT CTGATATTCC GCGCAGGAAT GTTGCAATGG
  	TTTTGGGCGA
  101	GTATTATGCT GTGGCTGGGC ATATCGGTTT TGGGGGCAAA
  	GCTGATGCCC
  151	GGCATATGGG GAATGACCCG CGCCGCGCCC TTGTTCATCC
  	CCCATTTTTA
  201	CCTGACTTTG GGCAGCATAT TTTTTTTCAT CGGGCATTGG
  	AACCGGAAAA
  251	CGGATGGAAA CGGATGGCAG GCAGACCCCG AACATCCTCT
  	GCTCGGGCTG
  301	TTTGCCGTCA GTAATGTATC GATGACGCTT GCTTTTGTCG
  	GAATATGTGC
  351	GTTGGTGCAT TATTGCTTTT CGNGAACGGT TCAAGTGTTT
  	GTGTTTGCGG
  401	CACTGCTCAA ACTTTATGCG CTGAAGCCGG TTTATTGGTT
  	CGTGTTGCAG
  451	TTTGTGCTGA TGGCGGTTGC CTATGTCCAC CGCTGCGGTA
  	TAGACCGGCA
  501	GCCGCCGTCA ACGTTCGGCG GNTCGCAGCT GCGACTCGGC
  	GGGTTGACGG
  551	CAGCGTTGAT GCAGNTCTCG GTACTGGTGC TGCTGCTTTC
  	AGAAATTGGA
  601	AGATAA

• This encodes a protein having amino acid sequence <SEQ ID 100>:

• 1	MILLHLDFLS ALLYAAVFLF LIFRAGMLQW FWASIMLWLG
  	ISVLGAKLMP
  51	GIWGMTRAAP LFIPHFYLTL GSIFFFI GHW NRKTDGNGWQ
  	ADPEHPLLGL
  101	FAVSNVSMTL AFVGICALVH YCFSXTVQVF VFAALLKLYA
  	LKPVYWFVLQ
  151	FVLMAVAYVH RCGIDRQPPS TFGGSQLRLG GLTAALMQXS
  	VLVLLLSEIG
  201	R*

• ORF18a and ORF18-1 show 99.0% identity in 201 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF18 shows 93.1% identity over a 116aa overlap with a predicted ORF (ORF18.ng) from N. gonorrhoeae:
• 
• The complete length ORF18ng nucleotide sequence is <SEQ ID 101>:

• 1	ATGATTTTGC TGCATTTGGA TTTTTTGTCT GCCTTACTGt
  	aTGCGGcggt
  51	tttTctgTTT CTGATATTCC GCGCAGGAAT GTTGCAATGG
  	TTTTGGGCGA
  101	GTATTGCGTT GTGGCTCGGC ATCTCGGTTT TAGGGGTAAA
  	GCTGATGCCG
  151	GGGATGTGGG GAATGACCCG CGCCGCGCCT TTGTTCATCC
  	CCCATTTTTA
  201	CCTGACTTTG GGCAGCATAT TTTTTTTCAT CGGGTATTGG
  	AACCGGAAAA
  251	CAGATGGAAA CGGATGGCAG GCAGACCCCG AACATCCGCT
  	GCTCGGGCTT
  301	TTTGCCGTCA GTAATGTATC GATGACGCTT GCTTTTGTCG
  	GAATATGTGC
  351	GTTGGTGCAT TATTGCTTTT CGGGAACGGT TCAAGTGTTT
  	GTGTTTGCGG
  401	CATTGCTCAA ACTTTATGCG CTGAAGCCGG TTTATTGGTT
  	CGTGTTGCAG
  451	TTTGTATTGA TGGCGGttgC CTATGTCCAC CGCTGCGGTA
  	TAGACCGGCA
  501	GCCGCCGTCA ACGTTCGGCG GTTCGCAGCT GCGACTCGGC
  	GTGTTGGCGG
  551	CGATGTTGAT GCAGGTTGCG GTAACGGCGA TGCTGCTTGC
  	CGAAATCGGC
  601	AGATGA

• This encodes a protein having amino acid sequence <SEQ ID 102>:

• 1	MILLHLDFLS ALLYAAVFLF LIFRAGMLQW FWASIALWLG
  	ISVLGVKLMP
  51	GMWGMTRAAP LFIPHFYLTL GSIFFFI GYW NRKTDGNGWQ
  	ADPEHPLLGL
  101	FAVSNVSMTL AFVGICALVH YCFSGTVQVF VFAALLKLYA
  	LKPVYWFVLQ
  151	FVLMAVAYVH RCGIDRQPPS TFGGSQLRLG VLAAMLMQVA
  	VTAMLLAEIG
  201	R*

• This ORF18ng protein sequence shows 94.0% identity in 201 aa overlap with ORF18-1:
• 
• Based on this analysis, including the presence of several putative transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 13
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 103>:

• 1	ATGAAAACCC CACTCCTCAA GCCTCTGCTN ATTACCTCGC
  	TTCCCGTTTT
  51	CGCCAGTGTT TTTACCGCCG CCTCCATCGT CTGGCAGCTA
  	GGCGAACCCA
  101	AGCTCGCCAT GCCCTTCGTA CTCGGCATCA TCGCCGGCGG
  	CCTTGTCGAT
  151	TTGGACAACC NCNTGACCGG ACGGCTNAAA AACATCATCA
  	CCACCGTCGC
  201	CCTGTTCACC CTCTCCTCGC TCACGGCACA AAGCACCCTC
  	GGCACAGGGC
  251	TGCCCTTCAT CCTCGCCATG ACCCTGATGA CTT.CG.CTT
  	CACCATTTTA
  301	GGCGCGGNCG ...

• This corresponds to the amino acid sequence <SEQ ID 104; ORF 19>:

• 1	MKTPLLKPLL ITSLPVFASV FTAASIVWQL GEPKLAMPFV
  	LGIIAGGLVD
  51	LDNXXTGRLK NIITTVALFT LSSLTAQSTL GTGLPFILAM
  	TLMTXXFTIL
  101	GAX...

• Further work revealed the complete nucleotide sequence <SEQ ID 105>:

• 1	ATGAAAACCC CACTCCTCAA GCCTCTGCTC ATTACCTCGC
  	TTCCCGTTTT
  51	CGCCAGTGTT TTTACCGCCG CCTCCATCGT CTGGCAGCTA
  	GGCGAACCCA
  101	AGCTCGCCAT GCCCTTCGTA CTCGGCATCA TCGCCGGCGG
  	CCTTGTCGAT
  151	TTGGACAACC GCCTGACCGG ACGGCTGAAA AACATCATCA
  	CCACCGTCGC
  201	CCTGTTCACC CTCTCCTCGC TCACGGCACA AAGCACCCTC
  	GGCACAGGGC
  251	TGCCCTTCAT CCTCGCCATG ACCCTGATGA CCTTCGGCTT
  	CACCATTTTA
  301	GGCGCGGTCG GGCTCAAATA CCGCACCTTC GCCTTCGGTG
  	CACTCGCCGT
  351	CGCCACCTAC ACCACACTTA CCTACACCCC CGAAACCTAC
  	TGGCTGACCA
  401	ACCCCTTCAT GATTTTATGC GGCACCGTAC TGTACAGCAC
  	CGCCATCCTC
  451	CTGTTCCAAA TCGTCCTGCC CCACCGCCCC GTCCAAGAAA
  	GCGTCGCCAA
  501	CGCCTACGAC GCACTCGGCG GCTACCTCGA AGCCAAAGCC
  	GACTTCTTCG
  551	ACCCCGATGA GGCAGCCTGG ATAGGCAACC GCCACATCGA
  	CCTCGCCATG
  601	AGCAACACCG GCGTCATCAC CGCCTTCAAC CAATGCCGTT
  	CCGCCCTGTT
  651	TTACCGCCTT CGCGGCAAAC ACCGCCACCC GCGCACCGCC
  	AAAATGCTGC
  701	GTTACTACTT TGCCGCCCAA GACATACACG AACGCATCAG
  	CTCCGCCCAC
  751	GTCGATTATC AGGAAATGTC CGAAAAATTC AAAAACACCG
  	ACATCATCTT
  801	CCGCATCCAC CGCCTGCTCG AAATGCAGGG ACAAGCCTGC
  	CGCAACACCG
  851	CCCAAGCCCT GCGCGCAAGC AAAGACTACG TTTACAGCAA
  	ACGCCTCGGC
  901	CGCGCCATCG AAGGCTGCCG CCAATCGCTG CGCCTCCTTT
  	CAGACAGCAA
  951	CGACAGTCCC GACATCCGCC ACCTGCGCCG CCTTCTCGAC
  	AACCTCGGCA
  1001	GCGTCGACCA GCAGTTCCGC CAACTCCAGC ACAACGGCCT
  	GCAGGCAGAA
  1051	AACGACCGCA TGGGCGACAC CCGCATCGCC GCCCTCGAAA
  	CCAGCAGCCT
  1101	CAAAAACACC TGGCAGGCAA TCCGTCCGCA GCTAAACCTC
  	GAATCAGGCG
  1151	TATTCCGCCA TGCCGTCCGC CTGTCCCTCG TCGTTGCCGC
  	CGCCTGCACC
  1201	ATCGTCGAAG CCCTCAACCT CAACCTCGGC TACTGGATAC
  	TACTGACCGC
  1251	CCTTTTCGTC TGCCAACCCA ACTACACCGC CACCAAAAGC
  	CGCGTCCGCC
  1301	AGCGCATCGC CGGCACCGTA CTCGGCGTAA TCGTCGGCTC
  	GCTCGTCCCC
  1351	TACTTCACCC CGTCTGTCGA AACCAAACTC TGGATTGTCA
  	TCGCCAGTAC
  1401	CACCCTCTTT TTCATGACCC GCACCTACAA ATACAGTTTC
  	TCCACCTTCT
  1451	TCATTACCAT TCAAGCCCTG ACCAGCCTCT CCCTCGCAGG
  	TTTGGACGTA
  1501	TACGCCGCCA TGCCCGTACG CATCATCGAC ACCATTATCG
  	GCGCATCCCT
  1551	TGCCTGGGCG GCAGTCAGCT ACCTGTGGCC AGACTGGAAA
  	TACCTCACGC
  1601	TCGAACGCAC CGCCGCCCTT GCCGTATGCA GCAACGGTGC
  	CTATCTCGAA
  1651	AAAATCACCG AACGCCTCAA AAGCGGCGAA ACCGGCGACG
  	ACGTCGAATA
  1701	CCGCGCCACC CGCCGCCGCG CCCACGAACA CACCGCCGCC
  	CTCAGCAGCA
  1751	CCCTTTCCGA CATGAGCAGC GAACCCGCAA AATTCGCCGA
  	CAGCCTGCAA
  1801	CCCGGCTTTA CCCTGCTCAA AACCGGCTAC GCCCTGACCG
  	GCTACATCTC
  1851	CGCCCTCGGC GCATACCGCA GCGAAATGCA CGAAGAATGC
  	AGCCCCGACT
  1901	TTACCGCACA GTTCCACCTC GCCGCCGAAC ACACCGCCCA
  	CATCTTCCAA
  1951	CACCTGCCCG AAACCGAACC CGACGACTTT CAGACAGCAC
  	TGGATACACT
  2001	GCGCGGCGAA CTCGACACCC TCCGCACCCA CAGCAGCGGA
  	ACACAAAGCC
  2051	ACATCCTCCT CCAACAGCTC CAACTCATCG CCCGACAGCT
  	CGAACCCTAC
  2101	TACCGCGCCT ACCGCCAAAT TCCGCACAGG CAGCCCCAAA
  	ATGCAGCCTG
  2151	A

• This corresponds to the amino acid sequence <SEQ ID 106; ORF19-1>:

• 1	MKTPLLKPLL ITSLPVFASV FTAASIVWQL GEPKLAMPFV
  	LGIIAGGLVD
  51	LDNRLTGRLK NIITTVALFT LSSLTAQSTL GTGLPFILAM
  	TLMTFGFTIL
  101	GAVGLKYRTF AFGALAVATY TTLTYTPETY WLTNPFMILC
  	GTVLYSTAIL
  151	LFQIVLPHRP VQESVANAYD ALGGYLEAKA DFFDPDEAAW
  	IGNRHIDLAM
  201	SNTGVITAFN QCRSALFYRL RGKHRHPRTA KMLRYYFAAQ
  	DIHERISSAH
  251	VDYQEMSEKF KNTDIIFRIH RLLEMQGQAC RNTAQALRAS
  	KDYVYSKRLG
  301	RAIEGCRQSL RLLSDSNDSP DIRHLRRLLD NLGSVDQQFR
  	QLQHNGLQAE
  351	NDRMGDTRIA ALETSSLKNT WQAIRPQLNL ESGVFRHAVR
  	LSLVVAAACT
  401	IVEALNLNLG YWILLTALFV CQPNYTATKS RVRQRIAGTV
  	LGVIVGSLVP
  451	YFTPSVETKL WIVIASTTLF FMTRTYKYSF STFFITIQAL
  	TSLSLAGLDV
  501	YAAMPVRIID TIIGASLAWA AVSYLWPDWK YLTLERTAAL
  	AVCSNGAYLE
  551	KITERLKSGE TGDDVEYRAT RRRAHEHTAA LSSTLSDMSS
  	EPAKFADSLQ
  601	PGFTLLKTGY ALTGYISALG AYRSEMHEEC SPDFTAQFHL
  	AAEHTAHIFQ
  651	HLPETEPDDF QTALDTLRGE LDTLRTHSSG TQSHILLQQL
  	QLIARQLEPY
  701	YRAYRQIPHR QPQNAA*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with Predicted Transmenbrane protein YHFK of H. influenzae (Accession Number P44289)
• ORF19 and YHFK proteins show 45% aa identity in 97 aa overlap:

• orf19	6	LKPLLITSLPVFASVFTAASIVWQLGEPKLAMPFVLGIIAGGLVDLDNXXTGRLKNIITT	65
  		L   +I+++PVF +V  AA  +W       +MP +LGIIAGGLVDLDN  TGRLKN+  T
  YHFK	5	LNAKVISTIPVFIAVNIAAVGIWFFDISSQSMPLILGIIAGGLVDLDNRLTGRLKNVFFT	64
  orf19	66	VALFTLSSLTAQSTLGTGLPFILAMTLMTXXFTILGA	102
  		+  F++SS   Q  +G  + +I+ MT++T  FT++GA
  YHFK	65	LIAFSISSFIVQLHIGKPIQYIVLMTVLTFIFTMIGA	101

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF19 shows 92.2% identity over a 102aa overlap with an ORF (ORF19a) from strain A of N. meningitidis:
• 
• The complete length ORF19a nucleotide sequence <SEQ ID 107> is:

• 1	ATGAAAACCC CACCCCTCAA GCCTCTGCTC ATTACCTCGC
  	TTCCCGTTTT
  51	CGCCAGTGTC TTTACCGCCG CCTCCATCGT CTGGCAGCTG
  	GGCGAACCCA
  101	AGCTCGCCAT GCCCTTCGTA CTCGGCATCA TCGCTGGCGG
  	CCTGGTCGAT
  151	TTGGACAACC GCCTGACCGG ACGGCTGAAA AACATCATCG
  	CCACCGTCGC
  201	CCTGTTCACC CTCTCCTCAC TTGTCGCGCA AAGCACCCTC
  	GGCACAGGTT
  251	TGCCATTCAT CCTCGCCATG ACCCTGATGA CTTTCGGCTT
  	TACCATCATG
  301	GGCGCGGTCG GGCTGAAATA CCGCACCTTC GCCTTCGGCG
  	CACTCGCCGT
  351	CGCCACCTAC ACCACACTTA CCTACACCCC CGAAACCTAC
  	TGGCTGACCA
  401	ACCCCTTTAT GATTCTGTGC GGAACCGTAC TGTACAGCAC
  	CGCCATCATC
  451	CTGTTCCAAA TCATCCTGCC CCACCGCCCC GTTCAAGAAA
  	ACGTCGCCAA
  501	CGCCTACGAA GCACTCGGCA GCTACCTCGA AGCCAAAGCC
  	GACTTTTTCG
  551	ATCCCGACGA AGCCGAATGG ATAGGCAACC GCCACATCGA
  	CCTCGCCATG
  601	AGCAACACCG GCGTCATCAC CGCCTTCAAC CAATGCCGTT
  	CCGCCCTGTT
  651	TTACCGCCTT CGCGGCAAAC ACCGCCACCC GCGCACCGCC
  	AAAATGCTGC
  701	GCTACTACTT CGCCGCCCAA GACATACACG AACGCATCAG
  	CTCCGCCCAC
  751	GTCGACTACC AAGAGATGTC CGAAAAATTC AAAAACACCG
  	ACATCATCTT
  801	CCGCATCCAC CGCCTGCTCG AAATGCAGGG ACAAGCCTGC
  	CGCAACACCG
  851	CCCAAGCCCT GCGCGCAAGC AAAGACTACG TTTACAGCAA
  	ACGCCTCGGC
  901	CGCGCCATCG AAGGCTGCCG CCAATCGCTG CGCCTCCTTT
  	CAGACAGCAA
  951	CGACAATCCC GACATCCGCC ACCTGCGCCG CCTTCTCGAC
  	AACCTCGGCA
  1001	GCGTCGACCA GCAGTTCCGC CAACTCCAGC ACAACGGCCT
  	GCAGGCAGAA
  1051	AACGACCGCA TGGGCGACAC CCGCATCGCC GCCCTCGAAA
  	CCGGCAGCCT
  1101	CAAAAACACC TGGCAGGCAA TCCGTCCGCA GCTAAACCTC
  	GAATCAGGCG
  1151	TATTCCGCCA TGCCGTCCGC CTGTCCCTTG TCGTTGCCGC
  	CGCCTGCACC
  1201	ATCGTCGAAG CCCTCAACCT CAACCTCGGC TACTGGATAC
  	TACTGACCGC
  1251	CCTTTTCGTC TGCCAACCCA ACTACACCGC CACCAAAAGC
  	CGCGTCCGCC
  1301	AGCGCATCGC CGGCACCGTA CTCGGCGTAA TCGTCGGCTC
  	GCTCGTCCCC
  1351	TACTTTACCC CCTCCGTCGA AACCAAACTC TGGATCGTCA
  	TCGCCAGTAC
  1401	CACCCTCTTT TTCATGACCC GCACCTACAA ATACAGCTTC
  	TCGACATTTT
  1451	TCATCACCAT TCAAGCCCTG ACCAGCCTCT CCCTCGCAGG
  	GTTGGACGTA
  1501	TACGCCGCCA TGCCCGTACG CATCATCGAC ACCATTATCG
  	GCGCATCCCT
  1551	TGCCTGGGCG GCAGTCAGCT ACCTGTGGCC AGACTGGAAA
  	TACCTCACGC
  1601	TCGAACGCAC CGCCGCCCTT GCCGTATGCA GCAACGGCGC
  	CTATCTCGAA
  1651	AAAATCACCG AACGCCTCAA AAGCGGCGAA ACCGGCGACG
  	ACGTCGAATA
  1701	CCGCGCCACC CGCCGCCGCG CCCACGAACA CACCGCCGCC
  	CTCAGCAGCA
  1751	CCCTTTCCGA CATGAGCAGC GAACCCGCAA AATTCGCCGA
  	CAGCCTGCAA
  1801	CCCGGCTTTA CCCTGCTCAA AACCGGCTAC GCCCTGACCG
  	GCTACATCTC
  1851	CGCCCTCGGC GCATACCGCA GCGAAATGCA CGAAGAATGC
  	AGCCCCGACT
  1901	TTACCGCACA GTTCCACCTC GCCGCCGAAC ACACCGCCCA
  	CATCTTCCAA
  1951	CACCTGCCCG AAACCGAACC CGACGACTTT CAGACAGCAC
  	TGGATACACT
  2001	GCGCGGCGAA CTCGACACCC TCCGCACCCA CAGCAGCGGA
  	ACACAAAGCC
  2051	ACATCCTCCT CCAACAGCTC CAACTCATCG CCCGGCAGCT
  	CGAACCCTAC
  2101	TACCGCGCCT ACCGACAAAT TCCGCACAGG CAGCCCCAAA
  	ACGCAGCCTG
  2151	A

• This encodes a protein having amino acid sequence <SEQ ID 108>:

• 1	MKTPPLKPLL ITSLPVFASV FTAASIVWQL GEPKLAMPFV
  	LGIIAGGLVD
  51	LDNRLTGRLK NIIATVALFT LSSLVAQSTL GTGLPFILAM
  	TLMTFGFTIM
  101	GAVGLKYRTF AFGALAVATY TTLTYTPETY WLTNPFMILC
  	GTVLYSTAII
  151	LFQIILPHRP VQENVANAYE ALGSYLEAKA DFFDPDEAEW
  	IGNRHIDLAM
  201	SNTGVITAFN QCRSALFYRL RGKHRHPRTA KMLRYYFAAQ
  	DIHERISSAH
  251	VDYQEMSEKF KNTDIIFRIH RLLEMQGQAC RNTAQALRAS
  	KDYVYSKRLG
  301	RAIEGCRQSL RLLSDSNDNP DIRHLRRLLD NLGSVDQQFR
  	QLQHNGLQAE
  351	NDRMGDTRIA ALETGSLKNT WQAIRPQLNL ESGVFRHAVR
  	LSLVVAAACT
  401	IVEALNLNLG YWILLTALFV CQPNYTATKS RVRQRIAGTV
  	LGVIVGSLVP
  451	YFTPSVETKL WIVIASTTLF FMTRTYKYSF STFFITIQAL
  	TSLSLAGLDV
  501	YAAMPVRIID TIIGASLAWA AVSYLWPDWK YLTLERTAAL
  	AVCSNGAYLE
  551	KITERLKSGE TGDDVEYRAT RRRAHEHTAA LSSTLSDMSS
  	EPAKFADSLQ
  601	PGFTLLKTGY ALTGYISALG AYRSEMHEEC SPDFTAQFHL
  	AAEHTAHIFQ
  651	HLPETEPDDF QTALDTLRGE LDTLRTHSSG TQSHILLQQL
  	QLIARQLEPY
  701	YRAYRQIPHR QPQNAA*

• ORF19a and ORF19-1 show 98.3% identity in 716 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF19 shows 95.1% identity over a 102aa overlap with a predicted ORF (ORF19.ng) from N. gonorrhoeae:
• 
• An ORF19ng nucleotide sequence <SEQ ID 109> is predicted to encode a protein having amino acid sequence <SEQ ID 110>:

• 1	MKTPLLKPLL ITSLPVFASV FTAASIVWQL GEPKLAMPFV
  	LGIIAGGLVD
  51	LDNRLTGRLK NIIATVALFT LSSLTAQSTL  GTGLPFILAM
  	TLMTFGFTIL
  101	GAVGLKYRTF AFGALAVATY TTLTYTPETY WLTNPFMILC
  	GTVLYSTAII
  151	LFQIILPHRP VQESVANAYE ALGGYLEAKA DFFDPDEAAW
  	IGNRHIDLAM
  201	SNTGVITAFN QCRSALFYRL RGKHRHPRTA KMLRYYFAAQ
  	DIHERISSAH
  251	VDYQEMSEKF KNTDIIFRIR RLLEMQGQAC RNTAQAIRSG
  	KDYVYSKRLG
  301	RAIEGCRQSL RLLSDGNDSP DIRHLSRLLD NLGSVDQQFR
  	QLRHSDSPAE
  351	NDRMGDTRIA ALETGSFKNT *

• Further work revealed the complete nucleotide sequence <SEQ ID 111>:

• 1	ATGAAAACCC CACTCCTCAA GCCTCTGCTC ATTACCTCGC
  	TTCCCGTTTT
  51	CGCCAGTGTC TTTACCGCCG CCTCCATCGT CTGGCAGCTA
  	GGCGAACCCA
  101	AGCTCGCCAT GCCCTTCGTA CTCGGCATCA TCGCCGGCGG
  	CCTGGTCGAT
  151	TTGGACAACC GCCTGACCGG ACGGCTGAAA AACATCATCG
  	CCACCGTCGC
  201	CCTGTTTACC CTCTCCTCGC TCACGGCGCA AAGCACCCTC
  	GGCACAGGGC
  251	TGCCCTTCAT CCTCGCCATG ACCCTGATGA CCTTCGGCTT
  	TACCATTTTA
  301	GGCGCGGTCG GGCTGAAATA CCGCACCTTC GCCTTCGGCG
  	CACTCGCCGT
  351	CGCCACCTAC ACCACGCTTA CCTACACCCC CGAAACCTAC
  	TGGCTGACCA
  401	ACCCCTTCAT GATTTTATGC GGCACCGTAC TGTACAGCAC
  	CGCCATCATC
  451	CTGTTCCAAA TCATCCTGCC CCACCGCCCC GTCCAAGAAA
  	GCGTCGCCAA
  501	TGCCTACGAA GCACTCGGCG GCTACCTCGA AGCCAAAGCC
  	GACTTCTTCG
  551	ACCCCGATGA GGCAGCCTGG ATAGGCAACC GCCACATCGA
  	CCTCGCCATG
  601	AGCAACACCG GCGTCATCAC CGCCTTCAAC CAATGCCGTT
  	CCGCCCTGTT
  651	TTACCGTTTG CGCGGCAAAC ACCGCCACCC GCGCACCGCC
  	AAAATGCTGC
  701	GCTACTACTT CGCCGCCCAA GACATCCACG AACGCATCAG
  	CTCCGCCCAC
  751	GTCGACTACC AAGAGATGTC CGAAAAATTC AAAAACACCG
  	ACATCATCTT
  801	CCGCATCCGC CGCCTGCTCG AAATGCAGGG GCAGGCGTGC
  	CGCAACACCG
  851	CCCAAGCCAT CCGGTCGGGC AAAGACTAcg tTTACAGCAA
  	ACGCCTCGGA
  901	CGCGCCATcg aaggctgCCG CCAGTCGCtg cgcctCCTTt
  	cagacggcaA
  951	CGACAGTCCC GACATCCGCC ACCTGAGccg CCTTCTCGAC
  	AACCTCGgca
  1001	GCGTcgacca gcagtTCcgc caactCCGAC ACAgcgactC
  	CCCCGCcgaa
  1051	Aacgaccgca tgggcgacaC CCGCATCGCC GCCCtcgaaa
  	ccggcagctT
  1101	caaaaaCAcc tggcaggCAA TCCGTCCGCa gctgaaCCTC
  	GAATCatgCG
  1151	TATTCCGCCA TGCCGTCCGC CTGTCCCTCG TCGTTGCCGC
  	CGCCTGCACC
  1201	ATCGTCgaag cCCTCAACCT CAACCTCGGC TACTGGATAC
  	TGCTGACCGC
  1251	CCTTTTCGTC TGCCAACCCA ACTACACCGC CACCAAAAGC
  	CGCGTGTACC
  1301	AACGCATCGC CGGCACCGTA CTCGGCGTAA TCGTCGGCTC
  	GCTCGTCCCC
  1351	TACTTCACCC CCTCCGTCGA AACCAAACTC TGGATTGTCA
  	TCGCCGGTAC
  1401	CACCCTGTTC TTCATGACCC GCACCTACAA ATACAGTTTC
  	TCCACCTTCT
  1451	TCATCACCAT TCAGGCACTG ACCAGCCTCT CCCTCGCAGG
  	TTTGGACGTA
  1501	TACGCCGCCA TGCCCGTGCG CATCATcgaC ACCATTATCG
  	GCGCATCCCT
  1551	TGCCTGGGCG GCGGTCAGCT ACCTGTGGCC AGACTGGAAA
  	TACCTCACGC
  1601	TCGAACGCAC CGCCGCCCTT GCCGTATGCA GCAGCGGCAC
  	ATACCTCCAA
  1651	AAAATTGCCG AACGCCTCAA AACCGGCGAA ACCGGCGACG
  	ACATAGAATA
  1701	CCGCATCACC CGCCGCCGCG CCCACGAACA CACCGCCGCC
  	CTCAGCAGCA
  1751	CCCTTTCCGA CATGAGCAGC GAACCCGCAA AATTCGCCGA
  	CAGCCTGCAA
  1801	CCCGGCTTTA CCCTGCTCAA AACCGGCTAC GCCCTGACCG
  	GCTACATCTC
  1851	CGCCCTCGGC GCATACCGCA GCGAAATGCA CGAAGAATGC
  	AGCCCCGACT
  1901	TTACCGCACA GTTCCACCTT GCCGCCGAAC ACACCGCCCA
  	CATCTTCCAA
  1951	CACCTGCCCG ACATGGGACC CGACGACTTT CAGACGGCAT
  	TGGATACACT
  2001	GCGCGGCGAA CTCGGCACCC TCCGCACCCG CAGCAGCGGA
  	ACACAAAGCC
  2051	ACATCCTCCT CCAACAGCTC CAACTCATCG CccgGCAACT
  	CGAACCCTAC
  2101	TACCGCGCCT ACCGACAAAT TCCGCACAGG CAGCCCCAAA
  	ACGCAGCCTG
  2151	A

• This corresponds to the amino acid sequence <SEQ ID 112; ORF19ng-1>:

• 1	MKTPLLKPLL ITSLPVFASV FTAASIVWQL GEPKLAMPFV
  	LGIIAGGLVD
  51	LDNRLTGRLK NIIATVALFT LSSLTAQSTL GTGLPFILAM
  	TLMTFGFTIL
  101	GAVGLKYRTF AFGALAVATY TTLTYTPETY WLTNPFMILC
  	GTVLYSTAII
  151	LFQIILPHRP VQESVANAYE ALGGYLEAKA DFFDPDEAAW
  	IGNRHIDLAM
  201	SNTGVITAFN QCRSALFYRL RGKHRHPRTA KMLRYYFAAQ
  	DIHERISSAH
  251	VDYQEMSEKF KNTDIIFRIR RLLEMQGQAC RNTAQAIRSG
  	KDYVYSKRLG
  301	RAIEGCRQSL RLLSDGNDSP DIRHLSRLLD NLGSVDQQFR
  	QLRHSDSPAE
  351	NDRMGDTRIA ALETGSFKNT WQAIRPQLNL ESCVFRHAVR
  	LSLVVAAACT
  401	IVEALNLNLG YWILLTALFV CQPNYTATKS RVYQRIAGTV
  	LGVIVGSLVP
  451	YFTPSVETKL WIVIAGTTLF FMTRTYKYSF STFFITIQAL
  	TSLSLAGLDV
  501	YAAMPVRIID TIIGASLAWA AVSYLWPDWK YLTLERTAAL
  	AVCSSGTYLQ
  551	KIAERLKTGE TGDDIEYRIT RRRAHEHTAA LSSTLSDMSS
  	EPAKFADSLQ
  601	PGFTLLKTGY ALTGYISALG AYRSEMHEEC SPDFTAQFHL
  	AAEHTAHIFQ
  651	HLPDMGPDDF QTALDTLRGE LGTLRTRSSG TQSHILLQQL
  	QLIARQLEPY
  701	YRAYRQIPHR QPQNAA*

• ORF19ng-1 and ORF19-1 show 95.5% identity in 716 aa overlap:
• 
• In addition, ORF19ng-1 shows significant homology to a hypothetical gonococcal protein previously entered in the databases:

• sp|O33369|YOR2_NEIGO HYPOTHETICAL 45.5 KD PROTEIN (ORF2) gnl|PID|e1154438
  (AJ002423) hypothetical protein [Neisseria gonorrh] Length = 417
  Score = 1512 (705.6 bits), Expect = 5.3e−203, P = 5.3e−203
  Identities = 301/326 (92%), Positives = 306/326 (93%)

  Query:	307	RQSLRLLSDGNDSPDIRHLSRLLDNLGSVDQQFRQLRHSDSPAENDRMGDTRIAALETGS	366
  		RQSLRLLSDGNDS DIRHLSRLLDNLGSVDQQFRQLRHSDSPAENDRMGDTRIAALETGS
  Sbjct:	1	RQSLRLLSDGNDSXDIRHLSRLLDNLGSVDQQFRQLRHSDSPAENDRMGDTRIAALETGS	60
  Query:	367	FKNTWQAIRPQLNLESCVFRHAVRLSLVVAAACTIVEALNLNLGYWILLTALFVCQPNYT	426
  		FKNTWQAIRPQLNLES VFRHAVRLSLVVAAACTIVEALNLNLGYWILLT LFVCQPNYT
  Sbjct:	61	FKNTWQAIRPQLNLESGVFRHAVRLSLVVAAACTIVEALNLNLGYWILLTRLFVCQPNYT	120
  Query:	427	ATKSRVYQRIAGTVLGVIVGSLVPYFTPSVETKLWIVIAGTTLFFMTRTYKYSFSTFFIT	486
  		ATKSRVYQRIAGTVLGVIVGSLVPYFTPSVETKLWIVIAGTTLFFMTRTYKYSFSTFFIT
  Sbjct:	121	ATKSRVYQRIAGTVLGVIVGSLVPYFTPSVETKLWIVIAGTTLFFMTRTYKYSFSTFFIT	180
  Query:	487	IQALTSLSLAGLDVYAAMPVRIIDTIIGASLAWAAVSYLWPDWKYLTLERTAALAVCSSG	546
  		IQALTSLSLAGLDVYAAMPVRIIDTIIGASLAWAAVSYLWPDWKYLTLERTAALAVCSSG
  Sbjct:	181	IQALTSLSLAGLDVYAAMPVRIIDTIIGASLAWAAVSYLWPDWKYLTLERTAALAVCSSG	240
  Query:	547	TYLQKIAERLKTGETGDDIEYRITRRRAHEHTAALSSTLSDMSSEPAKFADSLQPGFTLL	606
  		TYLQKIAERLKTGETGDDIEYRITRRRAHEHTAALSSTLSDMSSEPAKFAD+  P
  Sbjct:	241	TYLQKIAERLKTGETGDDIEYRITRRRAHEHTAALSSTLSDMSSEPAKFADTCNPALPCS	300
  Query:	607	KTGYALTGYISALGAYRSEMHEECSP	632
  		K   ALTGYISALG   ++  +  +P
  Sbjct:	301	KPATALTGYISALGHTAAKCTKNAAP	326

• Based on this analysis, including the presence of several putative transmembrane domains in the gonococcal protein (the first of which is also seen in the meningococcal protein), and on homology with the YHFK protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 14
• The following DNA sequence, believed to be complete, was identified in N. meningitidis <SEQ ID 113>:

• 1	ATGAATATGC TGGGAGCTTT GGCAAAAGTC GGCAGCCTGA
  	CGATGGTGTC
  51	GCGCGTTTTG GGATTTGTGC GCGATACGGT CATTGCGCGG
  	GCATTCGGCG
  101	CGGGTATGGC GACGGATGCG TTTTTTGTCG CGTTCAAACT
  	GCCCAACCTG
  151	CTTCGCCGCG TGTTTGCGGA GGGGGCGTTT GCCCAAGCGT
  	TTGTGCCGAT
  201	TTTGGCGGAA TACAAGGAAA CGCGTTCAAA AGAGGCGG.C
  	GAAGCCTTTA
  251	TCCGCCATGT GGCGGGGATG CTGTCGTTTG TACTGGTTAT
  	CGTTACCGCG
  301	CTGGGCATAC TTGCCGCGCC TTGGGTGATT TATGTTTCCG
  	CACCCGAGTT
  351	TTGCCCAAGA TGCCGACAAA TTTCAGCTCT CCATCGATTT
  	GCTGCGGATT
  401	ACGTTTCCTT ATATATTATT GATTTCCCTG TCTTCATTTG
  	TCGGCTCGGT
  451	ACTCAATTCT TATCATAAGT TCGGCATTCC GGCGTTTACG
  	CCAC.GTTTC
  501	TGAACGTGTC GTTTATCGTA TTCGCGCTGT TTTTCGTGCC
  	GTATTTCGAT
  551	CCGCCCGTTA CCGCGCyGGC GTGGGCGGTC TTTGTCGGCG
  	GCATTTTGCA
  601	ACTCGrmTTC CAACTGCCCT GGCTGGCGAA ACTGGGCTTT
  	TTGAAACTGC
  651	CCAAACtGAG TTTCAAAGAT GCGGCGGTCA ACCGCGTGAT
  	GAAACAGATG
  701	GCGCCTGCgA TTTTgGGCGT GAgCGTGGCG CAGGTTTCTT
  	TGGTGATCAA
  751	CACGATTTTc GCGTCTTATC TGCAATCGGG CAGCGTTTCA
  	TGGATGTATT
  801	ACGCCGACCG CATGATGGAG CTGCCCAGCG GCGTGCTGGG
  	GGCGGCACTC
  851	GGTACGATTT TGCTGCCGAC TTTGTCCAAA CACTCGGCAA
  	ACCaAGATAC
  901	GGaACAGTTT TCCGCCCTGC TCGACTGGGG TTTGCGCCTG
  	TGCATGCtgc
  951	TGACGCTGCC GGCGgcGGTC GGACTGGCGG TGTTGTCGTT
  	cCCgCtGGTG
  1001	GCGACGCTGT TTATGTACCG CGwATTTACG CTGTTTGACG
  	CGCAGATGAC
  1051	GCAACACGCG CTGATTGCCT ATTCTTTCGG TTTAATCGGC
  	TTAATCATGA
  1101	TTAAAGTGTT GGCACCCGGC TTCTATGCGC GGCAAAACAT
  	CAAwAmGCCC
  1151	GTCAAAATCG CCATCTTCAC GCTCATCTGC mCGCAGTTGA
  	TGAACCTTGs
  1201	CTTTAyCGGC CCACTrrAAC rCasTCGGAC TTTCGCTTGC
  	CATCGGTCTG
  1251	GGCGCGTGTA TCAATGCCGG ATTGTTGTTT TACCTGTTGC
  	GCAGACACGG
  1301	TATTTACCAA CCTGG.CAAG GGTTGGGCAG CGTTCTT.AG
  	CAAAAATGCT
  1351	GcTCTCGCTC GCCGTGA

• This corresponds to the amino acid sequence <SEQ ID 114; ORF20>:

• 1	MNMLGALAKV GSLTMVSRVL GFVRDTVIAR AFGAGMATDA
  	FFVAFKLPNL
  51	LRRVFAEGAF AQAFVPILAE YKETRSKEAX EAFIRHVAGM
  	LSFVLVIVTA
  101	LGILAAPWVI YVSAPSFAQD ADKFQLSIDL LRITFPYILL
  	ISLSSFVGSV
  151	LNSYHKFGIP AFTPXFLNVS FIVFALFFVP YFDPPVTAXA
  	WAVFVGGILQ
  201	LXFQLPWLAK LGFLKLPKLS FKDAAVNRVM KQMAPAILGV
  	SVAQVSLVIN
  251	TIFASYLQSG SVSWMYYADR MMELPSGVLG AALGTILLPT
  	LSKHSANQDT
  301	EQFSALLDWG LRLCMLLTLP AAVGLAVLSF PLVATLFMYR
  	XFTLFDAQMT
  351	QHALIAYSFG LIGLIMIKVL APGFYARQNI XXPVKIAIFT
  	LICXQLMNLX
  401	FXGPLXXIGL SLAIGLGACI NAGLLFYLLR RHGIYQPXQG
  	LGSVLXQKCC
  451	SRSP*

• These sequences were elaborated, and the complete DNA sequence <SEQ ID 115> is:

• 1	ATGAATATGC TGGGAGCTTT GGCAAAAGTC GGCAGCCTGA
  	CGATGGTGTC
  51	GCGCGTTTTG GGATTTGTGC GCGATACGGT CATTGCGCGG
  	GCATTCGGCG
  101	CGGGTATGGC GACGGATGCG TTTTTTGTCG CGTTCAAACT
  	GCCCAACCTG
  151	CTTCGCCGCG TGTTTGCGGA GGGGGCGTTT GCCCAAGCGT
  	TTGTGCCGAT
  201	TTTGGCGGAA TACAAGGAAA CGCGTTCAAA AGAGGCGGCG
  	GAGGCTTTTA
  251	TCCGCCATGT GGCGGGGATG CTGTCGTTTG TACTGGTTAT
  	CGTTACCGCG
  301	CTGGGCATAC TTGCCGCGCC TTGGGTGATT TATGTTTCCG
  	CACCCGGTTT
  351	TGCCCAAGAT GCCGACAAAT TTCAGCTCTC CATCGATTTG
  	CTGCGGATTA
  401	CGTTTCCTTA TATATTATTG ATTTCCCTGT CTTCATTTGT
  	CGGCTCGGTA
  451	CTCAATTCTT ATCATAAGTT CGGCATTCCG GCGTTTACGC
  	CCACGTTTCT
  501	GAACGTGTCG TTTATCGTAT TCGCGCTGTT TTTCGTGCCG
  	TATTTCGATC
  551	CGCCCGTTAC CGCGCTGGCG TGGGCGGTCT TTGTCGGCGG
  	CATTTTGCAA
  601	CTCGGCTTCC AACTGCCCTG GCTGGCGAAA CTGGGCTTTT
  	TGAAACTGCC
  651	CAAACTGAGT TTCAAAGATG CGGCGGTCAA CCGCGTGATG
  	AAACAGATGG
  701	CGCCTGCGAT TTTGGGCGTG AGCGTGGCGC AGGTTTCTTT
  	GGTGATCAAC
  751	ACGATTTTCG CGTCTTATCT GCAATCGGGC AGCGTTTCAT
  	GGATGTATTA
  801	CGCCGACCGC ATGATGGAGC TGCCCAGCGG CGTGCTGGGG
  	GCGGCACTCG
  851	GTACGATTTT GCTGCCGACT TTGTCCAAAC ACTCGGCAAA
  	CCAAGATACG
  901	GAACAGTTTT CCGCCCTGCT CGACTGGGGT TTGCGCCTGT
  	GCATGCTGCT
  951	GACGCTGCCG GCGGCGGTCG GACTGGCGGT GTTGTCGTTC
  	CCGCTGGTGG
  1001	CGACGCTGTT TATGTACCGC GAATTTACGC TGTTTGACGC
  	GCAGATGACG
  1051	CAACACGCGC TGATTGCCTA TTCTTTCGGT TTAATCGGCT
  	TAATCATGAT
  1101	TAAAGTGTTG GCACCCGGCT TCTATGCGCG GCAAAACATC
  	AAAACGCCCG
  1151	TCAAAATCGC CATCTTCACG CTCATCTGCA CGCAGTTGAT
  	GAACCTTGCC
  1201	TTTATCGGCC CACTGAAACA CGTCGGACTT TCGCTTGCCA
  	TCGGTCTGGG
  1251	CGCGTGTATC AATGCCGGAT TGTTGTTTTA CCTGTTGCGC
  	AGACACGGTA
  1301	TTTACCAACC TGGCAAGGGT TGGGCAGCGT TCTTAGCAAA
  	AATGCTGCTC
  1351	TCGCTCGCCG TGATGTGCGG CGGACTGTGG GCAGCGCAGG
  	CTTACCTGCC
  1401	GTTTGAATGG GCGCACGCCG GCGGAATGCG GAAAGCGGGG
  	CAGCTCTGCA
  1451	TCCTGATTGC CGTCGGCGGC GGACTGTATT TCGCATCACT
  	GGCGGCTTTG
  1501	GGCTTCCGTC CGCGCCATTT CAAACGCGTG GAAAACTGA

• This corresponds to the amino acid sequence <SEQ ID 116; ORF20-1>:

• 1	MNMLGALAKV GSLTMVSRVL GFVRDTVIAR AFGAGMATDA
  	FFVAFKLPNL
  51	LRRVFAEGAF AQAFVPILAE YKETRSKEAA EAFIRHVAGM
  	LSFVLVIVTA
  101	LGILAAPWVI YVSAPGFAQD ADKFQLSIDL LRITFPYILL
  	ISLSSFVGSV
  151	LNSYHKFGIP AFTPTFLNVS FIVFALFFVP YFDPPVTALA
  	WAVFVGGILQ
  201	LGFQLPWLAK LGFLKLPKLS FKDAAVNRVM KQMAPAILGV
  	SVAQVSLVIN
  251	TIFASYLQSG SVSWMYYADR MMELPSGVLG AALGTILLPT
  	LSKHSANQDT
  301	EQFSALLDWG LRLCMLLTLP AAVGLAVLSF PLVATLFMYR
  	EFTLFDAQMT
  351	QHALIAYSFG LIGLIMIKVL APGFYARQNI KTPVKIAIFT
  	LICTQLMNLA
  401	FIGPLKHVGL SLAIGLGACI NAGLLFYLLR RHGIYQPGKG
  	WAAFLAKMLL
  451	SLAVMCGGLW AAQAYLPFEW AHAGGMRKAG QLCILIAVGG
  	GLYFASLAAL
  501	GFRPRHFKRV EN*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with the MviN Virulence Factor of S. typhimurium (Accession Number P37169)
• ORF20 and MviN proteins show 63% aa identity in 440aa overlap:

• Orf20	1	MNMLGALAKVGSLTMVSRVLGFVRDTVIARAFGAGMATDAFFVAFKLPNLLRRVFAEGAF	60
  		MN+L +LA V S+TM SRVLGF RD ++AR FGAGMATDAFFVAFKLPNLLRR+FAEGAF
  MviN	14	MNLLKSLAAVSSMTMFSRVLGFARDAIVARIFGAGMATDAFFVAFKLPNLLRRIFAEGAF	73
  Orf20	61	AQAFVPILAEYKETRSKEAXEAFIRHVAGMLSFVLVIVTALGILAAPWVIYVSAPSFAQD	120
  		+QAFVPILAEYK  + +EA   F+ +V+G+L+  L +VT  G+LAAPWVI V+AP FA
  MviN	74	SQAFVPILAEYKSKQGEEATRIFVAYVSGLLTLALAVVTVAGMLAAPWVIMVTAPGFADT	133
  Orf20	121	ADKFQLSIDLLRITFPYILLISLSSFVGSVLNSYHKFGIPAFTPXFLNVSFIVFALFFVP	180
  		ADKF L+  LLRITFPYILLISL+S VG++LN++++F IPAF P FLN+S I FALF  P
  MviN	134	ADKFALTTQLLRITFPYILLISLASLVGAILNTWNRFSIPAFAPTFLNISMIGFALFAAP	193
  Orf20	181	YFDPPVTAXAWAVFVGGILQLXFQLPWLAKLGFLKLPKLSFKDAAVNRVMKQMAPAILGV	240
  		YF+PPV A AWAV VGG+LQL +QLP+L K+G L LP+++F+D    RV+KQM PAILGV
  MviN	194	YFNPPVLALAWAVTVGGVLQLVYQLPYLKKIGMLVLPRINFRDTGAMRVVKQMGPAILGV	253
  Orf20	241	SVAQVSLVINTIFASYLQSGSVSWMYYADRMMELPSGVLGAALGTILLPTLSKHSANQDT	300
  		SV+Q+SL+INTIFAS+L SGSVSWMYYADR+ME PSGVLG ALGTILLP+LSK  A+ +
  MviN	254	SVSQISLIINTIFASFLASGSVSWMYYADRLMEFPSGVLGVALGTILLPSLSKSFASGNH	313
  Orf20	301	EQFSALLDWGLRLCMLLTLPAAVGLAVLSFPLVATLFMYRXFTLFDAQMTQHALIAYSFG	360
  		+++  L+DWGLRLC LL LP+AV L +L+ PL  +LF Y  FT FDA MTQ ALIAYS G
  MviN	314	DEYCRLMDWGLRLCFLLALPSAVALGILAKPLTVSLFQYGKFTAFDAAMTQRALIAYSVG	373
  Orf20	361	LIGLIMIKVLAPGFYARQNIXXPVKIAIFTLICXQLMNLXFXXXXXXXXXXXXXXXXXCI	420
  		LIGLI++KVLAPGFY+RQ+I  PVKIAI TLI  QLMNL F                 C+
  MviN	374	LIGLIVVKVLAPGFYSRQDIKTPVKIAIVTLIMTQLMNLAFIGPLKHAGLSLSIGLAACL	433
  Orf20	421	NAGLLFYLLRRHGIYQPXQG	440
  		NA LL++ LR+  I+ P  G
  MviN	434	NASLLYWQLRKQNIFTPQPG	453

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF20 shows 93.5% identity over a 447aa overlap with an ORF (ORF20a) from strain A of N. meningitidis:
• 
• The complete length ORF20a nucleotide sequence <SEQ ID 117> is:

• 1	ATGAATATGC TGGGAGCTTT GGTAAAAGTC GGCAGCCTGA
  	CGATGGTGTC
  51	GCGCGTTTTG GGATTTGTGC GCGATACGGT CATTGCGCGC
  	GCATTCGGCG
  101	CAGGCATGGC GACGGATGCG TTCTTTGTCG CGTTCAAACT
  	GCCCAACCTG
  151	CTTCGCCGCG TGTTTGCGGA GGGGGCGTTT GCCCAAGCGT
  	TTGTGCCGAT
  201	TTTGGCGGAA TATAAGGAAA CGCGTTCTAA AGAGGCGACG
  	GAGGCTTTTA
  251	TCCGCCATGT GGCGGGGATG CTGTCGTTTG TACTGGTCAT
  	CGTTACCGCG
  301	CTGGGCATAC TTGCCGCGCC TTGGGTGATT TATGTTTCCG
  	CACCCGGTTT
  351	TGCCAAAGAT GCCGACAAAT TTCAGCTCTC TATCGATTTG
  	CTGCGGATTA
  401	CGTTTCCTTA TATCTTATTG ATTTCACTTT CCTCTTTTGT
  	CGGCTCGGTA
  451	CTCAATTCCT ATCATAAATT CAGCATTCCT GCGTTTACGC
  	CCACGTTCCT
  501	GAACGTGTCG TTTATCGTAT TCGCGCTGTT TTTCGTGCCG
  	TATTTCGATC
  551	CTCCCGTTAC CGCGCTGGCT TGGGCGGTTT TTGTCGGCGG
  	CATTTTGCAA
  601	CTCGGCTTCC AACTGCCCTG GCTGGCGAAA CTGGGTTTTT
  	TGAAACTGCC
  651	CAAACTGAGT TTCAAAGATG CGGCGGTCAA CCGCGTGATG
  	AAACAGATGG
  701	CGCCTGCGAT TTTGGGCGTG AGCGTGGCGC AGATTTCTTT
  	GGTGATCAAC
  751	ACGATTTTCG CGTCTTATCT GCAATCGGGC AGCGTTTCAT
  	GGATGTATTA
  801	CGCCGACCGC ATGATGGAAC TGCCCGGCGG CGTGCTGGGG
  	GCGGCACTCG
  851	GTACGATTTT GCTGCCGACT TTGTCCAAAC ACTCGGCAAA
  	CCAAGATACG
  901	GAACAGTTTT CCGCCCTGCT CGACTGGGGT TTGCGCNTGT
  	GCATGCTGCT
  951	GACGCTGCCG GCGGCGGTCG GAATGGCGGT GTTGTCGTTC
  	CCGCTGGTGG
  1001	CAACCTTGTT TATGTACCGA GAATTCACGC TGTTTGACGC
  	GCAGATGACG
  1051	CAACACGCGC TGATTGCCTA TTCTTTCGGT TTAATCGGTT
  	TAATCATGAT
  1101	TAAAGTGTTG GCGCCCGGCT TTTATGCGCG GCAAAACATC
  	AAAACGCCCG
  1151	TCAAAATCGC CATCTTCACG CTCATTTGCA CGCAGTTGAT
  	GAACCTTGCC
  1201	TTTATCGGCC CACTGAAACA CGTCGGACTT TCGCTTGCCA
  	TCGGTCTGGG
  1251	CGCGTGTATC AATGCCGGAT TGTTGTTTTA CCTGTTGCGC
  	AGACACGGTA
  1301	TTTACCAACC TGGCAAGGGT TGGGCAGCGT TCTTGGCAAA
  	AATGCTGCTC
  1351	TCGCTCGCCG TGATGGGAGG CGGCCTGTAT GCCGCCCAAA
  	TCTGGCTGCC
  1401	GTTCGACTGG GCACACGCCG GCGGAATGCA AAAGGCCGCC
  	CGGCTCTTCA
  1451	TCCTGATTGC CGTCGGCGGC GGACTGTATT TCGCATCACT
  	GGCGGCTTTG
  1501	GGCTTCCGTC CGCGCCATTT CAAACGCGTG GAAAGCTGA

• This encodes a protein having amino acid sequence <SEQ ID 118>:

• 1	MNMLGALVKV GSLTMVSRVL GFVRDTVIAR AFGAGMATDA
  	FFVAFKLPNL
  51	LRRVFAEGAF AQAFVPILAE YKETRSKEAT EAFIRHVAGM
  	LSFVLVIVTA
  101	LGILAAPWVI YVSAPGFAKD ADKFQLSIDL LRITFPYILL
  	ISLSSFVGSV
  151	LNSYHKFSIP AFTPTFLNVS FIVFALFFVP YFDPPVTALA
  	WAVFVGGILQ
  201	LGFQLPWLAK LGFLKLPKLS FKDAAVNRVM KQMAPAILGV
  	SVAQISLVIN
  251	TIFASYLQSG SVSWMYYADR MMELPGGVLG AALGTILLPT
  	LSKHSANQDT
  301	EQFSALLDWG LRXCMLLTLP AAVGMAVLSF PLVATLFMYR
  	EFTLFDAQMT
  351	QHALIAYSFG LIGLIMIKVL APGFYARQNI KTPVKIAIFT
  	LICTQLMNLA
  401	FIGPLKHVGL SLAIGLGACI NAGLLFYLLR RHGIYQPGKG
  	WAAFLAKMLL
  451	SLAVMGGGLY AAQIWLPFDW AHAGGMQKAA RLFILIAVGG
  	GLYFASLAAL
  501	GFRPRHFKRV ES*

• ORF20a and ORF20-1 show 96.5% identity in 512 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF20 shows 92.1% identity over a 454aa overlap with a predicted ORF (ORF20ng) from N. gonorrhoeae:
• 
• An ORF20ng nucleotide sequence <SEQ ID 119> was predicted to encode a protein having amino acid sequence <SEQ ID 120>:

• 1	MNMLGALAKV GSLTMVSRVL GFVRDTVIAR AFGAGMATDA
  	FFVAFKLPNL
  51	LRRVFAEGAF AQAFVPILAE YKETRSKEAT EAFIRHVAGM
  	LSFVLIVVTA
  101	LGILAAPWVI YVSAPGFTKD ADKFQLSISL LRITFPYILL
  	ISLSSFVGSI
  151	LNSYHKFGIP AFTPTFLNIS FIVFALFFVP YFDPPVTALA
  	WAVFVGGILQ
  201	LGFQLPWLAK LGFLKLPKLN FKDAAVNRVM KQMAPAILGV
  	SVAQISLVIN
  251	TIFASYLQSG SVSWMYYADR MMELPGGVLG AALGTILLPT
  	LSKHSANQDT
  301	EQFSALLDWG LRLCMLLTLP AAAGLAVLSF PLVATLFMYR
  	EFTLFDAQMT
  351	QHALIAYSFG LIGLIMIKVL ASGFYARQNI KTPVKIAIFT
  	LICTQLMNLA
  401	FIGPLKHAGL SLAIGLGACI NAGLLFFLFR KHGIYRPGQG
  	LGQPSWRKCC
  451	SRSP*

• Further DNA sequence analysis revealed the following DNA sequence <SEQ ID 121>:

• 1	ATGAATATGC TTGGAGCTTT GGCAAAAGTC GGCAGCCTGA
  	CGATGGTGTC
  51	GCGCGTTTTG GGATTTGTGC GCGATACGGT CATTGCGCGG
  	GCATTCGGCG
  101	CGGGTATGGC GACGGATGCG TTTTTTGTCG CGTTCAAACT
  	GCCCAACCTG
  151	CTTCGCCGCG TGTTTGCGGA GGGGGCGTTT GCCCAAGCGT
  	TTGTGCCGAT
  201	TTTGGCGGAA TATAAGGAAA CGCGTTCTAA AGAGGCGAcg
  	gAGGCTTTTA
  251	TCCGCCACGt tgcgggAatg CTGTCGTTTG TGCTGATcgt
  	cGttacCGCG
  301	CTGGGCATAC TTGCCGCgcc tTGGGTGATT TATGTTtccg
  	CgcccGGCTT
  351	TACCAAAGAC GCGGACAAGT TCCAACTTTC CATCAGCCTG
  	CTGCGGATTA
  401	CGTTTCCTTA TATATTATTG ATTTCTTTGT CTTCTTTTGT
  	CGGCTCGATA
  451	CTCAATTCCT ACCATAAGTT CGGCATTCCC GCGTTTACGC
  	CCACGTTTTT
  501	AAACATCTCT TTTATCGTAT TCGCACTGTT TTTCGTGCCG
  	TATTTCGATC
  551	CGCCCGTTAC CGCGCTGGCG TGGGCGGTTT TTGTCGGCGG
  	TATTTTGCAG
  601	CTCGGTTTCC AACTGCCGTG GCTGGCGAAA CTGGGCTTTT
  	TGAAACTGCC
  651	CAAACTGAAT TTCAAAGATG CGGCGGTCAA CCGCGTCATG
  	AAACAGATGG
  701	CGCCTGCGAT TTTGGGCGTG agcgTGGCGC AAATTTCTTT
  	GgttATCAAC
  751	ACGATTTTCG CGTCTTATCT GCAATCGGGC AGCGTTTCAT
  	GGATGTatta
  801	cgCCGACCGC ATGATGGAGc tgcgccGGGG CGTGCTGGGG
  	GCTGCACTCG
  851	GTACAATTTT GCTGCCGACT TTGTCCAAAC ACTCGGCAAA
  	CCAAGATACG
  901	GAACAGTTTT CCGCCCTGCT CGACTGGGGT TTGCGCCTGT
  	GCATGCTGCT
  951	GACGCTGCCG GCGGCGGccg GACTGGCGGT ATTGTCGTTC
  	CCGCTGGTGG
  1001	CGACGCTGTT TATGTACCGA GAATTCACGC TGTTTGACGC
  	ACAAATGACG
  1051	CAACACGCGC TGATTGCCTA TTCTTTCGGT TTAATCGGTT
  	TAATTATGAT
  1101	TAAAGTGTTG GCATCCGGCT TTTATGCGCG GCAAAACATC
  	AAAACGCCCG
  1151	TCAAAATCGC CATCTTCACG CTCATCTGCA CGCAGTTGAT
  	GAACCTCGCC
  1201	TTTATCGGTC CGTTGAAACA CGCCGGGCTT TCGCTCGCCA
  	TCGGCCTGGG
  1251	CGCGTGCATC AACGCCGGAT TGTTGTTCTT CCTGTTGCGC
  	AAACACGGTA
  1301	TTTACCGGCC cggcaggggt tgggcggcgt TCTTGGCGAA
  	AATGCTGCTC
  1351	GCGCTCGCCG TGATGTGCGG CGGACTGTGG GCGGCGCAGG
  	CTTGCCTGCC
  1401	GTTCGAATGG GCGCACGCCG GCGGAATGCG GAAAGCGGGG
  	CAGCTCTGCA
  1451	TCCTGATTGC CGTCGGCGGC GGACTGTATT TCGCATCTCT
  	GGCGGCTTTG
  1501	GGCTTCCGTC CGCGCCATTT CAAACGCGTG GAAAGCTGA

• This encodes the following amino acid sequence <SEQ ID 122; ORF20ng-1>:

• 1	MNMLGALAKV GSLTMVSRVL GFVRDTVIAR AFGAGMATDA
  	FFVAFKLPNL
  51	LRRVFAEGAF AQAFVPILAE YKETRSKEAT EAFIRHVAGM
  	LSFVLIVVTA
  101	LGILAAPWVI YVSAPGFTKD ADKFQLSISL LRITFPYILL
  	ISLSSFVGSI
  151	LNSYHKFGIP AFTPTFLNIS FIVFALFFVP YFDPPVTALA
  	WAVFVGGILQ
  201	LGFQLPWLAK LGFLKLPKLN FKDAAVNRVM KQMAPAILGV
  	SVAQISLVIN
  251	TIFASYLQSG SVSWMYYADR MMELRRGVLG AALGTILLPT
  	LSKHSANQDT
  301	EQFSALLDWG LRLCMLLTLP AAAGLAVLSF PLVATLFMYR
  	EFTLFDAQMT
  351	QHALIAYSFG LIGLIMIKVL ASGFYARQNI KTPVKIAIFT
  	LICTQLMNLA
  401	FIGPLKHAGL SLAIGLGACI NAGLLFFLLR KHGIYRPGRG
  	WAAFLAKMLL
  451	ALAVMCGGLW AAQACLPFEW AHAGGMRKAG QLCILIAVGG
  	GLYFASLAAL
  501	GFRPRHFKRV ES*

• ORF20ng-1 and ORF20-1 show 95.7% identity in 512 aa overlap:
• 
• In addition, ORF20ng-1 shows significant homology with a virulence factor of S. typhimurium:

• sp|P37169|MVIN_SALTY VIRULENCE FACTOR MVIN pir||S40271 mviN protein -
  Salmonella typhimurium gi|438252 (Z26133) mviB gene product
  [Salmonella typhimurium] gnl|PID|d1005521 (D25292) ORF2
  [Salmonella typhimurium] Length = 524
  Score = 1573 (750.1 bits), Expect = 1.1e−220, Sum P(2) = 1.1e−220
  Identities = 309/467 (66%), Positives = 368/467 (78%)

  Query:	1	MNMLGALAKVGSLTMVSRVLGFVRDTVIARAFGAGMATDAFFVAFKLPNLLRRVFAEGAF	60
  		MN+L +LA V S+TM SRVLGF RD ++AR FGAGMATDAFFVAFKLPNLLRR+FAEGAF
  Sbjct:	14	MNLLKSLAAVSSMTMFSRVLGFARDAIVARIFGAGMATDAFFVAFKLPNLLRRIFAEGAF	73
  Query:	61	AQAFVPILAEYKETRSKEATEAFIRHVAGMLSFVLIVVTALGILAAPWVIYVSAPGFTKD	120
  		+QAFVPILAEYK  + +EAT  F+ +V+G+L+  L VVT  G+LAAPWVI V+APGF
  Sbjct:	74	SQAFVPILAEYKSKQGEEATRIFVAYVSGLLTLALAVVTVAGMLAAPWVIMVTAPGFADT	133
  Query:	121	ADKFQLSISLLRITFPYILLISLSSFVGSILNSYHKFGIPAFTPTFLNISFIVFALFFVP	180
  		ADKF L+  LLRITFPYILLISL+S VG+ILN++++F IPAF PTFLNIS I FALF  P
  Sbjct:	134	ADKFALTTQLLRITFPYILLISLASLVGAILNTWNRFSIPAFAPTFLNISMIGFALFAAP	193
  Query:	181	YFDPPVTALAWAVFVGGILQLGFQLPWLAKLGFLKLPKLNFKDAAVNRVMKQMAPAILGV	240
  		YF+PPV ALAWAV VGG+LQL +QLP+L K+G L LP++NF+D    RV+KQM PAILGV
  Sbjct:	194	YFNPPVLALAWAVTVGGVLQLVYQLPYLKKIGMLVLPRINFRDTGAMRVVKQMGPAILGV	253
  Query:	241	SVAQISLVINTIFASYLQSGSVSWMYYADRMMELRRGVLGAALGTILLPTLSKHSANQDT	300
  		SV+QISL+INTIFAS+L SGSVSWMYYADR+ME   GVLG ALGTILLP+LSK  A+ +
  Sbjct:	254	SVSQISLIINTIFASFLASGSVSWMYYADRLMEFFSGVLGVALGTILLPSLSKSFASGNH	313
  Query:	301	EQFSALLDWGLRLCMLLTLPAAAGLAVLSFPLVATLFMYREFTLFDAQMTQHALIAYSFG	360
  		+++  L+DWGLRLC LL LP+A  L +L+ PL  +LF Y +FT FDA MTQ ALIAYS G
  Sbjct:	314	DEYCRLMDWGLRLCFLLALPSAVALGILAKPLTVSLFQYGKFTAFDAAMTQRALIAYSVG	373
  Query:	361	LIGLIMIKVLASGFYARQNIKTPVKIAIFTLICTQLMNLAFIGPLKHAGLSLAIGLGACI	420
  		LIGLI++KVLA GFY+RQ+IKTPVKIAI TLI TQLMNLAFIGPLKHAGLSL+IGL AC+
  Sbjct:	374	LIGLIVVKVLAPGFYSRQDIKTPVKIAIVTLIMTQLMNLAFIGPLKHAGLSLSIGLAACL	433
  Query:	421	NAGLLFFLLRKHGIYRPGRGWXXXXXXXXXXXXVMCGGLWAAQACLP	467
  		NA LL++ LRK  I+ P  GW            VM   L+     +P
  Sbjct:	434	NASLLYWQLRKQNIFTPQPGWMWFLMRLIISVLVMAAVLFGVLHIMP	480

  Score = 70 (33.4 bits), Expect = 1.1e−220, Sum P(2) = 1.1e−220
  Identities = 14/41 (34%), Positives = 23/41 (56%)

  Query:	469	EWAHAGGMRKAGQLCILIAVGGGLYFASLAALGFRPRHFKR	509
  		EW+    + +  +L  ++  G   YFA+LA LGF+ + F R
  Sbjct:	481	EWSQGSMLWRLLRLMAVVIAGIAAYFAALAVLGFKVKEFVR	521

• Based on this analysis, including the homology with a virulence factor from S. typhimurium, it is predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 15
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 123>:

• 1	atGATTAAAA TCAAAAAAGG TCTAAACCTG CCCATCGCGG
  	GCAGACCGGA
  51	GCAAGCCGTT tACGACGGCC CGGCCaTTAC CGAAGtCGCG
  	TTGCTTGGCG
  101	AAGAATATGC CGGTATGCGC CCCTCGATGA AAGTCAAGGA
  	AGGCGATGCC
  151	GTcAAAAAAG GCCAAGTGCT GTTTGAAGAC AAAAAGAATC
  	CGGGCGTGGT
  201	GTTTACTGCG CCGGCTTCAG GcAAAATCGC CGCGATTCAC
  	CGTGGCGAAA
  251	AGCGCGTACT TCAGTCAGTC GTGATTGCCG TTGAArGCAA
  	CGACGAAATC
  301	GAGTTTGAAC GCTACGCACC TGAAGCGCTG GCAAACTTAA
  	GCGGCGAAGA
  351	AGTGCGCCGC AACCTGATCC AATCCGGTTT GTGGACTGCG
  	CTGCGCACCC
  401	GTCCGTTCAG CAAAATTCCT GCCGTCGATG CCGAGCCGTT
  	CGCCATCTTC
  451	GTCAATGCGA tGGACACCAA TCCG..

• This corresponds to the amino acid sequence <SEQ ID 124; ORF22>:

• 1	MIKIKKGLNL PIAGRPEQAV YDGPAITEVA LLGEEYAGMR
  	PSMKVKEGDA
  51	VKKGQVLFED KKNPGVVFTA PASGKIAAIH RGEKRVLQSV
  	VIAVEXNDEI
  101	EFERYAPEAL ANLSGEEVRR NLIQSGLWTA LRTRPFSKIP
  	AVDAEPFAIF
  151	VNAMDTNP..

• Further work revealed the complete nucleotide sequence <SEQ ID 125>:

• 1	ATGATTAAAA TCAAAAAAGG TCTAAACCTG CCCATCGCGG
  	GCAGACCGGA
  51	GCAAGCCGTT TACGACGGCC CGGCCATTAC CGAAGTCGCG
  	TTGCTTGGCG
  101	AAGAATATGC CGGTATGCGC CCCTCGATGA AAGTCAAGGA
  	AGGCGATGCC
  151	GTCAAAAAAG GCCAAGTGCT GTTTGAAGAC AAAAAGAATC
  	CGGGCGTGGT
  201	GTTTACTGCG CCGGCTTCAG GCAAAATCGC CGCGATTCAC
  	CGTGGCGAAA
  251	AGCGCGTACT TCAGTCAGTC GTGATTGCCG TTGAAGGCAA
  	CGACGAAATC
  301	GAGTTTGAAC GCTACGCACC TGAAGCGCTG GCAAACTTAA
  	GCGGCGAAGA
  351	AGTGCGCCGC AACCTGATCC AATCCGGTTT GTGGACTGCG
  	CTGCGCACCC
  401	GTCCGTTCAG CAAAATTCCT GCCGTCGATG CCGAGCCGTT
  	CGCCATCTTC
  451	GTCAATGCGA TGGACACCAA TCCGCTGGCT GCCGACCCTA
  	CGGTCATTAT
  501	CAAAGAAGCC GCCGAGGATT TCAAACGCGG CCTGTTGGTA
  	TTGAGCCGTT
  551	TGACCGAACG CAAAATCCAT GTTTGTAAGG CAGCTGGCGC
  	AGACGTGCCG
  601	TCTGAAAATG CTGCCAACAT CGAAACACAT GAATTCGGCG
  	GCCCGCATCC
  651	TGCCGGTTTG AGTGGCACGC ACATTCATTT CATCGAGCCG
  	GTCGGCGCGA
  701	ATAAAACCGT GTGGACCATC AATTATCAAG ATGTAATTAC
  	CATTGGCCGT
  751	TTGTTTGCAA CAGGCCGTCT GAACACCGAG CGCGTGATTG
  	CCCTAGGTGG
  801	TTCTCAAGTC AACAAACCGC GCCTCTTGCG TACCGTTTTG
  	GGTGCGAAAG
  851	TATCGCAAAT TACTGCGGGC GAATTGGTTG ACACAGACAA
  	CCGCGTGATT
  901	TCCGGTTCGG TATTGAACGG CGCGATTACA CAAGGCGCGC
  	ACGATTATTT
  951	GGGACGCTAC CACAATCAGA TTTCCGTTAT CGAAGAAGGC
  	CGCAGCAAAG
  1001	AGCTGTTCGG CTGGGTTGCG CCGCAGCCGG ACAAATACTC
  	CATCACGCGT
  1051	ACAACCCTCG GCCATTTCCT GAAAAACAAA CTCTTCAAGT
  	TCAACACAGC
  1101	CGTCAACGGC GGCGACCGCG CCATGGTGCC GATTGGTACT
  	TACGAGCGCG
  1151	TGATGCCCTT GGATATCCTG CCCACCCTGC TTTTGCGCGA
  	TTTAATCGTC
  1201	GGCGATACCG ACAGCGCGCA GGCATTGGGT TGCTTGGAAT
  	TGGACGAAGA
  1251	AGACCTCGCT TTGTGCAGCT TCGTCTGCCC GGGCAAATAC
  	GAATACGGCC
  1301	CGCTGTTGCG CAAAGTGCTG GAAACCATTG AGAAGGAAGG
  	CTGA

• This corresponds to the amino acid sequence <SEQ ID 126; ORF22-1>:

• 1	MIKIKKGLNL PIAGRPEQAV YDGPAITEVA LLGEEYAGMR
  	PSMKVKEGDA
  51	VKKGQVLFED KKNPGVVFTA PASGKIAAIH RGEKRVLQSV
  	VIAVEGNDEI
  101	EFERYAPEAL ANLSGEEVRR NLIQSGLWTA LRTRPFSKIP
  	AVDAEPFAIF
  151	VNAMDTNPLA ADPTVIIKEA AEDFKRGLLV LSRLTERKIH
  	VCKAAGADVP
  201	SENAANIETH EFGGPHPAGL SGTHIHFIEP VGANKTVWTI
  	NYQDVITIGR
  251	LFATGRLNTE RVIALGGSQV NKPRLLRTVL GAKVSQITAG
  	ELVDTDNRVI
  301	SGSVLNGAIT QGAHDYLGRY HNQISVIEEG RSKELFGWVA
  	PQPDKYSITR
  351	TTLGHFLKNK LFKFNTAVNG GDRAMVPIGT YERVMPLDIL
  	PTLLLRDLIV
  401	GDTDSAQALG CLELDEEDLA LCSFVCPGKY EYGPLLRKVL
  	ETIEKEG*

• Further work identified the corresponding gene in strain A of N. meningitidis <SEQ ID 127>:

• 1	ATGATTAAAA TCAAAAAAGG TCTAAACCTG CCCATCGCGG
  	GCAGACCGGA
  51	GCAAGTCATT TATGACGGGC CCGTCATTAC CGAAGTCGCG
  	TTGCTTGGCG
  101	AAGAATATGC CGGTATGCGC CCCTNGATGA AAGTCAAGGA
  	AGGCGATGCC
  151	GTCAAAAAAG GCCAAGTGCT GTTTGAAGAC AAAAAGNATC
  	CGGGCGTGGT
  201	GTTTACCGCG CCNGTTTCAG GCAAAATCGC CGCCATCCAT
  	CGCGGCGAAA
  251	AGCGCGTACT TCAGTCGGTC GTGATTGCCG TTGAAGGCAA
  	CGACGAAATC
  301	GAGTTCGAAC GCTACGCGCC CGAAGCGTTG GCAAACTTAA
  	GCGGCGANGA
  351	ANTNNGNNGC AATCTGATCC AATCCGGTTT GTGGACTGCG
  	CTGCGTANCC
  401	GTCCGTTCAG CAAAATCCCT GCCGTCGATG CCGAGCCGTT
  	CGCCATCTTC
  451	GTCAATGCGA TGGACACCAA TCCGCTNGCG GCAGACCCTG
  	TGGTTGTGAT
  501	CAAAGAAGCC GNCGANGATT TCAGACGANG TNTGCTGGTA
  	TTGAGCCGTT
  551	TGACCGAGCG TAAAATCCAT GTGTGTAAGG CAGCTGGCGC
  	AGACGTGCCG
  601	TCTGAAAATG CTGCCAACAT CGAAACACAT GAATTCGGCG
  	GCCCGCATCC
  651	GGCCGGTTTG AGTGGCACGC ACATTCATTT CATTGAGCCG
  	GTCGGTGCAA
  701	ACAAAACCGT TTGGACCATC AATTATCAAG ATGTAATTGC
  	CATCGGACGT
  751	TTGTTTGCAA CAGGCCGTCT GAACACCGAG CGCGTGATTG
  	CTTTGGGTGG
  801	TTCTCAAGTC AACAAACCAC GCCTCTTGCG TACCGTTTTG
  	GGTGCGAAAG
  851	TATCGCAAAT TACTGCGGGC GAATTGGTTG ACGCAGACAA
  	CCGCGTGATT
  901	TCCGGTTCGG TATTGAACGG CGCGATTACA CAAGGCGCGC
  	ACGATTATTT
  951	GGGACGCTAC CACAATCAGA TTTCCGTTAT CGAAGAAGGC
  	CGCAGCAAAG
  1001	AGCTGTTCGG CTGGGTTGCG CCGCAGCCGG ACAAATACTC
  	CATCACGCGT
  1051	ACGACCCTCG GCCATTTCCT GAAAAACAAA CTCTTCAAGT
  	TCACGACAGC
  1101	CGTCAACGGT GGCGACCGCG CCATGGTGCC GATTGGTACT
  	TACGAGCGCG
  1151	TAATGCCGCT AGACATCCTG CCTACCCTGC TTTTGCGCGA
  	TTTAATCGTC
  1201	GGCGATACCG ACAGCGCGCA AGCATTGGGT TGCTTGGAAT
  	TGGACGAAGA
  1251	AGACCTCGCT TTGTGCAGCT TCGTCTGCCC GGGCAAATAC
  	GAATANGGCC
  1301	CGCTGTTGCG TAAGGTGCTG GAAACCNTTG AGAAGGAAGG
  	CTGA

• This encodes a protein having amino acid sequence <SEQ ID 128; ORF22a>:

• 1	MIKIKKGLNL PIAGRPEQVI YDGPVITEVA LLGEEYAGMR
  	PXMKVKEGDA
  51	VKKGQVLFED KKXPGVVFTA PVSGKIAAIH RGEKRVLQSV
  	VIAVEGNDEI
  101	EFERYAPEAL ANLSGXEXXX NLIQSGLWTA LRXRPFSKIP
  	AVDAEPFAIF
  151	VNAMDTNPLA ADPVVVIKEA XXDFRRXXLV LSRLTERKIH
  	VCKAAGADVP
  201	SENAANIETH EFGGPHPAGL SGTHIHFIEP VGANKTVWTI
  	NYQDVIAIGR
  251	LFATGRLNTE RVIALGGSQV NKPRLLRTVL GAKVSQITAG
  	ELVDADNRVI
  301	SGSVLNGAIT QGAHDYLGRY HNQISVIEEG RSKELFGWVA
  	PQPDKYSITR
  351	TTLGHFLKNK LFKFTTAVNG GDRAMVPIGT YERVMPLDIL
  	PTLLLRDLIV
  401	GDTDSAQALG CLELDEEDLA LCSFVCPGKY EXGPLLRKVL
  	ETXEKEG*

• The originally-identified partial strain B sequence (ORF22) shows 94.2% identity over a 158aa overlap with ORF22a:
• 
• The complete strain B sequence (ORF22-1) and ORF22a show 94.9% identity in 447 aa overlap:
• 
• Further work identified a partial gene sequence <SEQ ID 129> from N. gonorrhoeae, which encodes the following amino acid sequence <SEQ ID 130; ORF22ng>:

• 1	MIKIKKGLNL PIAGRPEQVI YDGPAITEVA LLGEEYVGMR
  	PSMKIKEGEA
  51	VKKGQVLFED KKNPGVVFTA PASGKIAAIH RGEKRVLQSV
  	VIAVEGNDEI
  101	EFERYVPEAL AKLSSEKVRR NLIQSGLWTA LRTRPFSKIP
  	AVDAEPFAIF
  151	VNAMDTNPLA ADPTVIIKEA AEDFKRGLLV LSRLTERKIH
  	VCKAAGADVP
  201	SENAANIETH EFGGPHPAGL SGTHIHFIEP VGANKTVWTI
  	NYQDVIAIGR
  251	LFVTGRLNTE RVVALGGLQV NKPRLLRTVL GAKVSQLTAG
  	ELVDADNRVI
  301	SGSVLNGAIA QGAHDYLGRY HN*

• Further work identified complete gonococcal gene <SEQ ID 131>:

• 1	ATGATTAAAA TCAAAAAAGG TCTAAATCTG CCCATCGCGG
  	GCAGACCGGA
  51	GCAAGTCATT TATGACGGCC CGGCCATTAC CGAAGTCGCG
  	TTGCTTGGCG
  101	AAGAATATGT CGGCATGCGC CCCTCGATGA AAATCAAGGA
  	AGGTGAAGCC
  151	GTCAAAAAAG GCCAAGTGCT GTTTGAAGAC AAAAAGAATC
  	CGGGCGTAGT
  201	ATTTACTGCG CCGGCTTCAG GCAAAATCGC CGCTATTCAC
  	CGTGGCGAAA
  251	AGCGCGTACT TCAGTCAGTC GTGATTGCCG TTGAAGGCAA
  	CGACGAAATC
  301	GAGTTCGAAC GCTACGTACC TGAAGCGCTG GCAAAATTGA
  	GCAGCGAAAA
  351	AGTGCGCCGC AACCTGATTC AATCAGGCTT ATGGACTGCG
  	CTTCGCACCC
  401	GTCCGTTCAG CAAAATCCCT GCCGTAGATG CCGAGCCGTT
  	CGCCATCTTC
  451	GTCAATGCGA TGGACACCAA TCCGCTGGCT GCCGACCCTA
  	CGGTCATCAT
  501	CAAAGAAGCC GCCGAAGACT TCAAACGCGG CCTGTTGGTA
  	TTGAGCCGCC
  551	TGACCGAACG TAAAATCCAT GTGTGTAAAG CAGCAGGCGC
  	AGACGTGCCG
  601	TCTGAAAATG CTGCCAATAT CGAAACACAT GAATTTGGCG
  	GCCCGCATCC
  651	TGCCGGCTTG AGTGGCACGC ACATTCATTT CATCGAGCCA
  	GTCGGCGCGA
  701	ATAAAACCGT GTGGACCATC AATTATCAAG ACGTGATTGC
  	TATCGGACGT
  751	TTGTTCGTAA CAGGCCGTCT GAATACCGAG CGCGTGGTTG
  	CCTTGGGCGG
  801	CCTGCAAGTC AACAAACCGC GCCTCTTGCG TACCGTTTTG
  	GGTGCGAAGG
  851	TGTCTCAACT TACCGCCGGC GAATTGGTTG ACGCGGACAA
  	CCGCGTGATT
  901	TCCGGTTCGG TATTGAACGG TGCGATTGCA CAAGGCGCGC
  	ATGATTATTT
  951	GGGACGCTAC CACAATCAGA TTTCCGTTAT CGAAGAAGGC
  	CGCAGCAAAG
  1001	AGCTGTTCGG CTGGGTTGCG CCGCAGCCGG ACAAATACTC
  	CATCACGCGC
  1051	ACCACTCTCG GCCATTTCCT AAAAAACAAA CTCTTCAAGT
  	TCACGACAGC
  1101	CGTCAACGGC GGCGACCGCG CCATGGTACC GATCGGCACT
  	TATGAGCGCG
  1151	TAATGCCGTT GGACATCCTG CCTACCTTGC TTTTGCGCGA
  	TTTAATCGTC
  1201	GGCGATACCG ACAGCGCGCA GGCTTTGGGT TGCTTGGAAT
  	TGGACGAAGA
  1251	AGACCTCGCT TTGTGCAGCT TCGTCTGCCC GGGCAAATAC
  	GAATACGGCC
  1301	CGCTGTTGCG CAAAGTGCTG GAAACCATTG AGAAGGAAGG
  	CTGA

• This encodes a protein having amino acid sequence <SEQ ID 132; ORF22ng-1>:

• 1	MIKIKKGLNL PIAGRPEQVI YDGPAITEVA LLGEEYVGMR
  	PSMKIKEGEA
  51	VKKGQVLFED KKNPGVVFTA PASGKIAAIH RGEKRVLQSV
  	VIAVEGNDEI
  101	EFERYVPEAL AKLSSEKVRR NLIQSGLWTA LRTRPFSKIP
  	AVDAEPFAIF
  151	VNAMDTNPLA ADPTVIIKEA AEDFKRGLLV LSRLTERKIH
  	VCKAAGADVP
  201	SENAANIETH EFGGPHPAGL SGTHIHFIEP VGANKTVWTI
  	NYQDVIAIGR
  251	LFVTGRLNTE RVVALGGLQV NKPRLLRTVL GAKVSQLTAG
  	ELVDADNRVI
  301	SGSVLNGAIA QGAHDYLGRY HNQISVIEEG RSKELFGWVA
  	PQPDKYSITR
  351	TTLGHFLKNK LFKFTTAVNG GDRAMVPIGT YERVMPLDIL
  	PTLLLRDLIV
  401	GDTDSAQALG CLELDEEDLA LCSFVCPGKY EYGPLLRKVL
  	ETIEKEG*

• The originally-identified partial strain B sequence (ORF22) shows 93.7% identity over a 158aa overlap with ORF22ng:
• 
• The complete sequences from strain B (ORF22-1) and gonococcus (ORF22ng) show 96.2% identity in 447 aa overlap:
• 
• Computer analysis of these sequences gave the following results:
• Homology with 48 kDa Outer Membrane Protein of Actinobacillus pleuropneumoniae (Accession Number U24492).
• ORF22 and this 48 kDa protein show 72% aa identity in 158aa overlap:

• Orf22	1	MIKIKKGLNLPIAGRPEQAVYDGPAITEVALLGEEYAGMRPSMKVKEGDAVKKGQVLFED	60
  		MI IKKGL+LPIAG P Q +++G  + EVA+LGEEY GMRPSMKV+EGD VKKGQVLFED
  48kDa	1	MITIKKGLDLPIAGTPAQVIHNGNTVNEVAMLGEEYVGMRPSMKVREGDVVKKGQVLFED	60
  orf22	61	KKNPGVVFTAPASGKIAAIHRGEKRVLQSVVIAVEXNDEIEFERYAPEALANLSGEEVRR	120
  		KKNPGVVFTAPASG +  I+RGEKRVLQSVVI VE +++I F RY    LA+LS E+V++
  48kDa	61	KKNPGVVFTAPASGTVVTINRGEKRVLQSVVIKVEGDEQITFTRYEAAQLASLSAEQVKQ	120
  orf22	121	NLIQSGLWTALRTRPFSKIPAVDAEPFAIFVNAMDTNP	158
  		NLI+SGLWTA RTRPFSK+PA+DA P +IFVNAMDTNP
  48kDa	121	NLIESGLWTAFRTRPFSKVPALDAIPSSIFVNAMDTNP	158

• ORF22a also shows homology to the 48 kDa Actinobacillus pleuropneumoniae protein:

• gi|1185395 (U24492) 48 kDa outer membrane protein [Actinobacillus
  pleuropneumoniae]
  Length = 449
  Score = 530 bits (1351), Expect = e−150
  Identities = 274/450 (60%), Positives = 323/450 (70%), Gaps = 4/450 (0%)

  Query:	1	MIKIKKGLNLPIAGRPEQVIYDGPVITEVALLGEEYAGMRPXMKVKEGDAVKKGQVLFED	60
  		MI IKKGL+LPIAG P QVI++G  + EVA+LGEEY GMRP MKV+EGD VKKGQVLFED
  Sbjct:	1	MITIKKGLDLPIAGTPAQVIHNGNTVNEVAMLGEEYVGMRPSMKVREGDVVKKGQVLFED	60
  Query:	61	KKXPGVVFTAPVSGKIAAIHRGEKRVLQSVVIAVEGNDEIEFERYAPEALANLSGXEXXX	120
  		KK PGVVFTAP SG +  I+RGEKRVLQSVVI VEG+++I F RY    LA+LS  +
  Sbjct:	61	KKNPGVVFTAPASGTVVTINRGEKRVLQSVVIKVEGDEQITFTRYEAAQLASLSAEQVKQ	120
  Query:	121	NLIQSGLWTALRXRPFSKIPAVDAEPFAIFVNAMDTNPLAADPVVVIKEAXXDFRRXXLV	180
  		NLI+SGLWTA R RPFSK+PA+DA P +IFVNAMDTNPLAADP VV+KE   DF+    V
  Sbjct:	121	NLIESGLWTAFRTRPFSKVPALDAIPSSIFVNAMDTNPLAADPEVVLKEYETDFKDGLTV	180
  Query:	181	LSRL--TERKIHVCKAAGADVP-SENAANIETHEFGGPHPAGLSGTHIHFIEPVGANKTV	237
  		L+RL   ++ +++CK A +++P S     I    F G HPAGL GTHIHF++PVGA K V
  Sbjct:	181	LTRLFNGQKPVYLCKDADSNIPLSPAIEGITIKSFSGVHPAGLVGTHIHFVDPVGATKQV	240
  Query:	238	WTINYQDVIAIGRLFATGRLNTERVIALGGSQVNKPRLLRTVLGAKVSQITAGELVDADN	297
  		W +NYQDVIAIG+LF TG L T+R+I+L G QV  PRL+RT LGA +SQ+TA EL   +N
  Sbjct:	241	WHLNYQDVIAIGKLFTTGELFTDRIISLAGPQVKNPRLVRTRLGANLSQLTANELNAGEN	300
  Query:	298	RVISGSVLNGAITQGAHDYLGRYHNQISVIEEGRSKELFGWVAPQPDKYSITRTTLGHFL	357
  		RVISGSVL+GA   G  DYLGRY  Q+SV+ EGR KELFGW+ P  DK+SITRT LGHF
  Sbjct:	301	RVISGSVLSGATAAGPVDYLGRYALQVSVLAEGREKELFGWIMPGSDKFSITRTVLGHFG	360
  Query:	358	KNKLFKFTTAVNGGDRAMVPIGTYERVMXXXXXXXXXXXXXXVGDTDSAQXXXXXXXXXX	417
  		K KLF FTTAV+GG+RAMVPIG YERVM               GDTDSAQ
  Sbjct:	361	K-KLFNFTTAVHGGERAMVPIGAYERVMPLDIIPTLLLRDLAAGDTDSAQNLGCLELDEE	419
  Query:	418	XXXXXSFVCPGKYEXGPLLRKVLETXEKEG	447
  		++VCPGK   GP+LR  LE  EKEG

• ORF22ng-1 also shows homology with the OMP from A. pleuropneumoniae:

• gi|1185395 (U24492) 48 kDa outer membrane protein [Actinobacillus
  pleuropneumoniae] Length = 449
  Score = 555 bits (1414), Expect = e−157
  Identities = 284/450 (63%), Positives = 337/450 (74%), Gaps = 4/450 (0%)

  Query:	27	MIKIKKGLNLPIAGRPEQVIYDGPAITEVALLGEEYVGMRPSMKIKEGEAVKKGQVLFED	86
  		MI IKKGL+LPIAG P QVI++G  + EVA+LGEEYVGMRPSMK++EG+ VKKGQVLFED
  Sbjct:	1	MITIKKGLDLPIAGTPAQVIHNGNTVNEVAMLGEEYVGMRPSMKVREGDVVKKGQVLFED	60
  Query:	87	KKNPGVVFTAPASGKIAAIHRGEKRVLQSVVIAVEGNDEIEFERYVPEALAKLSSEKVRR	146
  		KKNPGVVFTAPASG +  I+RGEKRVLQSVVI VEG+++I F RY    LA LS+E+V++
  Sbjct:	61	KKNPGVVFTAPASGTVVTINRGEKRVLQSVVIKVEGDEQITFTRYEAAQLASLSAEQVKQ	120
  Query:	147	NLIQSGLWTALRTRPFSKIPAVDAEPFAIFVNAMDTNPLAADPTVIIKEAAEDFKRGLLV	206
  		NLI+SGLWTA RTRPFSK+PA+DA P +IFVNAMDTNPLAADP V++KE   DFK GL V
  Sbjct:	121	NLIESGLWTAFRTRPFSKVPALDAIPSSIFVNAMDTNPLAADPEVVLKEYETDFKDGLTV	180
  Query:	207	LSRL--TERKIHVCKAAGADVP-SENAANIETHEFGGPHPAGLSGTHIHFIEPVGANKTV	263
  		L+RL   ++ +++CK A +++P S     I    F G HPAGL GTHIHF++PVGA K V
  Sbjct:	181	LTRLFNGQKPVYLCKDADSNIPLSPAIEGITIKSFSGVHPAGLVGTHIHFVDPVGATKQV	240
  Query:	264	WTINYQDVIAIGRLFVTGRLNTERVVALGGLQVNKPRLLRTVLGAKVSQLTAGELVDADN	323
  		W +NYQDVIAIG+LF TG L T+R+++L G QV  PRL+RT LGA +SQLTA EL   +N
  Sbjct:	241	WHLNYQDVIAIGKLFTTGELFTDRIISLAGPQVKNPRLVRTRLGANLSQLTANELNAGEN	300
  Query:	324	RVISGSVLNGAIAQGAHDYLGRYHNQISVIEEGRSKELFGWVAPQPDKYSITRTTLGHFL	383
  		RVISGSVL+GA A G  DYLGRY  Q+SV+ EGR KELFGW+ P  DK+SITRT LGHF
  Sbjct:	301	RVISGSVLSGATAAGPVDYLGRYALQVSVLAEGREKELFGWIMPGSDKFSITRTVLGHFG	360
  Query:	384	KNKLFKFTTAVNGGDRAMVPIGTYERVMXXXXXXXXXXXXXXVGDTDSAQXXXXXXXXXX	443
  		K KLF FTTAV+GG+RAMVPIG YERVM               GDTDSAQ
  Sbjct:	361	K-KLFNFTTAVHGGERAMVPIGAYERVMPLDIIPTLLLRDLAAGDTDSAQNLGCLELDEE	419
  Query:	444	XXXXXSFVCPGKYEYGPLLRKVLETIEKEG	473
  		++VCPGK  YGP+LR  LE IEKEG
  Sbjct:	420	DLALCTYVCPGKNNYGPMLRAALEKIEKEG	449

• Based on this analysis, including the homology with the outer membrane protein of Actinobacillus pleuropneumoniae, it was predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF22-1 (35.4 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 5A shows the results of affinity purification of the GST-fusion protein, and FIG. 5B shows the results of expression of the His-fusion in E. coli. Purified GST-fusion protein was used to immunise mice, whose sera were used for ELISA (positive result) and FACS analysis (FIG. 5C). These experiments confirm that ORF22-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 16
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 133>:

• 1	..GCGnCGnAAA TCATCCATCC CC..nACGTC GTAGGCCCTG
  	AAGCCAACTG
  51	GTTTTTTATG GTAGCCAGTA CGTTTGTGAT TGCTTTGATT
  	GGTTATTTTG
  101	TTACTGAAAA AATCGTCGAA CCGCAATTGG GCCCTTATCA
  	ATCAGATTTG
  151	TCACAAGAAG AAAAAGACAT TCGGCATTCC AATGAAATCA
  	CGCCTTTGGA
  201	ATATAAAGGA TTAATTTGGG CTGGCGTGGT GTTTGTTGCC
  	TTATCCGCCC
  251	TATTGGCTTG GAGCATCGTC CCTGCCGACG GTATTTTGCG
  	TCATCCTGAA
  301	ACAGGATTGG TTTCCGGTTC GCCGTTTTTA AAATCGATTG
  	TTGTTTTTAT
  351	TTTCTTGTTG TTTGCACTGC CGGGCATTGT TTATGGCCGG
  	GTAACCCGAA
  401	GTTTGCGCGG CGAACAGGAA GTCGTTAATG CGmyGGCCGA
  	ATCGATGAGT
  451	ACTCTGGsGC TTTmTTTGsw CAkcATCTTT TTTGCCGCAC
  	AGTTTGTCGC
  501	ATTTTTTAAT TGGACGAATA TTGGGCAATA TATTGCCGTT
  	AAAGGGGCGA
  551	CGTTCTTAAA AGAAGTCGGC TTGGGCGGCA GCGTGTTGTT
  	TATCGGTTTT
  601	ATTTTAATTT GTGCTTTTAT CAATCTGATG ATAGGCTCCG
  	CCTCCGCGCA
  651	ATGGGCGGTA ACTGCGCCGA TTTTCGTCCC TATGCTGATG
  	TTGGCCGGCT
  701	ACGCGCCCGA AGTCATTCAA GCCGCTTACC GCATCGGTGA
  	TTCCGTTACC
  751	AATATTATTA CGCCGATGAT GAGTTATTTC GGGCTGATTA
  	TGGCGACGGT
  801	GrkCmmmTAC AAAAAAGATG CGGGCGTGGG TaCGcTGATT
  	wCTATGATGT
  851	TGCCGTATTC CGCTTTCTTC TTGATTGCgT GGATTGCCTT
  	ATTCTGCATT
  901	TGGGTATTTg TTTTGGGCCT GCCCGTCGGT CCCGGCGCGC
  	CCACATTCTA
  951	TCCCGCACCT TAA

• This corresponds to the amino acid sequence <SEQ ID 134; ORF12>:

• 1	..AXXIIHPXXV VGPEANWFFM VASTFVIALI GYFVTEKIVE
  	PQLGPYQSDL
  51	SQEEKDIRHS NEITPLEYKG LIWAGVVFVA LSALLAWSIV
  	PADGILRHPE
  101	TGLVSGSPFL KSIVVFIFLL FALPGIVYGR VTRSLRGEQE
  	VVNAXAESMS
  151	TLXLXLXXIF FAAQFVAFFN WTNIGQYIAV KGATFLKEVG
  	LGGSVLFIGF
  201	ILICAFINLM IGSASAQWAV TAPIFVPMLM LAGYAPEVIQ
  	AAYRIGDSVT
  251	NIITPMMSYF GLIMATVXXY KKDAGVGTLI XMMLPYSAFF
  	LIAWIALFCI
  301	WVFVLGLPVG PGAPTFYPAP *

• Further sequence analysis revealed the complete DNA sequence <SEQ ID 135> to be:

• 1	ATGAGTCAAA CCGATACGCA ACGGGACGGA CGATTTTTAC
  	GCACAGTCGA
  51	ATGGCTGGGC AATATGTTGC CGCATCCGGT TACGCTTTTT
  	ATTATTTTCA
  101	TTGTGTTATT GCTGATTGCC TCTGCCGTCG GTGCGTATTT
  	CGGACTATCC
  151	GTCCCCGATC CGCGCCCTGT TGGTGCGAAA GGACGTGCCG
  	ATGACGGTTT
  201	GATTTACATT GTCAGCCTGC TCAATGCCGA CGGTTTTATC
  	AAAATCCTGA
  251	CGCATACCGT TAAAAATTTC ACCGGTTTCG CGCCGTTGGG
  	AACGGTGTTG
  301	GTTTCTTTAT TGGGCGTGGG GATTGCGGAA AAATCGGGCT
  	TGATTTCCGC
  351	ATTAATGCGC TTATTGCTCA CAAAATCGCC ACGCAAACTC
  	ACTACTTTTA
  401	TGGTTGTTTT TACAGGGATT TTATCTAATA CCGCTTCTGA
  	ATTGGGCTAT
  451	GTCGTCCTAA TCCCTTTGTC CGCCATCATC TTTCATTCCC
  	TCGGCCGCCA
  501	TCCGCTTGCC GGTCTGGCTG CGGCTTTCGC CGGCGTTTCG
  	GGCGGTTATT
  551	CGGCCAATCT GTTCTTAGGC ACAATCGATC CGCTCTTGGC
  	AGGCATCACC
  601	CAACAGGCGG CGCAAATCAT CCATCCCGAC TACGTCGTAG
  	GCCCTGAAGC
  651	CAACTGGTTT TTTATGGTAG CCAGTACGTT TGTGATTGCT
  	TTGATTGGTT
  701	ATTTTGTTAC TGAAAAAATC GTCGAACCGC AATTGGGCCC
  	TTATCAATCA
  751	GATTTGTCAC AAGAAGAAAA AGACATTCGG CATTCCAATG
  	AAATCACGCC
  801	TTTGGAATAT AAAGGATTAA TTTGGGCTGG CGTGGTGTTT
  	GTTGCCTTAT
  851	CCGCCCTATT GGCTTGGAGC ATCGTCCCTG CCGACGGTAT
  	TTTGCGTCAT
  901	CCTGAAACAG GATTGGTTTC CGGTTCGCCG TTTTTAAAAT
  	CGATTGTTGT
  951	TTTTATTTTC TTGTTGTTTG CACTGCCGGG CATTGTTTAT
  	GGCCGGGTAA
  1001	CCCGAAGTTT GCGCGGCGAA CAGGAAGTCG TTAATGCGAT
  	GGCCGAATCG
  1051	ATGAGTACTC TGGGGCTTTA TTTGGTCATC ATCTTTTTTG
  	CCGCACAGTT
  1101	TGTCGCATTT TTTAATTGGA CGAATATTGG GCAATATATT
  	GCCGTTAAAG
  1151	GGGCGACGTT CTTAAAAGAA GTCGGCTTGG GCGGCAGCGT
  	GTTGTTTATC
  1201	GGTTTTATTT TAATTTGTGC TTTTATCAAT CTGATGATAG
  	GCTCCGCCTC
  1251	CGCGCAATGG GCGGTAACTG CGCCGATTTT CGTCCCTATG
  	CTGATGTTGG
  1301	CCGGCTACGC GCCCGAAGTC ATTCAAGCCG CTTACCGCAT
  	CGGTGATTCC
  1351	GTTACCAATA TTATTACGCC GATGATGAGT TATTTCGGGC
  	TGATTATGGC
  1401	GACGGTGATC AAATACAAAA AAGATGCGGG CGTGGGTACG
  	CTGATTTCTA
  1451	TGATGTTGCC GTATTCCGCT TTCTTCTTGA TTGCGTGGAT
  	TGCCTTATTC
  1501	TGCATTTGGG TATTTGTTTT GGGCCTGCCC GTCGGTCCCG
  	GCGCGCCCAC
  1551	ATTCTATCCC GCACCTTAA

• This corresponds to the amino acid sequence <SEQ ID 136; ORF12-1>:

• 1	MSQTDTQRDG RFLRTVEWLG NMLPHPVTLF IIFIVLLLIA
  	SAVGAYFGLS
  51	VPDPRPVGAK GRADDGLIYI VSLLNADGFI KIL THTVKNF
  	TGFAPLGTVL
  101	VSLLGVGIAE KSGLISALMR LLLTKSPRKL TTFMVVFTGI
  	LSNTASELGY
  151	VVLIPLSAII FHSLGRHPLA GLAAAFAGVS GGYSANLFLG
  	TIDPLLAGIT
  201	QQAAQIIHPD YVVGPEANWF FMVASTFVIA LIGYFV TEKI
  	VEPQLGPYQS
  251	DLSQEEKDIR HSNEITPLEY KGLIWAGVVF VALSALLAWS
  	IVPADGILRH
  301	PETGLVSGSP FLKSIVVFIF LLFALPGIVY GRVTRSLRGE
  	QEVVNAMAES
  351	MSTLGLYLVI IFFAAQFVAF FNWTNIGQYI AVKGATFLKE
  	VGLGGSVLFI
  401	GFILICAFIN LMIGSASAQW AVTAPIFVPM LMLAGYAPEV
  	IQAAYRIGDS
  451	VTNIITPMMS YFGLIMATVI KYKKDAGVGT LISMMLPYSA
  	FFLIAWIALF
  501	CIWVFVLGLP VGPGAPTFYP AP*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF12 shows 96.3% identity over a 320aa overlap with an ORF (ORF12a) from strain A of N. meningitidis:
• 
• The complete length ORF12a nucleotide sequence <SEQ ID 137> is:

• 1	ATGAGTCAAA CCGATACGCA ACGGGACGGA CGATTTTTAC
  	GCACAGTCGA
  51	ATGGCTGGGC AATATGTTGC CGCACCCGGT TACGCTTTTT
  	ATTATTTTCA
  101	TTGTGTTATT GCTGATTGCC TCTGCCGCCG GTGCGTATTT
  	CGGACTATCC
  151	GTCCCCGATC CGCGCCCTGT TGGTGCGAAA GGACGTGCCG
  	ATGACGGTTT
  201	GATTCACGTT GTCAGCCTGC TCGATGCTGA CGGTTTGATC
  	AAAATCCTGA
  251	CGCATACCGT TAAAAATTTC ACCGGTTTCG CGCCGTTGGG
  	AACGGTGTTG
  301	GTTTCTTTAT TGGGCGTGGG GATTGCGGAA AAATCGGGCT
  	TGATTTCCGC
  351	ATTAATGCGC TTATTGCTCA CAAAATCTCC ACGCAAACTC
  	ACTACTTTTA
  401	TGGTTGTTTT TACAGGGATT TTATCTAATA CCGCTTCTGA
  	ATTGGGCTAT
  451	GTCGTCCTAA TCCCTTTGTC CGCCATCATC TTTCATTCCC
  	TCGGCCGCCA
  501	TCCGCTTGCC GGTCTGGCTG CGGCTTTCGC CGGCGTTTCG
  	GGCGGTTATT
  551	CGGCCAATCT GTTCTTAGGC ACAATCGATC CGCTCTTGGC
  	AGGCATCACC
  601	CAACAGGCGG CGCAAATCAT CCATCCCGAC TACGTCGTAG
  	GCCCTGAAGC
  651	CAACTGGTTT TTTATGGTAG CCAGTACGTT TGTGATTGCT
  	TTGATTGGTT
  701	ATTTTGTTAC TGAAAAAATC GTCGAACCGC AATTGGGCCC
  	TTATCAATCA
  751	GATTTGTCAC AAGAAGAAAA AGACATTCGA CATTCCAATG
  	AAATCACGCC
  801	TTTGGAATAT AAAGGATTAA TTTGGGCTGG CGTGGTGTTT
  	GTTGCCTTAT
  851	CCGCCCTATT GGCTTGGAGC ATCGTCCCTG CCGACGGTAT
  	TTTGCGTCAT
  901	CCTGAAACAG GATTGGTTTC CGGTTCGCCG TTTTTAAAAT
  	CAATTGTTGT
  951	TTTTATTTTC TTGTTGTTTG CACTGCCGGG CATTGTTTAT
  	GGCCGGGTAA
  1001	CCCGAAGTTT GCGCGGCGAA CAGGAAGTCG TTAATGCGAT
  	GGCCGAATCG
  1051	ATGAGTACTC TGGGGCTTTA TTTGGTCATC ATCTTTTTTG
  	CCGCACAGTT
  1101	TGTCGCATTT TTTAATTGGA CGAATATTGG GCAATATATT
  	GCCGTTAAAG
  1151	GGGCGACGTT CTTAAAAGAA GTCGGCTTGG GCGGCAGCGT
  	GTTGTTTATC
  1201	GGTTTTATTT TAATTTGTGC TTTTATCAAT CTGATGATAG
  	GCTCCGCCTC
  1251	CGCGCAATGG GCGGTAACTG CGCCGATTTT CGTCCCTATG
  	CTGATGTTGG
  1301	CCGGCTACGC GCCCGAAGTC ATTCAAGCCG CTTACCGCAT
  	CGGTGATTCC
  1351	GTTACCAATA TTATTACGCC GATGATGAGT TATTTCGGGC
  	TGATTATGGC
  1401	GACGGTGATC AAATACAAAA AAGATGCGGG CGTGGGTACG
  	CTGATTTCTA
  1451	TGATGTTGCC GTATTCCGCT TTCTTCTTGA TTGCGTGGAT
  	TGCCTTATTC
  1501	TGCATTTGGG TATTTGTTTT GGGCCTGCCC GTCGGTCCCG
  	GCGCGCCCAC
  1551	ATTCTATCCC GCACCTTAA

• This encodes a protein having amino acid sequence <SEQ ID 138>:

• 1	MSQTDTQRDG RFLRTVEWLG NMLPHPVTLF IIFIVLLLIA
  	SAAGAYFGLS
  51	VPDPRPVGAK GRADDGLIHV VSLLDADGLI KIL THTVKNF
  	TGFAPLGTVL
  101	VSLLGVGIAE KSGLISALMR LLLTKSPRKL TTFMVVFTGI
  	LSNTASELGY
  151	VVLIPLSAII FHSLGRHPLA GLAAAFAGVS GGYSANLFLG
  	TIDPLLAGIT
  201	QQAAQIIHPD YVVGPEANWF FMVASTFVIA LIGYFV TEKI
  	VEPQLGPYQS
  251	DLSQEEKDIR HSNEITPLEY KGLIWAGVVF VALSALLAWS
  	IVPADGILRH
  301	PETGLVSGSP FLKSIVVFIF LLFALPGIVY GRVTRSLRGE
  	QEVVNAMAES
  351	MSTLGLYLVI IFFAAQFVAF FNWTNIGQYI AVKGATFLKE
  	VGLGGSVLFI
  401	GFILICAFIN LMIGSASAQW AVTAPIFVPM LMLAGYAPEV
  	IQAAYRIGDS
  451	VTNIITPMMS YFGLIMATVI KYKKDAGVGT LISMMLPYSA
  	FFLIAWIALF
  501	CIWVFVLGLP VGPGAPTFYP AP*

• ORF12a and ORF12-1 show 99.0% identity in 522 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF12 shows 92.5% identity over a 320aa overlap with a predicted ORF (ORF12.ng) from N. gonorrhoeae:
• 
• The complete length ORF12ng nucleotide sequence <SEQ ID 139> is:

• 1	ATGAGTCAAA CCGACGCGCG TCGTAGCGGA CGATTTTTAC
  	GCACAGTCGA
  51	ATGGCTGGGC AATATGTTGC CGCACCCGGT TACGCTTTTT
  	ATTATTTTCA
  101	TTGTGTTATT GCTGATTGcc tctgCCGTCG GTGCGTATTT
  	CGGACTATCC
  151	GTCCCCGATC CGCGTCCTGT TGGGGCGAAA GGACGTGCCG
  	ATGACGGTTT
  201	GATTCACGTT GTCAGCCTGC TCGATGCCGA CGGTTTGATC
  	AAAATCCTGA
  251	CGCATACCGT TAAAAATTTC ACCGGTTTCG CGCCGTTGGG
  	AACGGTGTTG
  301	GTTTCTTTAT TGGGCGTGGG GATTGCGGAA AAATCGGGCT
  	TGATTTCCGC
  351	ATTAATGCGC TTATTGCTCA CAAAATCCCC ACGCAAACTC
  	ACTACTTTTA
  401	TGGTTGTTTT TACAGGGATT TTATCCAATA CGGCTTCTGA
  	ATTGGGCTAT
  451	GTCGTCCTAA TCCCTTTGTC CGCCGTCATC TTTCATTCGC
  	TCGGCCGCCA
  501	TCCGCTTGCC GGTTTGGCTG CGGCTTTCGC CGGCGTTTCG
  	GGCGGTTATT
  551	CGGCCAATCT GTTCTTAGGC ACAATCGATC CGCTCTTGGC
  	AGGCATCACC
  601	CAACAGGCGG CGCAAATCAT CCATCCCGAC TACGTCGTAG
  	GCCCTGAAGC
  651	CAACTGGTTT TTTATGGCAG CCAGTACGTT TGTGATTGCT
  	TTGATTGGTT
  701	ATTTTGTTAC TGAAAAAATC GTCGAACCGC AATTGGGCCC
  	TTATCAATCA
  751	GATTTGTCAC AAGAAGAAAA AGACATTCGG CATTCCAATG
  	AAATCACGCC
  801	TTTGGAATAT AAAGGATTAA TTTGGGCAGG CGTGGTGTTT
  	GTTGCCTTAT
  851	CCGCCCTATT GGCTTGGAGC ATCGTCCCTG CCGACGGTAT
  	TTTGCGTCAT
  901	CCTGAAACAG GATTGGTTGC CGGTTCGCCG TTTTTAAAAT
  	CGATTGTTGT
  951	TTTTATTTTC TTGTTGTTTG CGCTGCCGGG CATTGTTTAT
  	GGCCGGATAA
  1001	CCCGAAGTTT GCGCGGCGAA CGGGAAGTCG TTAATGCGAT
  	GGCCGAATCG
  1051	ATGAGTACTT TGGGACTTTA TTTGGTCATC ATCTTTTTTG
  	CCGCACAGTT
  1101	TGTCGCATTT TTTAATTGGA CGAATATTGG GCAATATATT
  	GCCGTTAAAG
  1151	GGGCGGTGTT CTTAAAAGAA GTCGGCTTGG GCGGCAGTGT
  	GTTGTTTATC
  1201	GGTTTTATTT TAATTTGTGC TTTTATCAAT CTGATGATAG
  	GCTCCGCCTC
  1251	CGCGCAATGG GCGGTAACTG CGCCGATTTT CGTCCCTATG
  	CTGATGTTGG
  1301	CCGGCTACGC GCCCGAAGTC ATTCAAGCCG CTTACCGCAT
  	CGGTGATTCC
  1351	GTTACCAATA TTATTACGCC GATGATGAGT TATTTCGGGC
  	TGATTATGGC
  1401	GACGGTAATC AAATACAAAA AAGATGCGGG CGTAGGCACG
  	CTGATTTCTA
  1451	TGATGTTGCC GTATTCCGCT TTCTTCTTAA TTGCATGGAT
  	CGCCTTATTC
  1501	TGCATTTGGG TATTTGTTTT GGGTCTGCCC GTCGGTCCCG
  	GCACACCCAC
  1551	ATTCTATCCG GTGCCTTAA

• This encodes a protein having amino acid sequence <SEQ ID 140>:

• 1	MSQTDARRSG RFLRTVEWLG NMLPHPVTLF IIFIVLLLIA
  	SAVGAYFGLS
  51	VPDPRPVGAK GRADDGLIHV VSLLDADGLI KIL THTVKNF
  	TGFAPLGTVL
  101	VSLLGVGIAE KSGLISALMR LLLTKSPRKL TTFMVVFTGI
  	LSNTASELGY
  151	VVLIPLSAVI FHSLGRHPLA GLAAAFAGVS GGYSANLFLG
  	TIDPLLAGIT
  201	QQAAQIIHPD YVVGPEANWF FMAASTFVIA LIGYFV TEKI
  	VEPQLGPYQS
  251	DLSQEEKDIR HSNEITPLEY KGLIWAGVVF VALSALLAWS
  	IVPADGILRH
  301	PETGLVAGSP FLKSIVVFIF LLFALPGIVY GRITRSLRGE
  	REVVNAMAES
  351	MSTLGLYLVI IFFAAQFVAF  FNWTNIGQYI AVKGAVFLKK
  	FRLGGSVLFI
  401	GFILICAFIN LMIGSASAQW AVTAPIFVPM LMLAGNAPQV
  	IQAAYRIGDS
  451	VTNIITPMMS YFGLIMATVI KYKKDAGVGT LISMMLPYSA
  	FFLIAWIALF
  501	CIWVFVLGLP VGPGTPTFYP VP*

• ORF12ng shows 97.1% identity in 522 aa overlap with ORF12-1:
• 
• In addition, ORF12ng shows significant homology with a hypothetical protein from E. coli:

• sp|P46133|YDAH_ECOLI HYPOTHETICAL 55.1 KD PROTEIN IN OGT-DBPA
  INTERGENIC REGION
  >gi|1787597 (AE000231) hypothetical protein in ogt 5′region
  [Escherichia coli]
  Length = 510
  Score = 329 bits (835), Expect = 2e−89
  Identities = 178/507 (35%), Positives = 281/507 (55%), Gaps = 15/507 (2%)

  Query:	8	RSGRFLRTVEWLGNMLPHPVTXXXXXXXXXXXASAVGAYFGLSVPDPRPVGAKGRADDGL	67
  		+SG+    VE +GN +PHP              +A+ + FG+S  +P         D
  Sbjct:	13	QSGKLYGWVERIGNKVPHPFLLFIYLIIVLMVTTAILSAFGVSAKNP--------TDGTP	64
  Query:	68	IHVVSLLDADGLIKILTHTVKNFTGFAPXXXXXXXXXXXXIAEKSGLISALMRLLLTKSP	127
  		+ V +LL  +GL   L + +KNF+GFAP            +AE+ GL+ ALM  + +
  Sbjct:	65	VVVKNLLSVEGLHWFLPNVIKNFSGFAPLGAILALVLGAGLAERVGLLPALMVKMASHVN	124
  Query:	128	RKLTTFMVVFTGILSNTASELGYVVLIPLSAVIFHSLGRHPLAGLAAAFAGVSGGYSANL	187
  		+  ++MV+F    S+ +S+   V++ P+ A+IF ++GRHP+AGL AA AGV  G++ANL
  Sbjct:	125	ARYASYMVLFIAFFSHISSDAALVIMPPMGALIFLAVGRHPVAGLLAAIAGVGCGFTANL	184
  Query:	188	FLGTIDPLLAGITQQAAQIIHPDYVVGPEANWFFMAASTFVIALIGYFVTEKIVEPQLGP	247
  		+ T D LL+GI+ +AA   +P   V    NW+FMA+S  V+ ++G  +T+KI+EP+LG
  Sbjct:	185	LIVTTDVLLSGISTEAAAAFNPQMHVSVIDNWYFMASSVVVLTIVGGLITDKIIEPRLGQ	244
  Query:	248	YQSDLSQEEKDIRHSNEITPLEYKGLIWAGVVFVALSALLAWSIVPADGILRHPETGLVA	307
  		+Q +  ++ + +  S         GL  AGVV +   A +A  ++P +GILR P    V
  Sbjct:	245	WQGNSDEKLQTLTESQRF------GLRIAGVVSLLFIAAIALMVIPQNGILRDPINHTVM	298
  Query:	308	GSPFLKSIVVFIFLLFALPGIVYGRITRSLRGEREVVNAMAESMSTLGLYLXXXXXXXXX	367
  		SPF+K IV  I L F +  + YG  TR++R + ++ + M E M  +  ++
  Sbjct:	299	PSPFIKGIVPLIILFFFVVSLAYGIATRTIRRQADLPHLMIEPMKEMAGFIVMVFPLAQF	358
  Query:	368	XXXXNWTNIGQYIAVKGAVFLKEVGLGGSVLFIGFILICAFINLMIGSASAQWAVTAPIF	427
  		NW+N+G++IAV     L+  GL G   F+G  L+ +F+ +I S SA W++ APIF
  Sbjct:	359	VAMFNWSNMGKFIAVGLTDILESSGLSGIPAFVGLALLSSFLCMFIASGSAIWSILAPIF	418
  Query:	428	VPMLMLAGYAPEVIQAAYRIGDSVTNIITPMMSYFGLIMATVIKYKKDAGVGTLISMMLP	487
  		VPM ML G+ P   Q  +RI DS    + P+  +  L +  + +YK DA +GT  S++LP
  Sbjct:	419	VPMFMLLGFHPAFAQILFRIADSSVLPLAPVSPFVPLFLGFLQRYKPDAKLGTYYSLVLP	478
  Query:	488	YSAFFLIAWIALFCIWVFVLGLPVGPG	514
  		Y   FL+ W+ +   W +++GLP+GPG
  Sbjct:	479	YPLIFLVVWLLMLLAW-YLVGLPIGPG	504

• Based on this analysis, including the presence of several putative transmembrane domains and the predicted actinin-type actin-binding domain signature (shown in bold) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 17
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 141>:

• 1	..ACAGCCGGCG CAGCAGGTTn CnCGGTCTTC GTTTTCGTAA
  	CGGACAGTCA
  51	GGTGGAGGTG TTCGGGAACA TCCAGACCGC AGTGGAAACA
  	GGTTTTTTTC
  101	ATGGCATTTC GGTTTCGTCT GTGTTTGGTG CGGCGGCACA
  	AGACTCGGCA
  151	ATgGCTTCGC GCAGTGCGTC TATACCGGTA TTTTCAGCAA
  	CGGAAATGCG
  201	GACGGcGgCA ATTTTTCCCG CAGCGTCGCG CCATATGCCC
  	GTGTTTTgTT
  251	CTTCAGACGG CAGCAGGTCG GTTTTGTTGT ACACCTTgAT
  	GCACGGAaTA
  301	TCGCCGGCAT GGATTTCTTG CAGTACGTTT TCCACGTCTT
  	CAATCTGCTG
  351	TCCGCTGTTC GGAGCGGCGG CATCGACGAC GTGCAGCAGC
  	ACATCgGcTT
  401	gCGCGGTTTC TTCCAGCGTG GCgGAAAAGG CGGAAATCAG
  	TTTgTGCGGC
  451	agATyGCTnA CGAATCCGAC GGTATCGGTC AGGATAATGC
  	TGCATTCGGG
  501	ACT..

• This corresponds to the amino acid sequence <SEQ ID 142; ORF14>:

• 1	..TAGAAGXXVF VFVTDSQVEV FGNIQTAVET GFFHGISVSS
  	VFGAAAQDSA
  51	MASRSASIPV FSATEMRTAA IFPAASRHMP VFCSSDGSRS
  	VLLYTLMHGI
  101	SPAWISCSTF STSSICCPLF GAAASTTCSS TSACAVSSSV
  	AEKAEISLCG
  151	RXLTNPTVSV RIMLHSG..

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF From N. meningitidis (Strain A)
• ORF14 shows 94.0% identity over a 167aa overlap with an ORF (ORF14a) from strain A of N. meningitidis:
• 
• The complete length ORF14a nucleotide sequence <SEQ ID 143> is:

• 1	ATGGAGGATT TGCAGGAAAT CGGGTTCGAT GTCGCCGCCG
  	TAAAGGTAGG
  51	TCGGCAGCGC GAACATCATC GTCTGCATCA TCCCCAGCCC
  	GGCAACGGCG
  101	AGGCGGACGA TGTATTGTTT GCGTTCTTTT TGGTTGGCGG
  	CTTCGATTTT
  151	TTGCGCGTCA TAGGGTGCGG CGGTGTAGCC TATCTGCCTG
  	ATTTTCAACA
  201	GAATGTCGGA AAGGCGGATT TTGCCGTCGT CCCAGACGAC
  	GCGGCAGCGG
  251	TGCGTGCTGT AATTGAGGTC GATGCGGACG ATGCCGTCTG
  	TACGCAAAAG
  301	CTGCTGTTCG ATCAGCCAGA CGCAGGCGGC GCAGGTGATG
  	CCGCCGAGCA
  351	TTAAAACCGC CTCGCGCGTG CCGCCGTGGG TTTCCACAAA
  	GTCGGACTGG
  401	ACTTCGGGCA GGTCGTACAG GCGGATTTGG TCGAGGATTT
  	CTTGGGGCGG
  451	CAGCTCGGTT TTTTGCGCGT CGGCGGTGCG TTGTTTGTAA
  	TAACTGCCCA
  501	AGCCCGCGTC AATAATGCTT TGTGCGACTG CCTGACAACC
  	GGCGCAGCAG
  551	GTTTCGCGGT CTTCGTTTTC GTAACGGACG GTCAGATGCA
  	GGTTTTCGGG
  601	AACGTCCAGC CCGCAGTGGA AACAGGTTTT TTTCATGGCA
  	TTTCGGTTTC
  651	GTCTGTGTTT GGTGCGGCGG CACAATACTC GGCAATGGCT
  	TCGCGCAGTG
  701	CGTCTATACC GGTATTTTCA GCAACGGAAA TGCGGACGGC
  	GGCAATTTTT
  751	CCCGCAGCGT CGCGCCATAT GCCCGTGTTT TGTTCTTCAG
  	ACGGCAGCAG
  801	GTCGGTTTTG TTGTACACCT TGATGCACGG AATATCGCCG
  	GCATGGATTT
  851	CTTGCAGTAC GTTTTCCACG TCTTCAATCT GCTGTCCGCT
  	GTTCGGAGCG
  901	GCGGCATCGA CGACGTGCAG CAGCACATCG GCTTGCGCGG
  	TTTCTTCCAG
  951	CGTGGCGGAA AAGGCGGAAA TCAGTTTGTG CGGCAGATCG
  	CTGACGAATC
  1001	CGACGGTATC GGTCAGGATA ATGCTGCATT CGGGACTGAT
  	GTACAGCCGC
  1051	CGCGCCGTCG TGTCGAGTGT GGCGAAAAGC TGGTCTTTCG
  	CATATATGCC
  1101	CGACTTGGTC AGCCGGTTGA ACAGACTGGA TTTGCCGACA
  	TTGGTATAG

• This encodes a protein having amino acid sequence <SEQ ID 144>:

• 1	MEDLQEIGFD VAAVKVGRQR EHHRLHHPQP GNGEADDVLF
  	AFFLVGGFDF
  51	LRVIGCGGVA YLPDFQQNVG KADFAVVPDD AAAVRAVIEV
  	DADDAVCTQK
  101	LLFDQPDAGG AGDAAEH*NR LARAAVGFHK VGLDFGQVVQ
  	ADLVEDFLGR
  151	QLGFLRVGGA LFVITAQARV NNALCDCLTT GAAGFAVFVF
  	VTDGQMQVFG
  201	NVQPAVETGF FHGISVSSVF GAAAQYSAMA SRSASIPVFS
  	ATEMRTAAIF
  251	PAASRHMPVF CSSDGSRSVL LYTLMHGISP AWISCSTFST
  	SSICCPLFGA
  301	AASTTCSSTS ACAVSSSVAE KAEISLCGRS LTNPTVSVRI
  	MLHSGLMYSR
  351	RAVVSSVAKS WSFAYMPDLV SRLNRLDLPT LV*

• It should be noted that this sequence includes a stop codon at position 118.
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF14 shows 89.8% identity over a 167aa overlap with a predicted ORF (ORF14.ng) from N. gonorrhoeae:
• 
• The complete length ORF14ng nucleotide sequence <SEQ ID 145> is predicted to encode a protein having amino acid sequence <SEQ ID 146>:

• 1	MEDLQEIGFD VAAVKVGRQR EHHRLHHTQS GNGKADDVLF
  	AFFLVGGFDF
  51	LRVIGCGGVA CLPDFQQNVG EADFAVVPDD AAAVRAVIEV
  	DADDAVCAQK
  101	LLFDQPDAGG AGNAAEHQHC FVRAIMGFHK VGLDFGQVVQ
  	ADLVEDFLGR
  151	QFGFFRVGGA SFVITAQAGI DDALCDCLTA DAAGFAVFAF
  	VADGQMQVFG
  201	NVQPAVETGF FHGISVSSVF GAAAQYSAMA SRSASIPVFS
  	ATEMRTAAIF
  251	PAASRHMPVF CSSDGSRSVL LYTLMHGISW AWISCSTFST
  	SSICCPLFRA
  301	AASTTCSSTS ACTVSSKVAE KAEISLCGRS LTNPTVSVRI
  	MLHAGLMYSR
  351	RAVVSRVAKS WSFAYMPDLV SRLNRLDLPT LV*

• Based on the putative transmembrane domain in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 18
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 147>:

• 1	..GGCCATTACT CCGACCGCAC TTGGAAGCCG CGTTTGGNCG
  	GCCGCCGTCT
  51	GCCGTATCTG CTTTATGGCA CGCTGATTGC GGTTATTGTG
  	ATGATTTTGA
  101	TGCCGAACTC GGGCAGCTTC GGTTTCGGCT ATGCGTCGCT
  	GGCGGCTTTG
  151	TCGTTCGGCG CGCTGATGAT TGCGCTGTTA GACGTGTCGT
  	CAAATATGGC
  201	GATGCAGCCG TTTAAGATGA TGGTCGGCGA CATGGTCAAC
  	GAGGAGCAGA
  251	AAA.NTACGC CTACGGGATT CAAAGTTTCT TAGCAAATAC
  	GGGCGCGGTC
  301	GTGGCGGCGA TTCTGCCGTT TGTGTTTGCG TATATCGGTT
  	TGGCGAACAC
  351	CGCCGANAAA GGCGTTGTGC CGCAGACCGT GGTCGTGGCG
  	TTTTATGTGG
  401	GTGCGGCGTT GCTGGTGATT ACCAGCGCGT TCACGATTTT
  	CAAAGTGAAG
  451	GAATACGANC CGGAAACCTA CGCCCGTTAC CACGGCATCG
  	ATGTCGCCGC
  501	GAATCAGGAA AAAGCCAACT GGATCGCACT CTTAAAA.CC
  	GCGC..

• This corresponds to the amino acid sequence <SEQ ID 148; ORF16>:

• 1	..GHYSDRTWKP RLXGRRLPYL LYGTLIAVIV MILMPNSGSF
  	GFGYASLAAL
  51	SFGALMIALL DVSSNMAMQP FKMMVGDMVN EEQKXYAYGI
  	QSFLANTGAV
  101	VAAILPFVFA YIGLANTAXK GVVPQTVVVA FYVGAALLVI
  	TSAFTIFKVK
  151	EYXPETYARY HGIDVAANQE KANWIALLKX A..

• Further work revealed the complete nucleotide sequence <SEQ ID 149>:

• 1	ATGTCGGAAT ATACGCCTCA AACAGCAAAA CAAGGTTTGC
  	CCGCGCTGGC
  51	AAAAAGCACG ATTTGGATGC TCAGTTTCGG CTTTCTCGGC
  	GTTCAGACGG
  101	CCTTTACCCT GCAAAGCTCG CAAATGAGCC GCATTTTTCA
  	AACGCTAGGC
  151	GCAGACCCGC ACAATTTGGG CTGGTTTTTC ATCCTGCCGC
  	CGCTGGCGGG
  201	GATGCTGGTG CAGCCGATTG TCGGCCATTA CTCCGACCGC
  	ACTTGGAAGC
  251	CGCGTTTGGG CGGCCGCCGT CTGCCGTATC TGCTTTATGG
  	CACGCTGATT
  301	GCGGTTATTG TGATGATTTT GATGCCGAAC TCGGGCAGCT
  	TCGGTTTCGG
  351	CTATGCGTCG CTGGCGGCTT TGTCGTTCGG CGCGCTGATG
  	ATTGCGCTGT
  401	TAGACGTGTC GTCAAATATG GCGATGCAGC CGTTTAAGAT
  	GATGGTCGGC
  451	GACATGGTCA ACGAGGAGCA GAAAGGCTAC GCCTACGGGA
  	TTCAAAGTTT
  501	CTTAGCAAAT ACGGGCGCGG TCGTGGCGGC GATTCTGCCG
  	TTTGTGTTTG
  551	CGTATATCGG TTTGGCGAAC ACCGCCGAGA AAGGCGTTGT
  	GCCGCAGACC
  601	GTGGTCGTGG CGTTTTATGT GGGTGCGGCG TTGCTGGTGA
  	TTACCAGCGC
  651	GTTCACGATT TTCAAAGTGA AGGAATACGA TCCGGAAACC
  	TACGCCCGTT
  701	ACCACGGCAT CGATGTCGCC GCGAATCAGG AAAAAGCCAA
  	CTGGATCGAA
  751	CTCTTGAAAA CCGCGCCTAA GGCGTTTTGG ACGGTTACTT
  	TGGTGCAATT
  801	CTTCTGCTGG TTCGCCTTCC AATATATGTG GACTTACTCG
  	GCAGGCGCGA
  851	TTGCGGAAAA CGTCTGGCAC ACCACCGATG CGTCTTCCGT
  	AGGTTATCAG
  901	GAGGCGGGTA ACTGGTACGG CGTTTTGGCG GCGGTGCAGT
  	CGGTTGCGGC
  951	GGTGATTTGT TCGTTTGTAT TGGCGAAAGT GCCGAATAAA
  	TACCATAAGG
  1001	CGGGTTATTT CGGCTGTTTG GCTTTGGGCG CGCTCGGCTT
  	TTTCTCCGTT
  1051	TTCTTCATCG GCAACCAATA CGCGCTGGTG TTGTCTTATA
  	CCTTAATCGG
  1101	CATCGCTTGG GCGGGCATTA TCACTTATCC GCTGACGATT
  	GTGACCAACG
  1151	CCTTGTCGGG CAAGCATATG GGCACTTACT TGGGCTTGTT
  	TAACGGCTCT
  1201	ATCTGTATGC CTCAAATCGT CGCTTCGCTG TTGAGTTTCG
  	TGCTTTTCCC
  1251	TATGCTGGGC GGCTTGCAGG CCACTATGTT CTTGGTAGGG
  	GGCGTCGTCC
  1301	TGCTGCTGGG CGCGTTTTCC GTGTTCCTGA TTAAAGAAAC
  	ACACGGCGGG
  1351	GTTTGA

• This corresponds to the amino acid sequence <SEQ ID 150; ORF16-1>:

• 1	MSEYTPQTAK QGLPALAKST IWMLSFGFLG VQTAFTLQSS
  	QMSRIFQTLG
  51	ADPHNLGWFF ILPPLAGMLV QPIVGHYSDR TWKPRLGGRR
  	LPYLLYGTLI
  101	AVIVMILMPN SGSFGFGYAS LAALSFGALM IALLDVSSNM
  	AMQPFKMMVG
  151	DMVNEEQKGY AYGIQSFLAN TGAVVAAILP FVFAYIGLAN
  	TAEKGVVPQT
  201	VVVAFYVGAA LLVITSAFTI FKVKEYDPET YARYHGIDVA
  	ANQEKANWIE
  251	LLKTAPKAFW TVTLVQFFCW FAFQYMWTYS AGAIAENVWH
  	TTDASSVGYQ
  301	EAGNWYGVLA AVQSVAAVIC SFVLAKVPNK YHKAGYFGCL
  	ALGALGFFSV
  351	FFIGNQYALV LSYTLIGIAW AGIITYPLTI VTNALSGKHM
  	GTYLGLFNGS
  401	ICMPQIVASL LSFVLFPMLG GLQATMFLVG GVVLLLGAFS
  	VFLIKETHGG
  451	V*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. Meningitidis (Strain A)
• ORF16 shows 96.7% identity over a 181 as overlap with an ORF (ORF16a) from strain A of N. meningitidis:
• 
• The complete length ORF16a nucleotide sequence <SEQ ID 151> is:

• 1	ATGTCGGAAT ATACGCCTCA AACAGCAAAA CAAGGTTTGC
  	CCGCGCTGGC
  51	AAAAAGCACG ATTTGGATGC TCAGTTTCGG CTTTCTCGGC
  	GTTCAGACGG
  101	CCTTTACCCT GCAAAGCTCG CAGATGAGCC GCATCTTCCA
  	GACGCTCGGT
  151	GCCGATCCGC ACAGCCTCGG CTGGTTCTTT ATCCTGCCGC
  	CGCTGGCGGG
  201	GATGCTGGTG CAGCCGATTG TCGGCCATTA CTCCGACCGC
  	ACTTGGAAGC
  251	CGCGTTTGGG CGGCCGCCGT CTGCCGTATC TGCTTTATGG
  	CACGCTGATT
  301	GCGGTTATTG TGATGATTTT GATGCCGAAC TCGGGCAGCT
  	TCGGTTTCGG
  351	CTATGCGTCG CTGGCGGCTT TGTCGTTCGG CGCGCTGATG
  	ATTGCGCTGT
  401	TAGACGTGTC GTCAAATATG GCGATGCAGC CGTTTAAGAT
  	GATGGTCGGC
  451	GACATGGTCA ACGAGGAGCA GAAAGGCTAC GCCTACGGGA
  	TTCAAAGTTT
  501	CTTAGCGAAT ACGGGCGCGG TCGTGGCGGC GATTCTGCCG
  	TTTGTGTTTG
  551	CGTATATCGG TTTGGCGAAC ACCGCCGAGA AAGGCGTTGT
  	GCCGCAGACC
  601	GTGGTCGTGG CGTTTTATGT GGGTGCGGCG TTGCTGGTGA
  	TTACCAGCGC
  651	GTTCACGATT TTCAAAGTGA AGGAATACAA TCCGGAAACC
  	TACGCCCGTT
  701	ACCACGGCAT CGATGTCGCC GCGAATCAGG AAAAAGCCAA
  	CTGGATCGAA
  751	CTCTTGAAAA CCGCGCCTAA GGCGTTTTGG ACGGTTACTT
  	TGGTGCAATT
  801	CTTCTGCTGG TTCGCCTTCC AATATATGTG GACTTACTCG
  	GCAGGCGCGA
  851	TTGCGGAAAA CGTCTGGCAC ACCACCGATG CGTCTTCCGT
  	AGGTTATCAG
  901	GAGGCGGGTA ACTGGTACGG CGTTTTGGCG GCGGTGCAGT
  	CGGTTGCGGC
  951	GGTGATTTGT TCGTTTGTAT TGGCGAAAGT GCCGAATAAA
  	TACCATAAGG
  1001	CGGGTTATTT CGGCTGTTTG GCTTTGGGCG CGCTCGGCTT
  	TTTCTCCGTT
  1051	TTCTTCATCG GCAACCAATA CGCGCTGGTG TTGTCTTATA
  	CCTTAATCGG
  1101	CATCGCTTGG GCGGGCATTA TCACTTATCC GCTGACGATT
  	GTGACCAACG
  1151	CCTTGTCGGG CAAGCATATG GGCACTTACT TGGGCCTGTT
  	TAACGGCTCT
  1201	ATCTGTATGC CGCAAATCGT CGCTTCGCTG TTGAGTTTCG
  	TGCTTTTCCC
  1251	TATGCTGGGC GGCTTGCAGG CCACTATGTT CTTGGTAGGG
  	GGCGTCGTCC
  1301	TGCTGCTGGG CGCGTTTTCC GTGTTCCTGA TTAAAGAAAC
  	ACACGGCGGG
  1351	GTTTGA

• This encodes a protein having amino acid sequence <SEQ ID 152>:

• 1	MSEYTPQTAK QGLPALAKST IWMLSFGFLG VQTAFTLQSS
  	QMSRIFQTLG
  51	ADPHSLGWFF ILPPLAGMLV QPIVGHYSDR TWKPRLGGRR
  	LPYLLYGTLI
  101	AVIVMILMPN SGSFGFGYAS LAALSFGALM IALLDVSSNM
  	AMQPFKMMVG
  151	DMVNEEQKGY AYGIQSFLAN TGAVVAAILP FVFAYIGLAN
  	TAEKGVVPQT
  201	VVVAFYVGAA LLVITSAFTI FKVKEYNPET YARYHGIDVA
  	ANQEKANWIE
  251	LLKTAPKAFW TVTLVQFFCW FAFQYMWTYS AGAIAENVWH
  	TTDASSVGYQ
  301	EAGNWYGVLA AVQSVAAVIC SFVLAKVPNK YHKAGYFGCL
  	ALGALGFFSV
  351	FFIGNQYALV LSYTLIGIAW AGIITYPLTI VTNALSGKHM
  	GTYLGLFNGS
  401	ICMPQIVASL LSFVLFPMLG GLQATMFLVG GVVLLLGAFS
  	VFLIKETHGG
  451	V*

• ORF16a and ORF16-1 show 99.6% identity in 451 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF16 shows 93.9% identity over a 181aa overlap with a predicted ORF (ORF16.ng) from N. gonorrhoeae:
• 
• The complete length ORF16ng nucleotide sequence <SEQ ID 153> is:

• 1	ATGATAGGGG ATCGCCGCGC CGGCAACCAT TTCGGATTTT
  	CCAAAGCAAA
  51	TACTTTTCAA ATCAAAAAAA AGGATTTACT TTATGTCGGA
  	ATATACGCCT
  101	CAAACAGCAA AACAAGGTTT GCCCGCGCCG GCAAAAAGCA
  	CGATTTGGAT
  151	GTTGAGCTTC GGCTATCTCG GCGTTCAGAC GGCCTTTACC
  	CTGCAAAGCT
  201	CGCAGATGAG CCGCATTTTT CAAACGCTAG GCGCAGACCC
  	GCACAATTTG
  251	GGCTGGTTTT TCATCCTGCC GCCGCTGGCG GGGATGCTGG
  	TTCAGCCGAT
  301	AGTGGCTACT ACTCAGACCG CACTTGGAAG CCGCGCTTGG
  	GCGGCCGCCG
  351	CCTGCCGTAT CTGCTTTACG GCACGCTGAT TGCGGTCATC
  	GTGATGATTT
  401	TGATGCCGAA CTCGGGCAGC TTCGGTTTCG GCTATGCGTC
  	GCTGGCGGCC
  451	TTGTCGTTCG GCGCGCTGAT GATTGCGCTG TTGGACGTGT
  	CGTCGAATAT
  501	GGCGATGCAG CCGTTTAAGA TGATGGTCGG CGATATGGTC
  	AACGAGGAGC
  551	AGAAAAGCTA CGCCTACGGG ATTCAAAGTT TCTTAGCGAA
  	TACGGACGCG
  601	GTTGTGGCAG CGATTCTGCC GTTTGTGTTC GCGTATATCG
  	GTTTGGCGAA
  651	CACTGCCGAG AAAGGCGTTG TGCCACAAAC CGTGGTCGTA
  	GCATTCTATG
  701	TGGGTGCGGC GTTACTGATT ATTACCAGTG CGTTCACAAT
  	CTCCAAAGTC
  751	AAAGAATACG ACCCGGAAAC CTACGCCCGT TACCACGGCA
  	TCGATGTCGC
  801	CGCGAATCAG GAAAAAGCCA ACTGGTTCGA ACTCTTAAAA
  	ACCGCGCCTA
  851	AAGTGTTTTG GACGGTTACT CCGGTACAGT TTTTCTGCTG
  	GTTCGCCTTC
  901	CGGTATATGT GGACTTACTC GGCAGGCGCG ATTGCAGAAA
  	ACGTCTGGCA
  951	CACTACCGAT GCGTCTTCCG TAGGCCATCA GGAGGCGGGC
  	AACCGGTACG
  1001	GCGTTTTGGC GGCGGTGTAG

• This encodes a protein having amino acid sequence <SEQ ID 154>:

• 1	MIGDRRAGNH FGFSKANTFQ IKKKDLLYVG IYASNSKTRF
  	ARAGKKHDLD
  51	VELRLSRRSD GLYPAKLADE PHFSNARRRP AQFGLVFHPA
  	AAGGDAGSAD
  101	SGYYSDRTWK PRLGGRRLPY LLYGTLIAVI VMILMPNSGS
  	FGFGYASLAA
  151	LSFGALMIAL LDVSSNMAMQ PFKMMVGDMV NEEQKSYAYG
  	IQSFLANTDA
  201	VVAAILPFVF AYIGLANTAE KGVVPQTVVV AFYVGAALLI
  	ITSAFTISKV
  251	KEYDPETYAR YHGIDVAANQ EKANWFELLK TAPKVFWTVT
  	PVQFFCWFAF
  301	RYMWTYSAGA IAENVWHTTD ASSVGHQEAG NRYGVLAAV*

• ORF16ng and ORF16-1 show 89.3% identity in 261 aa overlap:
• 
• Based on this analysis, including the presence of several putative transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 19
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 155>:

• 1	ATGTTGTTCC GTAAAACGAC CGCCGCCGTT TTGGCGCATA
  	CCTTGATGCT
  51	GAACGGCTGT ACGTTGATGT TGTGGGGAAT GAACAACCCG
  	GTCAGCGAAA
  101	CAATCACCCG NAAACACGTT GNCAAAGACC AAATCCGNGN
  	CTTCGGTGTG
  151	GTTGCCGAAG ACAATGCCCA ATTGGAAAAG GGCAGCCTGG
  	TGATGATGGG
  201	CGGAAAATAC TGGTTCGTCG TCAATCCCGA AGATTCGGCG
  	AA.NTGACGG
  251	GNATTTTGAN GGCAGGGCTG GACAAACCCT TCCAAATAGT
  	TNAGGATACC
  301	CCGAGCTATG C.TGCCACCA AGCCCTGCCG GTCAAACTCG
  	GATCGNCTGG
  351	CAGCCAGAAT...

• This corresponds to the amino acid sequence <SEQ ID 156; ORF28>:

• 1	MLFRKTTAAV LAHTLMLNGC TLMLWGMNNP VSETITRKHV
  	XKDQIRXFGV
  51	VAEDNAQLEK GSLVMMGGKY WFVVNPEDSA XXTGILXAGL
  	DKPFQIVXDT
  101	PSYXCHQALP VKLGSXGSQN...

• Further work revealed the complete nucleotide sequence <SEQ ID 157>:

• 1	ATGTTGTTCC GTAAAACGAC CGCCGCCGTT TTGGCGGCAA
  	CCTTGATGCT
  51	GAACGGCTGT ACGTTGATGT TGTGGGGAAT GAACAACCCG
  	GTCAGCGAAA
  101	CAATCACCCG CAAACACGTT GACAAAGACC AAATCCGCGC
  	CTTCGGTGTG
  151	GTTGCCGAAG ACAATGCCCA ATTGGAAAAG GGCAGCCTGG
  	TGATGATGGG
  201	CGGAAAATAC TGGTTCGTCG TCAATCCCGA AGATTCGGCG
  	AAGCTGACGG
  251	GCATTTTGAA GGCAGGGCTG GACAAACCCT TCCAAATAGT
  	TGAGGATACC
  301	CCGAGCTATG CTCGCCACCA AGCCCTGCCG GTCAAACTCG
  	AATCGCCTGG
  351	CAGCCAGAAT TTCAGTACCG AAGGCCTTTG CCTGCGCTAC
  	GATACCGACA
  401	AGCCTGCCGA CATCGCCAAG CTGAAACAGC TCGGGTTTGA
  	AGCGGTCAAA
  451	CTCGACAATC GGACCATTTA CACGCGCTGC GTATCCGCCA
  	AAGGCAAATA
  501	CTACGCCACA CCGCAAAAAC TGAACGCCGA TTACCATTTT
  	GAGCAAAGTG
  551	TGCCTGCCGA TATTTATTAC ACGGTTACTG AAGAACATAC
  	CGACAAATCC
  601	AAGCTGTTTG CAAATATCTT ATATACGCCC CCCTTTTTGA
  	TACTGGATGC
  651	GGCGGGCGCG GTACTGGCCT TGCCTGCGGC GGCTCTGGGT
  	GCGGTCGTGG
  701	ATGCCGCCCG CAAATGA

• This corresponds to the amino acid sequence <SEQ ID 158; ORF28-1>:

• 1	MLFRKTTAAV LAATLMLNGC TLMLWGMNNP VSETITRKHV
  	DKDQIRAFGV
  51	VAEDNAQLEK GSLVMMGGKY WFVVNPEDSA KLTGILKAGL
  	DKPFQIVEDT
  101	PSYARHQALP VKLESPGSQN FSTEGLCLRY DTDKPADIAK
  	LKQLGFEAVK
  151	LDNRTIYTRC VSAKGKYYAT PQKLNADYHF EQSVPADIYY
  	TVTEEHTDKS
  201	KLFANILYTP PFLILDAAGA VLALPAAALG AVVDAARK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF28 shows 79.2% identity over a 120aa overlap with an ORF (ORF28a) from strain A of N. meningitidis:
• 
• The complete length ORF28a nucleotide sequence <SEQ ID 159> is:

• 1	ATGTTGTTCC GTAAAACGAC CGCCGCCGTT TTGGCGGCAA
  	CCTTGATGTT
  51	GAACGGCTGT ACGGTAATGA TGTGGGGTAT GAACAGCCCG
  	TTCAGCGAAA
  101	CGACCGCCCG CAAACACGTT GACAAGGACC AAATCCGCGC
  	CTTCGGTGTG
  151	GTTGCCGAAG ACAATGCCCA ATTGGAAAAG GGCAGCCTGG
  	TGATGATGGG
  201	CGGGAAATAC TGGTTCGTCG TCAATCCTGA AGATTCGGCG
  	AAGCTGACGG
  251	GCATTTTGAA GGCCGGGTTG GACAAGCAGT TTCAAATGGT
  	TGAGCCCAAC
  301	CCGCGCTTTG CCTACCAAGC CCTGCCGGTC AAACTCGAAT
  	CGCCCGCCAG
  351	CCAGAATTTC AGTACCGAAG GCCTTTGCCT GCGCTACGAT
  	ACCGACAGAC
  401	CTGCCGACAT CGCCAAGCTG AAACAGCTTG AGTTTGAAGC
  	GGTCGAACTC
  451	GACAATCGGA CCATTTACAC GCGCTGCGTC TCCGCCAAAG
  	GCAAATACTA
  501	CGCCACACCG CAAAAACTGA ACGCCGATTA TCATTTTGAG
  	CAAAGTGTGC
  551	CTGCCGATAT TTATTACACG GTTACGAAAA AACATACCGA
  	CAAATCCAAG
  601	TTGTTTGAAA ATATTGCATA TACGCCCACC ACGTTGATAC
  	TGGATGCGGT
  651	GGGCGCGGTG CTGGCCTTGC CTGTCGCGGC GTTGATTGCA
  	GCCACGAATT
  701	CCTCAGACAA ATGA

• This encodes a protein having amino acid sequence <SEQ ID 160>:

• 1	MLFRKTTAAV LAATLMLNGC TVMMWGMNSP FSETTARKHV
  	DKDQIRAFGV
  51	VAEDNAQLEK GSLVMMGGKY WFVVNPEDSA KLTGILKAGL
  	DKQFQMVEPN
  101	PRFAYQALPV KLESPASQNF STEGLCLRYD TDRPADIAKL
  	KQLEFEAVEL
  151	DNRTIYTRCV SAKGKYYATP QKLNADYHFE QSVPADIYYT
  	VTKKHTDKSK
  201	LFENIAYTPT TLILDAVGAV LALPVAALIA ATNSSDK*

• ORF28a and ORF28-1 show 86.1% identity in 238 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF28 shows 84.2% identity over a 120aa overlap with a predicted ORF (ORF28.ng) from N. gonorrhoeae:
• 
• The complete length ORF28ng nucleotide sequence <SEQ ID 161> is

• 1	ATGTTGTTCC GTAAAACGAC CGCCGCCGTT TTGGCGGCAA
  	CCTTGATACT
  51	GAACGGCTGT ACGATGATGT TGCGGGGGAT GAACAACCCG
  	GTCAGCCAAA
  101	CAATCACCCG CAAACACGTT GACAAAGACC AAATCCGCGC
  	CTTCGGTGTG
  151	GTTGCCGAAG ACAATGCCCA ATTGGAAAAG GGCAGCCTGG
  	TGATGATGGG
  201	CGGGAAATAC TGGTTCGCCG TCAATCCCGA AGATTCGGCG
  	AAGCTGACGG
  251	GCCTTTTGAA GGCCGGGTTG GACAAGCCCT TCCAAATAGT
  	TGAGGATACC
  301	CCGAGCTATG CCCGCCACCA AGCCCTGCCG GTCAAATTCG
  	AAGCGCCCGG
  351	CAGCCAGAAT TTCAGTACCG GAGGTCTTTG CCTGCGCTAT
  	GATACCGGCA
  401	GACCTGACGA CATCGCCAAG CTGAAACAGC TTGAGTTTAA
  	AGCGGTCAAA
  451	CTCGACAATC GGACCATTTA CACGCGCTGC GTATCCGCCA
  	AAGGCAAATA
  501	CTACGCCACG CCGCAAAAAC TGAACGCCGA TTATCATTTT
  	GAGCAAAGTG
  551	TGCCCGCCGA TATTTATTAT ACGGTTACTG AAAAACATAC
  	CGACAAATCC
  601	AAGCTGTTTG GAAATATCTT ATATACGCCC CCCTTGTTGA
  	TATTGGATGC
  651	GGCGGCCGCG GTGCTGGTCT TGCCTATGGC TCTGATTGCA
  	GCCGCGAATT
  701	CCTCAGACAA ATGA

• This encodes a protein having amino acid sequence <SEQ ID 162>:

• 1	MLFRKTTAAV LAATLILNGC TMMLRGMNNP VSQTITRKHV
  	DKDQIRAFGV
  51	VAEDNAQLEK GSLVMMGGKY WFAVNPEDSA KLTGLLKAGL
  	DKPFQIVEDT
  101	PSYARHQALP VKFEAPGSQN FSTGGLCLRY DTGRPDDIAK
  	LKQLEFKAVK
  151	LDNRTIYTRC VSAKGKYYAT PQKLNADYHF EQSVPADIYY
  	TVTEKHTDKS
  201	KLFGNILYTP PLLILDAAAA VLVLPMALIA AANSSDK*

• ORF28ng and ORF28-1 share 90.0% identity in 231 aa overlap:
• 
• Based on this analysis, including the presence of a putative transmembrane domain in the gonococcal protein, it was predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF28-1 (24 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 6A shows the results of affinity purification of the GST-fusion protein, and FIG. 6B shows the results of expression of the His-fusion in E. coli. Purified GST-fusion protein was used to immunise mice, whose sera were used for ELISA, which gave a positive result. These experiments confirm that ORF28-1 is a surface-exposed protein, and that it may be a useful immunogen.
Example 20
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 163>:

• 1	..GTCAGTCCTG TACTGCCTAT TACACACGAA CGGACAGGGT
  	TTGAAGGTGT
  51	TATCGGTTAT GAAACCCATT TTTCAGGGCA CGGACATGAA
  	GTACACAGTC
  101	CGTTCGATCA TCATGATTCA AAAAGCACTT CTGATTTCAG
  	CGGCGGTGTA
  151	GACGGCGGTT TTACTGTTTA CCAACTTCAT CGAACATGGT
  	CGGAAATCCA
  201	TCCGGAGGAT GAATATGACG GGCCGCAAGC AGCG.ATTAT
  	CCGCCCCCCG
  251	GAGGAGCAAG GGATATATAC AGCTATTATG TCAAAGGAAC
  	TTCAACAAAA
  301	ACAAAGACTA GTATTGTCCC TCAAGCCCCA TTTTCAGACC
  	GTTGGCTAGA
  351	AGAAAATGCC GGTGCCGCCT CTGGT..

• This corresponds to the amino acid sequence <SEQ ID 164; ORF29>:

• 1	..VSPVLPITHE RTGFEGVIGY ETHFSGHGHE VHSPFDHHDS
  	KSTSDFSGGV
  51	DGGFTVYQLH RTWSEIHPED EYDGPQAAXY PPPGGARDIY
  	SYYVKGTSTK
  101	TKTSIVPQAP FSDRWLEENA GAASG..

• Further work revealed the complete nucleotide sequence <SEQ ID 165>:

• 1	ATGAATTTGC CTATTCAAAA ATTCATGATG CTGTTTGCAG
  	CAGCAATATC
  51	GTTGCTGCAA ATCCCCATTA GTCATGCGAA CGGTTTGGAT
  	GCCCGTTTGC
  101	GCGATGATAT GCAGGCAAAA CACTACGAAC CGGGTGGTAA
  	ATACCATCTG
  151	TTTGGTAATG CTCGCGGCAG TGTTAAAAAG CGGGTTTACG
  	CCGTCCAGAC
  201	ATTTGATGCA ACTGCGGTCA GTCCTGTACT GCCTATTACA
  	CACGAACGGA
  251	CAGGGTTTGA AGGTGTTATC GGTTATGAAA CCCATTTTTC
  	AGGGCACGGA
  301	CATGAAGTAC ACAGTCCGTT CGATCATCAT GATTCAAAAA
  	GCACTTCTGA
  351	TTTCAGCGGC GGTGTAGACG GCGGTTTTAC TGTTTACCAA
  	CTTCATCGAA
  401	CAGGGTCGGA AATCCATCCG GAGGATGGAT ATGACGGGCC
  	GCAAGGCAGC
  451	GATTATCCGC CCCCCGGAGG AGCAAGGGAT ATATACAGCT
  	ATTATGTCAA
  501	AGGAACTTCA ACAAAAACAA AGACTAATAT TGTCCCTCAA
  	GCCCCATTTT
  551	CAGACCGTTG GCTAAAAGAA AATGCCGGTG CCGCCTCTGG
  	TTTTTTCAGC
  601	CGTGCGGATG AAGCAGGAAA ACTGATATGG GAAAGCGACC
  	CCAATAAAAA
  651	TTGGTGGGCT AACCGTATGG ATGATGTTCG CGGCATCGTC
  	CAAGGTGCGG
  701	TTAATCCTTT TTTAATGGGT TTTCAAGGAG TAGGGATTGG
  	GGCAATTACA
  751	GACAGTGCAG TAAGCCCGGT CACAGATACA GCCGCGCAGC
  	AGACTCTACA
  801	AGGTATTAAT GATTTAGGAA AATTAAGTCC GGAAGCACAA
  	CTTGCTGCCG
  851	CGAGCCTATT ACAGGACAGT GCTTTTGCGG TAAAAGACGG
  	TATCAACTCT
  901	GCCAAACAAT GGGCTGATGC CCATCCAAAT ATAACAGCTA
  	CTGCCCAAAC
  951	TGCCCTTTCC GCAGCAGAGG CCGCAGGTAC GGTTTGGAGA
  	GGTAAAAAAG
  1001	TAGAACTTAA CCCGACTAAA TGGGATTGGG TTAAAAATAC
  	CGGTTATAAA
  1051	AAACCTGCTG CCCGCCATAT GCAGACTTTA GATGGGGAGA
  	TGGCAGGTGG
  1101	GAATAAACCT ATTAAATCTT TACCAAACAG TGCCGCTGAA
  	AAAAGAAAAC
  1151	AAAATTTTGA GAAGTTTAAT AGTAACTGGA GTTCAGCAAG
  	TTTTGATTCA
  1201	GTGCACAAAA CACTAACTCC CAATGCACCT GGTATTTTAA
  	GTCCTGATAA
  1251	AGTTAAAACT CGATACACTA GTTTAGATGG AAAAATTACA
  	ATTATAAAAG
  1301	ATAACGAAAA CAACTATTTT AGAATCCATG ATAATTCACG
  	AAAACAGTAT
  1351	CTTGATTCAA ATGGTAATGC TGTGAAAACC GGTAATTTAC
  	AAGGTAAGCA
  1401	AGCAAAAGAT TATTTACAAC AACAAACTCA TATCAGGAAC
  	TTAGACAAAT
  1451	GA

• This corresponds to the amino acid sequence <SEQ ID 166; ORF29-1>:

• 1	MNLPIQKFMM LFAAAISLLQ IPISHANGLD ARLRDDMQAK
  	HYEPGGKYHL
  51	FGNARGSVKK RVYAVQTFDA TAVSPVLPIT HERTGFEGVI
  	GYETHFSGHG
  101	HEVHSPFDHH DSKSTSDFSG GVDGGFTVYQ LHRTGSEIHP
  	EDGYDGPQGS
  151	DYPPPGGARD IYSYYVKGTS TKTKTNIVPQ APFSDRWLKE
  	NAGAASGFFS
  201	RADEAGKLIW ESDPNKNWWA NRMDDVRGIV QGAVNPFLMG
  	FQGVGIGAIT
  251	DSAVSPVTDT AAQQTLQGIN DLGKLSPEAQ LAAASLLQDS
  	AFAVKDGINS
  301	AKQWADAHPN ITATAQTALS AAEAAGTVWR GKKVELNPTK
  	WDWVKNTGYK
  351	KPAARHMQTL DGEMAGGNKP IKSLPNSAAE KRKQNFEKFN
  	SNWSSASFDS
  401	VHKTLTPNAP GILSPDKVKT RYTSLDGKIT IIKDNENNYF
  	RIHDNSRKQY
  451	LDSNGNAVKT GNLQGKQAKD YLQQQTHIRN LDK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF29 shows 88.0% identity over a 125aa overlap with an ORF (ORF29a) from strain A of N. meningitidis:
• 
• The complete length ORF29a nucleotide sequence <SEQ ID 167> is:

• 1	ATGAATTNGC CTATTCAAAA ATTCATGATG CTGTTTGCAG
  	CAGCAATATC
  51	GTNGCTGCAA ATCCCNATTA GTCATGCGAA CGGTTTGGAT
  	GCCCGTTTGC
  101	GCGATGATAT GCAGGCAAAA CACTACGAAC CGGGTGGTAA
  	ATACCATCTG
  151	TTTGGTAATG CTCGCGGCAG TGTTAAAAAT CGGGTTTACG
  	CCGTCCAAAC
  201	ATTTGATGCA ACTGCGGTCG GCCCCATACT GCCTATTACA
  	CACGAACGGA
  251	CAGGATTTGA AGGCATTATC GGTTATGAAA CCCATTTTTC
  	AGGACATGGA
  301	CATGAAGTAC ACAGTCCGTT CGATAATCAT GATTCAAAAA
  	GCACTTCTGA
  351	TTTCAGCGGC GGCGTAGACG GTGGTTTTAC CGTTTACCAA
  	CTTCATCGGA
  401	CAGGGTCGGA AATCCATCCG GAGGATGGAT ATGACGGGCC
  	GCAAGGCAGC
  451	GATTATCCGC CCCCCGGAGG AGCAAGGGAT ATATACANNT
  	ANTATGTCAA
  501	AGGAACTTCA ACAAAAACAA AGAGTAATAT TGTTCCCCGA
  	GCCCCATTTT
  551	CAGACCGCTG GCTAAAAGAA AATGCCGGTG CCGCCTCTGG
  	TTTTTTCAGC
  601	CGTGCTGATG AAGCAGGAAA ACTGATATGG GAAAGCGACC
  	CCAATAAAAA
  651	TTGGTGGGCT AACCGTATGG ATGATATTCG CGGCATCGTC
  	CAAGGTGCGG
  701	TTAATCCTTT TTTAATGGGT TTTCAAGGAG TAGGGATTGG
  	GGCAATTACA
  751	GACAGTGCAG TAAGCCCGGT CACAGATACA GCCGCGCAGC
  	AGACTCTACA
  801	AGGTATNAAT CATTTAGGAA ANTTAAGTCC CGAAGCACAA
  	CTTGCGGCTG
  851	CAACCGCATT ACAAGACAGT GCTTTTGCGG TAAAAGACGG
  	TATCAATTCC
  901	GCCAGACAAT GGGCTGATGC CCATCCGAAT ATAACTGCAA
  	CAGCCCAAAC
  951	TGCCCTTGCC GTAGCAGANG CCGCAACTAC GGTTTGGGGC
  	GGTAAAAAAG
  1001	TAGAACTTAA CCCGACCAAA TGGGATTGGG TTAAAAATAC
  	NGGCTATAAN
  1051	ACACCTGCTG TTCGCACCAT GCATACTTTG GATGGGGAAA
  	TGGCCGGTGG
  1101	GAATAGACCG CCTAAATCTA TAACGTCCAA CAGCAAAGCA
  	GATGCTTCCA
  1151	CACAACCGTC TTTACAAGCG CAACTAATTG GAGAACAAAT
  	TANNNNNGGG
  1201	CATGCTTATA ACAAGCATGT CATAAGACAA CAAGAATTTA
  	CGGATTTAAA
  1251	TATCAATTCA CCAGCAGATT TTGCTCGGCA TATTGAAAAT
  	ATTGTTAGCC
  1301	ATCCANCAAA TATGAAAGAG TTACCTCGCG GTAGAACTGC
  	GTATTGGGAT
  1351	NATAAAACAG GGACNATAGT TATCCGAGAT AAAAATTCTG
  	ACGATGGAGG
  1401	TACAGCATTT AGACCAACAT CAGGTAAAAA ATATTATGAT
  	GATTTATAG

• This encodes a protein having amino acid sequence <SEQ ID 168>:

• 1	MNXPIQKFMM LFAAAISXLQ IPISHANGLD ARLRDDMQAK
  	HYEPGGKYHL
  51	FGNARGSVKN RVYAVQTFDA TAVGPILPIT HERTGFEGII
  	GYETHFSGHG
  101	HEVHSPFDNH DSKSTSDFSG GVDGGFTVYQ LHRTGSEIHP
  	EDGYDGPQGS
  151	DYPPPGGARD IYXXYVKGTS TKTKSNIVPR APFSDRWLKE
  	NAGAASGFFS
  201	RADEAGKLIW ESDPNKNWWA NRMDDIRGIV QGAVNPFLMG
  	FQGVGIGAIT
  251	DSAVSPVTDT AAQQTLQGXN HLGXLSPEAQ LAAATALQDS
  	AFAVKDGINS
  301	ARQWADAHPN ITATAQTALA VAXAATTVWG GKKVELNPTK
  	WDWVKNTGYX
  351	TPAVRTMHTL DGEMAGGNRP PKSITSNSKA DASTQPSLQA
  	QLIGEQIXXG
  401	HAYNKHVIRQ QEFTDLNINS PADFARHIEN IVSHPXNMKE
  	LPRGRTAYWD
  451	XKTGTIVIRD KNSDDGGTAF RPTSGKKYYD DL*

• ORF29a and ORF29-1 show 90.1% identity in 385 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF29 shows 88.8% identity over a 125aa overlap with a predicted ORF (ORF29.ng) from N. gonorrhoeae:
• 
• The complete length ORF29ng nucleotide sequence <SEQ ID 169> is predicted to encode a protein having amino acid sequence <SEQ ID 170>:

• 1 MNLPIQKFMM LFAAAISLLQ IPISHANGLD ARLRDDMQAK HYEPGGKYHL
  51 FGNARGSVKN RVCAVQTFDA TAVGPILPIT HERTGFEGVI GYETHFSGHG
  101 HEVHSPFDNH DSKSTSDFSG GVDGGFTVYQ LHRTGSEIHP EDGYDGPQGG
  151 GYPPPGGARD IYSYHIKGTS TKTKINTVPQ APFSDRWLKE NAGAASGFLS
  201 RADEAGKLIW ENDPDKNWRA NRMDDIRGIV QGAVNPFLTG FQGLGVGAIT
  251 DSAVSPVTYA AARKTLQGIH NLGNLSPEAQ LAAATALQDS AFAVKDSINS
  301 ARQWADAHPN ITATAQTALA VTEAATTVWG GKKVELNPAK WDWVKNTGYK
  351 KPAARHMQTV DGEMAGGNKP LESKNTVTTN NFFENTGYTE KVLRQASNGD
  401 YHGFPQSVDA FSENGTVIQI VGGDNIVRHK LYIPGSYKGK DGNFEYIREA
  451 DGKINHRLFV PNQQLPEK*

• In a second experiment, the following DNA sequence <SEQ ID 171> was identified:

• 1 atgAATTTGC CTATTCAAAA ATTCATGATG ctgttggcAg cggcaatatc
  51 gatgctGCat ATCCCCATTA GTCATGCGAA CGGTTTGGAT GCCCGTTTGC
  101 GCGATGATAT GCAGGCAAAA CACTACGAAC CGGGTGGCAA ATACCATCTG
  151 TTTGGTAATG CTCGCGGCAG TGTTAAAAAT CGGGTTTGCG CCGTCCAAAC
  201 ATTTGATGCA ACTGCGGTCG GCCCCATACT GCCTATTACA CACGAACGGA
  251 CAGGATTTGA AGGTGTTATC GGCTATGAAA CCCATTTTTC AGGACACGGA
  301 CACGAAGTAC ACAGTCCGTT CGATAATCAT GATTCAAAAA GCACTTCTGA
  351 TTTCAGCGGC GGCGTAGACG GCGGTTTTAC CGTTTACCAA CTTCATCGGA
  401 CAGGGTCGGA AATACATCCC GCAGACGGAT ATGACGGGCC TCAAGGCGGC
  451 GGTTATCCGG AACCACAAGG GGCAAGGGAT ATATACAGCT ACCATATCAA
  501 AGGAACTTCA ACCAAAACAA AGATAAACAC TGTTCCGCAA GCCCCTTTTT
  551 CAGACCGCTG GCTAAAAGAA AATGCCGGTG CCGCTTCCGG TTTTCTCAGC
  601 CGTGCGGATG AAGCAGGAAA ACTGATATGG GAAAACGACC CCGATAAAAA
  651 TTGGCGGGCT AACCGTATGG ATGATATTCG CGGCATCGTC CAAGGTGCGG
  701 TTAATCCTTT TTTAACGGGT TTTCAAGGGG TAGGGATTGG GGCAATTACA
  751 GACAGTGCGG TAAGCCCGGT CACAGATACA GCCGCTCAGC AGACTCTACA
  801 AGGTATTAAT GATTTAGGAA ATTTAAGTCC GGAAGCACAA CTTGCCGCCG
  851 CGAGCCTATT ACAGGACAGT GCCTTTGCGG TAAAAGACGG CATCAATTCC
  901 GCCAGACAAT GGGCTGATGC CCATCCGAAT ATAACAGCAA CAGCCCAAAC
  951 TGCCCTTGCC GTAGCAGAGG CCGCAGGTAC GGTTTGGCGC GGTAAAAAAG
  1001 TAGAACTTAA CCCGACCAAA TGGGATTGGG TTAAAAATAC CGGCTATAAA
  1051 AAACCTGCTG CCCGCCATAT GCAGACTGTA GATGGGGAGA TGGCAGGGGG
  1101 GAATAGACCG CCTAAATCTA TAACGTCGGA AGGAAAAGCT AATGCTGCAA
  1151 CCTATCCTAA GTTGGTTAAT CAGCTAAATG AGCAAAACTT AAATAACATT
  1201 GCGGCTCAAG ATCCAAGATT GAGTCTAGCT ATTCATGAGG GTAAAAAAAA
  1251 TTTTCCAATA GGAACTGCAA CTTATGAAGA GGCAGATAGA CTAGGTAAAA
  1301 TTTGGGTTGG TGAGGGTGCA AGACAAACTA GTGGAGGCGG ATGGTTAAGT
  1351 AGAGATGGCA CTCGACAATA TCGGCCACCA ACAGAAAAAA AATCACAATT
  1401 TGCAACTACA GGTATTCAAG CAAATTTTGA AACTTATACT ATTGATTCAA
  1451 ATGAAAAAAG AAATAAAATT AAAAATGGAC ATTTAAATAT TAGGTAA

• This encodes a protein having amino acid sequence <SEQ ID 172; ORF29ng-1>:

• 1 MNLPIQKFMM LLAAAISMLH IPISHANGLD ARLRDDMQAK HYEPGGKYHL
  51 FGNARGSVKN RVCAVQTFDA TAVGPILPIT HERTGFEGVI GYETHFSGHG
  101 HEVHSPFDNH DSKSTSDFSG GVDGGFTVYQ LHRTGSEIHP ADGYDGPQGG
  151 GYPEPQGARD IYSYHIKGTS TKTKINTVPQ APFSDRWLKE NAGAASGFLS
  201 RADEAGKLIW ENDPDKNWRA NRMDDIRGIV QGAVNPFLTG FQGVGIGAIT
  251 DSAVSPVTDT AAQQTLQGIN DLGNLSPEAQ LAAASLLQDS AFAVKDGINS
  301 ARQWADAHPN ITATAQTALA VAEAAGTVWR GKKVELNPTK WDWVKNTGYK
  351 KPAARHMQTV DGEMAGGNRP PKSITSEGKA NAATYPKLVN QLNEQNLNNI
  401 AAQDPRLSLA IHEGKKNFPI GTATYEEADR LGKIWVGEGA RQTSGGGWLS
  451 RDGTRQYRPP TEKKSQFATT GIQANFETYT IDSNEKRNKI KNGHLNIR*

• ORF29ng-1 and ORF29-1 show 86.0% identity in 401 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 21
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 173>:

• 1 ATGAAAAAAC AAATCACCGC AGCCGTAATG ATGCTGTCTA TGATTGCCCC
  51 CGCAATGGCA AACGGCTTGG ACAATCAGGC ATTTGAAGAC CAAATGTTCC
  101 ACACGCGGGC AGATGCACCG ATGCAG...

• This corresponds to the amino acid sequence <SEQ ID 174; ORF30>:

• 1 MKKQITAAVM MLSMIAPAMA NGLDNQAFED QMFHTRADAP MQ..

• Further work revealed the complete nucleotide sequence <SEQ ID 175>:

• 1 ATGAAAAAAC AAATCACCGC AGCCGTAATG ATGCTGTCTA TGATTGCCCC
  51 CGCAATGGCA AACGGCTTGG ACAATCAGGC ATTTGAAGAC CAAGTGTTCC
  101 ACACGCGGGC AGATGCACCG ATGCAGTTGG CGGAGCTTTC TCAAAAGGAG
  151 ATGAAGGAGA CAGAGGGGGC GTTTCTTCCA TTGGCTATCT TGGGTGGTGC
  201 TGCCATTGGT ATGTGGACAC AGCATGGTTT TAGTTATGCA ACGACAGGCA
  251 GACCAGCTTC TGTTAGAGAT GTTGCTATTG CTGGCGGATT AGGCGCAATT
  301 CCTGGTGGTG TAGGCGCCGC AGGAAAGGTT GTTTCCTTTG CTAAATATGG
  351 ACGTGAGATT AAAATCGGCA ATAATATGCG GATAGCCCCT TTCGGTAATA
  401 GAACAGGTCA TCCTATTGGA AAATTTCCCC ATTATCATCG TCGAGTTACG
  451 GATAATACGG GCAAGACTTT GCCTGGACAG GGAATTGGTC GTCATCGCCC
  501 TTGGGAATCA AAATCTACGG ACAGATCATG GAAAAACCGC TTCTAA

• This corresponds to the amino acid sequence <SEQ ID 176; ORF30-1>:

• 1 MKKQITAAVM MLSMIAPAMA NGLDNQAFED QVFHTRADAP MQLAELSQKE
  51 MKETEGAFLP LAILGGAAIG MW TQHGFSYA TTGRPASVRD VAIAGGLGAI
  101 PGGVGAAGKV VSFAKYGREI KIGNNMRIAP FGNRTGHPIG KFPHYHRRVT
  151 DNTGKTLPGQ GIGRHRPWES KSTDRSWKNR F*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF30 shows 97.6% identity over a 42aa overlap with an ORF (ORF30a) from strain A of N. meningitidis:
• 
• The complete length ORF30a nucleotide sequence <SEQ ID 177> is:

• 1 ATGAAAAAAC AAATCACCGC AGCCGTAATG ATGCTGTCTA TGATTGCCCC
  51 CGCAATGGCA AACGGCTTGG ACAATCAGGC ATTTGAAGAC CAAGTGTTCC
  101 ACACGCGGGC AGATGCACCG ATGCAGTTGG CGGAGCTTTC TCAAAAGGAG
  151 ATGAAGGANA CAGNGGGGGC GTTTCTTCCA TTGGNTATCT TGGGTGGTGC
  201 TGCCATTGGT ATGTGGACAC AGCATGGTTT TAGTTATGCA ACGACAGGCA
  251 GACCAGCTTC TGTTAGAGAT GTTGCTATTG CTGGCGGATT AGGCGCAATT
  301 CCTGGTGNTG TAGGCGCCGC AGGAAAGGTT GTTTCCTTTG CTAAATATGG
  351 ACGTGAGATT AAAATCGGCA ATAATATGCG GATAGCCCCT TTCGGTAATA
  401 GAACAGGTCA TCCTATTGGN AAATTTCCCC ATTATCATCG TCGAGTTACG
  451 GATAATACGG GCAAGACTTT GCCTGGACAG GGAATTGGTC GTCATCGCCC
  501 TTGGGAATCA AAATCTACGG ACAGATCATG GAAAAACCGC TTCTAA

• This encodes a protein having amino acid sequence <SEQ ID 178>:

• 1 MKKQITAAVM MLSMIAPAMA NGLDNQAFED QVFHTRADAP MQLAELSQKE
  51 MKXTXGAFLP LXILGGAAIG MW TQHGFSYA TTGRPASVRD VAIAGGLGAI
  101 PGXVGAAGKV VSFAKYGREI KIGNNMRIAP FGNRTGHPIG KFPHYHRRVT
  151 DNTGKTLPGQ GIGRHRPWES KSTDRSWKNR F*

• ORF30a and ORF30-1 show 97.8% identity in 181 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF30 shows 97.6% identity over a 42aa overlap with a predicted ORF (ORF30.ng) from N. gonorrhoeae:
• 
• The complete length ORF30ng nucleotide sequence <SEQ ID 179> is

• 1 ATGAAAAAAC AAATCACCGC AGCCGTAATG ATGCTGTCTA TGATCGCCCC
  51 CGCAATGGCA AACGGATTGG ACAATCAGGC ATTTGAAGAC CAAGTGTTCC
  101 ACACGCGGGC AGATGCGCCG ATGCAGTTGG CGGAGCTTTC TCAGAAGGAG
  151 ATGAAGGAGA CTGAAGGGGC TTTTCTTCCA TTGGCTATCT TGGGTGGTGC
  201 TGCCATTGGT ATGTGGACAC AGCATGGTTT TAGTTATGCA ACGACAGGCA
  251 GACCAGCTTC TGTTAGAGAT GTTGCTGGCG GATTAGGCGC AATTCCTGGT
  301 GATGTAGGTG CTGCAGGAAA GGTTGTTTCC TTTGCTAAAT ATGGACGTGA
  351 GATTAAAATC GGCAATAATA TGCGGATAGC CCCTTTCGGT AATAGAACAG
  401 GTCATCCTAT TGGAAAATTT CCCCATTATC ATCGTCGAGT TACGGATAAT
  451 ACGGGCAAGA CTTTGCCTGG ACAGGGAATT GGTCGTCATC GCCCTTGGGA
  501 ATCAAAATCT ACGGACAGAT CATGGAAAAA CCGCTTCTAA

• This encodes a protein having amino acid sequence <SEQ ID 180>:

• 1 MKKQITAAVM MLSMIAPAMA NGLDNQAFED QVFHTRADAP MQLAELSQKE
  51 MKETEGAFLP LAILGGAAIG MWTQHGFSYA TTGRPASVRD VAGGLGAIPG
  101 DVGAAGKVVS FAKYGREIKI GNNMRIAPFG NRTGHPIGKF PHYHRRVTDN
  151 TGKTLPGQGI GRHRPWESKS TDRSWKNRF*

• ORF30ng and ORF30-1 show 98.3% identity in 181 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 22
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 181>:

• 1 ATGAATAAAA CTCTCTATCG TGTAATTTTC AACCGCAAAC GTGGGGCTGT
  51 GrTAGCCGTT GCTGAAACTA CCAAGCGCGA AGGTAAAAGC TGTGCCGATA
  101 GTGATTCAGG CAGCGCTCAT GTGAAATCTG TTCCTTTTGG TACTACTCAT
  151 GCACCTGTTT GTg.CGTTaC AAATATCTTT TCTTTTTCTT TATTGGGCTT
  201 TTCTTTATGT TTGGCTGTAG GtacGGyCAA TATTGCTTTT GCTGATGGCA
  251 TT..

• This corresponds to the amino acid sequence <SEQ ID 182; ORF31>:

• 1 MNKTLYRVIF NRKRGAVXAV AETTKREGKS CADSDSGSAH VKSVPFGTTH
  51 APVCXVTNIF SFSLLGFSLC LAVGTXNIAF ADGI..

• Further work revealed a further partial nucleotide sequence <SEQ ID 183>:

• 1 ATGAATAAAA CTCTCTATCG TGTAATTTTC AACCGCAAAC GTGGGGCTGT
  51 GGTAGCCGTT GCTGAAACTA CCAAGCGCGA AGGTAAAAGC TGTGCCGATA
  101 GTGATTCAGG CAGCGCTCAT GTGAAATCTG TTCCTTTTGG TACTACTCAT
  151 GCACCTGTTT GTCGTTCAAA TATCTTTTCT TTTTCTTTAT TGGGCTTTTC
  201 TTTATGTTTG GCTGTAGGTA CGGCCAATAT TGCTTTTGCT GATGGCATT..

• This corresponds to the amino acid sequence <SEQ ID 184; ORF31-1>:
• • 1 MNKTLYRVIF NRKRGAVVAV AETTKREGKS CADSDSGSAH VKSVPFGTTH
  • 51 APVCRSNIFS FSLLGFSLCL AVGTANIAFA DGI . . . .
• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF31 shows 76.2% identity over a 84aa overlap with a predicted ORF (ORF31.ng) from N. gonorrhoeae:
• 
• The complete length ORF31ng nucleotide sequence <SEQ ID 185> is:

• 1 ATGAACAAAA CCCTCTATCG TGTGATTTTC AACCGCAAAC GCGGTGCTGT
  51 GGTAGCTGTT GCCGAAACCA CCAAGCGCGA AGGTAAAAGC TGTGCCGATA
  101 GTGGTTCGGG CAGCGTTTAT GTGAAATCCG TTTCTTTCAT TCCTACTCAT
  151 TCCAAAGCCT TTTGTTTTTC TGCATTAGGC TTTTCTTTAT GTTTGGCTTT
  201 GGGTACGGTC AATATTGCTT TTGCTGACGG CATTATTACT GATAAAGCTG
  251 CTCCTAAAAC CCAACAAGCC ACGATTCTGC AAACAGGTaa cGGCATACCG
  301 CAAGTCAATA TTCAAACCCC TACTTCGGCA GGGGTTTCTG TTAATCAATA
  351 TGCCCAGTTT GATGTGGGTA ATCGCGGGGC GATTTTAAAC AACAGTCGCA
  401 GCAACACCCA AACACAGCTA GGCGGTTGGA TTCAAGGCAA TCCTTGGTTG
  451 ACAAGGGGCG AAGCACGTGT GGTTGTAAAC CAAATCAACA GCAGCCATCC
  501 TTCACAACTG AATGGCTATA TTGAAGTGGG TGGACGACGT GCAGAAGTCG
  551 TTATTGCCAA TCCGGCAGGG ATTGCAGTCA ATGGTGGTGG TTTTATCAAT
  601 GCTTCCCGTG CCACTTTGAC GACAGGCCAA CCGCAATATC AAGCAGGAGA
  651 CTTTAGCGGC TTTAAGATAA GGCAAGGCAA TGCTGTAATC GCCGGACACG
  701 GTTTGGATGC CCGTGATACC GATTTCACAC GTATTCTTGT ATGCCAACAA
  751 AATCACCTTG ATCAGTACGG CCGAACAAGC AGGCATTCGT AA

• This encodes a protein having amino acid sequence <SEQ ID 186>:

• 1	MNKTLYRVIF NRKRGAVVAV AETTKREGKS CADSGSGSVY
  	VKSVSFIPTH
  51	SKAFCFSALG FSLCLALGTV NIAFADGIIT DKAAPKTQQA
  	TILQTGNGIP
  101	QVNIQTPTSA GVSVNQYAQF DVGNRGAILN NSRSNTQTQL
  	GGWIQGNPWL
  151	TRGEARVVVN QINSSHPSQL NGYIEVGGRR AEVVIANPAG
  	IAVNGGGFIN
  201	ASRATLTTGQ PQYQAGDFSG FKIRQGNAVI AGHGLDARDT
  	DFTRILVCQQ
  251	NHLDQYGRTS RHS*

• This gonococcal protein shares 50% identity over a 149aa overlap with the pore-forming hemolysins-like HecA protein from Erwinia chrysanthemi (accession number L39897):

• orf31ng	96	GNGIPQVNIQTPTSAGVSVNQYAQFDVGNRGAILNNSRSN-TQTQLGGWIQGNPWLTRGE	154
  		GNG+P VNI TP ++G+S N+Y  F+V NRG ILNN  +  T +QLGG IQ NP L
  HecA	45	GNGVPVVNIATPDASGLSHNRYHDFNVDNRGLILNNGTARLTPSQLGGLIQNNPNLNGRA	104
  Orf31ng	155	ARVVVNQINSSHPSQLNGYIEVGGRRAEVVIANPAGIAVNGGGFINASRATLTTGQPQYQ	214
  		A  ++N++ S + S+L GY+EV G+ A VV+ANP GI  +G GF+N  R TLTTG PQ+
  HecA	105	AAAILNEVVSPNRSRLAGYLEVAGQAANVVVANPYGITCSGCGFLNTPRLTLTTGTPQFD	164
  Orf31ng	215	-AGDFSGFKIRQGNAVIAGHGLDARDTDF	242
  		AG  SG  +R G+ +I G GLDA  +D+
  HecA	165	AAGGLSGLDVRGGDILIDGAGLDASRSDY	193

• Furthermore, ORF31ng and ORF31-1 show 79.5% identity in 83 aa overlap:
• 
• On this basis, including the homology with hemolysins, and also with adhesins, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 23
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 187>:

• 1	ATGAATACTC CTCCTTTTGT CTGTTGGATT TTTTGCAAGG
  	TCATCGACAA
  51	TTTCGGCGAC ATCGGCGTTT CGTGGCGGCT CGCCCGTGTT
  	TTGCACCGCG
  101	AACTCGGTTG GCAGGTGCAT TTGTGGACGG ACGATGTGTC
  	CGCCTTGCGT
  151	GCGCTTTGCC CTGATTTGCC CGATGTTCCC TGCGTTCATC
  	AGGATATTCA
  201	TGTCCGCACT TGGCATTCCG ATGCGGCAGA TATTGATACC
  	GCG..

• This corresponds to the amino acid sequence <SEQ ID 188; ORF32>:

• 1	MNTPPFVCWI FCKVIDNFGD IGVSWRLARV LHRELGWQVH
  	LWTDDVSALR
  51	ALCPDLPDVP CVHQDIHVRT WHSDAADIDT A..

• Further work revealed the complete nucleotide sequence <SEQ ID 189>:

• 1	ATGAATACTC CTCCTTTTGT CTGTTGGATT TTTTGCAAGG
  	TCATCGACAA
  51	TTTCGGCGAC ATCGGCGTTT CGTGGCGGCT CGCCCGTGTT
  	TTGCACCGCG
  101	AACTCGGTTG GCAGGTGCAT TTGTGGACGG ACGATGTGTC
  	CGCCTTGCGT
  151	GCGCTTTGCC CTGATTTGCC CGATGTTCCC TGCGTTCATC
  	AGGATATTCA
  201	TGTCCGCACT TGGCATTCCG ATGCGGCAGA TATTGATACC
  	GCGCCTGTTC
  251	CCGATGTCGT CATCGAAACT TTTGCCTGCG ACCTGCCCGA
  	AAATGTGCTG
  301	CACATTATCC GCCGACACAA GCCGCTTTGG CTGAATTGGG
  	AATATTTGAG
  351	CGCGGAGGAA AGCAATGAAA GGCTGCATCT GATGCCTTCG
  	CCGCAGGAGG
  401	GTGTTCAAAA ATATTTTTGG TTTATGGGTT TCAGCGAAAA
  	AAGCGGCGGG
  451	TTGATACGCG AACGTGATTA CTGCGAAGCC GTCCGTTTCG
  	ATACTGAAGC
  501	CCTGCGAGAG CGGCTGATGC TGCCCGAAAA AAACGCCTCC
  	GAATGGCTGC
  551	TTTTCGGCTA TCGGAGCGAT GTTTGGGCAA AGTGGCTGGA
  	AATGTGGCGA
  601	CAGGCAGGCA GCCCGATGAC ACTGTTGCTG GCGGGGACGC
  	AAATCATCGA
  651	CAGCCTCAAA CAAAGCGGCG TTATTCCGCA AGATGCCCTG
  	CAAAACGACG
  701	GCGATGTTTT TCAGACGGCA TCCGTCCGCC TCGTCAAAAT
  	CCCTTTCGTG
  751	CCGCAACAGG ACTTCGACCA ACTGCTGCAC CTTGCCGACT
  	GCGCCGTCAT
  801	CCGCGGCGAA GACAGTTTCG TGCGCGCCCA GCTTGCGGGC
  	AAACCCTTCT
  851	TTTGGCACAT CTACCCGCAA GACGAGAATG TCCATCTCGA
  	CAAACTCCAC
  901	GCCTTTTGGG ATAAGGCACA CGGTTTCTAC ACGCCCGAAA
  	CCGTGTCGGC
  951	ACACCGCCGT CTTTCGGACG ACCTCAACGG CGGAGAGGCT
  	TTATCCGCAA
  1001	CACAACGCCT CGAATGTTGG CAAACCCTGC AACAACATCA
  	AAACGGCTGG
  1051	CGGCAAGGCG CGGAGGATTG GAGCCGTTAT CTTTTCGGGC
  	AGCCGTCAGC
  1101	TCCTGAAAAA CTCGCTGCCT TTGTTTCAAA GCATCAAAAA
  	ATACGCTAG

• This corresponds to the amino acid sequence <SEQ ID 190; ORF32-1>:

• 1	MNTPPFVCWI FCKVIDNFGD IGVSWRLARV LHRELGWQVH
  	LWTDDVSALR
  51	ALCPDLPDVP CVHQDIHVRT WHSDAADIDT APVPDVVIET
  	FACDLPENVL
  101	HIIRRHKPLW LNWEYLSAEE SNERLHLMPS PQEGVQKYFW
  	FMGFSEKSGG
  151	LIRERDYCEA VRFDTEALRE RLMLPEKNAS EWLLFGYRSD
  	VWAKWLEMWR
  201	QAGSPMTLLL AGTQIIDSLK QSGVIPQDAL QNDGDVFQTA
  	SVRLVKIPFV
  251	PQQDFDQLLH LADCAVIRGE DSFVRAQLAG KPFFWHIYPQ
  	DENVHLDKLH
  301	AFWDKAHGFY TPETVSAHRR LSDDLNGGEA LSATQRLECW
  	QTLQQHQNGW
  351	RQGAEDWSRY LFGQPSAPEK LAAFVSKHQK IR*w

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF32 shows 93.8% identity over a 81 as overlap with an ORF (ORF32a) from strain A of N. meningitidis:
• 
• The complete length ORF32a nucleotide sequence <SEQ ID 191> is:

• 1	ATGAATACTC CTCCTTTTTC TGCTGGANTT TTTTGCAAGG
  	TCATCGACAA
  51	TTTCGGCGAC ATCGGCGTTT CGTGGCGGCT TGCCCGTGTT
  	TTGCACCGCG
  101	AACTCGGTTG GCAGGTGCAT TTGTGGACGG ACGATGTGTC
  	CGCCTTGCGT
  151	GCGCTTTGCC CTGATTTGCC CGATGTTCNC TGCGTTCATC
  	AGGATATTCA
  201	TGTCCGCACT TGGCATTCCG ATGCGGCAGA TATTGATACC
  	GCGCCTGTTC
  251	NCGATGTCGT CATCGAAACT TTTGCCTGCG ACCTGCCCGA
  	AAATGTGCTG
  301	CACATCATCC GCCGACACAA GCCGCTTTGG CTGAANTGGG
  	AATATTTGAG
  351	CGCGGAGGAN AGCAATGAAA GGCTGCACNT GATGCCTTCG
  	CCGCAGGAGA
  401	GTGTTCNAAA ATANTTTTGG TTTATGGGTT TCAGCGAANN
  	NAGCGGCGGA
  451	CTGATACGCG AACGCGATTA CTGCGAAGCC GTCCGTTTCG
  	ATAGCGGAGC
  501	CTTGCGCAAG AGGCTGATGC TTCCCGAAAA AAACGNCCCC
  	GAATGGCTGC
  551	TTTTCGGCTA TCGGAGCGAT GTTTGGGCAA AGTGGCTGGA
  	AATGTGGCGA
  601	CAGGCAGGCA GTCCGTTGAC ACTTTTGCTG GCNGGGGCGC
  	ANATTATCGA
  651	CAGCCTCAAA CAAAACGGCG TTATTCCGCA AGATGCCCTG
  	CAAAACGACG
  701	GCGATGTTTT TCAGACGGCA TCCGTCCGCC TCGTCAAAAT
  	CCCTTTCGTG
  751	CCGCAACAGG ACTTCGACAA ACTGCTGCAC CTTGCCGACT
  	GCGCCGTCAT
  801	CCGCGGCGAA GACAGTTTCG TGCGCGCCCA GCTTGCGGGC
  	AAACCCTTCT
  851	TTTGGCACAT CTACCCGCAA GATGAGAATG TCCATCTCGA
  	CAAACTCCAC
  901	GCCTTTTGGG ATAAGGCACA CGGTTTCTAC ACGCCCGAAA
  	CCGCATCGGC
  951	ACACCGCCGC CTTTCAGACG ACCTCAACGG CGGAGAGGCT
  	TTATCCGCAA
  1001	CACAACGCCT CGAATGTTGG CAAATCCTGC AACAACATCA
  	AAACGGCTGG
  1051	CGGCAAGGCG CGGAGGATTG GAGCCGTTAT CTTTTTGGGC
  	AGCCTTCCGC
  1101	ATCCGAAAAA CTCGCCGCCT TTGTTTCAAA GCATCAAAAA
  	ATACGCTAG

• This encodes a protein having amino acid sequence <SEQ ID 192>:

• 1	MNTPPFSAGX FCKVIDNFGD IGVSWRLARV LHRELGWQVH
  	LWTDDVSALR
  51	ALCPDLPDVX CVHQDIHVRT WHSDAADIDT APVXDVVIET
  	FACDLPENVL
  101	HIIRRHKPLW LXWEYLSAEX SNERLHXMPS PQESVXKXFW
  	FMGFSEXSGG
  151	LIRERDYCEA VRFDSGALRK RLMLPEKNXP EWLLFGYRSD
  	VWAKWLEMWR
  201	QAGSPLTLLL AGAXIIDSLK QNGVIPQDAL QNDGDVFQTA
  	SVRLVKIPFV
  251	PQQDFDKLLH LADCAVIRGE DSFVRAQLAG KPFFWHIYPQ
  	DENVHLDKLH
  301	AFWDKAHGFY TPETASAHRR LSDDLNGGEA LSATQRLECW
  	QILQQHQNGW
  351	RQGAEDWSRY LFGQPSASEK LAAFVSKHQK IR*

• ORF32a and ORF32-1 show 93.2% identity in 382 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF32 shows 95.1% identity over a 82aa overlap with a predicted ORF (ORF32.ng) from N. gonorrhoeae:
• 
• An ORF32ng nucleotide sequence <SEQ ID 193> was predicted to encode a protein having amino acid sequence <SEQ ID 194>:

• 1	MVMNTYAFPV CWIFCKVIDN FGDIGVSWRL ARVLHRELGW
  	QVHLWTDDVS
  51	ALRALCPDLP DVPFVHQDIH VRTWHSDAAD IDTAPVPDAV
  	IETFACDLPE
  101	NVLNIIRRHK PLWLNWEYLS AEESNERLHL MPSPQEGVQK
  	YFWFMGFSEK
  151	SGGLIRERDY REAVRFDTEA LRRRLVLPEK NAPEWLLFGY
  	RGDVWAKWLD
  201	MWQQAGSLMT LLLAGAQIID SLKQSGVIPQ NALQNEGGVF
  	QTASVRLVKI
  251	PFVPQQDFDK LLHLADCAVI RGEDSFVRTQ LAGKPFFWHI
  	YPQDENVHLD
  301	KLHAFWDKAY GFYTPETASV HRLLSDDLNG GEALSATQRL
  	ECGVL*

• Further sequencing revealed the following DNA sequence <SEQ ID 195>:

• 1	ATGAATACAT ACGCTTTTCC TGTCTGTTGG ATTTTTTGCA
  	AGGTCATCGA
  51	CAATTTCGGC GACATCGGCG TTTCGTGGCG GCTCGCCCGT
  	GTTTTGCACC
  101	GCGAACTCGG TTGGCAGGTG CATTTGTGGA CGGACGACGT
  	GTCCGCCTTG
  151	CGCGCGCTTT GTCCCGATTT GCCCGATGTT CCCTTCGTTC
  	ATCAGGATAT
  201	TCATGTCCGC ACTTGGCATT CCGATGCGGC AGACATTGAT
  	ACCGCGCCCG
  251	TTCCCGATGC CGTTATCGAA ACTTTTGCCT GCGACCTGCC
  	CGAAAATGTG
  301	CTGAACATCA TCCGCCGACA CAAACCGCTT TGGCTGAATT
  	GGGAATATTT
  351	GAGCGCGGAG GAAAGCAATG AAAGGCTGCA CCTGATGCCT
  	TCGCCGCAGG
  401	AGGGCGTTCA AAAATATTTT TGGTTTATGG GTTTCAGCGA
  	AAAAAGCGGC
  451	GGGTTGATAC GCGAACGCGA TTACCGCGAA GCCGTCCGTT
  	TCGATACCGA
  501	AGCCCTGCGC CGGCGGCTGG TGCTGCCCGA AAAAAACGCC
  	CCCGAATGGC
  551	TGCTTTTCGG CTATCGGGGC GATGTTTGGG CAAAGTGGCT
  	GGACATGTGG
  601	CAACAGGCAG GCAGCCTGAT GACCCTACTG CTGGCGGGGG
  	CGCAAATTAT
  651	CGACAGCCTC AAACAAAGCG GCGTTATTCC GCAAAACGCC
  	CTGCAAAAtg
  701	aaggcgGTGT CTTTCagacG gcatccgTcC gccttGTCAA
  	AAtcCCGTTC
  751	GTGCcGCAAC AGGAcTTCGA CAAATTGCTG CAcctcgcCG
  	ACTGCGCCGT
  801	GATACGCGGC GAAGACAGTT TCGTGCGTAC CCAGCTTGCC
  	GGAAAACCCT
  851	TTTTTTGGCA CATCTACCCG CAAGACGAGA ATGTCCATCT
  	CGACAAACTC
  901	CACGCCTTTT GGGATAAGGC ATACGGCTTC TACACGCCCG
  	AAACCGCATC
  951	GGTGCACCGC CTCCTTTCGG ACGACCTCAA CGGCGGAGAG
  	GCTTTATCCG
  1001	CAACACAACG CCTCGAATGT TGGCAAACCC TGCAACAACA
  	TCAAAACGGC
  1051	TGGCGGCAAG GCGCGGAGGA TTGGAGCCGT TATCTTTTCG
  	GGCAGCCTTC
  1101	CGCATCCGAA AAACTCGCCG CCTTTGTTTC AAAGCATCAA
  	AAAATACGCT
  1151	AG

• This encodes a protein having amino acid sequence <SEQ ID 196; ORF32ng-1>:

• 1	MNTYAFPVCW IFCKVIDNFG DIGVSWRLAR VLHRELGWQV
  	HLWTDDVSAL
  51	RALCPDLPDV PFVHQDIHVR TWHSDAADID TAPVPDAVIE
  	TFACDLPENV
  101	LNIIRRHKPL WLNWEYLSAE ESNERLHLMP SPQEGVQKYF
  	WFMGFSEKSG
  151	GLIRERDYRE AVRFDTEALR RRLVLPEKNA PEWLLFGYRG
  	DVWAKWLDMW
  201	QQAGSLMTLL LAGAQIIDSL KQSGVIPQNA LQNEGGVFQT
  	ASVRLVKIPF
  251	VPQQDFDKLL HLADCAVIRG EDSFVRTQLA GKPFFWHIYP
  	QDENVHLDKL
  301	HAFWDKAYGF YTPETASVHR LLSDDLNGGE ALSATQRLEC
  	WQTLQQHQNG
  351	WRQGAEDWSR YLFGQPSASE KLAAFVSKHQ KIR*

• ORF32ng-1 and ORF32-1 show 93.5% identity in 383 aa overlap:
• 
• On this basis, including the RGD sequence in the gonococcal protein, characteristic of adhesins, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF32-1 (42 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 7A shows the results of affinity purification of the His-fusion protein, and FIG. 7B shows the results of expression of the GST-fusion in E. coli. Purified His-fusion protein was used to immunise mice, whose sera were used for ELISA, giving a positive result. These experiments confirm that ORF32-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 24
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 197>:

• 1	..TTGTTCCTGC GTGTNAAAGT GGGGCGTTTT TTCAGCAGTC
  	CGGCGACGTG
  51	GTTTCGGGNC AAAGACCCTG TAAATCAGGC GGTGTTGCGG
  	CTGTATNCGG
  101	ACGAGTGGCG GCA.ACTTCG GTACGTTGGA AAATAGNCGC
  	AACGTCGCAC
  151	AGCCTGTGGC TCTGCACGCT GCTCGGAATG CTGGTGTCGG
  	TATTGTTGCT
  201	GCTTTTGGTG CGGCAATATA CGTTCAACTG GGAAAGCACG
  	CTGTTGAGCA
  251	ATGCCGCTTC GGTACGCGCG GTGGAAATGT TGGCATGGCT
  	GCCGTCGAAA
  301	CTCGGTTTCC CTGTCCCCGA TGCGCGGTCG GTCATCGAAG
  	GCCGTCTGAA
  351	CGGCAATATT GCCGATGCGC GGGCTTGGTC GGGGCTGCTG
  	GTCGNCAGTA
  401	TCGCCTGCTA NGGCATCCTG CCGCGCCTG..

• This corresponds to the amino acid sequence <SEQ ID 198; ORF33>:

• 1	..LFLRVKVGRF FSSPATWFRX KDPVNQAVLR LYXDEWRXTS
  	VRWKIXATSH
  51	SLWLCTLLGM LVSVLLLLLV RQYTFNWEST LLSNAASVRA
  	VEMLAWLPSK
  101	LGFPVPDARS VIEGRLNGNI ADARAWSGLL VXSIACXGIL
  	PRL..

• Further work revealed the complete nucleotide sequence <SEQ ID 199>:

• 1	ATGTTGAATC CATCCCGAAA ACTGGTTGAG CTGGTCCGTA
  	TTTTGGACGA
  51	AGGCGGTTTT ATTTTCAGCG GCGATCCCGT ACAGGCGACG
  	GAGGCTTTGC
  101	GCCGCGTGGA CGGCAGTACG GAGGAAAAAA TCATCCGTCG
  	GGCGGAGATG
  151	ATTGACAGGA ACCGTATGCT GCGGGAGACG TTGGAACGTG
  	TGCGTGCGGG
  201	GTCGTTCTGG TTGTGGGTGG TGGCGGCGAC GTTTGCATTT
  	TTTACCGGTT
  251	TTTCAGTCAC TTATCTTCTA ATGGACAATC AGGGTCTGAA
  	TTTCTTTTTG
  301	GTTTTGGCGG GCGTGTTGGG CATGAATACG CTGATGCTGG
  	CAGTATGGTT
  351	GGCAATGTTG TTCCTGCGTG TGAAAGTGGG GCGTTTTTTC
  	AGCAGTCCGG
  401	CGACGTGGTT TCGGGGCAAA GACCCTGTAA ATCAGGCGGT
  	GTTGCGGCTG
  451	TATGCGGACG AGTGGCGGCA ACCTTCGGTA CGTTGGAAAA
  	TAGGCGCAAC
  501	GTCGCACAGC CTGTGGCTCT GCACGCTGCT CGGAATGCTG
  	GTGTCGGTAT
  551	TGTTGCTGCT TTTGGTGCGG CAATATACGT TCAACTGGGA
  	AAGCACGCTG
  601	TTGAGCAATG CCGCTTCGGT ACGCGCGGTG GAAATGTTGG
  	CATGGCTGCC
  651	GTCGAAACTC GGTTTCCCTG TCCCCGATGC GCGGGCGGTC
  	ATCGAAGGCC
  701	GTCTGAACGG CAATATTGCC GATGCGCGGG CTTGGTCGGG
  	GCTGCTGGTC
  751	GGCAGTATCG CCTGCTACGG CATCCTGCCG CGCCTGCTGG
  	CTTGGGTAGT
  801	GTGTAAAATC CTTTTGAAAA CAAGCGAAAA CGGATTGGAT
  	TTGGAAAAGC
  851	CCTATTATCA GGCGGTCATC CGCCGCTGGC AGAACAAAAT
  	CACCGATGCG
  901	GATACGCGTC GGGAAACCGT GTCCGCCGTT TCACCGAAAA
  	TCATCTTGAA
  951	CGATGCGCCG AAATGGGCGG TCATGCTGGA GACCGAGTGG
  	CAGGACGGCG
  1001	AATGGTTCGA GGGCAGGCTG GCGCAGGAAT GGCTGGATAA
  	GGGCGTTGCC
  1051	ACCAATCGGG AACAGGTTGC CGCGCTGGAG ACAGAGCTGA
  	AGCAGAAACC
  1101	GGCGCAACTG CTTATCGGCG TGCGCGCCCA AACTGTGCCG
  	GACCGCGGCG
  1151	TGTTGCGGCA GATTGTCCGA CTCTCGGAAG CGGCGCAGGG
  	CGGCGCGGTG
  1201	GTGCAGCTTT TGGCGGAACA GGGGCTTTCA GACGACCTTT
  	CGGAAAAGCT
  1251	GGAACATTGG CGTAACGCGC TGGCCGAATG CGGCGCGGCG
  	TGGCTTGAGC
  1301	CTGACAGGGC GGCGCAGGAA GGGCGTTTGA AAGACCAATA
  	A

• This corresponds to the amino acid sequence <SEQ ID 200; ORF33-1>:

• 1	MLNPSRKLVE LVRILDEGGF IFSGDPVQAT EALRRVDGST
  	EEKIIRRAEM
  51	IDRNRMLRET LERVRAGSFW LWVVAATFAF FTGFS VTYLL
  	MDNQGLNFFL
  101	VLAGVLGMNT LMLAVWLAML FLRVKVGRFF SSPATWFRGK
  	DPVNQAVLRL
  151	YADEWRQPSV RWKIGATSHS LWLCTLLGML VSVLLLLLVR
  	QYTFNWESTL
  201	LSNAASVRAV EMLAWLPSKL GFPVPDARAV IEGRLNGNIA
  	DARAWSGLLV
  251	GSIACYGILP RLLAWVVCKI LLKTSENGLD LEKPYYQAVI
  	RRWQNKITDA
  301	DTRRETVSAV SPKIILNDAP KWAVMLETEW QDGEWFEGRL
  	AQEWLDKGVA
  351	TNREQVAALE TELKQKPAQL LIGVRAQTVP DRGVLRQIVR
  	LSEAAQGGAV
  401	VQLLAEQGLS DDLSEKLEHW RNALAECGAA WLEPDRAAQE
  	GRLKDQ*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF33 shows 90.9% identity over a 143aa overlap with an ORF (ORF33a) from strain A of N. meningitidis:
• 
• The complete length ORF33a nucleotide sequence <SEQ ID 201> is:

• 1	ATGTTGAATC CATCCCGAAA ACTGGTTGAG CTGGTCCGTA
  	TTTTGGAAGA
  51	AGGCGGCTTT ATTTTCAGCG GCGATCCCGT GCAGGCGACG
  	GAGGCTTTGC
  101	GCCGCGTGGA CGGCAGTACG GAGGAAAAAA TCATCCGTCG
  	GGCGAAGATG
  151	ATCGACAGGA ACCGTATGCT GCGGGAGACG TTGGAACGTG
  	TGCGTGCGGG
  201	GTCGTTCTGG TTGTGGGTGG CGGCGGCGAC GTTTGCGTTT
  	NTTACCGNTT
  251	TTTCAGTTAC TTATCTTCTA ATGGACAATC AGGGTCTGAA
  	TTTCTTTTTG
  301	GTTTTGGCGG GCGTGNTGGG CATGAATACG CTGATGCTGG
  	CAGTATGGTT
  351	GGCAATGTTG TTCCTGCGCG TGAAAGTGGG GCGTTTTTTC
  	AGCAGTCCGG
  401	CGACGTGGTT TCGGGGCAAA GACCCTGTCA ATCAGGCGGT
  	GTTGCGGCTG
  451	TATGCGGACG AGTGGCGGCN ACCTTCGGTA CGTTGGAAAA
  	TAGGCGCAAC
  501	GTCGCACAGC CTGTGGCTCT GCACGCTGCT CGGAATGCTG
  	GTGTCGGTAT
  551	TGTTGCTGCT TTTGGTGCGG CAATATACGT TCAACTGGGA
  	AAGCACGCTG
  601	TTGGGCGATT CGTCTTCGGT ACGGCTGGTG GAAATGTTGG
  	CATGGCTGCC
  651	TGCGAAACTG GGTTTTCCCG TGCCTGATGC GCGGGCGGTC
  	ATCGAAGGTC
  701	GTCTGAACGG CAATATTGCC GATGCGCGGG CTTGGTCGGG
  	GCTGCTGGTC
  751	GGCAGTATCG CCTGCTACGG CATCCTGCCG CGCCTCTTGG
  	CTTGGGCGGT
  801	ATGCAAAATC CTTNTGNAAA CAAGCGAAAA CGGCTTGGAT
  	TTGGAAAAGC
  851	NCNNNNNTCN NNCGNTCATC CGCCGCTGGC AGAACAAAAT
  	CACCGATGCG
  901	GATACGCGTC GGGAAACCGT GTCCGCCGTT TCGCCGAAAA
  	TCGTCTTGAA
  951	CGATGCGCCG AAATGGGCGG TCATGCTGGA GACCGAATGG
  	CAGGACGGCG
  1001	AATGGTTCGA GGGCAGGCTG GCGCAGGAAT GGCTGGATAA
  	GGGCGTTGCC
  1051	GCCAATCGGG AACAGGTTGC CGCGCTGGAG ACAGAGCTGA
  	AGCAGAAACC
  1101	GGCGCAACTG CTTATCGGCG TGCGCGCCCA AACTGTGCCC
  	GACCGCGGCG
  1151	TGTTGCGGCA GATCGTCCGA CTTTCGGAAG CGGCGCAGGG
  	CGGCGCGGTG
  1201	GTGCANCTTT TGGCGGAACA GGGGCTTTCA GACGACCTTT
  	CGGAAAAGCT
  1251	GGAACATTGG CGTAACGCGC TGACCGAATG CGGCGCGGCG
  	TGGCTGGAAC
  1301	CCGACAGAGC GGCGCAGGAA GGCCGTCTGA AAACCAACGA
  	CCGCACTTGA

• This encodes a protein having amino acid sequence <SEQ ID 202>:

• 1	MLNPSRKLVE LVRILEEGGF IFSGDPVQAT EALRRVDGST
  	EEKIIRRAKM
  51	IDRNRMLRET LERVRAGSFW LWVAAATFAF XTXFS VTYLL
  	MDNQGLNFFL
  101	VLAGVXGMNT LMLAVWLAML FLRVKVGRFF SSPATWFRGK
  	DPVNQAVLRL
  151	YADEWRXPSV RWKIGATSHS LWLCTLLGML VSVLLLLLVR
  	QYTFNWESTL
  201	LGDSSSVRLV EMLAWLPAKL GFPVPDARAV IEGRLNGNIA
  	DARAWSGLLV
  251	GSIACYGILP RLLAWAVCKI LXXTSENGLD LEKXXXXXXI
  	RRWQNKITDA
  301	DTRRETVSAV SPKIVLNDAP KWAVMLETEW QDGEWFEGRL
  	AQEWLDKGVA
  351	ANREQVAALE TELKQKPAQL LIGVRAQTVP DRGVLRQIVR
  	LSEAAQGGAV
  401	VXLLAEQGLS DDLSEKLEHW RNALTECGAA WLEPDRAAQE
  	GRLKTNDRT*

• ORF33a and ORF33-1 show 94.1% identity in 444 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF33 shows 91.6% identity over a 143aa overlap with a predicted ORF (ORF33.ng) from N. gonorrhoeae:
• 
• An ORF33ng nucleotide sequence <SEQ ID 203> was predicted to encode a protein having amino acid sequence <SEQ ID 204>:

• 1	MIDRDRMLRD TLERVRAGSF WLWVVVASMM FTAGFSGTYL
  	LMDNQGLNFF
  51	LVLAGVLGMN TLMLAVWLAT LFLRVKVGRF FSSPATWFRG
  	KGPVNQAVLR
  101	LYADQWRQPS VRWKIGATAH SLWLCTLLGM LVSVLLLLLV
  	RQYTFNWEST
  151	LLSNAASVRA VEMLAWLPSK LGFPVPDARA VIEGRLNGNI
  	ADARAWSGLL
  201	VGSIVCYGIL PRLLAWVVCK ILLKTSENGL DLEKTYYQAV
  	IRRWQNKITD
  251	ADTRRETVSA VSPKIVLNDA PKWALMLETE WQDGQWFEGR
  	LAQEWLDKGV
  301	AANREQVAAL ETELKQKPAQ LLIGVRAQTV PDRGVLRQIV
  	RLSEAAQGGA
  351	VVQLLAEQGL SDDLSEKLEH WRNALTECGA AWLEPDRVAQ
  	EGRLKDQ*

• Further sequence analysis revealed the following DNA sequence <SEQ ID 205>:

• 1	ATGTTGaatC CATCCCgaAA ACTGgttgag ctGgTCCgtA
  	Ttttgaataa
  51	agggggtTTT attttcagcg gcgatcctgt gcaggcgacg
  	gaggctttgc
  101	gccgcgtgga cggcAGTACG GAggAaaaaa tcttccgtcg
  	GGCGGAGAtg
  151	atcgACAGGg accgtatgtt gcgggACaCg TtggaacGTG
  	TGCGTGCggg
  201	gtcgtTctgG TTATGGGTGG TggtggCAtC gATGATGTtt
  	aCCGCCGGAT
  251	TTTCAGgcac ttatCttCTG ATGGACaatC AGGGGCtGAA
  	TtTCTTTTTA
  301	GTTTTggcgG GAGTGTtggG CATGaatacG ctgATGCTGG
  	CAGTATGGtt
  351	gGCAACGTTG TTCCTGCGCG TGAAAGTGGG ACGGTTTTTC
  	AGCAGTCCGG
  401	CGACGTGGTT TCGGGGCAAA GGCCCTGTAA ATCAGGCGGT
  	GTTGCGGCTG
  451	TATGCGGACC AGTGGCGGCA ACCTTCGGTA CGATGGAAAA
  	TAGGCGCAAC
  501	GGCGCACAGC TTGTGGCTCT GCACGCTGCT CGGAATGCTG
  	GTGTCGGTAT
  551	TGCTGCTGCT TTTGGTGCGG CAATATACGT TCAACTGGGA
  	AAGCACGCTG
  601	TTGAGCAATG CCGCTTCGGT ACGCGCGGTG GAAATGTTGG
  	CATGGCTGCC
  651	GTCGAAACTC GGTTTCCCTG TCCCCGATGC GCGGGCGGTC
  	ATCGAAGGTC
  701	GTCTGAACGG CAATATTGCC GATGCGCGGG CTTGGTCGGG
  	GCTGCTGGTC
  751	GGCAGTATCG TCTGCTACGG CATCCTGCCG CGCCTCTTGG
  	CTTGGGTAGT
  801	GTGTAAAATC CTTTTGAAAA CAAGCGAAAA CGGattgGAT
  	TTGGAAAAAA
  851	CCTATTATCA GGCGGTCATC CGCCGCTGGC AGAACAAAAT
  	CACCGATGCG
  901	GATACGCGTC GGGAAACCGT GTCCGCCGTT TCGCcgaAAA
  	TCGTCTTGAA
  951	CGATGCGCCG AAATGGGCGC TCATGCTGGA GACCGAGTGG
  	CAGGACGGCC
  1001	AATGGTTCGA GGGCAGGCTG GCGCAGGAAT GGCTGGATAA
  	GGGCGTTGCC
  1051	GCCAATCGGG AACAGGTTGC CGCGCTGGAG ACAGAGCTGA
  	AGCAGAAACC
  1101	GGCGCAACTG CTTATCGGCG TACGCGCCCA AACTGTGCCG
  	GACCGGGGCG
  1151	TGCTGCGGCA GATTGTGCGG CTTTCGGAAG CGGCGCAGGG
  	CGGCGCGGTG
  1201	GTGCAGCTTT TGGCGGAACA GGGGCTTTCA GACGACCTTT
  	CGGAAAAGCT
  1251	GGAACATTGG CGTAACGCGC TGACCGAATG CGGCGCGGCG
  	TGGCTTGAGC
  1301	CTGACAGGGT GGCGCAGGAA GGCCGTTTGA AAGACCAATA
  	A

• This encodes a protein having amino acid sequence <SEQ ID 206; ORF33ng-1>:

• 1	MLNPSRKLVE LVRILNKGGF IFSGDPVQAT EALRRVDGST
  	EEKIFRRAEM
  51	IDRDRMLRDT LERVRAGSFW LWVVVASMMF TAGFS GTYLL
  	MDNQGLNFFL
  101	VLAGVLGMNT LMLAVWLATL FLRVKVGRFF SSPATWFRGK
  	GPVNQAVLRL
  151	YADQWRQPSV RWKIGATAHS LWLCTLLGML VSVLLLLLVR
  	QYTFNWESTL
  201	LSNAASVRAV EMLAWLPSKL GFPVPDARAV IEGRLNGNIA
  	DARAWSGLLV
  251	GSIVCYGILP RLLAWVVCKI LLKTSENGLD LEKTYYQAVI
  	RRWQNKITDA
  301	DTRRETVSAV SPKIVLNDAP KWALMLETEW QDGQWFEGRL
  	AQEWLDKGVA
  351	ANREQVAALE TELKQKPAQL LIGVRAQTVP DRGVLRQIVR
  	LSEAAQGGAV
  401	VQLLAEQGLS DDLSEKLEHW RNALTECGAA WLEPDRVAQE
  	GRLKDQ*

• ORF33ng-1 and ORF33-1 show 94.6% identity in 446 aa overlap:
• 
• Based on the presence of several putative transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 25
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 207>:

• 1	..CAGAAGAGTT TGTCGAGAAT TTCTTTATGG GGTTTGGGCG
  	GCGTGTTTTT
  51	CGGGGTGTCC GGTCTGGTAT GGTTTTCTTT GGGCGTTTCT
  	TT.GAGTGCG
  101	CCTGTTTTTC GGGTGTTTCT TTTCGGGGTT CGGGACGGGG
  	GACGTTTGTG
  151	GGCAGTACGG GGGTTTCTTT GAGTGTGTTT TCAGCTTGTG
  	TTCC.GGCGT
  201	CGTCCGGCTG CCTGTCGGTT TGAGCTGTGT CGGCAGGTTG
  	CG..GTTTGA
  251	CCCGGTTTTT CTTGGGTGCG GCAGGGGACG TCATTCTCCT
  	GCCGCTTTCG
  301	TCTGTGCCGT CCGGCTGTGC GGGTTCGGAT GAGGCGGCGT
  	GGTGGTGTTC
  351	GGGTTGGGCG GCATCTTGTT CCGACTACGC CGTTTGGCAG
  	CCAGAATTCG
  401	GTTTCGCGGG GGCTGTCGGT GTGTTGCGGT TCGGCTTGAA
  	GGGTTTTGTC
  451	GTCC..

• This corresponds to the amino acid sequence <SEQ ID 208; ORF34>:

• 1	..QKSLSRISLW GLGGVFFGVS GLVWFSLGVS XECACFSGVS
  	FRGSGRGTFV
  51	GSTGVSLSVF SACVXGVVRL PVGLSCVGRL XXLTRFFLGA
  	AGDVILLPLS
  101	SVPSGCAGSD EAAWWCSGWA ASCPTTPFGS QNSVSRGLSV
  	CCGSA*RVLS
  151	S..

• Further work revealed the complete nucleotide sequence <SEQ ID 209>:

• 1	ATGATGATGC CGTTCATAAT GCTTCCTTGG ATTGCkGGTG
  	TGCCTGCCGT
  51	GCCGGGTCAG AATAGGTTGT CCAGAATTTC TTTATGGGGT
  	TTGGGCGGCG
  101	TGTTTTTCGG GGTGTCCGGT TTGGTATGGT TTTCTTTGGG
  	CGTTTCTTTG
  151	GGCTGCGCCT GTTTTTCGGG TGTTTCTTTT CGGGGTTCGG
  	GACGGGGGAC
  201	GTTTGTGGGC AGTACGGGGG TTTCTTTGAG TGTGTTTTCA
  	GCTTGTGTTC
  251	CGGCGTCGTC CGGCTGCCTG TCGGTTTGAG CTGTGTCGGC
  	AGGTTGCGGT
  301	TTGACCCGGT TTTTCTTGGG TGCGGCAGGG GACGGCAGTC
  	CGCTGCCGCT
  351	TTCGTCTGTG CCGTCCGGCT GTGCGGGTTC GGATGAGGCG
  	GCGTGGTGGT
  401	GTTCGGGTTG GGCGGCATCT TGTCCGACTA CGCCGTTTGG
  	CAGCCAGAAT
  451	TCGGTTTCGC GGGGGCTGTC GGTGTGTTGC GGTTCGGCTT
  	GAAGGGTTTT
  501	GTCGCCGTTC GGGTTGAATG TGCTGACGAT GCCTATTGCC
  	AATGCGCCGA
  551	TGGCGGCGAT ACAGATGAGC AATACGGCGC GTATCAGGAG
  	TTTGGGGGTC
  601	AGCCTGAAGG GTTTGTTCGG TTTTTTTGCC ATTTTGATTG
  	TGCTTTTGGG
  651	GTGTCGGGCA ATGCCGTCTG AAGGCGGTTC AGACGGCATT
  	GCCGAGTCAG
  701	CGTTGGACGT AGTTTTGGTA GAGGGTGATG ACTTTTTGTA
  	CGCCGACGGT
  751	GGTGCTGACT TTTTGGGTAA TCTGCGCCTG TTCTTCGGGG
  	GTGAGGATGC
  801	CCATAACGTA GGTTACGTTG CCGTAGGTAA CGATTTTGAC
  	GCGCGCCTGT
  851	GTGGCGGGGC TGATGCCCAA CAGCGTGGCG CGGACTTTGG
  	ATGTGTTCCA
  901	AGTGTCGCCG GCGATGTCGC CGGCAGTGCG CGGCAGGGAG
  	GCGACGGTAA
  951	TATAGTTGTA CACGCCTTCG GCGGCCTGTT CGGAACGTGC
  	AATCTGACCG
  1001	ACGAACTGTT TTTCGCCTTC GGTGGCGACT TGTCCGAGCA
  	GCAGCAGGTG
  1051	GCGGTTGTAG CCGACGACGG AGATTTGGGG CGTGTAGCCT
  	TTGGTTTGGT
  1101	TGTTTTGGCG CAGATAGGAA CGGGCGGTGG TTTCGATACG
  	CAACGCCATA
  1151	ACGTTGTCGT CGGTTTGCGC GCCGGTGGTT CGGCGGTCGA
  	CGGCGGATTT
  1201	CGCGCCGACG GCGGCGCTTC CGATTACTGC GCTGACGCAG
  	CCGCTAAGGG
  1251	CAAGGCTGAA AATGGCGGCA ATCAGGGTGC GGACGGTGTG
  	CGGTTTGGGT
  1301	TTCATCGGGT GCTTCCTTTC TTGGGCGTTT CAGACGGCAT
  	TGCTTTGCGC
  1351	CATGCCGTCT GA

• This corresponds to the amino acid sequence <SEQ ID 210; ORF34-1>:

• 1	MMMPFIMLPW IAGVPAVPGQ NRLSRISLWG LGGVFFGVSG
  	LVWFSLGVSL
  51	GCACFSGVSF RGSGRGTFVG STGVSLSVFS ACVPASSGCL
  	SV*AVSAGCG
  101	LTRFFLGAAG DGSPLPLSSV PSGCAGSDEA AWWCSGWAAS
  	CPTTPFGSQN
  151	SVSRGLSVCC GSA*RVLSPF GLNVLTMPIA NAPMAAIQMS
  	NTARIRSLGV
  201	SLKGLFGFFA ILIVLLGCRA MPSEGGSDGI AESALDVVLV
  	EGDDFLYADG
  251	GADFLGNLRL FFGGEDAHNV GYVAVGNDFD ARLCGGADAQ
  	QRGADFGCVP
  301	SVAGDVAGSA RQGGDGNIVV HAFGGLFGTC NLTDELFFAF
  	GGDLSEQQQV
  351	AVVADDGDLG RVAFGLVVLA QIGTGGGFDT QRHNVVVGLR
  	AGGSAVDGGF
  401	RADGGASDYC ADAAAKGKAE NGGNQGADGV RFGFHRVLPF
  	LGVSDGIALR
  451	HAV*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF34 shows 73.3% identity over a 161aa overlap with an ORF (ORF34a) from strain A of N. meningitidis:
• 
• The complete length ORF34a nucleotide sequence <SEQ ID 211> is:

• 1	ATGATGATNC CGTTNATAAT GCTTCCTTGG ATTGCGGGTG
  	TGCCTGCCGT
  51	GCCGGGTCAG AAGAGGTTGT CGAGAANTTC TTTATGGGGT
  	TTAGGCGGCN
  101	TGTTTTTCGG GGTGTCCGGT TTGGTATGGT TTTCTTTGGG
  	CGTTTCTNTT
  151	TCTTTGGGTG TTTCTNTGGG CTGTGCCTGT TTTTCGGGTG
  	TTTCTTTTCG
  201	GGGTTCGGGA CGGGGGACGT TTGTGGGCAG TACNGGGGTT
  	TCTTTGAGTG
  251	TGTTTTCAGC TTGTGCTCCG GCGTCGTCCG GCTGCCTGTC
  	GGTTTNAGCT
  301	GTGTCGGCAG GTTGCGGTTT GACCCGGNTT TTCTTNGGTG
  	CGGCAGGGGA
  351	CGGCAGTCCG CTGCCGCTTT CGTCTGTGCC GTCCGGCTGT
  	GCGGGTGCGG
  401	ATGAGGAGGC GTNGTNGTGT TCGGGTTGGG CGGCATCTTG
  	TCCGACTACG
  451	CCGTTTGGCA GCCAGAATTC GGTTTCGCGG GGGCTGTCGG
  	TGTGTTGCGG
  501	TTCGGTNTGG AGGGTTTTGT CNCCGTTCGG GTNGAATGTG
  	CTGACGATGC
  551	CTATTGCCAA TGCGCCGATG GCGGTGATAC AGATGAGCAA
  	TACGGCGCGT
  601	ATCAGGAGTT TGGGGGTCAG CCTGAAGGGT TTGTTCNGTT
  	TTTTTGCCAT
  651	TTTGATTGTG CTTTTGGGGT GTCGGGCAAT GCCGTCTGAA
  	GGCGGTTCAG
  701	ACGGCATTGC CGAGTCAGCG TTGGACGTAG TTTNGGTAGA
  	GGGTGATGAC
  751	TTTTTGTACG CCGACGGTGG TGCTGACTTT TTGGGTAATC
  	TGCGCCTGTT
  801	CTTCGGGGGT GAGGATGCCC ATAACGTAGG TTACGTTGCC
  	GTAGGTAACG
  851	ATTTTGACGC GCGCCTGTGT GGCGGGGCTG ATGCCCAACA
  	GCGTGGCGCG
  901	GACTTTGGAT GTGTTCCAAG TGTCGCCGGC GATGTCGCCG
  	GCAGTGCGCG
  951	GCAGGGAGGC GACGGTAATG TANTTGTACA CGCCTTCGGC
  	GGCCTGTTCG
  1001	GAACGTGCAA TCTGACCGAC GAACTGTTTC TCGCCTTCGG
  	TGGCGACTTG
  1051	TCCGAGCAGC AGCAGGTGGC GGTTGTAGCC GACAACGGAG
  	ATTTGGGGCG
  1101	TGTANCCTTT GGTTTGGTTG TTTTGGCGCA GATAGGAGCG
  	GGCGGTGGTT
  1151	TCGATACGCA GCGCCATTAC GTTGTCGTCG GTTNGCGCGC
  	CGGTGGTTCG
  1201	GCGGTCGACG GCGGATTTCG CGCCGACCGC CGCGCCGCCG
  	ACGACTGCGC
  1251	TGACGCAGCC GCCGAGGGCA AGGCTGAGGA CGGCGGCAGT
  	CAGGGTGCGG
  1301	ACGGTGTGCG GTTTGGGTTT CATCGGGTGC TTCCTTTCTT
  	GGGCGTTTCA
  1351	GACGGCATTG CTTTGCGCCA TGCCGTCTGA

• This encodes a protein having amino acid sequence <SEQ ID 212>:

• 1	MMXPXIMLPW IAGVPAVPGQ KRLSRXSLWG LGGXFFGVSG
  	LVWFSLGVSX
  51	SLGVSXGCAC FSGVSFRGSG RGTFVGSTGV SLSVFSACAP
  	ASSGCLSVXA
  101	VSAGCGLTRX FXGAAGDGSP LPLSSVPSGC AGADEEAXXC
  	SGWAASCPTT
  151	PFGSQNSVSR GLSVCCGSVW RVLSPFGXNV LTMPIANAPM
  	AVIQMSNTAR
  201	IRSLGVSLKG LFXFFAILIV LL GCRAMPSE GGSDGIAESA
  	LDVVXVEGDD
  251	FLYADGGADF LGNLRLFFGG EDAHNVGYVA VGNDFDARLC
  	GGADAQQRGA
  301	DFGCVPSVAG DVAGSARQGG DGNVXVHAFG GLFGTCNLTD
  	ELFLAFGGDL
  351	SEQQQVAVVA DNGDLGRVXF GLVVLAQIGA GGGFDTQRHY
  	VVVGXRAGGS
  401	AVDGGFRADR RAADDCADAA AEGKAEDGGS QGADGVRFGF
  	HRVLPFLGVS
  451	DGIALRHAV*

• ORF34a and ORF34-1 show 91.3% identity in 459 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF34 shows 77.6% identity over a 161 as overlap with a predicted ORF (ORF34.ng) from N. gonorrhoeae:
• 
• The complete length ORF34ng nucleotide sequence <SEQ ID 213> is:

• 1	ATGATGATGC CGTTCATAAT GCTTCCTTGG ATTGCGGGTG
  	TGCCTGCCGT
  51	GCCGGGTCAA AAGAGGTTGT CGAGAATCTC TTTATGGGGT
  	TTGGCCGGCG
  101	TGTTTTTCGG GGTGTCCGGT TTGGTATGGT TTTCTTTGGG
  	CGTTTCTTTT
  151	TCTTTGGGTG TTTCTTTGGG CTGCGCCTGT TTTTCGGGTG
  	TTTCTTTTCG
  201	GGGTTCGGGA TGGGGGGCGT TTGTGGGCAG TACGGGGGTT
  	TCTTTGAGTG
  251	TGTTTTCAGC TTGTGTTCCG GTGCCGGTTA ACGAATCGGC
  	TGCCCGGGCC
  301	GCATCCGAAG GGCGCGGTTT gACCCGGTTT TTCTTGGGTG
  	CGGCAGGGGA
  351	CGGCAGTCCG CTGCCGCTTT CTTCTGTGCC GTCCGGCTGT
  	GCGGGTTCGG
  401	ATGAGGCGGC GTGGTGGTGT TCGGGTTGGG CGGCATCTTG
  	TCCGACGGCG
  451	CCGTTTGGCA GCCAGAATTC GGTTTCGCGG GGGCTGTCGG
  	TGTGTTGCGG
  501	TTCGGTTTGG AGGGTTTTGT CGCCGTTCGG GTTGAATGTG
  	CTGACGATGC
  551	CTACTGCCAA TGCGCCGATG GCGGTGATAC AGATGAGCAA
  	TACGGCGCGT
  601	ATCAGGAGTT TGGGGGTCAG CCTGAAGGGT TTGTTCGGTT
  	TTTTTGCCAT
  651	TTTGATTGTG CTTTTGGGGT GTCGGGCAAT GCCGTCTGAA
  	GGCGGTTCAG
  701	ACGGCATTGC CGAGTCAGCG TTGGACGTAG TTTTGGTAGA
  	GGGTAATGAC
  751	TTTTTGTACG CCGAcggTGG TGCTGACTTT TTGGGTAATC
  	TGCGCCTGTT
  801	CTTCGGGGGT GAGGATGCCC ATAACGTAGG TTACATTGCC
  	GTAGGTAATG
  851	ATTTTGACGC GCGCCTGTGT AGCGGGGCTG ATGCCCAGCA
  	GcgtgGCGCG
  901	GACTTTGGAC GTGTTCCAAG TGTCGCCGGC GATGTCGCCC
  	GCAGTGCGCG
  951	GCAGGGAGGC GACGGTAATG TAGTTGTATA CGCCTTCGGC
  	GGCCTGTTCG
  1001	GAACGTGCAA TCTGACCGAC GAACTGTTTT TCGCCTTCGG
  	TGGCGACTTG
  1051	TCCGAGCAGC AGCAGGTGGC GGTTGTAGCC GACGACGGAG
  	ATTTGGGGCG
  1101	TGTAGCCTTT GGTTTGGTTG TTTTGGCGCA GGTAGGAACG
  	GGCGGTGGTT
  1151	TCGATACGCA ACGCCATAAC GTtgtCATCG GTTtgcgcgc
  	CGGTGGTTcg
  1201	gCGGTCGATG ACGGATTTTG CGCCGACGGC GGCCCCGCCG
  	ACGACTGCGC
  1251	TGAAGCAGCC GCCGAGGGCA AGGCTGAGGA CGGCGGCAAT
  	CAGGGTGCGG
  1301	ACGGTGTGTG GTTTGGGTTT CATCGGGGAC TTCCTTTCTT
  	GGGCGTTTCA
  1351	GACGGCATTG CTTTGCGCCA TGCCGTCTGA

• This encodes a protein having amino acid sequence <SEQ ID 214>:

• 1	MMMPFIMLPW IAGVPAVPGQ KRLSRISLWG LAGVFFGVSG
  	LVWFSLGVSF
  51	SLGVSLGCAC FSGVSFRGSG WGAFVGSTGV SLSVFSACVP
  	VPVNESAARA
  101	ASEGRGLTRF FLGAAGDGSP LPLSSVPSGC AGSDEAAWWC
  	SGWAASCPTA
  151	PFGSQNSVSR GLSVCCGSVW RVLSPFGLNV LTMPTANAPM
  	AVIQMSNTAR
  201	IRSLGVSLKG LFGFFAILIV LL GCRAMPSE GGSDGIAESA
  	LDVVLVEGND
  251	FLYADGGADF LGNLRLFFGG EDAHNVGYIA VGNDFDARLC
  	SGADAQQRGA
  301	DFGRVPSVAG DVARSARQGG DGNVVVYAFG GLFGTCNLTD
  	ELFFAFGGDL
  351	SEQQQVAVVA DDGDLGRVAF GLVVLAQVGT GGGFDTQRHN
  	VVIGLRAGGS
  401	AVDDGFCADG GPADDCAEAA AEGKAEDGGN QGADGVWFGF
  	HRGLPFLGVS
  451	DGIALRHAV*

• ORF34ng and ORF34-1 show 90.0% identity in 459 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence (double-underlined) and several putative transmembrane domains (single-underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 26
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 215>:

• 1	ATGAAAACCT TCTTCAAAAC CCTTTCCGCC GCCGCACTCG
  	CGCTCATCCT
  51	CGCCGCCTGC GGATT.CAAA AAGACAGCGC GCCCGCCGCA
  	TCCGCTTCTG
  101	CCGCCGCCGA CAACGGCGCG GCGTAAAAAA GAAATCGTCT
  	TCGGCACGAC
  151	CGTCGGCGAC TTCGGCGATA TGGTCAAAGA ACAAATCCAA
  	GCCGAGCTGG
  201	AGAAAAAAGG CTACACCGTC AAACTGGTCG AGTTTACCGA
  	CTATGTACGC
  251	CCGAATCTGG CATTGGCTGA GGGCGAGTTG

• This corresponds to the amino acid sequence <SEQ ID 216; ORF4>:

• 1	MKTFFKTLSA AALALILAAC G.QKDSAPAA SASAAADNGA
  	AKKEIVFGTT
  51	VGDFGDMVKE QIQAELEKKG YTVKLVEFTD YVRPNLALAE
  	GEL

• Further sequence analysis revealed the complete nucleotide sequence <SEQ ID 217>:

• 1	ATGAAAACCT TCTTCAAAAC CCTTTCCGCC GCCGCACTCG
  	CGCTCATCCT
  51	CGCCGCCTGC GGCGGTCAAA AAGACAGCGC GCCCGCCGCA
  	TCCGCTTCTG
  101	CCGCCGCCGA CAACGGCGCG GCGAAAAAAG AAATCGTCTT
  	CGGCACGACC
  151	GTCGGCGACT TCGGCGATAT GGTCAAAGAA CAAATCCAAG
  	CCGAGCTGGA
  201	GAAAAAAGGC TACACCGTCA AACTGGTCGA GTTTACCGAC
  	TATGTACGCC
  251	CGAATCTGGC ATTGGCTGAG GGCGAGTTGG ACATCAACGT
  	CTTCCAACAC
  301	AAACCCTATC TTGACGACTT CAAAAAAGAA CACAATCTGG
  	ACATCACCGA
  351	AGTCTTCCAA GTGCCGACCG CGCCTTTGGG ACTGTACCCG
  	GGCAAGCTGA
  401	AATCGCTGGA AGAAGTCAAA GACGGCAGCA CCGTATCCGC
  	GCCCAACGAC
  451	CCGTCCAACT TCGCCCGCGT CTTGGTGATG CTCGACGAAC
  	TGGGTTGGAT
  501	CAAACTCAAA GACGGCATCA ATCCGTTGAC CGCATCCAAA
  	GCGGACATCG
  551	CCGAGAACCT GAAAAACATC AAAATCGTCG AGCTTGAAGC
  	CGCGCAACTG
  601	CCGCGTAGCC GCGCCGACGT GGATTTTGCC GTCGTCAACG
  	GCAACTACGC
  651	CATAAGCAGC GGCATGAAGC TGACCGAAGC CCTGTTCCAA
  	GAACCGAGCT
  701	TTGCCTATGT CAACTGGTCT GCCGTCAAAA CCGCCGACAA
  	AGACAGCCAA
  751	TGGCTTAAAG ACGTAACCGA GGCCTATAAC TCCGACGCGT
  	TCAAAGCCTA
  801	CGCGCACAAA CGCTTCGAGG GCTACAAATC CCCTGCCGCA
  	TGGAATGAAG
  851	GCGCAGCCAA ATAA

• This corresponds to the amino acid sequence <SEQ ID 218; ORF4-1>:

• 1	MKTFFKTLSA AALALILAAC GGQKDSAPAA SASAAADNGA
  	AKKEIVFGTT
  51	VGDFGDMVKE QIQAELEKKG YTVKLVEFTD YVRPNLALAE
  	GELDINVFQH
  101	KPYLDDFKKE HNLDITEVFQ VPTAPLGLYP GKLKSLEEVK
  	DGSTVSAPND
  151	PSNFARVLVM LDELGWIKLK DGINPLTASK ADIAENLKNI
  	KIVELEAAQL
  201	PRSRADVDFA VVNGNYAISS GMKLTEALFQ EPSFAYVNWS
  	AVKTADKDSQ
  251	WLKDVTEAYN SDAFKAYAHK RFEGYKSPAA WNEGAAK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF4 shows 93.5% identity over a 93aa overlap with an ORF (ORF4a) from strain A of N. meningitidis:
• 
• The complete length ORF4a nucleotide sequence <SEQ ID 219> is:

• 1	ATGAAAACCT TCTTCAAAAC CCTTTCCGCC GCCGCACTCG
  	CGCTCATCCT
  51	CGCCGCCTGC GGCGGTCAAA AAGATAGCGC GCCCGCCGCA
  	TCCGCTTCTG
  101	CCGCCGCCGA CAACGGCGCG GCGAANAAAG AAATCGTCTT
  	CGGCACGACC
  151	GTCGGCGACT TCGGCGATAT GGTCAAAGAA CANATCCAAC
  	CCGAGCTGGA
  201	GAAAAAAGGC TACACCGTCA AACTGGTCGA GTNTACCGAC
  	TATGTGCGCN
  251	CGAATCTGGC ATTGGCTGAG GGCGAGTTGG ACATCAACGT
  	CTTNCAACAC
  301	ANACNCTATC TTGACGACTN CAAAAAANAA CACAATCTGG
  	ACATCACCNN
  351	AGTCTTNCAA GTGCCGACCG CGCCTTTGGG ACTGTACCCG
  	GGCAAGCTGA
  401	AATCGCTGGA NNAAGTCAAA GANGGCAGCA CCGTATCCGC
  	GCCCAACGAC
  451	CCGTNNNACT TCGNCCGCGT CTTGGTGATG CTCGACGAAC
  	TGGGTTNGAT
  501	CAAACTCAAA GACNGCATCA NNNNGNNGNN NNNANCNANA
  	NNNGANANNN
  551	NNNNANNNNT NNNNNNNNNN NNNNNCNNCG NNNNNNNANN
  	NNNNNNNNNN
  601	NCGNNTNNNN NNGCNNNNNT NNANNNTNNN NNCNNCNNNN
  	NNNNNTNNNN
  651	NANNANNAGC GGCATGAAGC TGACCGAAGC CCTGTTCCAA
  	GAACCGAGCT
  701	TTGCCTATGT CAACTGGTCT GCCGTCAAAA CCGCCGACAA
  	AGACAGCCAA
  751	TGGCTTAAAG ACGTAACCGA GGCCTATAAC TCCGACGCGT
  	TCAAAGCCTA
  801	CGCGCACAAA CGCTTCGAGG GCTACAAATC CCCTGCCGCA
  	TGGAATGAAG
  851	GCGCAGCCAA ATAA

• This is predicted to encode a protein having amino acid sequence <SEQ ID 220>:

• 1	MKTFFKTLSA AALALILAAC GGQKDSAPAA SASAAADNGA
  	AXKEIVFGTT
  51	VGDFGDMVKE XIQPELEKKG YTVKLVEXTD YVRXNLALAE
  	GELDINVXQH
  101	XXYLDDXKKX HNLDITXVXQ VPTAPLGLYP GKLKSLXXVK
  	XGSTVSAPND
  151	PXXFXRVLVM LDELGXIKLK DXIXXXXXXX XXXXXXXXXX
  	XXXXXXXXXX
  201	XXXXAXXXXX XXXXXXXXXS GMKLTEALFQ EPSFAYVNWS
  	AVKTADKDSQ
  251	WLKDVTEAYN SDAFKAYAHK RFEGYKSPAA WNEGAAK*

• A leader peptide is underlined.
• Further analysis of these strain A sequences revealed the complete DNA sequence <SEQ ID 221>:

• 1	ATGAAAACCT TCTTCAAAAC CCTTTCCGCC GCCGCACTCG
  	CGCTCATCCT
  51	CGCCGCCTGC GGCGGTCAAA AAGATAGCGC GCCCGCCGCA
  	TCCGCTTCTG
  101	CCGCCGCCGA CAACGGCGCG GCGAAAAAAG AAATCGTCTT
  	CGGCACGACC
  151	GTCGGCGACT TCGGCGATAT GGTCAAAGAA CAAATCCAAC
  	CCGAGCTGGA
  201	GAAAAAAGGC TACACCGTCA AACTGGTCGA GTTTACCGAC
  	TATGTGCGCC
  251	CGAATCTGGC ATTGGCTGAG GGCGAGTTGG ACATCAACGT
  	CTTCCAACAC
  301	AAACCCTATC TTGACGACTT CAAAAAAGAA CACAATCTGG
  	ACATCACCGA
  351	AGTCTTCCAA GTGCCGACCG CGCCTTTGGG ACTGTACCCG
  	GGCAAGCTGA
  401	AATCGCTGGA AGAAGTCAAA GACGGCAGCA CCGTATCCGC
  	GCCCAACGAC
  451	CCGTCCAACT TCGCCCGCGT CTTGGTGATG CTCGACGAAC
  	TGGGTTGGAT
  501	CAAACTCAAA GACGGCATCA ATCCGCTGAC CGCATCCAAA
  	GCGGACATTG
  551	CCGAAAACCT GAAAAACATC AAAATCGTCG AGCTTGAAGC
  	CGCGCAACTG
  601	CCGCGTAGCC GCGCCGACGT GGATTTTGCC GTCGTCAACG
  	GCAACTACGC
  651	CATAAGCAGC GGCATGAAGC TGACCGAAGC CCTGTTCCAA
  	GAACCGAGCT
  701	TTGCCTATGT CAACTGGTCT GCCGTCAAAA CCGCCGACAA
  	AGACAGCCAA
  751	TGGCTTAAAG ACGTAACCGA GGCCTATAAC TCCGACGCGT
  	TCAAAGCCTA
  801	CGCGCACAAA CGCTTCGAGG GCTACAAATC CCCTGCCGCA
  	TGGAATGAAG
  851	GCGCAGCCAA ATAA

• This encodes a protein having amino acid sequence <SEQ ID 222; ORF4a-1>:

• 1	MKTFFKTLSA AALALILAAC GGQKDSAPAA SASAAADNGA
  	AKKEIVFGTT
  51	VGDFGDMVKE QIQPELEKKG YTVKLVEFTD YVRPNLALAE
  	GELDINVFQH
  101	KPYLDDFKKE HNLDITEVFQ VPTAPLGLYP GKLKSLEEVK
  	DGSTVSAPND
  151	PSNFARVLVM LDELGWIKLK DGINPLTASK ADIAENLKNI
  	KIVELEAAQL
  201	PRSRADVDFA VVNGNYAISS GMKLTEALFQ EPSFAYVNWS
  	AVKTADKDSQ
  251	WLKDVTEAYN SDAFKAYAHK RFEGYKSPAA WNEGAAK*

• ORF4a-1 and ORF4-1 show 99.7% identity in 287 aa overlap:
• 
• Homology with an Outer Membrane Protein of Pasteurella haemolitica (Accession q08869).
• ORF4 and this outer membrane protein show 33% aa identity in 91aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF4 shows 93.6% identity over a 94aa overlap with a predicted ORF (ORF4.ng) from N. gonorrhoeae:
• 
• The complete length ORF4ng nucleotide sequence <SEQ ID 223> was predicted to encode a protein having amino acid sequence <SEQ ID 224>:

• 1	MKTFFKTLST ASLALILAAC GGQKDSAPAA SAAAPSADNG
  	AAKKEIVFGT
  51	TVGDFGDMVK EQIQAELEKK GYTVKLVEFT DYVRPNLALA
  	EGELDINVFQ
  101	HKPYLDDFKK EHNLDITEAF QVPTAPLGLY PGKLKSLEEV
  	KDGSTVSAPN
  151	DPSNFARALV MLNELGWIKL KDGINPLTAS KADIAENLKN
  	IKIVELEAAQ
  201	LPRSRADVDF AVVNGNYAIS SGMKLTEALF QEPSFAYVNW
  	SAVKTADKDS
  251	QWLKDVTEAY NSDAFKAYAH KRFEGYKYPA AWNEGAAK*

• Further analysis revealed the complete length ORF4ng DNA sequence <SEQ ID 225> to be:

• 1	atgAAAACCT TCTTCAAAAC cctttccgcc gccgcaCTCG
  	CGCTCATCCT
  51	CGCAGCCTGc ggCggtcaAA AAGACAGCGC GCCCgcagcc
  	tctgcCGCCG
  101	CCCCTTCTGC CGATAACGgc gCgGCGAAAA AAGAAAtcgt
  	ctTCGGCACG
  151	Accgtgggcg acttcggcgA TAtggTCAAA GAACAAATCC
  	AagcCGAgct
  201	gGAGAAAAAA GgctACACcg tcAAattggt cgaatttacc
  	gactatgtGC
  251	gCCCGAATCT GGCATTGGCG GAGGGCGAGT TGGACATCAA
  	CGTCTTCCAA
  301	CACAAACCCT ATCTTGACGA TTTCAAAAAA GAACACAACC
  	TGGACATCAC
  351	CGAAGCCTTC CAAGTGCCGA CCGCGCCTTT GGGACTGTAT
  	CCGGGCAAAC
  401	TGAAATCGCT GGAAGAAGTC AAAGACGGCA GCACCGTATC
  	CGCGCCCAac
  451	gACccgTCCA ACTTCGCACG CGCCTTGGTG ATGCTGAACG
  	AACTGGGTTG
  501	GATCAAACTC AAAGACGGCA TCAATCCGCT GACCGCATCC
  	AAAGCCGACA
  551	TCGCGGAAAA CCTGAAAAAC ATCAAAATCG TCGAGCTTGA
  	AGCCGCACAA
  601	CTGCCGCGCA GCCGCGCCGA CGTGGATTTT GCCGTCGTCA
  	ACGGCAACTA
  651	CGCCATAAGC AGCGGCATGA AGCTGACCGA AGCCCTGTTC
  	CAAGAGCCGA
  701	GCTTTGCCTA TGTCAACTGG TCTGCCgtcA AAACCGCCGA
  	CAAAGACAGC
  751	CAATGGCTTA AAGACGTAAC CGAGGCCTAT AACTCCGACG
  	CGTTCAAAGC
  801	CTACGCGCAC AAACGCTTCG AGGGCTACAA ATACCCTGCC
  	GCATGGAATG
  851	AAGGCGCAGC CAAATAA

• This encodes a protein having amino acid sequence <SEQ ID 226; ORF4ng-1>:

• 1	MKTFFKTLSA AALALILAAC GGQKDSAPAA SAAAPSADNG
  	AAKKEIVFGT
  51	TVGDFGDMVK EQIQAELEKK GYTVKLVEFT DYVRPNLALA
  	EGELDINVFQ
  101	HKPYLDDFKK EHNLDITEAF QVPTAPLGLY PGKLKSLEEV
  	KDGSTVSAPN
  151	DPSNFARALV MLNELGWIKL KDGINPLTAS KADIAENLKN
  	IKIVELEAAQ
  201	LPRSRADVDF AVVNGNYAIS SGMKLTEALF QEPSFAYVNW
  	SAVKTADKDS
  251	QWLKDVTEAY NSDAFKAYAH KRFEGYKYPA AWNEGAAK*

• This shows 97.6% identity in 288 aa overlap with ORF4-1:
• 
• In addition, ORF4ng-1 shows significant homology with an outer membrane protein from the database:
• 
• Based on this analysis, including the homology with the outer membrane protein of Pasteurella haemolitica, and on the presence of a putative prokaryotic membrane lipoprotein lipid attachment site in the gonococcal protein, it was predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF4-1 (30 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIGS. 8A and 8B show, respectively, the results of affinity purification of the His-fusion and GST-fusion proteins. Purified His-fusion protein was used to immunise mice, whose sera were used for ELISA (positive result), Western blot (FIG. 8C), FACS analysis (FIG. 8D), and a bactericidal assay (FIG. 8E). These experiments confirm that ORF4-1 is a surface-exposed protein, and that it is a useful immunogen.
• FIG. 8F shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF4-1.
Example 27
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 227>:

• 1	CCTCGTCGTC CTCGGCATGC TCCAGTTTCA AGGGGCGATT
  	TACTCCAAGG
  51	CGGTGGAACG TATGCTCGGC ACGGTCATCG GGCTGGGCGC
  	GGGTTTGGGC
  101	GTTTTATGGC TGAACCAGCA TTATTTCCAC GGCAACCTCC
  	TCTTCTACCT
  151	CACCGTCGGC ACGGCAAGCG CACTGGCCGG CTGGGCGGCG
  	GTCGGCAAAA
  201	ACGGCTACGT CCCTmTGCTG GCAGGGCTGA CGATGTGTAT
  	GCTCATCGGC
  251	GACAACGGCA GCGAATGGCT CGACAGCGGA CTCATGCGCG
  	CCATGAACGT
  301	CCTCATCGGC GyGGCCATCG CCATCGCCGC CGCCAAACTG
  	CTGCCGCTGA
  351	AATCCACACT GATGTGGCGT TTCATGCTTG CCGACAACCT
  	GGCCGACTGC
  401	AGCAAAATGA TTGCCGAAAT CAGCAACGGC AGGCGCATGA
  	CCCGCGAACG
  451	CCTCGAGGAG AACATGGCGA AAATGCGCCA AATCAACGCA
  	CGCATGGTCA
  501	AAAGCCGCAG CCATCTCGCC GCCACATCGG GCGAAAGCTG
  	CATCAGCCCC
  551	GCCATGATGG AAGCCATGCA GCACGCCCAC CGTAAAATCG
  	TCAACACCAC
  601	CGAGCTGCTC CTGACCACCG CCGCCAAGCT GCAATCTCCC
  	AAACTCAACG
  651	GCAGCGAAAT CCGGCTGCTT GACCGCCACT TCACACTGCT
  	CCAAAC....
  701	............................. GC AGACACGCCC
  	GCCGCATCCG
  751	CATCGACACC GCCATCAACC CCGAACTGGA AGCCCTCGCC
  	GAACACCTCC
  801	ACTACCAATG GCAGGGCTTC CTCTGGCTCA GCACCGATAT
  	GCGTCAGGAA
  851	ATTTCCGCCC TCGTCATCCT GCTGCAACGC ACCCGCCGCA
  	AATGGCTGGA
  901	TGCCCACGAA CGCCAACACC TGCGCCAAAG CCTGCTTGA

• This corresponds to the amino acid sequence <SEQ ID 228; ORF8>:

• 1	......PRRP RHAPVSRGDL LQGGGTYARH GHRAGRGFGR
  	FMAEPALFPR
  51	QPPLLPHRRH GKRTGRLGGG RQKRLRPXAG RADDVYAHRR
  	QRQRMARQRT
  101	HARHERPHRR GHRHRRRQTA AAEIHTDVAF HACRQPGRLQ
  	QNDCRNQQRQ
  151	AHDPRTPRGE HGENAPNQRT HGQKPQPSRR HIGRKLHQPR
  	HDGSHAARPP
  201	XNRQHHRAAP DHRRQAAISQ TQRQRNPAAX PPLHTAPN..
  	.........Q
  251	TRPPHPHRHR HQPRTGSPRR TPPLPMAGLP LAQHRYASGN
  	FRPRHPAATH
  301	PPQMAGCPRT PTPAPKPA*

• Computer analysis of this amino acid sequence gave the following results:
Sequence Motifs
• ORF8 is proline-rich and has a distribution of proline residues consistent with a surface localization. Furthermore the presence of an RGD motif may indicate a possible role in bacterial adhesion events.
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF8 shows 86.5% identity over a 312aa overlap with a predicted ORF (ORF8.ng) from N. gonorrhoeae:
• 
• The complete length ORF8ng nucleotide sequence <SEQ ID 229> is predicted to encode a protein having amino acid sequence <SEQ ID 230>:

• 1	MDRDDRLRRP RHAPVPRRDL LQRGGTYARY GHRAGRGFGR
  	FMAEPALFPR
  51	QPPLLPDHRH GKRTGRLGGG RQKRLRPYVG GADDVHAHRR
  	QRQRMARQRP
  101	DARDERPHRR RHRHCRRQTA AAEIHTDVAF HACRQPGRLQ
  	QNDCRNQQRQ
  151	AYDARTFGAE YGQNAPNQRT HGQKPQPPRR HIGRKPHQPL
  	HDGSHAARPP
  201	QNRQHHRAAP DHRRQAAISQ TQRQRNPAAR PPLHTAPNRP
  	ATNRRPHQRQ
  251	TRPPHPHRHR HQPRTGSPRR TPPLPMAGFP LAQHQYASGN
  	FRPRHPPATH
  301	PPQMAGCPRT PTPAPKPA*

• Based on the sequence motifs in these proteins, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 28
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 231>:

• 1	..GAAATCAGCC TGCGGTCCGA CNACAGGCCG GTTTCCGTGN
  	CGAAGCGGCG
  51	GGATTCGGAA CGTTTTCTGC TGTTGGACGG CGGCAACAGC
  	CGGCTCAAGT
  101	GGGCGTGGGT GGAAAACGGC ACGTTCGCAA CCGTCGGTAG
  	CGCGCCGTAC
  151	CGCGATTTGT CGCCTTTGGG CGCGGAGTGG GCGGAAAAGG
  	CGGATGGAAA
  201	TGTCCGCATC GTCGGTTGCG CTGTGTGCGG AGAATTCAAA
  	AAGGCACAAG
  251	TGCAGGAACA GCTCGCCCGA AAAATCGAGT GGCTGCCGTC
  	TTCCGCACAG
  301	GCTTT.GGCA TACGCAACCA CTACCGCCAC CCCGAAGAAC
  	ACGGTTCCGA
  351	CCGCTGGTTC AACGCCTTGG GCAGCCGCCG CTTCAGCCGC
  	AACGCCTGCG
  401	TCGTCGTCAG TTGCGGCACG GCGGTAACGG TTGACGCGCT
  	CACCGATGAC
  451	GGACATTATC TCGGAGA.GG AACCATCATG CCCGGTTTCC
  	ACCTGATGAA
  501	AGAATCGCTC GCCGTCCGAA CCGCCAACCT CAACCGGCAC
  	GCCGGTAAGC
  551	GTTATCCTTT CCCGACCGG..

• This corresponds to the amino acid sequence <SEQ ID 232; ORF61>:

• 1	..EISLRSDXRP VSVXKRRDSE RFLLLDGGNS RLKWAWVENG
  	TFATVGSAPY
  51	RDLSPLGAEW AEKADGNVRI VGCAVCGEFK KAQVQEQLAR
  	KIEWLPSSAQ
  101	AXGIRNHYRH PEEHGSDRWF NALGSRRFSR NACVVVSCGT
  	AVTVDALTDD
  151	GHYLGXGTIM PGFHLMKESL AVRTANLNRH AGKRYPFPT..

• Further work revealed the complete nucleotide sequence <SEQ ID 233>:

• 1	ATGACGGTTT TGAAGCTTTC GCACTGGCGG GTGTTGGCGG
  	AGCTTGCCGA
  51	CGGTTTGCCG CAACACGTCT CGCAACTGGC GCGTATGGCG
  	GATATGAAGC
  101	CGCAGCAGCT CAACGGTTTT TGGCAGCAGA TGCCGGCGCA
  	CATACGCGGG
  151	CTGTTGCGCC AACACGACGG CTATTGGCGG CTGGTGCGCC
  	CATTGGCGGT
  201	TTTCGATGCC GAAGGTTTGC GCGAGCTGGG GGAAAGGTCG
  	GGTTTTCAGA
  251	CGGCATTGAA GCACGAGTGC GCGTCCAGCA ACGACGAGAT
  	ACTGGAATTG
  301	GCGCGGATTG CGCCGGACAA GGCGCACAAA ACCATATGCG
  	TGACCCACCT
  351	GCAAAGTAAG GGCAGGGGGC GGCAGGGGCG GAAGTGGTCG
  	CACCGTTTGG
  401	GCGAGTGTCT GATGTTCAGT TTTGGCTGGG TGTTTGACCG
  	GCCGCAGTAT
  451	GAGTTGGGTT CGCTGTCGCC TGTTGCGGCA GTGGCGTGTC
  	GGCGCGCCTT
  501	GTCGCGTTTA GGTTTGGATG TGCAGATTAA GTGGCCCAAT
  	GATTTGGTTG
  551	TCGGACGCGA CAAATTGGGC GGCATTCTGA TTGAAACGGT
  	CAGGACGGGC
  601	GGCAAAACGG TTGCCGTGGT CGGTATCGGC ATCAATTTTG
  	TCCTGCCCAA
  651	GGAAGTAGAA AATGCCGCTT CCGTGCAATC GCTGTTTCAG
  	ACGGCATCGC
  701	GGCGGGGCAA TGCCGATGCC GCCGTGCTGC TGGAAACGCT
  	GTTGGTGGAA
  751	CTGGACGCGG TGTTGTTGCA ATATGCGCGG GACGGATTTG
  	CGCCTTTTGT
  801	GGCGGAATAT CAGGCTGCCA ACCGCGACCA CGGCAAGGCG
  	GTATTGCTGT
  851	TGCGCGACGG CGAAACCGTG TTCGAAGGCA CGGTTAAAGG
  	CGTGGACGGA
  901	CAAGGCGTTT TGCACTTGGA AACGGCAGAG GGCAAACAGA
  	CGGTCGTCAG
  951	CGGCGAAATC AGCCTGCGGT CCGACGACAG GCCGGTTTCC
  	GTGCCGAAGC
  1001	GGCGGGATTC GGAACGTTTT CTGCTGTTGG ACGGCGGCAA
  	CAGCCGGCTC
  1051	AAGTGGGCGT GGGTGGAAAA CGGCACGTTC GCAACCGTCG
  	GTAGCGCGCC
  1101	GTACCGCGAT TTGTCGCCTT TGGGCGCGGA GTGGGCGGAA
  	AAGGCGGATG
  1151	GAAATGTCCG CATCGTCGGT TGCGCTGTGT GCGGAGAATT
  	CAAAAAGGCA
  1201	CAAGTGCAGG AACAGCTCGC CCGAAAAATC GAGTGGCTGC
  	CGTCTTCCGC
  1251	ACAGGCTTTG GGCATACGCA ACCACTACCG CCACCCCGAA
  	GAACACGGTT
  1301	CCGACCGCTG GTTCAACGCC TTGGGCAGCC GCCGCTTCAG
  	CCGCAACGCC
  1351	TGCGTCGTCG TCAGTTGCGG CACGGCGGTA ACGGTTGACG
  	CGCTCACCGA
  1401	TGACGGACAT TATCTCGGGG GAACCATCAT GCCCGGTTTC
  	CACCTGATGA
  1451	AAGAATCGCT CGCCGTCCGA ACCGCCAACC TCAACCGGCA
  	CGCCGGTAAG
  1501	CGTTATCCTT TCCCGACCAC AACGGGCAAT GCCGTCGCCA
  	GCGGCATGAT
  1551	GGATGCGGTT TGCGGCTCGG TTATGATGAT GCACGGGCGT
  	TTGAAAGAAA
  1601	AAACCGGGGC GGGCAAGCCT GTCGATGTCA TCATTACCGG
  	CGGCGGCGCG
  1651	GCAAAAGTTG CCGAAGCCCT GCCGCCTGCA TTTTTGGCGG
  	AAAATACCGT
  1701	GCGCGTGGCG GACAACCTCG TCATTTACGG GTTGTTGAAC
  	ATGATTGCCG
  1751	CCGAAGGCAG GGAATATGAA CATATTTAA

• This corresponds to the amino acid sequence <SEQ ID 234; ORF61-1>:

• 1	MTVLKLSHWR VLAELADGLP QHVSQLARMA DMKPQQLNGF
  	WQQMPAHIRG
  51	LLRQHDGYWR LVRPLAVFDA EGLRELGERS GFQTALKHEC
  	ASSNDEILEL
  101	ARIAPDKAHK TICVTHLQSK GRGRQGRKWS HRLGECLMFS
  	FGWVFDRPQY
  151	ELGSLSPVAA VACRRALSRL GLDVQIKWPN DLVVGRDKLG
  	GILIETVRTG
  201	GKTVAVVGIG INFVLPKEVE NAASVQSLFQ TASRRGNADA
  	AVLLETLLVE
  251	LDAVLLQYAR DGFAPFVAEY QAANRDHGKA VLLLRDGETV
  	FEGTVKGVDG
  301	QGVLHLETAE GKQTVVSGEI SLRSDDRPVS VPKRRDSERF
  	LLLDGGNSRL
  351	KWAWVENGTF ATVGSAPYRD LSPLGAEWAE KADGNVRIVG
  	CAVCGEFKKA
  401	QVQEQLARKI EWLPSSAQAL GIRNHYRHPE EHGSDRWFNA
  	LGSRRFSRNA
  451	CVVVSCGTAV TVDALTDDGH YLGGTIMPGF HLMKESLAVR
  	TANLNRHAGK
  501	RYPFPTTTGN AVASGMMDAV CGSVMMMHGR LKEKTGAGKP
  	VDVIITGGGA
  551	AKVAEALPPA FLAENTVRVA DNLVIYGLLN MIAAEGREYE
  	HI*

• FIG. 9 shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF61-1. Further computer analysis of this amino acid sequence gave the following results:
• Homology with the Baf Protein of B. pertussis (Accession Number U12020).
• ORF61 and baf protein show 33% aa identity in 166aa overlap:

• orf61  23 LLLDGGNSRLKWAWVE-NGTFATVGSAPYR----DLSPLGAEWAEKADGNVRIVGCAVCG  77
  +L+D GNSRLK  W + +   A    AP      DL  LG   A      R +G V   G
  baf     3 ILIDSGNSRLKVGWFDPDAPQAAREPAPVAFDNLDLDALGRWLATLPRRPQRALGVNVAG  62
  orf61  78 EFKKAQVQEQLAR---KIEWLPSSAQAXGIRNHYRHPEEHGSDRW---FNALGSRRFSRN 131
  +   +   L      I WL +   A G+RN YR+P++ G+DRW      L  +
  baf    63 LARGEAIAATLRAGGCDIRWLRAQPLAMGLRNGYRNPDQLGADRWACMVGVLARQPSVHP 122
  orf61 132 ACVVVSCGTAVTVDALTDDGHYLGXGTIMPGFHLMKESLAVRTANL               177
  +V S GTA T+D + D   + G G I+PG  +M+ +LA  TA+L
  baf   123 PLLVASFGTATTLDTIGPDNVFPG-GLILPGPAMMRGALAYGTAHL               167

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF61 shows 97.4% identity over a 189aa overlap with an ORF (ORF61a) from strain A of N. meningitidis:
• 
• The complete length ORF61a nucleotide sequence <SEQ ID 235> is:

• 1	ATGACGGTTT TGAAGCCTTC GCACTGGCGG GTGTTGGCGG
  	AGCTTGCCGA
  51	CGGTTTGCCG CAACACGTCT CGCAACTGGC GCGTATGGCG
  	GATATGAAGC
  101	CGCAGCAGCT CAACGGTTTT TGGCAGCAGA TGCCGGCGCA
  	CATACGCGGG
  151	CTGTTGCGCC AACACGACGG CTATTGGCGG CTGGTGCGCC
  	CATTGGCGGT
  201	TTTCGATGCC GAAGGTTTGC GCGAGCTGGG GGAAAGGTCG
  	GGTTTTCAGA
  251	CGGCATTGAA GCACGAGTGC GCGTCCAGCA ACGACGAGAT
  	ACTGGAATTG
  301	GCGCGGATTG CGCCGGACAA GGCGCACAAA ACCATATGTG
  	TGACCCACCT
  351	GCAAAGTAAG GGCAGGGGGC GGCAGGGGCG GAAGTGGTCG
  	CACCGTTTGG
  401	GCGAGTGTCT GATGTTCAGT TTTGGCTGGG TGTTTGACCG
  	GCCGCAGTAT
  451	GAGTTGGGTT CGCTGTCGCC TGTTGCGGCA GTGGCGTGCC
  	GGCGCGCCTT
  501	GTCGCGTTTG GGTTTGAAAA CGCAAATCAA GTGGCCAAAC
  	GATTTGGTCG
  551	TCGGACGCGA CAAATTGGGC GGCATTCTGA TTGAAACGGT
  	CAGGACGGGC
  601	GGCAAAACGG TTGCCGTGGT CGGTATCGGC ATCAATTTCG
  	TGCTGCCCAA
  651	GGAAGTGGAA AACGCCGCTT CCGTGCAATC GCTGTTTCAG
  	ACGGCATCGC
  701	GGCGGGGAAA TGCCGATGCC GCCGTGTTGC TGGAAACGCT
  	GTTGGCGGAA
  751	CTTGATGCGG TGTTGTTGCA ATATGCGCGG GACGGATTTG
  	CGCCTTTTGT
  801	GGCGGAATAT CAGGCTGCCA ACCGCGACCA CGGCAAGGCG
  	GTATTGCTGT
  851	TGCGCGACGG CGAAACCGTG TTCGAAGGCA CGGTTAAAGG
  	CGTGGACGGA
  901	CAAGGCGTTC TGCACTTGGA AACGGCAGAG GGCAAACAGA
  	CGGTCGTCAG
  951	CGGCGAAATC AGCCTGCGGT CCGACGACAG GCCGGTTTCC
  	GTGCCGAAGC
  1001	GGCGGGATTC GGAACGTTTT CTGCTGTTGG ACGGCGGCAA
  	CAGCCGGCTC
  1051	AAGTGGGCGT GGGTGGAAAA CGGCACGTTC GCAACCGTCG
  	GTAGCGCGCC
  1101	GTACCGCGAT TTGTCGCCTT TGGGCGCGGA GTGGGCGGAA
  	AAGGTGGATG
  1151	GAAATGTCCG CATCGTCGGT TGCGCCGTGT GCGGAGAATT
  	CAAAAAGGCA
  1201	CAAGTGCAGG AACAGCTCGC CCGAAAAATC GAGTGGCTGC
  	CGTCTTCCGC
  1251	ACAGGCTTTG GGCATACGCA ACCACTACCG CCACCCCGAA
  	GAACACGGTT
  1301	CCGACCGCTG GTTCAACGCC TTGGGCAGCC GCCGCTTCAG
  	CCGCAACGCC
  1351	TGCGTCGTCG TCAGTTGCGG CACGGCGGTA ACGGTTGACG
  	CGCTCACCGA
  1401	TGACGGACAT TATCTCGGGG GAACCATCAT GCCCGGTTTC
  	CACCTGATGA
  1451	AAGAATCGCT CGCCGTCCGA ACCGCCAACC TCAACCGGCA
  	CGCCGGTAAG
  1501	CGTTATCCTT TCCCGACCAC AACGGGCAAT GCCGTCGCCA
  	GCGGCATGAT
  1551	GGATGCGGTT TGCGGCTCGG TTATGATGAT GCACGGGCGT
  	TTGAAAGAAA
  1601	AAACCGGGGC GGGCAAGCCT GTCGATGTCA TCATTACCGG
  	CGGCGGCGCG
  1651	GCAAAAGTTG CCGAAGCCCT GCCGCCTGCA TTTTTGGCGG
  	AAAATACCGT
  1701	GCGCGTGGCG GACAACCTCG TCATTCACGG GCTGCTGAAC
  	CTGATTGCCG
  1751	CCGAAGGCGG GGAATCGGAA CATACTTAA

• This encodes a protein having amino acid sequence <SEQ ID 236>:

• 1	MTVLKPSHWR VLAELADGLP QHVSQLARMA DMKPQQLNGF
  	WQQMPAHIRG
  51	LLRQHDGYWR LVRPLAVFDA EGLRELGERS GFQTALKHEC
  	ASSNDEILEL
  101	ARIAPDKAHK TICVTHLQSK GRGRQGRKWS HRLGECLMFS
  	FGWVFDRPQY
  151	ELGSLSPVAA VACRRALSRL GLKTQIKWPN DLVVGRDKLG
  	GILIETVRTG
  201	GKTVAVVGIG INFVLPKEVE NAASVQSLFQ TASRRGNADA
  	AVLLETLLAE
  251	LDAVLLQYAR DGFAPFVAEY QAANRDHGKA VLLLRDGETV
  	FEGTVKGVDG
  301	QGVLHLETAE GKQTVVSGEI SLRSDDRPVS VPKRRDSERF
  	LLLDGGNSRL
  351	KWAWVENGTF ATVGSAPYRD LSPLGAEWAE KVDGNVRIVG
  	CAVCGEFKKA
  401	QVQEQLARKI EWLPSSAQAL GIRNHYRHPE EHGSDRWFNA
  	LGSRRFSRNA
  451	CVVVSCGTAV TVDALTDDGH YLGGTIMPGF HLMKESLAVR
  	TANLNRHAGK
  501	RYPFPTTTGN AVASGMMDAV CGSVMMMHGR LKEKTGAGKP
  	VDVIITGGGA
  551	AKVAEALPPA FLAENTVRVA DNLVIHGLLN LIAAEGGESE
  	HT*

• ORF61a and ORF61-1 show 98.5% identity in 591 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF61 shows 94.2% identity over a 189aa overlap with a predicted ORF (ORF61.ng) from N. gonorrhoeae:
• 
• An ORF61ng nucleotide sequence <SEQ ID 237> was predicted to encode a protein having amino acid sequence <SEQ ID 238>:

• 1	MFSFGWAFDR PQYELGSLSP VAALACRRAL GCLGLETQIK
  	WPNDLVVGRD
  51	KLGGILIETV RAGGKTVAVV GIGINFVLPK EVENAASVQS
  	LFQTASRRGN
  101	ADAAVLLETL LAELGAVLEQ YAEEGFAPFL NEYETANRDH
  	GKAVLLLRDG
  151	ETVCEGTVKG VDGRGVLHLE TAEGEQTVVS GEISLRPDNR
  	SVSVPKRPDS
  201	ERFLLLEGGN SRLKWAWVEN GTFATVGSAP YRDLSPLGAE
  	WAEKADGNVR
  251	IVGCAVCGES KKAQVKEQLA RKIEWLPSSA QALGIRNHYR
  	HPEEHGSDRW
  301	FNALGSRRFS RNACVVVSCG TAVTVDALTD DGHYLGGTIM
  	PGFHLMKESL
  351	AVRTANLNRP AGKRYPFPTT TGNAVASGMM DAVCGSIMMM
  	HGRLKEKNGA
  401	GKPVDVIITG GGAAKVAEAL PPAFLAENTV RVADNLVIHG
  	LLNLIAAEGG
  451	ESEHA*

• Further analysis revealed the complete gonococcal DNA sequence <SEQ ID 239> to be:

• 1	ATGACGGTTT TGAAGCCTTC GCATTGGCGG GTGTTGGCGG
  	AGCTTGCCGA
  51	CGGTTTGCCG CAACACGTAT CGCAATTGGC GCGTGAGGCG
  	GACATGAAGC
  101	CGCAGCAGCT CAACGGTTTT TGGCAGCAGA TGCCGGCGCA
  	TATACGCGGG
  151	CTGTTGCGCC AACACGACGG CTATTGGCGG CTGGTGCGCC
  	CCTTGGCGGT
  201	TTTCGATGCC GAAGGTTTGC GCGATCTGGG GGAAAGGTCG
  	GGTTTTCAGA
  251	CGGCATTGAA GCACGAGTGC GCGTCCAGCA ACGACGAGAT
  	ACTGGAATTG
  301	GCGCGGATTG CGCCGGACAA GGCGCACAAA ACCATATGCG
  	TGACCCACCT
  351	GCAAAGTAAG GGCAGGGGGC GGCAGGGGCG GAAGTGGTCG
  	CACCGTTTGG
  401	GCGAGTGCCT GATGTTCAGT TTCGGCTGGG CGTTTGACCG
  	GCCGCAGTAT
  451	GAGTTGGGTT CGCTGTCGCC TGTTGCGGCA CTTGCGTGCC
  	GGCGCGCTTT
  501	GGGGTGTTTG GGTTTGGAAA CGCAAATCAA GTGGCCAAAC
  	GATTTGGTCG
  551	TCGGACGCGA CAAATTGGGC GGCATTCTGA TTGAAACAGT
  	CAGGGCGGGC
  601	GGTAAAACGG TTGCCGTGGT CGGTATCGGC ATCAATTTCG
  	TGCTGCCCAA
  651	GGAAGTGGAA AACGCCGCTT CCGTGCAGTC GCTGTTTCAG
  	ACGGCATCGC
  701	GGCGGGGCAA TGCCGATGCC GCCGTATTGC TGGAAACATT
  	GCTTGCGGAA
  751	CTGGGCGCGG TGTTGGAACA ATATGCGGAA GAAGGGTTCG
  	CGCCATTTTT
  801	AAATGAGTAT GAAACGGCCA ACCGCGACCA CGGCAAGGCG
  	GTATTGCTGT
  851	TGCGCGACGG CGAAACCGTG TGCGAAGGCA CGGTTAAAGG
  	CGTGGACGGA
  901	CGAGGCGTTC TGCACTTGGA AACGGCAgaa ggcgaACAGa
  	cggtcgtcag
  951	cggcgaaaTC AGcctGCggc ccgacaacaG GTCGGtttcc
  	gtgccgaagc
  1001	ggccggatTC GgaacgtTTT tTGCtgttgg aaggcgggaa
  	cagccgGCTC
  1051	AAGTGGGCGT GggtggAAAa cggcacgttc gcaaccgtgg
  	gcagcgcgCc
  1101	gtaCCGCGAT TTGTCGCCTT TGGGCGCGGA GTGGGCGGAA
  	AAGGCGGATG
  1151	GAAATGTCCG CATCGTCGGT TGCGCCGTGT GCGGAGAATC
  	CAAAAAGGCA
  1201	CAAGTGAAGG AACAGCTCGC CCGAAAAATC GAGTGGCTGC
  	CGTCTTCCGC
  1251	ACAGGCTTTG GGCATACGCA ACCACTACCG CCACCCCGAA
  	GAACACGGTT
  1301	CCGACCGTTG GTTCAACGCC TTGGGCAGCC GCCGCTTCAG
  	CCGCAACGCC
  1351	TGCGTCGTCG TCAGTTGCGG CACGGCGGTA ACGGTTGACG
  	CGCTCACCGA
  1401	TGACGGACAT TATCTCGGCG GAACCATCAT GCCCGGCTTC
  	CACCTGATGA
  1451	AAGAATCGCT CGCCGTCCGA ACCGCCAACC TCAACCGCCC
  	CGCCGGCAAA
  1501	CGTTACCCTT TCCCGACCAC AACGGGCAAC GCCGTCGCAA
  	GCGGCATGAT
  1551	GGACGCGGTT TGCGGCTCGA TAATGATGAT GCACGGCCGT
  	TTGAAAGAAA
  1601	AAAACGGCGC GGGCAAGCCT GTCGATGTCA TCATTACCGG
  	CGGCGGCGCG
  1651	GCGAAAGTCG CCGAAGCCCT GCCGCCTGCA TTTTTGGCGG
  	AAAATACCGT
  1701	GCGCGTGGCG GACAACCTCG TCATCCACGG GCTGCTGAAC
  	CTGATTGCCG
  1751	CCGAAGGCGG GGAATCGGAA CACGCTTAA

• This corresponds to the amino acid sequence <SEQ ID 240; ORF61ng-1>:

• 1	MTVLKPSHWR VLAELADGLP QHVSQLAREA DMKPQQLNGF
  	WQQMPAHIRG
  51	LLRQHDGYWR LVRPLAVFDA EGLRDLGERS GFQTALKHEC
  	ASSNDEILEL
  101	ARIAPDKAHK TICVTHLQSK GRGRQGRKWS HRLGECLMFS
  	FGWAFDRPQY
  151	ELGSLSPVAA LACRRALGCL GLETQIKWPN DLVVGRDKLG
  	GILIETVRAG
  201	GKTVAVVGIG INFVLPKEVE NAASVQSLFQ TASRRGNADA
  	AVLLETLLAE
  251	LGAVLEQYAE EGFAPFLNEY ETANRDHGKA VLLLRDGETV
  	CEGTVKGVDG
  301	RGVLHLETAE GEQTVVSGEI SLRPDNRSVS VPKRPDSERF
  	LLLEGGNSRL
  351	KWAWVENGTF ATVGSAPYRD LSPLGAEWAE KADGNVRIVG
  	CAVCGESKKA
  401	QVKEQLARKI EWLPSSAQAL GIRNHYRHPE EHGSDRWFNA
  	LGSRRFSRNA
  451	CVVVSCGTAV TVDALTDDGH YLGGTIMPGF HLMKESLAVR
  	TANLNRPAGK
  501	RYPFPTTTGN AVASGMMDAV CGSIMMMHGR LKEKNGAGKP
  	VDVIITGGGA
  551	AKVAEALPPA FLAENTVRVA DNLVIHGLLN LIAAEGGESE
  	HA*

• ORF61ng-1 and ORF61-1 show 93.9% identity in 591 aa overlap:
• 
• Based on this analysis, including the homology with the baf protein of B. pertussis and the presence of a putative prokaryotic membrane lipoprotein lipid attachment site, it is predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 29
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 241>:

• 1	ATGTTTTACC AAATCCTTGC CCTGATTATC TGGAGCAGCT
  	CGTTTATTGC
  51	CGCCAAATAT GTCTATGGCG GCATCGATCC CGCATTGATG
  	GTCGGCGTGC
  101	GCCTGCTAAT TGCCGCGCTG CCTGCACTGC CCGCCTGCCG
  	CCGTCATGTC
  151	GGCAAGATTC CGCGTGAGGA ATGGAAGCCG TTGCTGATTG
  	TGTCGTTCGT
  201	CAACTATGTG CTGACCCTGC TGCTTCAGTT TGTCGGGTTG
  	AAATACACTT
  251	CCGCCGCCAG CGCATCGGTC ATTGTCGGAC TCGAGCCGCT
  	GCTGATGGTG
  301	TTTGTCGGAC ACTTTTTCTT CAACGACAAA GCGCGTGCCT
  	ACCACTGGAT
  351	ATGCGGCGCG GCGGCATTTG CCGGTGTCGC GCTGCTGATG
  	GCGGGCGGTG
  401	CGGaAGAGGG CGGCGaAGTC GGCTGGTTCG GCTGCCTGCT
  	GGTGTTGTTG
  451	GCGGGCGCGG GCTTTTGTGC CGCTATGCGT CCGACGCAAA
  	GGCTGATTGC
  501	ACGCATCGGC GCACCGGCAT TCACATCTGT TTCCATTGCC
  	GCCGCATCGT
  551	TGATGTGCCT GCCGTTTTCG CTTGCTTTGG CGCAAAGTTA
  	TACCGTGGAC
  601	TGGAGCGTCG GGATGGTATT GTCGCTGCTG TATTTGGGTT
  	TGGGGTGC..

• This corresponds to the amino acid sequence <SEQ ID 242; ORF62>:

• 1	MFYQILALII WSSSFIAAKY VYGGIDPALM VGVRLLIAAL
  	PALPACRRHV
  51	GKIPREEWKP LLIVSFVNYV LTLLLQFVGL KYTSAASASV
  	IVGLEPLLMV
  101	FVGHFFFNDK ARAYHWICGA AAFAGVALLM AGGAEEGGEV
  	GWFGCLLVLL
  151	AGAGFCAAMR PTQRLIARIG APAFTSVSIA AASLMCLPFS
  	LALAQSYTVD
  201	WSVGMVLSLL YLGLGC..

• Further work revealed the complete nucleotide sequence <SEQ ID 243>:

• 1	ATGTTTTACC AAATCCTTGC CCTGATTATC TGGAGCAGCT
  	CGTTTATTGC
  51	CGCCAAATAT GTCTATGGCG GCATCGATCC CGCATTGATG
  	GTCGGCGTGC
  101	GCCTGCTAAT TGCCGCGCTG CCTGCACTGC CCGCCTGCCG
  	CCGTCATGTC
  151	GGCAAGATTC CGCGTGAGGA ATGGAAGCCG TTGCTGATTG
  	TGTCGTTCGT
  201	CAACTATGTG CTGACCCTGC TGCTTCAGTT TGTCGGGTTG
  	AAATACACTT
  251	CCGCCGCCAG CGCATCGGTC ATTGTCGGAC TCGAGCCGCT
  	GCTGATGGTG
  301	TTTGTCGGAC ACTTTTTCTT CAACGACAAA GCGCGTGCCT
  	ACCACTGGAT
  351	ATGCGGCGCG GCGGCATTTG CCGGTGTCGC GCTGCTGATG
  	GCGGGCGGTG
  401	CGGAAGAGGG CGGCGAAGTC GGCTGGTTCG GCTGCCTGCT
  	GGTGTTGTTG
  451	GCGGGCGCGG GCTTTTGTGC CGCTATGCGT CCGACGCAAA
  	GGCTGATTGC
  501	ACGCATCGGC GCACCGGCAT TCACATCTGT TTCCATTGCC
  	GCCGCATCGT
  551	TGATGTGCCT GCCGTTTTCG CTTGCTTTGG CGCAAAGTTA
  	TACCGTGGAC
  601	TGGAGCGTCG GGATGGTATT GTCGCTGCTG TATTTGGGTT
  	TGGGGTGCGG
  651	CTGGTACGCC TATTGGCTGT GGAACAAGGG GATGAGCCGT
  	GTTCCTGCCA
  701	ATGTTTCGGG ACTGTTGATT TCGCTCGAAC CCGTCGTCGG
  	CGTGCTGCTG
  751	GCGGTTTTGA TTTTGGGCGA ACACCTGTCG CCCGTGTCCG
  	CCTTGGGCGT
  801	GTTTGTCGTC ATCGCCGCCA CCTTGGTTGC CGGCCGGCTG
  	TCGCATCAAA
  851	AATAA

• This corresponds to the amino acid sequence <SEQ ID 244; ORF62-1>:

• 1	MFYQILALII WSSSFIAAKY VYGGIDPALM VGVRLLIAAL
  	PALPACRRHV
  51	GKIPREEWKP LLIVSFVNYV LTLLLQFV GL KYTSAASASV
  	IVGLEPLLMV
  101	FVGHFFFNDK ARAYHWICGA AAFAGVALLM AGGAEEGGEV
  	GWFGCLLVLL
  151	AGAGFCAAMR PTQRLIARIG APAFTSVSIA AASLMCLPFS
  	LALAQSYTVD
  201	WSVGMVLSLL YLGLGCGWYA YWLWNKGMSR VPANVSGLLI
  	SLEPVVGVLL
  251	AVLILGEHLS PVSALGVFVV IAATLVAGRL SHQK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with Hypothetical Transmembrane Protein H10976 of H. influenzae (Accession Number Q57147)
• ORF62 and HI0976 show 50% aa identity in 114aa overlap:

• Orf62	1	MFYQILALIIWSSSFIAAKYVYGGIDPALMVGVRXXXXXXXXXXXCRRHVGKIPREEWKP	60
  		M YQILAL+IWSSS I  K  Y  +DP L+V VR             R   KI +   K
  HI0976	1	MLYQILALLIWSSSLIVGKLTYSMMDPVLVVQVRLIIAMIIVMPLFLRRWKKIDKPMRKQ	60
  Orf62	61	LLIVSFVNYVLTLLLQFVGLKYTSAASASVIVGLEPLLMVFVGHFFFNDKARAY	114
  		L  ++F NY    LLQF+GLKYTSA+SA  ++GLEPLL+VFVGHFFF  K   +
  HI0976	61	LWWLAFFNYTAVFLLQFIGLKYTSASSAVTMIGLEPLLVVFVGHFFFKTKQNGF	114

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF62 shows 99.5% identity over a 216aa overlap with an ORF (ORF62a) from strain A of N. meningitidis:
• 
• The complete length ORF62a nucleotide sequence <SEQ ID 245> is:

• 1	ATGTTTTACC AAATCCTTGC CCTGATTATC TGGAGCAGCT
  	CGTTTATTGC
  51	CGCCAAATAT GTCTATGGCG GCATCGATCC CGCATTGATG
  	GTCGGCGTGC
  101	GCCTGCTGAT TGCTGCGCTG CCTGCACTGC CCGCCTGCCG
  	CCGTCATGTC
  151	GGCAAGATTC CGCGTGAGGA ATGGAAGCCG TTGCTGATTG
  	TGTCGTTCGT
  201	CAACTATGTG CTGACCCTGC TACTTCAGTT TGTCGGGTTG
  	AAATACACTT
  251	CCGCCGCCAG CGCATCGGTC ATTGTCGGAC TCGAGCCACT
  	GCTGATGGTG
  301	TTTGTCGGAC ACTTTTTCTT CAACGACAAA GCGCGTGCCT
  	ACCACTGGAT
  351	ATGCGGCGCG GCGGCATTTG CCGGTGTCGC GCTGCTGATG
  	GCGGGCGGTG
  401	CGGAAGAGGG CGGCGAAGTC GGCTGGTTCG GCTGCCTGCT
  	GGTGTTGTTG
  451	GCGGGCGCGG GCTTTTGTGC CGCTATGCGT CCGACGCAAA
  	GGCTGATTGC
  501	ACGCATCGGC GCACCGGCAT TCACATCTGT TTCCATTGCC
  	GCCGCATCGT
  551	TGATGTGCCT GCCGTTTTCG CTTGCTTTGG CGCAAAGTTA
  	TACCGTGGAC
  601	TGGAGCGTCG GAATGGTATT GTCGCTGCTG TATTTGGGCG
  	TGGGGTGCAG
  651	CTGGTACGCC TATTGGCTGT GGAACAAGGG GATGAGCCGT
  	GTTCCTGCCA
  701	ACGTTTCGGG ACTGTTGATT TCGCTCGAAC CCGTCGTCGG
  	CGTGCTGCTG
  751	GCGGTTTTGA TTTTGGGCGA ACACCTGTCG CCCGTGTCCG
  	TCTTGGGCGT
  801	GTTTGTCGTC ATCGCCGCCA CCTTGGTTGC CGGCCGGCTG
  	TCGCATCAAA
  851	AATAA

• This encodes a protein having amino acid sequence <SEQ ID 246>:

• 1	MFYQILALII WSSSFIAAKY VYGGIDPALM VGVRLLIAAL
  	PALPACRRHV
  51	GKIPREEWKP LLIVSFVNYV LTLLLQFV GL KYTSAASASV
  	IVGLEPLLMV
  101	FVGHFFFNDK ARAYHWICGA AAFAGVALLM AGGAEEGGEV
  	GWFGCLLVLL
  151	AGAGFCAAMR PTQRLIARIG APAFTSVSIA AASLMCLPFS
  	LALAQSYTVD
  201	WSVGMVLSLL YLGVGCSWYA YWLWNKGMSR VPANVSGLLI
  	SLEPVVGVLL
  251	AVLILGEHLS PVSVLGVFVV IAATLVAGRL SHQK*

• ORF62a and ORF62-1 show 98.9% identity in 284 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF62 shows 99.5% identity over a 216aa overlap with a predicted ORF (ORF62.ng) from N. gonorrhoeae:
• 
• The complete length ORF62ng nucleotide sequence <SEQ ID 247> is:

• 1	ATGTTTTACC AAATCCTTGC CCTGATTATC TGGGGCAGCT
  	CGTTTATTGC
  51	CGCCAAATAT GTCTATGGCG GCATCGATCC CGCATTGATG
  	GTCGGCGTGC
  101	GCCTGCTGAT TGCCGCGCTG CCTGCACTGC CCGCCTGCCG
  	CCGTCATGTC
  151	GGCAAGATTC CGCGTGAGGA ATGGAAGCCG TTGCTGATTG
  	TGTCGTTCGT
  201	CAACTATGTG CTGACCCTGC TGCTTCAGTT TGTCGGGTTG
  	AAATACACTT
  251	CCGCCGCCAG CGCATCGGTC ATTGTCGGAC TCGAGCCGCT
  	GCTGATGGTG
  301	TTTGTCGGAC ACTTTTTCTT CAACGACAAA GCGCGTGCCT
  	ACCACTGGAT
  351	ATGCGGCGCG GCGGCATTTG CCGGTGTCGC GCTGCTGATG
  	GCGGGCGGTG
  401	CGGAAGAGGG CGGCGAAGTC GGCTGGTTCG GCTGCCTGCT
  	GGTGTTGTTG
  451	GCGGGCGCGG GCTTTTGTGC CGCTATGCGT CCGACGCAAA
  	GGCTGATTGC
  501	CCGCATCGGC GCACCGGCAT TCACATCTGT TTCCATTGCC
  	GCCGCATCGT
  551	TGATGTGCCT GCCGTTTTCG CTTGCTTTGG CGCAAAGTTA
  	TACCGTGGAC
  601	TGGAGCGTCG GGATGGTATT GTCGCTGTTG TATTTGGGTT
  	TGGGGTGCGG
  651	CTGGTACGCC TATTGGCTGT GGAACAAGGG GATGAGCCGT
  	GTTCCTGCCA
  701	ACGCGTCGGG ACTGTTGATT TCGCTCGAAC CCGTCGTCGG
  	CGTGCTGTTG
  751	GCGGTTTTGA TTTTGGGCGA ACATTTATCG CCCGTGTCCG
  	CCTTGGGCGT
  801	GTTTGTCGTC ATCGCCGCCA CTTTCGCCGC CGGCCGGCTG
  	TCGCGCAGGG
  851	ACGCGCAAAA CGGCAATGCC GTCTGA

• This encodes a protein having amino acid sequence <SEQ ID 248>:

• 1	MFYQILALII WGSSFIAAKY VYGGIDPALM VGVRLLIAAL
  	PALPACRRHV
  51	GKIPREEWKP LLIVSFVNYV LTLLLQFV GL KYTSAASASV
  	IVGLEPLLMV
  101	FVGHFFFNDK ARAYHWICGA AAFAGVALLM AGGAEEGGEV
  	GWFGCLLVLL
  151	AGAGFCAAMR PTQRLIARIG APAFTSVSIA AASLMCLPFS
  	LALAQSYTVD
  201	WSVGMVLSLL YLGLGCGWYA YWLWNKGMSR VPANASGLLI
  	SLEPVVGVLL
  251	AVLILGEHLS PVSALGVFVV IAATFAAGRL SRRDAQNGNA
  	V*

• ORF62ng and ORF62-1 show 97.9% identity in 283 aa overlap:
• 
• Furthermore, ORF62ng shows significant homology to a hypothetical H. influenzae protein:

• sp|Q57147|Y976_HAEIN HYPOTHETICAL PROTEIN HI0976 >gi|1074589|pir||B64163
  hypothetical protein HI0976 - Haemophilus influenzae (strain Rd KW20)
  >gi|1574004 (U32778) hypothetical [Haemophilus influenzae] Length = 128
  Score = 106 bits (262), Expect = 2e−22
  Identities = 56/114 (49%), Positives = 68/114 (59%)

  Query:	1	MFYQILALIIWGSSFIAAKYVYGGIDPALMVGVRXXXXXXXXXXXCRRHVGKIPREEWKP	60
  		M YQILAL+IW SS I  K  Y  +DP L+V VR             R   KI +   K
  Sbjct:	1	MLYQILALLIWSSSLIVGKLTYSMMDPVLVVQVRLIIAMIIVMPLFLRRWKKIDKPMRKQ	60
  Query:	61	LLIVSFVNYVLTLLLQFVGLKYTSAASASVIVGLEPLLMVFVGHFFFNDKARAY	114
  		L  ++F NY    LLQF+GLKYTSA+SA  ++GLEPLL+VFVGHFFF  K   +
  Sbjct:	61	LWWLAFFNYTAVFLLQFIGLKYTSASSAVTMIGLEPLLVVFVGHFFFKTKQNGF	114

• Based on this analysis, including the homology with the transmembrane protein of H. influenzae and the putative leader sequence and several transmembrane domains in the gonococcal protein, it is predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 30
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 249>:

• 1	ATGCGCCGTT TTCTACCGAT CGCAGCCATA TGCGCmGwms
  	TCCTGkkGTA
  51	sGGACTGACG GCGGCAACCG GCAGCACCAG TTCGCTGGCG
  	GATTATTTCT
  101	GGTGGATTGT TGCGTTCAGC GCAATGCTGC TGCTGGTGTT
  	GTCCGCCGTT
  151	TTGGCACGTT ATGTCATATT GCTGTTGAAA GACAGGCGCG
  	ACGGCGTATT
  201	CGGTTCGCtA srTyGCCAAA gsGCCTgkks TGGG.ATGTT
  	TACGCTGGTT
  251	GCCGkACTGC CCGGCGTGTT TCTGTTCGGC TTTCCCGCAC
  	AGTTCATCAA
  301	CGGCACGATT AATTCGTGGT TCGGCAACGA TACCCACGAG
  	GCGCTTGAAC
  351	GCAGCCTCAA TTTGAGCAAG TCCGCATTGA ATTTGGCGGC
  	AGACAACGCC
  401	CTCGGCAACG CCGTCCCCGT GCAGATAGAC CTCATCGGCG
  	CGGCTTCCCT
  451	GCCCGGGGAT ATGGGCAGGG TGCTGGAACA TTACGCCGGC
  	AGCGGTTTTG
  501	CCCAGCTTGC CCTGTACAAy ksCGCAAGCG GCAAAATCGA
  	AAAAAGCATC
  551	AACCCGCACA AGCTCGATCA GCCGTTTCCA GGTAAGGCGC
  	GTTGGGAaAa
  601	AATCCaACGG GCGGGTTCGG TCAGGGATTT GGAAAGCATA
  	GGCGGCGTAT
  651	TGTaCGCGCA GGGCTGGCTG TCGGCGGGTA CGCACwACGG
  	GCGCGATTAC
  701	GCCTTGTTTT TCCGTCAGCC GGTTCCCAAA GGCGTGGCAG
  	AGGATGCCGT
  751	yTTAATCGAA AAGGCAAGGG CGAAATATGC TGAGTTGAGT
  	TACAGCAAAA
  801	AAGGTTTGCA GACCTTTTTC CTGGCAACCC TGCTGATTGC
  	CTCGCTGCTG
  851	TCGATTTTTC TTGCACTGGT CATGGCACTG TATTTCGCCC
  	GCCGTTTCGT
  901	CGAACCCGTC CTATCGCTTG CCGAGGGGGC GAAGGCGGTG
  	GCGCAAGGCG
  951	ATTTCAGCCA GACGCGCCCC GTGTTGCGCA ACGACGAGTT
  	CGGACGCTTG
  1001	ACCArGTTGT TCAACCACAT GACCGAGCAG CTTTCCATCG
  	CCAAAGATGC
  1051	AGACGAGCGC AACCGCCGGC GCGAGGAAGC CGCCAGGCAT
  	TATCTTGAAT
  1101	GCGTGTTGGA GGGGCTGACC ACGGGCGTGG TGGTGTTTGA
  	CGAACAAGGC
  1151	TGTCTGAAAA CCTTCAACAA AGCGGCGGGT ACC..

• This corresponds to the amino acid sequence <SEQ ID 250; ORF64>:

• 1	MRRFLPIAAI CAXXLXXGLT AATGSTSSLA DYFWWIVAFS
  	AMLLLVLSAV
  51	LARYVILLLK DRRDGVFGSX XAKXPXXXMF TLVAXLPGVF
  	LFGFPAQFIN
  101	GTINSWFGND THEALERSLN LSKSALNLAA DNALGNAVPV
  	QIDLIGAASL
  151	PGDMGRVLEH YAGSGFAQLA LYNXASGKIE KSINPHKLDQ
  	PFPGKARWEK
  201	IQRAGSVRDL ESIGGVLYAQ GWLSAGTHXG RDYALFFRQP
  	VPKGVAEDAV
  251	LIEKARAKYA ELSYSKKGLQ TFFLATLLIA SLLSIFLALV
  	MALYFARRFV
  301	EPVLSLAEGA KAVAQGDFSQ TRPVLRNDEF GRLTXLFNHM
  	TEQLSIAKDA
  351	DERNRRREEA ARHYLECVLE GLTTGVVVFD EQGCLKTFNK
  	AAGT..

• Further work revealed the complete nucleotide sequence <SEQ ID 251>:

• 1	ATGCGCCGTT TTCTACCGAT CGCAGCCATA TGCGCCGTCG
  	TCCTGTTGTA
  51	CGGACTGACG GCGGCAACCG GCAGCACCAG TTCGCTGGCG
  	GATTATTTCT
  101	GGTGGATTGT TGCGTTCAGC GCAATGCTGC TGCTGGTGTT
  	GTCCGCCGTT
  151	TTGGCACGTT ATGTCATATT GCTGTTGAAA GACAGGCGCG
  	ACGGCGTATT
  201	CGGTTCGCAG ATTGCCAAAC GCCTTTCTGG GATGTTTACG
  	CTGGTTGCCG
  251	TACTGCCCGG CGTGTTTCTG TTCGGCGTTT CCGCACAGTT
  	CATCAACGGC
  301	ACGATTAATT CGTGGTTCGG CAACGATACC CACGAGGCGC
  	TTGAACGCAG
  351	CCTCAATTTG AGCAAGTCCG CATTGAATTT GGCGGCAGAC
  	AACGCCCTCG
  401	GCAACGCCGT CCCCGTGCAG ATAGACCTCA TCGGCGCGGC
  	TTCCCTGCCC
  451	GGGGATATGG GCAGGGTGCT GGAACATTAC GCCGGCAGCG
  	GTTTTGCCCA
  501	GCTTGCCCTG TACAATGCCG CAAGCGGCAA AATCGAAAAA
  	AGCATCAACC
  551	CGCACAAGCT CGATCAGCCG TTTCCAGGTA AGGCGCGTTG
  	GGAAAAAATC
  601	CAACGGGCGG GTTCGGTCAG GGATTTGGAA AGCATAGGCG
  	GCGTATTGTA
  651	CGCGCAGGGC TGGCTGTCGG CGGGTACGCA CAACGGGCGC
  	GATTACGCCT
  701	TGTTTTTCCG TCAGCCGGTT CCCAAAGGCG TGGCAGAGGA
  	TGCCGTCTTA
  751	ATCGAAAAGG CAAGGGCGAA ATATGCTGAG TTGAGTTACA
  	GCAAAAAAGG
  801	TTTGCAGACC TTTTTCCTGG CAACCCTGCT GATTGCCTCG
  	CTGCTGTCGA
  851	TTTTTCTTGC ACTGGTCATG GCACTGTATT TCGCCCGCCG
  	TTTCGTCGAA
  901	CCCGTCCTAT CGCTTGCCGA GGGGGCGAAG GCGGTGGCGC
  	AAGGCGATTT
  951	CAGCCAGACG CGCCCCGTGT TGCGCAACGA CGAGTTCGGA
  	CGCTTGACCA
  1001	AGTTGTTCAA CCACATGACC GAGCAGCTTT CCATCGCCAA
  	AGAAGCAGAC
  1051	GAGCGCAACC GCCGGCGCGA GGAAGCCGCC AGGCATTATC
  	TTGAATGCGT
  1101	GTTGGAGGGG CTGACCACGG GCGTGGTGGT GTTTGACGAA
  	CAAGGCTGTC
  1151	TGAAAACCTT CAACAAAGCG GCGGAACAGA TTTTGGGGAT
  	GCCGCTTACC
  1201	CCCCTGTGGG GCAGCAGCCG GCACGGTTGG CACGGCGTTT
  	CGGCGCAGCA
  1251	GTCCCTGCTT GCCGAAGTGT TTGCCGCCAT CGGCGCGGCG
  	GCAGGTACGG
  1301	ACAAACCGGT CCATGTGAAA TATGCCGCGC CGGACGATGC
  	CAAAATCCTG
  1351	CTGGGCAAGG CAACCGTCCT GCCCGAAGAC AACGGCAACG
  	GCGTGGTAAT
  1401	GGTGATTGAC GACATCACCG TTTTGATACA CGCGCAAAAA
  	GAAGCCGCGT
  1451	GGGGCGAAGT GGCGAAGCGG CTGGCACACG AAATCCGCAA
  	TCCGCTCACG
  1501	CCCATCCAGC TTTCCGCCGA ACGGCTGGCG TGGAAATTGG
  	GCGGGAAGCT
  1551	GGATGAGCAG GATGCGCAAA TCCTGACGCG TTCGACCGAC
  	ACCATCGTCA
  1601	AACAGGTGGC GGCATTGAAG GAAATGGTCG AAGCATTCCG
  	CAATTATGCG
  1651	CGTTCCCCTT CGCTCAAATT GGAAAATCAG GATTTGAACG
  	CCTTAATCGG
  1701	CGATGTGTTG GCATTGTATG AAGCCGGTCC GTGCCGGTTT
  	GCGGCGGAGC
  1751	TTGCCGGCGA ACCGCTGACG GTGGCGGCGG ATACGACCGC
  	CATGCGGCAG
  1801	GTGCTGCACA ATATTTTCAA AAATGCCGCC GAAGCGGCGG
  	AAGAAGCCGA
  1851	TGTGCCCGAA GTCAGGGTAA AATCGGAAAC AGGGCAGGAC
  	GGTCGGATTG
  1901	TCCTGACGGT TTGCGACAAC GGCAAAGGGT TCGGCAGGGA
  	AATGCTGCAC
  1951	AACGCCTTCG AGCCGTATGT AACGGACAAA CCGGCGGGAA
  	CGGGATTGGG
  2001	TCTGCCTGTG GTGAAAAAAA TCATTGAAGA ACACGGCGGC
  	CGCATCAGCC
  2051	TGAGCAATCA GGATGCGGGT GGCGCGTGTG TCAGAATCAT
  	CTTGCCAAAA
  2101	ACGGTAAAAA CTTATGCGTA G

• This corresponds to the amino acid sequence <SEQ ID 252; ORF64-1>:

• 1	MRRFLPIAAI CAVVLLYGLT AATGSTSSLA DYFWWIVAFS
  	AMLLLVLSAV
  51	LARYVILLLK DRRDGVFGSQ IAKRLSGMFT LVAVLPGVFL
  	FGVSAQFING
  101	TINSWFGNDT HEALERSLNL SKSALNLAAD NALGNAVPVQ
  	IDLIGAASLP
  151	GDMGRVLEHY AGSGFAQLAL YNAASGKIEK SINPHKLDQP
  	FPGKARWEKI
  201	QRAGSVRDLE SIGGVLYAQG WLSAGTHNGR DYALFFRQPV
  	PKGVAEDAVL
  251	IEKARAKYAE LSYSKKGLQT FFLATLLIAS LLSIFLALVM
  	ALYFARRFVE
  301	PVLSLAEGAK AVAQGDFSQT RPVLRNDEFG RLTKLFNHMT
  	EQLSIAKEAD
  351	ERNRRREEAA RHYLECVLEG LTTGVVVFDE QGCLKTFNKA
  	AEQILGMPLT
  401	PLWGSSRHGW HGVSAQQSLL AEVFAAIGAA AGTDKPVHVK
  	YAAPDDAKIL
  451	LGKATVLPED NGNGVVMVID DITVLIHAQK EAAWGEVAKR
  	LAHEIRNPLT
  501	PIQLSAERLA WKLGGKLDEQ DAQILTRSTD TIVKQVAALK
  	EMVEAFRNYA
  551	RSPSLKLENQ DLNALIGDVL ALYEAGPCRF AAELAGEPLT
  	VAADTTAMRQ
  601	VLHNIFKNAA EAAEEADVPE VRVKSETGQD GRIVLTVCDN
  	GKGFGREMLH
  651	NAFEPYVTDK PAGTGLGLPV VKKIIEEHGG RISLSNQDAG
  	GACVRIILPK
  701	TVKTYA*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF64 shows 92.6% identity over a 392aa overlap with an ORF (ORF64a) from strain A of N. meningitidis:
• 
• The complete length ORF64a nucleotide sequence <SEQ ID 253> is:

• 1	ATGCGCCGTT TTCTACCGAT CGCAGCCATA TGCGCCGTCG
  	TCCTGTTGTA
  51	CGGACTGACG GCGGCAACCG GCAGCACCAG TTCGCTGGCG
  	GATTATTTCT
  101	GGTGGATTGT TGCGTTCAGC GCAATGCTGC TGCTGGTGTT
  	GTCCGCCGTT
  151	TTGGCACGTT ATGTCATATT GCTGTTGAAA GACAGGCGCG
  	ACGGCGTATT
  201	CGGTTCGCAG ATTGCCAAAC GCCTTTCCGG GATGTTTACG
  	CTGGTTGCCG
  251	TACTGCCCGG CGTGTTTCTG TTCGGCGTTT CCGCACAGTT
  	TATCAACGGC
  301	ACGATTAATT CGTGGTTCGG CAACGATACC CACGAGGCGC
  	TTGAACGCAG
  351	CCTCAATTTG AGCAAGTCCG CATTGAATCT GGCGGCAGAC
  	AACGCCCTTG
  401	GCAACGCCAT CCCCGTGCAG ATAGACNTCA TCGGCGCGGC
  	TTCCCTGCCC
  451	NGGGATATGG GCAGGGTGCT GGAACATTAC GCCGGCAGCG
  	GTTTTGCCCA
  501	GCTTGCCCTG TACAATGCCG CAAGCGGCAA AATCGAAAAA
  	AGCATCAACC
  551	CGCACAAGCT CGATCAGCCG TTTCCAGGTA AGGCGCGTTG
  	GGAAAAAATC
  601	CAACAGGCGG GTTCGGTCAG GGATNNGGAA AGCATAGGCG
  	GCGTATTGTA
  651	CGCGCANGGC TGGCTGTCGG CAGNNACGCA CAACGGGCGC
  	GATTACGCCT
  701	TGTTTTTCCG TCAGCCGGTT CCCAAAGGCG TGGCAGAGGA
  	TGCCGTCTTA
  751	ATCGAAAAGG CAAGGGCGNA ANANNNTNAG TTGAGTTACA
  	GCAAAAAAGG
  801	TTTGCAGACC TTTTTCCTNG CAACCCTGCT GATTGCCTCN
  	CTGCTGTCGA
  851	TTTTTCTTGC ACTGGTCATG GCACTGTATT TCGCCCGCCG
  	TTTCGTCGAA
  901	CCCGTCCTAT CGCTTGCCGA GGGGGCGAAG GCGGTGGCGC
  	AAGGCGATTT
  951	CAGCCAGACG CGCCCCGTGT TGCGCAACGA CGAGTTCGGA
  	CGCTTGACCA
  1001	AGTTGTTCAA CCACATGACC GAGCAGCTTT CCATCGCCAA
  	AGAAGCAGAC
  1051	GAGCGCAACC GCCGGCGCGA GGAAGCCGCC AGACATTATC
  	TCGAATGCGT
  1101	GTTGGAGGGG CTGACCACGG GCGTGGTGGT GTTTGACGAA
  	CAAGGCTGTC
  1151	TGAAAACCTT CAACAAAGCG GCGGAACAGA TTTTGGGGAT
  	GCCGCTTACC
  1201	CCCCTGTGGG GCAGCAGCCG GCACGGTTGG CACGGCGTTT
  	CGGCGCAGCA
  1251	GTCCCTGCTT GCCGAAGTGT TTGCCGCCAT CGGCGCGGCG
  	GCAGGTACGG
  1301	ACAAACCGGT CCATGTGAAA TATGCCGCGC CGGACGATGC
  	CAAAATCCTG
  1351	CTGGGCAAGG CAACCGTCCT GCCCGAAGAC AACNGCAACG
  	GCGTGGTAAT
  1401	GGTGATTGAC GACATCACCG TTTTGATACA CGCGCAAAAA
  	GAAGCCGCGT
  1451	GGGGCGAAGT GGCAAAACGG CTGGCACACG AAATCCGCAA
  	TCCGCTCACG
  1501	CCCATCCAGC TTTCTGCCGA ACGGCTGGCG TGGAAATTGG
  	GCGGGAAGCT
  1551	GGACGAGCAN GACGCGCAAA TCCTGACACG TTCGACCGAC
  	ACCATCATCA
  1601	AACAAGTGGC GGCATTAAAA GAAATGGTCG AGGCATTCCG
  	CAATTACNCG
  1651	CGTTCCCCTT CGNCTCAATT GGAAAATCAG GATTTGAACG
  	CCTTAATCGG
  1701	CGATGTGTTG GCATTGTACG AAGCTGGTCC GTGCCGGTTT
  	GCGGCGGAAC
  1751	TTGCCGGCGA ACCGCTGATG ATGGCGGCGG ATACGACCGC
  	CATGCGGCAG
  1801	GTGCTGCACA ATATTTTCAA AAATGCCGCC GAAGCGGCGG
  	AAGAAGCCGA
  1851	TGTGCCCGAA GTCAGGGTAA AATCGGAAGC GGGGCAGGAC
  	GGACGGATTG
  1901	TCCTGACAGT TTGCGACAAC GGCAAGGGGT TCGGCAGGGA
  	AATGCTGCAC
  1951	AATGCCTTCG AGCCGTATGT AACGGACAAA CCGGCTGGAA
  	CGGGATTGNG
  2001	ACTGCCCGTG GTGAAAAAAA TCATTGAAGA ACACGGCGGC
  	CNCATCAGCC
  2051	TGAGCAATCA GGATGCGGGC GGCGCGTNTG TCAGAATCAT
  	CTTGCCAAAA
  2101	ACGGTAGAAA CTTATGCGTA G

• This encodes a protein having amino acid sequence <SEQ ID 254>:

• 1	MRRFLPIAAI CAVVLLYGLT AATGSTSSLA DYFWWIVAFS
  	AMLLLVLSAV
  51	LARYVILLLK DRRDGVFGSQ IAKRLSGMFT LVAVLPGVFL
  	FGVSAQFING
  101	TINSWFGNDT HEALERSLNL SKSALNLAAD NALGNAIPVQ
  	IDXIGAASLP
  151	XDMGRVLEHY AGSGFAQLAL YNAASGKIEK SINPHKLDQP
  	FPGKARWEKI
  201	QQAGSVRDXE SIGGVLYAXG WLSAXTHNGR DYALFFRQPV
  	PKGVAEDAVL
  251	IEKARAXXXX LSYSKKGLQT FFLATLLIAS LLSIFLALVM
  	ALYFARRFVE
  301	PVLSLAEGAK AVAQGDFSQT RPVLRNDEFG RLTKLFNHMT
  	EQLSIAKEAD
  351	ERNRRREEAA RHYLECVLEG LTTGVVVFDE QGCLKTFNKA
  	AEQILGMPLT
  401	PLWGSSRHGW HGVSAQQSLL AEVFAAIGAA AGTDKPVHVK
  	YAAPDDAKIL
  451	LGKATVLPED NXNGVVMVID DITVLIHAQK EAAWGEVAKR
  	LAHEIRNPLT
  501	PIQLSAERLA WKLGGKLDEX DAQILTRSTD TIIKQVAALK
  	EMVEAFRNYX
  551	RSPSXQLENQ DLNALIGDVL ALYEAGPCRF AAELAGEPLM
  	MAADTTAMRQ
  601	VLHNIFKNAA EAAEEADVPE VRVKSEAGQD GRIVLTVCDN
  	GKGFGREMLH
  651	NAFEPYVTDK PAGTGLXLPV VKKIIEEHGG XISLSNQDAG
  	GAXVRIILPK
  701	TVETYA*

• ORF64a and ORF64-1 show 96.6% identity in 706 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF64 shows 86.6% identity over a 387aa overlap with a predicted ORF (ORF64.ng) from N. gonorrhoeae:
• 
• An ORF64ng nucleotide sequence <SEQ ID 255> was predicted to encode a protein having amino acid sequence <SEQ ID 256>:

• 1	MRRFLPIAAI CAVVLLYGLT AATGSTSSLA DYFWWIVSFS
  	AMLLLVLSAV
  51	LARYVILLLK DRRNGVFGSQ IAKRLSGMFT LVAVLPGLFL
  	FGISAQFING
  101	TINSWFGNDT HEALERSLNL SKSALDLAAD NAVSNAVPVQ
  	IDLIGTASLS
  151	GNMGSVLEHY AGSGFAQLAL YNAASGKIEK SINPHQFDQP
  	LPDKEHWEQI
  201	QQTGSVRSLE SIGGVLYAQG WLSAGTHNGR DYALFFRQPI
  	PENVAQDAVL
  251	IEKARAKYAE LSYSKKGLQT FFLVTLLIAS LLSIFLALVM
  	ALYFARRFVE
  301	PILSLAEGAK AVAQGDFSQT RPVLRNDEFG RLTKLFNHMT
  	EQLSIAKEAD
  351	ERNRRREEAA RHYLECVLDG LTTGVVVSYP LSCCRTAVFS
  	TCHSSPLSYF*

• Further work revealed the complete gonococcal DNA sequence <SEQ ID 257>:

• 1	ATGCGCCGCT TCCTACCGAT CGCAGCCATA TGCGCCGTCG
  	TCCTGCTGTA
  51	CGGATTGACG GCGGCGACCG GCAGCACCAG TTCGCTGGCG
  	GATTATTTCT
  101	GGTGGATAGT CTCGTTCAGC GCAATGCTGC TGCTGGTGTT
  	GTCCGCCGTT
  151	TTGGCACGTT ATGTCATATT GCTGTTGAAA GACAGGCGCA
  	ACGGCGTGTT
  201	CGGTTCGCAG ATTGCCAAAC GCCTTTCCGG GATGTTCACG
  	CTGGTCGCCG
  251	TACTGCCCGG CTTGTTCCTG TTCGGCATTT CCGCGCAGTT
  	TATCAACGGC
  301	ACGATTAATT CGTGGTTCGG CAACGACACC CACGAAGCCC
  	TCGAACGCAG
  351	CCTTAATTTG AGCAAGTCCG CACTGGATTT GGCGGCAGAC
  	AATGCCGTCA
  401	GCAACGCCGT TCCCGTACAG ATAGACCTCA TCGGCACCGC
  	CTCCCTGTCG
  451	GGCAATATGG GCAGTGTGCT GGAACACTAC GCCGGCAGCG
  	GTTTTGCCCA
  501	GCTTGCCCTG TACAATGCCG CAAGCGGGAA AATCGAAAAA
  	AGCATCAATC
  551	CGCACCAATT CGACCAGCCG CTTCCCGACA AAGAACATTG
  	GGAACAGATT
  601	CAGCAGACCG GTTCGGTTCG GAGTTTGGAA AGCATAGGCG
  	GCGTATTGTA
  651	CGCGCAGGGA TGGTTGTCGG CAGGTACGCA CAACGGGCGC
  	GATTACGCGC
  701	TGTTCTTCCG CCAGCCGATT CCCGAAAATG TGGCACAGGA
  	TGCCGTTCTG
  751	ATTGAAAAGG CGCGGGCGAA ATATGCCGAA TTGAGTTACA
  	GCAAAAAAGG
  801	TTTGCAGACC TTTTTTCTGG TAACCCTGCT GATTGCCTCG
  	CTGCTGTCGA
  851	TTTTTCTTGC GCTGGTAATG GCACTGTATT TTGCCCGCCG
  	TTTCGTCGAA
  901	CCCATTCTGT CGCTTGCCGA GGGCGCAAAG GCGGTGGCGC
  	AGGGTGATTT
  951	CAGCCAGACG CGCCCCGTAT TGCGCAACGA CGAGTTCGGA
  	CGTTTGACCA
  1001	AGCTGTTCAA CCATATGACC GAGCAGCTTT CCATCGCCAA
  	AGAAGCAGAC
  1051	GAACGCAACC GCCGGCGCGA GGAAGCCGCC CGTCACTACC
  	TCGAGTGCGT
  1101	GTTGGATGGG TTGACTACCG GTGTGGTGGT GTTTGACGAA
  	AAAGGCCGTT
  1151	TGAAAACCTT CAACAAGGCG GCGGAACAGA TTTTGGGGAT
  	GCCGCTCGCC
  1201	CCCCTGTGGG GCAGCAGCCG GCACGGTTGG CACGGCGTTT
  	CGGCGCAGCA
  1251	GTCCCTGCTT GCCGAAGTGT TtgccgccAT CGGTGCGGCG
  	GCAGGTACGG
  1301	ACAAACCGGT CCAGGTGGAA TATGCCGCGC CGGACGATGC
  	CAAAATCCTG
  1351	CTGGGCAAGG CGACGGTATT GCCCGAAGAC AACGGCAACG
  	GCGTGGTGAT
  1401	GGTGATTGAC GACATCACCG TGCTGATACG CGCGCAAAAA
  	GAAGCCGCGT
  1451	GGGGTGAAGT GGCGAAGCGG CTGGCACACG AAATCCGCAA
  	TCCGCTCACG
  1501	CCCATCCAGC TTTCCGCCGA ACGGCTGGCG TGGAAATTGG
  	GCGGGAAGCT
  1551	GGACGATCAG GACGCGCAAA TCCTGACGCG TtcgACCGAC
  	ACCATCATCA
  1601	AACAGgtggc gGCGTTAAAA GAAATGGTCG AGGCATTCCG
  	CAATTACGCG
  1651	CGCGCCCCTT CGCTCAAACT GGAAAATCAG GATTTGAACG
  	CCTTAATCGG
  1701	CGATGTTTTG GCCCTGTACG AAGCCGGCCC GTGCCGGTTT
  	GAGGCGGAAC
  1751	TTGCCGGCGA ACCGCTGATG ATGGCGGCGG ATACGACCGC
  	CATGCGGCAG
  1801	GTGCTGCACA ATATTTTCAA AAATGCCGCC GAAGCGGCGG
  	AAGAAGCCGA
  1851	TATGCCCGAA GTCAGGGTAA AATCGGAAAC GGGGCAGGAC
  	GGACGGATTG
  1901	TCCTGACGGT TTGCGACAAC GGCAAGGGAT TCGGCAAGGA
  	AATGCTGCAC
  1951	AATGCTTTCG AGCCGTATGT GACGGATAAG CCGGCGGGAA
  	CGGGACTGGG
  2001	TCTGCCTGTA GTGAAAAAAA TCATTGGAGA ACACGGCGGC
  	CGCATCAGCC
  2051	TGAGCAATCA GGATGCGGGT GGGGCGTGTG TCAGAATCAT
  	CTTGCCAAAA
  2101	ACGGTAGAAA CTTATGCGTA G

• This corresponds to the amino acid sequence <SEQ ID 258; ORF64ng-1>:

• 1	MRRFLPIAAI CAVVLLYGLT AATGSTSSLA DYFWWIVSFS
  	AMLLLVLSAV
  51	LARYVILLLK DRRNGVFGSQ IAKRLSGMFT LVAVLPGLFL
  	FGISAQFING
  101	TINSWFGNDT HEALERSLNL SKSALDLAAD NAVSNAVPVQ
  	IDLIGTASLS
  151	GNMGSVLEHY AGSGFAQLAL YNAASGKIEK SINPHQFDQP
  	LPDKEHWEQI
  201	QQTGSVRSLE SIGGVLYAQG WLSAGTHNGR DYALFFRQPI
  	PENVAQDAVL
  251	IEKARAKYAE LSYSKKGLQT FFLVTLLIAS LLSIFLALVM
  	ALYFARRFVE
  301	PILSLAEGAK AVAQGDFSQT RPVLRNDEFG RLTKLFNHMT
  	EQLSIAKEAD
  351	ERNRRREEAA RHYLECVLDG LTTGVVVFDE KGRLKTFNKA
  	AEQILGMPLA
  401	PLWGSSRHGW HGVSAQQSLL AEVFAAIGAA AGTDKPVQVE
  	YAAPDDAKIL
  451	LGKATVLPED NGNGVVMVID DITVLIRAQK EAAWGEVAKR
  	LAHEIRNPLT
  501	PIQLSAERLA WKLGGKLDDQ DAQILTRSTD TIIKQVAALK
  	EMVEAFRNYA
  551	RAPSLKLENQ DLNALIGDVL ALYEAGPCRF EAELAGEPLM
  	MAADTTAMRQ
  601	VLHNIFKNAA EAAEEADMPE VRVKSETGQD GRIVLTVCDN
  	GKGFGKEMLH
  651	NAFEPYVTDK PAGTGLGLPV VKKIIGEHGG RISLSNQDAG
  	GACVRIILPK
  701	TVETYA*

• ORF64ng-1 and ORF64-1 show 93.8% identity in 706 aa overlap:
• 
• Furthermore, ORF64ng-1 shows significant homology to a protein from A. caulinodans:

• sp|Q04850|NTRY_AZOCA NITROGEN REGULATION PROTEIN NTRY
  >gi|77479|pir||S18624 ntrY protein - Azorhizobium caulinodans >gi|38737
  (X63841) NtrY gene product [Azorhizobium caulinodans] Length = 771
  Score = 218 bits (550), Expect = 7e−56
  Identities = 195/720 (27%), Positives = 320/720 (44%), Gaps = 58/720 (8%)

  Query:	7	IAAICAVVLLYGLTAATGSTSSLADYFWWIXXXXXXXXXXXXXXXXRYVILLLKDRRNGV	66
  		I+A+   ++L GLT    +   +      +                R +  + K R  G
  Sbjct:	35	ISALATFLILMGLTPVVPTHQVVIS----VLLVNAAAVLILSAMVGREIWRIAKARARGR	90
  Query:	67	FGSQIAKRLSGMFTLVAVLPGLFLFGISAQFINGTINSWFGNDTHEALERSLNLSKSALD	126
  		+++  R+ G+F +V+V+P + +  +++  ++  ++ WF   T E +  S++++++ +
  Sbjct:	91	AAARLHIRIVGLFAVVSVVPAILVAVVASLTLDRGLDRWFSMRTQEIVASSVSVAQTYVR	150
  Query:	127	LAADNAVSNAVPVQIDLIGTASLSGNMGSVLEHYAG--SGFAQLALYNAASGKIEKSINP	184
  		A N   + + +  DL    S+          Y G  S F Q+    AA   +  ++
  Sbjct:	151	EHALNIRGDILAMSADLTRLKSV----------YEGDRSRFNQILTAQAALRNLPGAMLI	200
  Query:	185	HQFDQPLPDKEHWEQIQQTGSVRSLESIGGVLYAQGWLSAGTHNGRDYA-----------	233
  		+ D  + ++ +   I +   V +  +IG     Q  +     N  DY
  Sbjct:	201	RR-DLSVVERAN-VNIGREFIVPANLAIGDATPDQPVIYLP--NDADYVAAVVPLKDYDD	256
  Query:	234	--LFFRQPIPENVAQDAVLIEKARAKYAELSYSKKGLQTFFLVTXXXXXXXXXXXXXVMA	291
  		L+  + I   V       ++  A Y  L   + G+Q  F +               +
  Sbjct:	257	LYLYVARLIDPRVIGYLKTTQETLADYRSLEERRFGVQVAFALMYAVITLIVLLSAVWLG	316
  Query:	292	LYFARRFVEPILSLAEGAKAVAQGDFSQTRPVLRND-EFGRLTKLFNHMTEQLSIXXXXX	350
  		L F++  V PI  L   A  VA+G+     P+ R + +   L + FN MT +L
  Sbjct:	317	LNFSKWLVAPIRRLMSAADHVAEGNLDVRVPIYRAEGDLASLAETFNKMTHELRSQREAI	376
  Query:	351	XXXXXXXXXXXHYLECVLDGLTTGVVVFDEKGRLKTFNKAAEQILGMPLAPLWGSSRHGW	410
  		+ E VL G+  GV+  D + R+   N++AE++LG  L+ +    RH
  Sbjct:	377	LTARDQIDSRRRFTEAVLSGVGAGVIGLDSQERITILNRSAERLLG--LSEVEALHRHLA	434
  Query:	411	HGVSAQQSLLAEVFXXXXXXXXTDKPVQVEYAAPDDAKILLGKATVLPEDNG---NGVVM	467
  		V     LL E            + VQ       D +  +    V  E +    +G V+
  Sbjct:	435	EVVPETAGLLEEA------EHARQRSVQGNITLTRDGRERVFAVRVTTEQSPEAEHGWVV	488
  Query:	468	VIDDITVLIRAQKEAAWGEVAKRLAHEIRNPLTPIQLSAERLAWKLGGKLDDQDAQILTR	527
  		+DDIT LI AQ+ +AW +VA+R+AHEI+NPLTPIQLSAERL  K G  +  QD +I  +
  Sbjct:	489	TLDDITELISAQRTSAWADVARRIAHEIKNPLTPIQLSAERLKRKFGRHV-TQDREIFDQ	547
  Query:	528	STDTIIKQVAALKEMVEAFRNYARAPSLKLENQDLNALIGDVLALYEAGPCRFEAELAGE	587
  		TDTII+QV  +  MV+ F ++AR P   +++QD++ +I   + L   G      +
  Sbjct:	548	CTDTIIRQVGDIGRMVDEFSSFARMPKPVVDSQDMSEIIRQTVFLMRVGHPEVVFDSEVP	607
  Query:	588	PLMMAA-DTTAMRQVLHNIFKNXXXXXXXXDMPEVRVK-------SETGQDGRIVLTVCD	639
  		P M A  D   + Q L NI KN          P+VR +       +  G+D  +V+ + D
  Sbjct:	608	PAMPARFDRRLVSQALTNILKNAAEAIEAVP-PDVRGQGRIRVSANRVGED--LVIDIID	664
  Query:	640	NGKGFGKEMLHNAFEPYVTDKPAGTGLGLPVVKKIIGEHGGRISLSNQDAG-GACVRIIL	698
  		NG G  +E  +   EPYVT +  GTGLGL +V KI+ EHGG I L++   G GA +R+ L
  Sbjct:	665	NGTGLPQESRNRLLEPYVTTREKGTGLGLAIVGKIMEEHGGGIELNDAPEGRGAWIRLTL	724

• Based on this analysis, including the presence of a putative leader sequence (double-underlined) and several putative transmembrane domains (single-underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 31
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 259>:

• 1	ATGTACGCAT TTACCGCCGC ACAGCAACAG AAGGCACTCT
  	TCCGGCTGGT
  51	GCTTTTTCAT ATCCTCATCA TCGCCGCCAG CAACTATCTG
  	GTGCAGTTCC
  101	CTTTCCAAAT TTTCGGCATC CACACCACTT GGGGCGCATT
  	TTCCTTTCCC
  151	TTCATCTTCC TTGCCACCGA CCTGACCGTC CGCATTTTCG
  	GTTCTCACTT
  201	GGCACGGCGG ATTATCTTTT GGGTGATGTT CCCCGCCCTT
  	TTGCTTTCCT
  251	ACGTCTTTTC CGTTTTGTTC CACAACGGCA GTTGGACAGG
  	CTTGGGCGCG
  301	CTGTCCGAAT TCAACACCTT TGTCGGACGC ATCGCCTTAG
  	CCAGCTTTGC
  351	CGCCTACGCG ATCGGACAAA TCCTTGATAT TTTTGTATTC
  	AACAAATTAC
  401	GCCGTCTGAA AGCGTGGTGG ATTGCACCGA ACGCATCAAC
  	CGTCATCGGG
  451	CACGCGTTGG ATACG...

• This corresponds to the amino acid sequence <SEQ ID 260; ORF66>:

• 1	MYAFTAAQQQ KALFRLVLFH ILIIAASNYL VQFPFQIFGI
  	HTTWGAFSFP
  51	FIFLATDLTV RIFGSHLARR IIFWVMFPAL LLSYVFSVLF
  	HNGSWTGLGA
  101	LSEFNTFVGR IALASFAAYA IGQILDIFVF NKLRRLKAWW
  	IAPNASTVIG
  151	HALDT...

• Further work revealed the complete nucleotide sequence <SEQ ID 261>:

• 1	ATGTACGCAT TTACCGCCGC ACAGCAACAG AAGGCACTCT
  	TCCGGCTGGT
  51	GCTTTTTCAT ATCCTCATCA TCGCCGCCAG CAACTATCTG
  	GTGCAGTTCC
  101	CTTTCCAAAT TTTCGGCATC CACACCACTT GGGGCGCATT
  	TTCCTTTCCC
  151	TTCATCTTCC TTGCCACCGA CCTGACCGTC CGCATTTTCG
  	GTTCTCACTT
  201	GGCACGGCGG ATTATCTTTT GGGTGATGTT CCCCGCCCTT
  	TTGCTTTCCT
  251	ACGTCTTTTC CGTTTTGTTC CACAACGGCA GTTGGACAGG
  	CTTGGGCGCG
  301	CTGTCCGAAT TCAACACCTT TGTCGGACGC ATCGCCTTAG
  	CCAGCTTTGC
  351	CGCCTACGCG ATCGGACAAA TCCTTGATAT TTTTGTATTC
  	AACAAATTAC
  401	GCCGTCTGAA AGCGTGGTGG ATTGCACCGA CCGCATCAAC
  	CGTCATCGGC
  451	AACGCCTTGG ATACGCTGGT ATTTTTCGCC GTTGCCTTCT
  	ACGCAAGCAG
  501	CGATGGATTT ATGGCGGCAA ACTGGCAGGG CATCGCTTTT
  	GTCGATTACC
  551	TGTTCAAACT TACCGTCTGC ACCCTCTTCT TCCTGCCCGC
  	CTACGGCGTG
  601	ATACTGAATC TGCTGACGAA AAAACTGACA ACCCTGCAAA
  	CCAAACAGGC
  651	GCAAGACCGC CCCGCGCCCT CGCTGCAAAA TCCGTAA

• This corresponds to the amino acid sequence <SEQ ID 262; ORF66-1>:

• 1	MYAFTAAQQQ KALFRLVLFH ILIIAASNYL VQFPFQIFGI
  	HTTWGAFSFP
  51	FIFLATDLTV RIFGSHLARR IIFWVMFPAL LLSYVFSVLF
  	HNGSWTGLGA
  101	LSEFNTFVGR IALASFAAYA IGQILDIFVF NKLRRLKAWW
  	IAPTASTVIG
  151	NALDTLVFFA VAFYASSDGF MAANWQGIAF VDYLFKLTVC
  	TLFFLPAYGV
  201	ILNLLTKKLT TLQTKQAQDR PAPSLQNP*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with the Hypothetical Protein o221 of E. coli (Accession Number P37619)
• ORF66 and o221 protein show 67% aa identity in 155aa overlap:

• orf66	1	MYAFTAAQQQKALFRLVLFHILIIAASNYLVQFPFQIFGIHTTWGAFSFPFIFLATDLTV	60
  		M  F+  Q+ KALF L LFH+L+I +SNYLVQ P  I G HTTWGAFSFPFIFLATDLTV
  o221	1	MNVFSQTQRYKALFWLSLFHLLVITSSNYLVQLPVSILGFHTTWGAFSFPFIFLATDLTV	60
  orf66	61	RIFGSHLARRIIFWVMFPALLLSYVFSVLFHNGSWTGLGALSEFNTFVGRIALASFAAYA	120
  		RIFG+ LARRIIF VM PALL+SYV S LF+ GSW G GAL+ FN FV RIA ASF AYA
  o221	61	RIFGAPLARRIIFAVMIPALLISYVISSLFYMGSWQGFGALAHFNLFVARIATASFMAYA	120
  orf66	121	IGQILDIFVFNKLRRLKAWWIAPNASTVIGHALDT	155
  		+GQILD+ VFN+LR+ + WW+AP AST+ G+  DT
  o221	121	LGQILDVHVFNRLRQSRRWWLAPTASTLFGNVSDT	155

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF66 shows 96.1% identity over a 155aa overlap with an ORF (ORF66a) from strain A of N. meningitidis:
• 
• The complete length ORF66a nucleotide sequence <SEQ ID 263> is:

• 1	ATGTACGCAT TTACCGCCGC ACAGCAACAG AAGGCACTCT
  	TCTGGCTGGT
  51	GCTTTTTCAT ATCCTCATCA TCGCCGCCAG CAACTATCTG
  	GTGCAGTTCC
  101	CCTTCCAAAT TTCCGGCATC CACACCACTT GGGGCGCGTT
  	TTCCTTTCCC
  151	TTCATCTTCC TCGCCACCGA CCTGACCGTC CGCATTTTCG
  	GTTCGCACTT
  201	GGCACGGCGG ATTATCTTTT GGGTCATGTT CCCCGCCCTT
  	TTGCTTTCCT
  251	ACGTCTTTTC CGTTTTGTTC CACAACGGCA GTTGGACGGG
  	CTTGGGCGCG
  301	CTGTCCGAAT TCAACACCTT TGTCGGACGC ATCGCGCTGG
  	CAAGTTTTGC
  351	CGCCTACGCG CTCGGACAAA TCCTTGATAT TTTTGTGTTC
  	AACAAATTAC
  401	GCCGTCTGAA AGCGTGGTGG GTTGCCCCGA CTGCATCAAC
  	CGTCATCGGC
  451	AACGCCTTAG ATACGTTGGT ATTTTTCGCC GTTGCCTTCT
  	ACGCAAGCAG
  501	CGATGGATTT ATGGCGGCAA ACTGGCAGGG CATCGCTTTT
  	GTCGATTACC
  551	TGTTCAAACT CACCGTCTGC GGTCTGTTTT TCCTGCCCGC
  	CTACGGCGTG
  601	ATTCTGAATC TGCTGACGAA AAAACTGACG ACCCTGCAAA
  	CCAAACAGGC
  651	GCAAGACCGC CCCGCGCCCT CGCTGCAAAA TCCGTAA

• This encodes a protein having amino acid sequence <SEQ ID 264>:

• 1	MYAFTAAQQQ KALFWLVLFH ILIIAASNYL VQFPFQISGI
  	HTTWGAFSFP
  51	FIFLATDLTV RIFGSHLARR IIFWVMFPAL LLSYVFSVLF
  	HNGSWTGLGA
  101	LSEFNTFVGR IALASFAAYA LGQILDIFVF NKLRRLKAWW
  	VAPTASTVIG
  151	NALDTLVFFA VAFYASSDGF MAANWQGIAF VDYLFKLTVC
  	GLFFLPAYGV
  201	ILNLLTKKLT TLQTKQAQDR PAPSLQNP*

• ORF66a and ORF66-1 show 97.8% identity in 228 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF66 shows 94.2% identity over a 155aa overlap with a predicted ORF (ORF66.ng) from N. gonorrhoeae:
• 
• The complete length ORF66ng nucleotide sequence <SEQ ID 265> is:

• 1	ATGTACGCAT TGACCGCCGC ACAGCAACAG AAGGCACTCT
  	TCCGGCTGGT
  51	GCTTTTCCAT ATCCTCATCA TCGCCGCCAG CAACTATCTG
  	GTGCAGTTCC
  101	CCTTCCGGAT TTTCGGCATC CACACCACTT GGGGCGCGTT
  	TTCCTTTCCC
  151	TTCATCTTCC TCGCCACCGA CCTGACCGTC CGCATTTTCG
  	GTTCGCACTT
  201	GGCGCGGCGG ATTATCTTTT GGGTGATGTT CCCCGCCCTT
  	ttgCTTTcat
  251	aCGTCTTTTC CGTTTTGTTC CACAACGGCA GTTGGACGGG
  	CTTGGGCGCG
  301	ctgTCCCAAT TCAACACCTT TGTCGGACGC ATCGCGCTGG
  	CAAGTTTTGC
  351	CGCCTACGCG CTCGGACAAA TCCTTGATAT TTTCGTATTC
  	GACAAATTAC
  401	GCCGTCTGAA AGCGTGGTGG ATTGCCCCGG CCGCATCAAC
  	CGTCATCGGC
  451	AATGCACTGG ACACGTTAGT ATTTTTTGCC GTTGCCTTTT
  	ACGCAAGCAG
  501	CGATGAATTT ATGGCGGCAA ACTGGCAGGG CATCGCTTTT
  	GTCGATTACC
  551	TGTTCAAACT TACCGTCTGC ACCCTCTTCT TCCTGCCCGC
  	CTACGGCGTG
  601	ATACTGAATC TGCTGACGAA AAAACTGACG GCCCTGCAAA
  	CCAAACAGGC
  651	GCAAGACCGC CCCGTGCCCT CGCTGCAAAA TCCGTAA

• This encodes a protein having amino acid sequence <SEQ ID 266>:

• 1	MYALTAAQQQ KALFRLVLFH ILIIAASNYL VQFPFRIFGI
  	HTTWGAFSFP
  51	FIFLATDLTV RIFGSHLARR IIFWVMFPAL SLSYVFSVLF
  	HNGSWTGLGA
  101	PSQFNTFVGR IALASFAAYA LGQILDIFVF DKLRRLKAWW
  	IAPAASTVIG
  151	NALDTLVFFA VAFYASSDEF MAANWQGIAF VDYLFKLTVC
  	TLFFLPAYGV
  201	ILNLLTKKLT ALQTKQAQDR PVPSLQNP*

• An alternative annotated sequence is:

• 1	MYALTAAQQQ KALFRLVLFH ILIIAASNYL VQFPFRIFGI
  	HTTWGAFSFP
  51	FIFLATDLTV RIFGSHLARR  IIFWVMFPAL LLSYVFS VLF
  	HNGSWTGLGA
  101	LSQFNTFVGR IALASFAAYA LGQILDIFVF DKLRRLKAWW
  	IAPAASTVIG
  151	NALDTLVFFA VAFYASSDEF MAANWQGIAF VDYLFKLTVC
  	TLFFLPAYGV
  201	ILNLLTKKLT ALQTKQAQDR PVPSLQNP*

• ORF66ng and ORF66-1 show 96.1% identity in 228 aa overlap:
• 
• Furthermore, ORF66ng shows significant homology with an E. coli ORF:

• sp|P37619|YHHQ_ECOLI HYPOTHETICAL 25.3 KD PROTEIN IN FTSY-NIKA INTERGENIC
  REGION (O221)
  >gi|1073495|pir||S47690 hypothetical protein o221 - Escherichia coli
  >gi|466607 (U00039) No definition line found [Escherichia coli]
  >gi|1789882 (AE000423) hypothetical 25.3 kD protein in ftsY-nikA
  intergenic region [Escherichia coli]
  Length = 221
  Score = 273 bits (692), Expect = 5e−73
  Identities = 132/203 (65%), Positives = 155/203 (76%)

  Query:	1	MYALTAAQQQKALFRLVLFHILIIAASNYLVQFPFRIFGIHTTWGAFSFPFIFLATDLTV	60
  		M   +  Q+ KALF L LFH+L+I +SNYLVQ P  I G HTTWGAFSFPFIFLATDLTV
  Sbjct:	1	MNVFSQTQRYKALFWLSLFHLLVITSSNYLVQLPVSILGFHTTWGAFSFPFIFLATDLTV	60
  Query:	61	RIFGSHLARRIIFWVMFPALLLSYVFSVLFHNGSWTGLGALSQFNTFVGRIALASFAAYA	120
  		RIFG+ LARRIIF VM PALL+SYV S LF+ GSW G GAL+ FN FV RIA ASF AYA
  Sbjct:	61	RIFGAPLARRIIFAVMIPALLISYVISSLFYMGSWQGFGALAHFNLFVARIATASFMAYA	120
  Query:	121	LGQILDIFVFDKLRRLKAWWIAPAASTVIGNALDTLVFFAVAFYASSDEFMAANWQGIAF	180
  		LGQILD+ VF++LR+ + WW+AP AST+ GN  DTL FF +AF+ S D FMA +W  IA
  Sbjct:	121	LGQILDVHVFNRLRQSRRWWLAPTASTLFGNVSDTLAFFFIAFWRSPDAFMAEHWMEIAL	180
  Query:	181	VDYLFKLTVCTLFFLPAYGVILN	203
  		VDY FK+ +  +FFLP YGV+LN
  Sbjct:	181	VDYCFKVLISIVFFLPMYGVLLN	203

• Based on this analysis, including the homology with the E. coli protein and the presence of several putative transmembrane domains in the gonococcal protein, it is predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 32
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 267>:

• 1	ATGGTCATAA AATATACAAA TTTGAATTTT GCGAAATTGT
  	CGATAATTGC
  51	AATTTTGATG ATGTATTCGT TTGAAGCGAA TGCAAAyGCA
  	GTmwrAATAT
  101	CTGAAACTGT TTCAGTTGAT ACCGGACAAG GTGCGAAAAT
  	TCATAAGTTT
  151	GTACCTAAAA ATAGTAAAAC TTATTCATCT GATTTAATAA
  	AAACGGTAGA
  201	TTTAACACAC AyyCCTACGG GCGCAAAAGC CCGAATCAAC
  	GCCAAAATAA
  251	CCGCCAGCGT ATCCCGCGCC GGCGTATTGG CGGGGGTCGG
  	CAAACTTGCC
  301	CGCTTAGgCG CGAAATTCAG CACAAGGGCG GTtCCCTATG
  	TCGGAACAGC
  351	CcTTTTAGCC CACGACGTAT ACGAAAcTTT CAAAGAAGAC
  	ATACAGGCAC
  401	GAGGCTACCA ATACGACCCC GAAACCGACA AATTTGTAAA
  	AGGCTACGAA
  451	TATAGTAATT GCCTTTGGTA CGAAGACAAA AGACGTATTA
  	ATAGAACCTA
  501	TGGCTGCTAC GGCGTTGAT..

• This corresponds to the amino acid sequence <SEQ ID 268; ORF72>:

• 1	MVIKYTNLNF AKLSIIAILM MYSFEANANA VXISETVSVD
  	TGQGAKIHKF
  51	VPKNSKTYSS DLIKTVDLTH XPTGAKARIN AKITASVSRA
  	GVLAGVGKLA
  101	RLGAKFSTRA VPYVGTALLA HDVYETFKED IQARGYQYDP
  	ETDKFVKGYE
  151	YSNCLWYEDK RRINRTYGCY GVD..

• Further work revealed the complete nucleotide sequence <SEQ ID 269>:

• 1	ATGGTCATAA AATATACAAA TTTGAATTTT GCGAAATTGT
  	CGATAATTGC
  51	AATTTTGATG ATGTATTCGT TTGAAGCGAA TGCAAATGCA
  	GTAAAAATAT
  101	CTGAAACTGT TTCAGTTGAT ACCGGACAAG GTGCGAAAAT
  	TCATAAGTTT
  151	GTACCTAAAA ATAGTAAAAC TTATTCATCT GATTTAATAA
  	AAACGGTAGA
  201	TTTAACACAC ATCCCTACGG GCGCAAAAGC CCGAATCAAC
  	GCCAAAATAA
  251	CCGCCAGCGT ATCCCGCGCC GGCGTATTGG CGGGGGTCGG
  	CAAACTTGCC
  301	CGCTTAGGCG CGAAATTCAG CACAAGGGCG GTTCCCTATG
  	TCGGAACAGC
  351	CCTTTTAGCC CACGACGTAT ACGAAACTTT CAAAGAAGAC
  	ATACAGGCAC
  401	GAGGCTACCA ATACGACCCC GAAACCGACA AATTTGCAAA
  	GGTCTCAGGC
  451	TAA

• This corresponds to the amino acid sequence <SEQ ID 270; ORF72-1>:

• 1	MVIKYTNLNF AKLSIIAILM MYSFEANANA VKISETVSVD
  	TGQGAKIHKF
  51	VPKNSKTYSS DLIKTVDLTH IPTGAKARIN AKITASVSRA
  	GVLAGVGKLA
  101	RLGAKFSTRA VPYVGTALLA HDVYETFKED IQARGYQYDP
  	ETDKFAKVSG
  151	*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF72 shows 98.0% identity over a 147aa overlap with an ORF (ORF72a) from strain A of N. meningitidis:
• 
• The complete length ORF72a nucleotide sequence <SEQ ID 271> is:

• 1	ATGGTCATAA AATATACAAA TTTGAATTTT GCGAAATTGT
  	CGATAATTGC
  51	AATTTTGATG ATGTATTCGT TTGAAGCGAA TGCAAATGCA
  	GTAAAAATAT
  101	CTGAAACTGT TTCAGTTGAT ACCGGACAAG GTGCGAAAAT
  	TCATAAGTTT
  151	GTACCTAAAA ATAGTAAAAC TTATTCATCT GATTTAATAA
  	AAACGGTAGA
  201	TTTAACACAC ATCCCTACGG GCGCAAAAGC CCGAATCAAC
  	GCCAAAATAA
  251	CCGCCAGCGT ATCCCGCGCC GGCGTATTGG CGGGGGTCGG
  	CAAACTTGCC
  301	CGCTTAGGCG CGAAATTCAG CACAAGGGCG GTTCCCTATG
  	TCGGAACAGC
  351	CCTTTTAGCC CACGACGTAT ACGAAACTTT CAAAGAAGAC
  	ATACAGGCAC
  401	GAGGCTACCA ATACGACCCC GAAACCGACA AATTTGCAAA
  	GGTCTCAGGC
  451	TAA

• This encodes a protein having amino acid sequence <SEQ ID 272>:

• 1	MVIKYTNLNF AKLSIIAILM MYSFEANANA VKISETVSVD
  	TGQGAKIHKF
  51	VPKNSKTYSS DLIKTVDLTH IPTGAKARIN AKITASVSRA
  	GVLAGVGKLA
  101	RLGAKFSTRA VPYVGTALLA HDVYETFKED IQARGYQYDP
  	ETDKFAKVSG
  151	*

• ORF72a and ORF72-1 show 100.0% identity in 150 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF72 shows 89% identity over a 173aa overlap with a predicted ORF (ORF72.ng) from N. gonorrhoeae:
• 
• An ORF72ng nucleotide sequence <SEQ ID 273> was predicted to encode a protein having amino acid sequence <SEQ ID 274>:

• 1	MVTKHTNLNF AKLSIIAILM MYSFEANANA VKISETLSVD
  	TGQGAKVHKF
  51	VPKSSNIYSS DLTKAVDLTH IPTGAKARIN AKITASVSRA
  	GVLSGVGKLV
  101	RQGAKFGTRA VPYVGTALLA HDVYETFKED IQARGCRYDP
  	ETDKFVKGYE
  151	YANCLWYEDE RRINRTYGCY GVDSSIMRLM PDRSRFPEVK
  	QLMESQMYRL
  201	ARPFWNWRKE ELNKLSSLDW NNFVLNRCTF DWNGGGCAVN
  	KGDDFRAGAS
  251	FSLGRNPKYK EEMDAKKPEE ILSLKVDADP DKYIEATGYP
  	GYSEKVEVAP
  301	GTKVNMGPVT DRNGNPVQVA ATFGRDAQGN TTADVQVIPR
  	PDLTPASAEA
  351	PHAQPLPEVS PAENPANNPD PDENPGTRPN PEPDPDLNPD
  	ANPDTDGQPG
  401	TSPDSPAVPD RPNGRHRKER KEGEDGGLSC DYFPEILACQ
  	EMGKPSDRMF
  451	HDISIPQVTD DKTWSSHNFL PSNGVCPQPK TFHVFGRQYR
  	ASYEPLCVFA
  501	EKIRFAVLLA FIIMSAFVVF GSLGGE*

• After further analysis, the following gonococcal DNA sequence <SEQ ID 275> was identified:

• 1	ATGGTCACAA AACATACAAA TTTGAATTTT GCGAAATTGT
  	CGATAATTGC
  51	AATTTTGATG ATGTATTCGT TTGAAGCGAA TGCAAATGCA
  	GTAAAAATAT
  101	CTGAAACTCT TTCGGTTGAT ACCGGACAAG GCGCGAAAGT
  	TCATAAGTTC
  151	GTTCCTAAAT CAAGTAATAT TTATTCATCT GATTTAACAA
  	AAGCGGTAGA
  201	TTTAACGCAT ATCCCCACGG GCGCAAAAGC CCGAATCAAC
  	GCCAAAATAA
  251	CCGCCAGCGT ATCCCGCGCC GGCGTATTGT CGGGGGTCGG
  	CAAACTTGTC
  301	CGCCAAGGCG CGAAATTCGG CACAAGGGCG GTTCCCTATG
  	TCGGAACAGC
  351	CCTTTTAGCC CACGACGTAT ACGAAACTTT CAAAGAAGAC
  	ATACAGGCAC
  401	GAGGCTGCCG ATACGATCCC GAAACCGACA AATTT

• This corresponds to the amino acid sequence <SEQ ID 276; ORF72ng-1>:

• 1	MVTKHTNLNF AKLSIIAILM MYSFEANANA VKISETLSVD
  	TGQGAKVHKF
  51	VPKSSNIYSS DLTKAVDLTH IPTGAKARIN AKITASVSRA
  	GVLSGVGKLV
  101	RQGAKFGTRA VPYVGTALLA HDVYETFKED IQARGCRYDP
  	ETDKF

• ORF72ng-1 and ORF721-1 show 89.7% identity in 145 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence and transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 33
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 277>:

• 1	ATGAGATTTT TCGGTATCGG TTTTTTGGTG CTGCTGTTTT
  	TGGAGATTAT
  51	GTCGATTGTG TGGGTTGCCG ATTGGCTGGG CGGCGGCTGG
  	ACGTTGTTTT
  101	TGATGGCGGC AGGTTTTGCC GCCGGCGTGC TGATGCTCAG
  	GCAAACCGGG
  151	GCTGACCGGT CTTTTATTGG CGGGCGCGGC AATGAGAAGC
  	GGCGGGAAGG
  201	TATCCGTTTA TCAGATGTTG TGGCCTATC..

• This corresponds to the amino acid sequence <SEQ ID 278; ORF73>:

• 1	MRFFGIGFLV LLFLEIMSIV WVADWLGGGW TLFLMAAGFA
  	AGVLMLRQTG
  51	LTGLLLAGAA MRSGGKVSVY QMLWPI..

• Further work revealed the complete nucleotide sequence <SEQ ID 279>:

• 1	ATGAGATTTT TCGGTATCGG TTTTTTGGTG CTGCTGTTTT
  	TGGAGATTAT
  51	GTCGATTGTG TGGGTTGCCG ATTGGCTGGG CGGCGGCTGG
  	ACGTTGTTTT
  101	TGATGGCGGC AGGTTTTGCC GCCGGCGTGC TGATGCTCAG
  	GCATACGGGG
  151	CTGTCCGGTC TTTTATTGGC GGGCGCGGCA ATGAGAAGCG
  	GCGGGAGGGT
  201	ATCCGTTTAT CAGATGTTGT GGCCTATCCG TTATACGGTG
  	GCGGCTGTGT
  251	GTCTGATGAG TCCGGGATTC GTATCCTCGG TGTTGGCGGT
  	ATTGCTGCTG
  301	CTGCCGTTTA AGGGAGGGGC AGTGTTGCAG GCAGGAGGTG
  	CGGAAAATTT
  351	TTTCAACATG AACCAATCGG GCAGAAAAGA GGGCTTTTCC
  	CGCGATGACG
  401	ATATTATCGA GGGAGAATAT ACGGTTGAAG AGCCTTACGG
  	CGGCAATCGT
  451	TCCCGAAACG CCATCGAACA CAAAAAAGAC GAATAA

• This corresponds to the amino acid sequence <SEQ ID 280; ORF73-1>:

• 1	MRFFGIGFLV LLFLEIMSIV WVADWLGGGW TLFLMAAGFA
  	AGVLMLRHTG
  51	LSGLLLAGAA MRSGGRVSVY QMLWPIRYTV AAVCLMSPGF
  	VSSVLAVLLL
  101	LPFKGGAVLQ AGGAENFFNM NQSGRKEGFS RDDDIIEGEY
  	TVEEPYGGNR
  151	SRNAIEHKKD E*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF73 shows 90.8% identity over a 76aa overlap with an ORF (ORF73a) from strain A of N. meningitidis:
• 
• The complete length ORF73a nucleotide sequence <SEQ ID 281> is:

• 1	ATGAGATTTT TCGGTATCGG TTTTTTGGTG CTGCTGTTTT
  	TGGAGATTAT
  51	GTCGATTGTG TGGGTTGCCG ATTGGTTGGG CGGCGGTTGG
  	ACGCTGTTTC
  101	TAATGGCGGC AACCTTTGCC GCCGGCGTGG TGATGCTCAG
  	GCATACGGGG
  151	CTGTCCGGTC TTTTATTGGC GGGCGCGGCA ATGAGAAGCG
  	GCGGGAGGGT
  201	ATCCGTTTAT CANATGTTGT GGCNTATCCG TTATACGGTG
  	GCGGCGGTGT
  251	GTCNGATGAG TCCGGGATTC GTATCCTCGG TGTNGGCGGT
  	ATTGCTGNTG
  301	CTNCCGTTTA AGGGAGGTGC AGTGTTGCAG GCAGGAGGTG
  	CGGAAAATTT
  351	TTTCAACATG AACCANTCGG GCAGAAAAGA NGGCNTTTCC
  	CGCGATGACG
  401	ATATTATCGA GGGGGAATAT ACGGTTGAAG ANCCTTACGG
  	CGGCANTCGT
  451	TTCCGAAACG CCNTNGAACA CAAAAAAGAC GAATAA

• This encodes a protein having amino acid sequence <SEQ ID 282>:

• 1	MRFFGIGFLV LLFLEIMSIV WVADWLGGGW TLFLMAATFA
  	AGVVMLRHTG
  51	LSGLLLAGAA MRSGGRVSVY XMLWXIRYTV AAVCXMSPGF
  	VSSVXAVLLX
  101	LPFKGGAVLQ AGGAENFFNM NXSGRKXGXS RDDDIIEGEY
  	TVEXPYGGXR
  151	FRNAXEHKKD E*

• ORF73a and ORF73-1 show 91.3% identity in 161 aa overlap
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF73 shows 92.1% identity over a 76aa overlap with a predicted ORF (ORF73.ng) from N. gonorrhoeae:
• 
• The complete length ORF73ng nucleotide sequence <SEQ ID 283> is:

• 1	ATGAGATTTT TCGGTATCGG TTTTTTGGTG CTGCTGTTTT
  	TGGAAATTAT
  51	GTCGATTGTG TGGGTTGCCG ATTGGCTGGG CGGCGGTTGG
  	AcgcTGTTTC
  101	TAATGGCGGC AACCTTTGCC GCCGGTGTGC TGATGCTCAG
  	GCATAcggGG
  151	CTGTCCGGTC TTTTATTGGC TGGCGCGGCG GTAAAAagta
  	gtgGGAAGGT
  201	ATCTGTTTAT CagatgtTGT GGCCTATCCG TTATAcggtg
  	gcggcggtgT
  251	GTCTGatgag tCcggGATTC GTATCCTccg tgttggCGGT
  	ATTGCTGCTG
  301	CTGCcgttta aggGaggGgc agtgttgcag gcaggaggtg
  	cggaaaATTT
  351	TTTCAACATg aaCcaatcgg gcagaaAaga gggatttttc
  	cacgatgacg
  401	atattatcga gggagaatat acggttgaaa aacctgacgg
  	cggcaatcgt
  451	tcccgaAAcg ccatcgaaca cgaaaAagac gaataA

• This encodes a protein having amino acid sequence <SEQ ID 284>:

• 1	MRFFGIGFLV LLFLEIMSIV WVADWLGGGW TLFLMAATFA
  	AGVLMLRHTG
  51	LSGLLLAGAA VKSSGKVSVY QMLWPIRYTV AAVCLMSPGF
  	VSSVLAVLLL
  101	LPFKGGAVLQ AGGAENFFNM NQSGRKEGFF HDDDIIEGEY
  	TVEKPDGGNR
  151	SRNAIEHEKD E*

• ORF73ng and ORG73-1 show 93.8% identity in 161 aa overlap
• 
• Based on this analysis, including the presence of a putative leader sequence and putative transmembrane domain in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 34
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 285>:

• 1	ATGTTTGTTT TTCAGACGGC ATTCTT.ATG TTTCAGAAAC
  	ATTTGCAGAA
  51	AGCCTCCGAC AGCGTCGTCG GAGGGACATT ATACGTGGTT
  	GCCACGCCCA
  101	TCGGCAATTT GGCGGACATT ACCCTGCGCG CTTTGGCGGT
  	ATTGCAAAAG
  151	GCG....... .....GCCGA AGACACGCGC GTTACCGCAC
  	AGCTTTTGAG
  201	CGCGTACGGC ATTCAGGGCA AACTCGTCAG TGTGCGCGAA
  	CACAACGAAC
  251	GGCAGATGGC GGACAAGATT GTCGGCTATC TTTCAGACGG
  	CATGGTTGTG
  301	GCACAGGTTT CCGATGCGGG TACGCCGGCC GTGTGCGACC
  	CGGGCGCGAA
  351	ACTCGCCCGC CGCGTGCGTG AGGCCGGGTT TAAAGTCGTT
  	CCCGTCGTGG
  401	GCGCAAC.GC GGTGATGGCG GCTTTGAGCG TGGCCGGTGT
  	GGAAGGATCC
  451	GATTTTTATT TCAACGGTTT TGTACCGCCG AAATCGGGAG
  	AACGCAGGAA
  501	ACTGTTTGCC AAATGGGTGC GGGCGGCGTT TCCTATCGTC
  	ATGTTTGAAA
  551	CGCCGCACCG CATCGGTGCA GCGCTTGCCG ATATGGCGGA
  	ACTGTTCCCC
  601	GAACGCCGAT TAATGCTGGC GCGCGAAATT ACGAAAACGT
  	TTGAAACGTT
  651	CTTAAGCGGC ACGGTTGGGG AAATTCAGAC GGCATTGTCT
  	GCCGACGGCG
  701	ACCAATCGCG CGGCGAGATG GTGTTGGTGC TTTATCCGGC
  	GCAGGATGAA
  751	AAACACGAAG GCTTGTCCGA GTCCGCGCAA AACATCATGA
  	AAATCCTCAC
  801	AGCCGAGCTG CCGACCAAAC AGGCGGCGGA GCTTGCTGCC
  	AAAATCACGG
  851	GCGAGGGAAA GAAAGCTTTG TACGAT..

• This corresponds to the amino acid sequence <SEQ ID 286; ORF75>:

• 1	MFVFQTAFXM FQKHLQKASD SVVGGTLYVV ATPIGNLADI
  	TLRALAVLQK
  51	A....AEDTR VTAQLLSAYG IQGKLVSVRE HNERQMADKI
  	VGYLSDGMVV
  101	AQVSDAGTPA VCDPGAKLAR RVREAGFKVV PVVGAXAVMA
  	ALSVAGVEGS
  151	DFYFNGFVPP KSGERRKLFA KWVRAAFPIV MFETPHRIGA
  	ALADMAELFP
  201	ERRLMLAREI TKTFETFLSG TVGEIQTALS ADGDQSRGEM
  	VLVLYPAQDE
  251	KHEGLSESAQ NIMKILTAEL PTKQAAELAA KITGEGKKAL
  	YD..

• Further work revealed the complete nucleotide sequence <SEQ ID 287>:

• 1	ATGTTTCAGA AACATTTGCA GAAAGCCTCC GACAGCGTCG
  	TCGGAGGGAC
  51	ATTATACGTG GTTGCCACGC CCATCGGCAA TTTGGCGGAC
  	ATTACCCTGC
  101	GCGCTTTGGC GGTATTGCAA AAGGCGGACA TCATCTGTGC
  	CGAAGACACG
  151	CGCGTTACCG CACAGCTTTT GAGCGCGTAC GGCATTCAGG
  	GCAAACTCGT
  201	CAGTGTGCGC GAACACAACG AACGGCAGAT GGCGGACAAG
  	ATTGTCGGCT
  251	ATCTTTCAGA CGGCATGGTT GTGGCACAGG TTTCCGATGC
  	GGGTACGCCG
  301	GCCGTGTGCG ACCCGGGCGC GAAACTCGCC CGCCGCGTGC
  	GTGAGGCCGG
  351	GTTTAAAGTC GTTCCCGTCG TGGGCGCAAG CGCGGTGATG
  	GCGGCTTTGA
  401	GCGTGGCCGG TGTGGAAGGA TCCGATTTTT ATTTCAACGG
  	TTTTGTACCG
  451	CCGAAATCGG GAGAACGCAG GAAACTGTTT GCCAAATGGG
  	TGCGGGCGGC
  501	GTTTCCTATC GTCATGTTTG AAACGCCGCA CCGCATCGGT
  	GCGACGCTTG
  551	CCGATATGGC GGAACTGTTC CCCGAACGCC GATTAATGCT
  	GGCGCGCGAA
  601	ATTACGAAAA CGTTTGAAAC GTTCTTAAGC GGCACGGTTG
  	GGGAAATTCA
  651	GACGGCATTG TCTGCCGACG GCAACCAATC GCGCGGCGAG
  	ATGGTGTTGG
  701	TGCTTTATCC GGCGCAGGAT GAAAAACACG AAGGCTTGTC
  	CGAGTCCGCG
  751	CAAAACATCA TGAAAATCCT CACAGCCGAG CTGCCGACCA
  	AACAGGCGGC
  801	GGAGCTTGCT GCCAAAATCA CGGGCGAGGG AAAGAAAGCT
  	TTGTACGATC
  851	TGGCTCTGTC TTGGAAAAAC AAATAG

• This corresponds to the amino acid sequence <SEQ ID 288; ORF75-1>:

• 1	MFQKHLQKAS DSVVGGTLYV VATPIGNLAD ITLRALAVLQ
  	KADIICAEDT
  51	RVTAQLLSAY GIQGKLVSVR EHNERQMADK IVGYLSDGMV
  	VAQVSDAGTP
  101	AVCDPGAKLA RRVREAGFKV VPVVGASAVM AALSVAGVEG
  	SDFYFNGFVP
  151	PKSGERRKLF AKWVRAAFPI VMFETPHRIG ATLADMAELF
  	PERRLMLARE
  201	ITKTFETFLS GTVGEIQTAL SADGNQSRGE MVLVLYPAQD
  	EKHEGLSESA
  251	QNIMKILTAE LPTKQAAELA AKITGEGKKA LYDLALSWKN
  	K*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF75 shows 95.8% identity over a 283aa overlap with an ORF (ORF75a) from strain A of N. meningitidis:
• 
• The complete length ORF75a nucleotide sequence <SEQ ID 289> is:

• 1	ATGTTTCAGA AACATTTGCA GAAAGCCTCC GACAGCGTCG
  	TCGGAGGGAC
  51	ATTATACGTG GTTGCCACGC CCATCGGCAA TTTGGCGGAC
  	ATTACCCTGC
  101	GCGCTTTGGC GGTATTGCAA AAGGCGGACA TCATCTGTGC
  	CGAAGACACG
  151	CGCGTTACCG CGCAGCTTTT GAGCGCGTAC GGCATTCAGG
  	GCAAACTCGT
  201	CAGCGTGCGC GAACACAACG AACGGCAGAT GGCGGACAAG
  	ATTGTCGGCT
  251	ATCTTTCAGA CGGCATGGTT GTGGCACAGG TTTCCGATGC
  	GGGTACGCCG
  301	GCCGTGTGCG ACCCGGGCGC GAAACTCGCC CGCCGCGTGC
  	GTGAGGTCGG
  351	GTTTAAAGTT GTCCCTGTTG TCGGCGCAAG CGCGGTGATG
  	GCGGCTTTGA
  401	GTGTGGCTGG TGTGGCGGGA TCCGATTTTT ATTTCAACGG
  	TTTTGTACCG
  451	CCGAAATCGG GCGAACGTAG GAAATTGTTT GCCAAATGGG
  	TGCGGGTGGC
  501	GTTTCCCGTC GTGATGTTTG AAACGCCGCA CCGCATCGGG
  	GCGACGCTTG
  551	CCGATATGGC GGAACTGTTC CCCGAACGCC GATTAATGCT
  	GGCGCGCGAA
  601	ATCACGAAAA CGTTTGAAAC GTTCTTAAGC GGCACGGTTG
  	GGGAAATTCA
  651	GACGGCATTG GCGGCGGACG GCAACCAATC GCGCGGCGAG
  	ATGGTGTTGG
  701	TGCTTTATCC GGCGCAGGAT GAAAAACACG AAGGCTTGTC
  	CGAGTCCGCG
  751	CAAAACATCA TGAAAATCCT CACAGCCGAG CTGCCGACCA
  	AACAGGCGGC
  801	GGAGCTTGCC GCCAAAATCA CGGGCGAGGG AAAAAAAGCT
  	TTGTACGATC
  851	TGGCACTGTC TTGGAAAAAC AAATGA

• This encodes a protein having amino acid sequence <SEQ ID 290>:

• 1	MFQKHLQKAS DSVVGGTLYV VATPIGNLAD ITLRALAVLQ
  	KADIICAEDT
  51	RVTAQLLSAY GIQGKLVSVR EHNERQMADK IVGYLSDGMV
  	VAQVSDAGTP
  101	AVCDPGAKLA RRVREVGFKV VPVVGASAVM AALSVAGVAG
  	SDFYFNGFVP
  151	PKSGERRKLF AKWVRVAFPV VMFETPHRIG ATLADMAELF
  	PERRLMLARE
  201	ITKTFETFLS GTVGEIQTAL AADGNQSRGE MVLVLYPAQD
  	EKHEGLSESA
  251	QNIMKILTAE LPTKQAAELA AKITGEGKKA LYDLALSWKN
  	K*

• ORF75a and ORF75-1 show 98.3% identity in 291 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF75 shows 93.2% identity over a 292aa overlap with a predicted ORF (ORF75.ng) from N. gonorrhoeae:
• 
• An ORF75ng nucleotide sequence <SEQ ID 291> was predicted to encode a protein having amino acid sequence <SEQ ID 292>:

• 1	MSVFQTAFFM FQKHLQKASD SVVGGTLYVV ATPIGNLADI
  	TLRALAVLQK
  51	ADIICAEDTR VTAQLLSAYG IQGRLVSVRE HNERQMADKV
  	IGFLSDGLVV
  101	AQVSDAGTPA VCDPGAKLAR RVREAGFKVV PVVGASAVMA
  	ALSVAGVAES
  151	DFYFNGFVPP KSGERRKLFA KWVRAAFPVV MFETPHRIGA
  	TLADMAELFP
  201	ERRLMLAREI TKTFETFLSG TVGEIQTALA ADGNQSRGEM
  	VLVLYPAQDE
  251	KHEGLSESAQ NAMKILAAEL PTKQAAELAA KITGEGKKAL
  	YDLALSWKNK
  301	*

• After further analysis, the following gonococcal DNA sequence <SEQ ID 293> was identified:

• 1	ATGTTTCAGA AACACTTGCA GAAAGCCTCC GACAGCGTCG
  	TCGGAGGGAC
  51	ATTATACGTG GTTGCCACGC CCATCGGCAA TTTGGCAGAC
  	ATTACCCTGC
  101	GCGCTTTGGC GGTATTGCAA AAGGCGGACA TCATTTGTGC
  	CGAAGACACG
  151	CGCGTTACTG CGCAGCTTTT GAGCGCGTAC GGCATTCAGG
  	GCAGGTTGGT
  201	CAGTGTGCGC GAACACAACG AGCGGCAGAT GGCGGACAAG
  	GTAATCGGTT
  251	TCCTTTCAGA CGGCCTGGTT GTGGCGCAGG TTTCCGATGC
  	GGGTACGCCG
  301	GCCGTGTGCG ACCCGGGCGC GAAACTCGCC CGCCGCGTGC
  	GCGAAGCAGG
  351	GTTCAAAGTC GTTCCCGTCG TGGGCGCAAG CGCGGTAATG
  	GCGGCGTTGA
  401	GTGTGGCCGG TGTGGCGGAA TCCGATTTTT ATTTCAACGG
  	TTTTGTACCG
  451	CCGAAATCGG GCGAACGTAG GAAATTGTTT GCCAAATGGG
  	TGCGGGCGGC
  501	ATTTCCTGTC GTCATGTTTG AAACGCCGCA CCGAATCGGG
  	GCAACGCTTG
  551	CCGATATGGC GGAATTGTTC CCCGAACGCC GTCTGATGCT
  	GGCGCGCGAA
  601	ATCACGAAAA CGTTTGAAAC GTTCTTAAGC GGCACGGTTG
  	GGGAAATTCA
  651	GACGGCATTG GCGGCGGACG GCAACCAATC GCGCGGCGAG
  	ATGGTGTTGG
  701	TGCTTTATCC GGCGCAGGAT GAAAAACACG AAGGCTTGTC
  	CGAGTCTGCG
  751	CAAAATGCGA TGAAAATCCT TGCGGCCGAG CTGCCGACCA
  	AGCAGGCGGC
  801	GGAGCTTGCC GCCAAGATTA CAGGTGAGGG CAAAAAGGCT
  	TTGTACGATT
  851	TGGCACTGTC GTGGAAAAAC AAATGA

• This corresponds to the amino acid sequence <SEQ ID 294; ORF75ng-1>:

• 1	MFQKHLQKAS DSVVGGTLYV VATPIGNLAD ITLRALAVLQ
  	KADIICAEDT
  51	RVTAQLLSAY GIQGRLVSVR EHNERQMADK VIGFLSDGLV
  	VAQVSDAGTP
  101	AVCDPGAKLA RRVREAGFKV VPVVGASAVM AALSVAGVAE
  	SDFYFNGFVP
  151	PKSGERRKLF AKWVRAAFPV VMFETPHRIG ATLADMAELF
  	PERRLMLARE
  201	ITKTFETFLS GTVGEIQTAL AADGNQSRGE MVLVLYPAQD
  	EKHEGLSESA
  251	QNAMKILAAE LPTKQAAELA AKITGEGKKA LYDLALSWKN
  	K*

• ORF75ng-1 and ORF75-1 show 96.2% identity in 291 aa overlap:
• 
• Furthermore, ORG75ng-1 shows significant homology to a hypothetical E. coli protein:

• sp|P45528|YRAL_ECOLI HYPOTHETICAL 31.3 KD PROTEIN IN AGAI-MTR INTERGENIC
  REGION (F286)
  >gi|606086 (U18997) ORF_f286 [Escherichia coli]
  >gi|1789535 (AE000395) hypothetical 31.3 kD protein in agai-mtr
  intergenic region [Escherichia coli] Length = 286
  Score = 218 bits (550), Expect = 3e−56
  Identities = 128/284 (45%), Positives = 171/284 (60%), Gaps = 4/284 (1%)

  Query:	4	KHLQKASDSVVGGTLYVVATPIGNLADITLRALAVLQKADIICAEDTRVTAQLLSAYGIQ	63
  		K  Q A +S   G LY+V TPIGNLADIT RAL VLQ  D+I AEDTR T  LL  +GI
  Sbjct:	2	KQHQSADNSQ--GQLYIVPTPIGNLADITQRALEVLQAVDLIAAEDTRHTGLLLQHFGIN	59
  Query:	64	GRLVSVREHNERQMADKVIGFLSDGLVVAQVSDAGTPAVCDPGAKLARRVREAGFKVVPV	123
  		RL ++ +HNE+Q A+ ++  L +G  +A VSDAGTP + DPG  L R  REAG +VVP+
  Sbjct:	60	ARLFALHDHNEQQKAETLLAKLQEGQNIALVSDAGTPLINDPGYHLVRTCREAGIRVVPL	119
  Query:	124	VGASAVMAALSVAGVAESDFYFNGFVPPKSGERRKLFAKWVRAAFPVVMFETPHRIGATL	183
  		G  A + ALS AG+    F + GF+P KS  RR            ++ +E+ HR+  +L
  Sbjct:	120	PGPCAAITALSAAGLPSDRFCYEGFLPAKSKGRRDALKAIEAEPRTLIFYESTHRLLDSL	179
  Query:	184	ADMAELFPERR-LMLAREITKTFETFLSGTVGEIQTALAADGNQSRGEMVLVLYPAQDEK	242
  		D+  +  E R ++LARE+TKT+ET     VGE+   +  D N+ +GEMVL++      +
  Sbjct:	180	EDIVAVLGESRYVVLARELTKTWETIHGAPVGELLAWVKEDENRRKGEMVLIV-EGHKAQ	238
  Query:	243	HEGLSESAQNAMKILAAELPTKQAAELAAKITGEGKKALYDLAL	286
  		E L   A   + +L AELP K+AA LAA+I G  K ALY  AL
  Sbjct:	239	EEDLPADALRTLALLQAELPLKKAAALAAEIHGVKKNALYKYAL	282

• Based on this analysis, including the presence of a putative transmembrane domain in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 35
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 295>:
• 
• This corresponds to the amino acid sequence <SEQ ID 296; ORF76>:
• 
• Further work revealed the complete nucleotide sequence <SEQ ID 297>:

• 1	ATGAAACAGA AAAAAACCGC TGCCGCAGTT ATTGCTGCAA
  	TGTTGGCAGG
  51	TTTTGCGGCA GCCAAAGCAC CCGAAATCGA CCCGGCTTTG
  	GTGGATACGC
  101	TGGTGGCGCA GATCATGCAG CAGGCAGACC GGCATGCGGA
  	GCAGTCCCAA
  151	AAACCGGACG GGCAGGCAAT CCGAAACGAT GCCGTCCGCC
  	GGCTACAAAC
  201	TTTGGAAGTT TTGAAAAACA GGGCATTGAA GGAAGGTTTG
  	GATAAGGATA
  251	AGGATGTCCA AAACCGCTTT AAAATCGCCG AAGCGTCTTT
  	TTATGCCGAG
  301	GAGTACGTCC GTTTTCTGGA ACGTTCGGAA ACGGTTTCCG
  	AAGACGAGCT
  351	GCACAAGTTT TACGAACAGC AAATCCGCAT GATCAAATTG
  	CAGCAGGTCA
  401	GCTTCGCAAC CGAAGAGGAG GCGCGTCAGG CGCAGCAGCT
  	CCTGCTCAAA
  451	GGGCTGTCTT TTGAAGGGCT GATGAAGCGT TATCCGAACG
  	ACGAGCAGGC
  501	TTTTGACGGT TTCATTATGG CGCAGCAGCT TCCCGAGCCG
  	CTGGCTTCGC
  551	AGTTTGCCGC GATGAATCGG GGCGACGTTA CCCGCGATCC
  	GGTCAAATTG
  601	GGCGAACGCT ATTATCTGTT CAAACTCAGC GAGGTCGGGA
  	AAAACCCCGA
  651	CGCGCAGCCT TTCGAGTTGG TCAGAAACCA GTTGGAGCAG
  	GGTTTGAGAC
  701	AGGAAAAAGC CCGCTTGAAA ATCGATGCCC TTTTGGAAGA
  	AAACGGTGTC
  751	AAACCGTAA

• This corresponds to the amino acid sequence <SEQ ID 298; ORF76-1>:

• 1	MKQKKTAAAV IAAMLAGFAA AKAPEIDPAL VDTLVAQIMQ
  	QADRHAEQSQ
  51	KPDGQAIRND AVRRLQTLEV LKNRALKEGL DKDKDVQNRF
  	KIAEASFYAE
  101	EYVRFLERSE TVSEDELHKF YEQQIRMIKL QQVSFATEEE
  	ARQAQQLLLK
  151	GLSFEGLMKR YPNDEQAFDG FIMAQQLPEP LASQFAAMNR
  	GDVTRDPVKL
  201	GERYYLFKLS EVGKNPDAQP FELVRNQLEQ GLRQEKARLK
  	IDALLEENGV
  251	KP*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF76 shows 96.7% identity over a 30aa overlap and 96.8% identity over a 31aa overlap with an ORF (ORF76a) from strain A of N. meningitidis:
• 
• The complete length ORF76a nucleotide sequence <SEQ ID 299> is:

• 1	ATGAAACAGA AAAAAACCGC TGCCGCAGTT ATTGCTGCAA
  	TGTTGGCAGG
  51	TTTTGCGGCA GCCAAAGCAC CCGAAATCGA CCCGGCTTTG
  	GTGGATACGC
  101	TGGTGGCGCA GATCATGCAG CAGGCAGACC GGCATGCGGA
  	GCAGTCCCAA
  151	AAACCGGACG GGCAGGCAAT CCGAAACGAT GCCGTCCGTC
  	GGCTGCAAAC
  201	TTTGGAAGTT TTGAAAAACA GGGCATTGAA GGAAGGTTTG
  	GATAAGGATA
  251	AGGATGTCCA AAACCGCTTT AAAATCGCCG AAGCGTCTTT
  	TTATGCCGAG
  301	GAGTACGTCC GTTTTCTGGA ACGTTCGGAA ACGGTTTCCG
  	AAAGCGCACT
  351	GCGTCAGTTT TATGAGCGGC AAATCCGCAT GATCAAATTG
  	CAGCAGGTCA
  401	GCTTCGCAAC CGAAGAGGAG GCGCGTCAGG CGCAGCAGCT
  	CCTGCTCAAA
  451	GGGCTGTCTT TTGAAGGGCT GATGAAGCGT TATCCGAACG
  	ACGAGCAGGC
  501	TTTTGACGGT TTCATTATGG CGCAGCAGCT TCCCGAGCCG
  	CTGGCTTCGC
  551	AGTTTGCAGC GATGAATCGG GGCGACGTTA CCCGCGATCC
  	GGTCAAATTG
  601	GGCGAACGCT ATTATCTGTT CAAACTCAGC GAGGTCGGGA
  	AAAACCCCGA
  651	CGCGCAGCCT TTCGAGTTGG TCAGAAACCA GTTGGAACAA
  	GGTTTGAGAC
  701	AGGAAAAAGC CCGCTTGAAA ATCGATGCCA TTTTGGAAGA
  	AAACGGTGTC
  751	AAACCGTAA

• This encodes a protein having amino acid sequence <SEQ ID 300>:

• 1	MKQKKTAAAV IAAMLAGFAA AKAPEIDPAL VDTLVAQIMQ
  	QADRHAEQSQ
  51	KPDGQAIRND AVRRLQTLEV LKNRALKEGL DKDKDVQNRF
  	KIAEASFYAE
  101	EYVRFLERSE TVSESALRQF YERQIRMIKL QQVSFATEEE
  	ARQAQQLLLK
  151	GLSFEGLMKR YPNDEQAFDG FIMAQQLPEP LASQFAAMNR
  	GDVTRDPVKL
  201	GERYYLFKLS EVGKNPDAQP FELVRNQLEQ GLRQEKARLK
  	IDAILEENGV
  251	KP*

• ORF76a and ORF76-1 show 97.6% identity in 252 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• The aligned aa sequences of ORF76 and a predicted ORF (ORF76.ng) from N. gonorrhoeae of the N- and C-termini show 96.7% and 100% identity in 30 and 31 overlap, respectively:
• 
• The complete length ORF76ng nucleotide sequence <SEQ ID 301> is:

• 1	ATGAAACAGA AAAAGACCGC TGCCGCAGTT ATTGCTGCAA
  	TGTTGGCAGG
  51	TTTTGCGGCA GCCAAAGCAC CCGAAATCGA CCCGGCTTTG
  	GTGGATACGC
  101	TGGTGGCGCA GATCATGCAG CAGGCAGACC GGCATGCGGA
  	GCAGTCCCAA
  151	AGACCGGACG GGCAGGCAAT CCGAAACGAT GCCGTCCGCC
  	GGCTGCAAAC
  201	TTTGGAAGTT TTGAAAAACA GGGCATTGAA GGAAGGTTTG
  	GATAAGGATA
  251	AGGATGTCCA AAACCGCTTT AAAATCGCCG AAGCGTCTTT
  	TTATGCCGAG
  301	GAGTACGTCC GTTTTCTGGA ACGTTCGGAA ACGGTTTCCG
  	AAAGCGCACT
  351	GCGTCAGTTT TATGAGCGGC AAATCCGCAT GATCAAATTG
  	CAGCAGGTCA
  401	GCTTCGCAAC CGAAGAGGAG GCGCGTCAGG CGCAGCAGCT
  	CCTGCTCAAA
  451	GGGCTGTCTT TTGAAGGGCT GATGAAGCGT TATCCGAACG
  	ACGAGCAGGC
  501	GTTCGACGGT TTCATTATGG CGCAGCAGCT TCCCGAGCCG
  	CTGGCTTcgc
  551	agtttgCCGG TATGAACCGT GGCGACGTTA CCCGCAATCC
  	GGTCAAATTG
  601	GGCGAACGCT ATTACCTGTT CAAACTCGGC GCGGTCGGGA
  	AAAACCCCGA
  651	CGCGCAGCCT TTCGAGTTGG TCAGAAACCA GTTGGAACAA
  	GGTTTGAGGC
  701	AGGAAAAAGC CCGCTTGAAA ATCGATGCCC TTTTGGAaga
  	Aaacggtgtc
  751	AaacCGTAA

• This encodes a protein having amino acid sequence <SEQ ID 302>:

• 1	MKQKKTAAAV IAAMLAGFAA AKAPEIDPAL VDTLVAQIMQ
  	QADRHAEQSQ
  51	RPDGQAIRND AVRRLQTLEV LKNRALKEGL DKDKDVQNRF
  	KIAEASFYAE
  101	EYVRFLERSE TVSESALRQF YERQIRMIKL QQVSFATEEE
  	ARQAQQLLLK
  151	GLSFEGLMKR YPNDEQAFDG FIMAQQLPEP LASQFAGMNR
  	GDVTRNPVKL
  201	GERYYLFKLG AVGKNPDAQP FELVRNQLEQ GLRQEKARLK
  	IDALLEENGV
  251	KP*

• ORF76ng and ORF76-1 show 96.0% identity in 252 aa overlap
• 
• Furthermore, ORF76ng shows significant homology to a B. subtilis export protein precursor:

• sp|P24327|PRSA_BACSU PROTEIN EXPORT PROTEIN PRSA
  PRECURSOR >gi|98227|pir||S15269 33K lipoprotein - Bacillus subtilis
  >gi|39782 (X57271) 33 kDa lipoprotein [Bacillus subtilis]
  >gi|2226124|gnl|PID|e325181 (Y14077) 33 kDa lipoprotein
  [Bacillus subtilis]
  >gi|2633331|gnl|PID|e1182997 (Z99109) molecular chaperonin
  [Bacillus subtilis]
  Length = 292
  Score = 50.4 bits (118), Expect = 1e−05
  Identities = 48/199 (24%), Positives = 82/199 (41%), Gaps = 32/199 (16%)

  Query:	70	VLKNRALKEGLDK-----DKDVQNRFKIAEASF----------YAEEYVRFLERSETVSE	114
  		VL     ++ LDK     DK++ N+ K  +             Y ++Y++   + E +++
  Sbjct:	53	VLTQLVQEKVLDKKYKVSDKEIDNKLKEYKTQLGDQYTALEKQYGKDYLKEQVKYELLTQ	112
  Query:	115	SA-----------LRQFYERQIRMIKLQQVSFATEEEARQAQQLLLKGLSFEGLMKRYPN	163
  		A           +++++E     I+   +  A ++ A + ++ L KG  FE L K Y
  Sbjct:	113	KAAKDNIKVTDADIKEYWEGLKGKIRASHILVADKKTAEEVEKKLKKGEKFEDLAKEYST	172
  Query:	164	DEQAFDG-----FIMAQQLPEPLASQFAAMNRGDVTRDPVKLGERYYLFKLSEVGKNPDA	218
  		D  A  G     F    Q+ E  +     +  G+V+ DPVK    Y++ K +E     D
  Sbjct:	173	DSSASKGGDLGWFAKEGQMDETFSKAAFKLKTGEVS-DPVKTQYGYHIIKKTEERGKYDD	231
  Query:	219	QPFELVRNQLEQGLRQEKA	237
  		EL    LEQ L    A
  Sbjct:	232	MKKELKSEVLEQKLNDNAA	250

• Based on this analysis, including the presence of a putative leader sequence and a RGD motif in the gonococcal protein, it was predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF76-1 (27.8 kDa) was cloned in the pET vector and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 10A shows the results of affinity purification of the His-fusion protein, Purified His-fusion protein was used to immunise mice, whose sera were used for Western blot (FIG. 10B), ELISA (positive result), and FACS analysis (FIG. 10C). These experiments confirm that ORF76-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 36
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 303>:
• 
• This corresponds to the amino acid sequence <SEQ ID 304; ORF81>:
• 
• Further work revealed the complete nucleotide sequence <SEQ ID 305>:

• 1	ATGAAAAAAT CTTTCCTTAC GCTTGTTCTG TATTCGTCTT
  	TACTTACCGC
  51	CAGCGAAATT GCCTATCGCT TTGTATTTGG GATTGAAACC
  	TTACCGGCGG
  101	CAAAAATTGC GGAAACGTTT GCGCTGACAT TTGTGATTGC
  	TGCGCTGTAT
  151	CTGTTTGCGC GTTATAAGGT GACGCGTTTG TTGATTGCGG
  	TGTTTTTTGC
  201	GTTCAGCATT ATTGCCAACA ATGTGCATTA CGCGGTTTAT
  	CAAAGCTGGA
  251	TGACGGGCAT CAATTATTGG CTGATGCTGA AAGAGGTTAC
  	CGAAGTCGGC
  301	AGCGCGGGTG CGTCGATGTT GGATAAGTTG TGGCTGCCTG
  	TGTTGTGGGG
  351	CGTGTTGGAA GTCATGTTGT TTTGCAGCCT TGCCAAGTTC
  	CGCCGTAAGA
  401	CGCATTTTTC TGCCGATATA CTGTTTGCCT TCCTAATGCT
  	GATGATTTTC
  451	GTGCGTTCGT TCGACACGAA ACAAGAGCAC GGTATTTCGC
  	CCAAACCGAC
  501	ATACAGCCGC ATCAAAGCCA ATTATTTCAG CTTCGGTTAT
  	TTTGTCGGAC
  551	GCGTGTTGCC GTATCAGTTG TTTGATTTAA GCAGGATTCC
  	CGCCTTTAAG
  601	CAGCCTGCTC CAAGCAAAAT CGGGCAGGGC AGTGTTCAAA
  	ATATCGTCCT
  651	GATTATGGGC GAAAGCGAAA GCGCGGCGCA TTTGAAGCTG
  	TTTGGCTACG
  701	GACGCGAAAC TTCGCCGTTT TTAACCCGGC TGTCGCAAGC
  	CGATTTTAAG
  751	CCGATTGTGA AACAAAGTTA TTCCGCAGGC TTTATGACTG
  	CAGTGTCCCT
  801	GCCCAGTTTT TTCAATGCGA TACCGCACGC CAACGGCTTG
  	GAACAAATCA
  851	GCGGCGGCGA TACCAATATG TTCCGCCTCG CCAAAGAGCA
  	GGGCTATGAA
  901	ACGTATTTTT ACAGCGCGCA GGCGGAAAAC GAGATGGCGA
  	TTTTGAACTT
  951	AATCGGTAAG AAATGGATAG ACCATCTGAT TCAGCCGACG
  	CAACTTGGCT
  1001	ACGGCAACGG CGACAATATG CCCGATGAGA AGCTGCTGCC
  	GTTGTTCGAC
  1051	AAAATCAATT TGCAGCAGGG CAAGCATTTT ATCGTGTTGC
  	ACCAACGCGG
  1101	TTCGCACGCC CCATACGGCG CATTGTTGCA GCCTCAAGAT
  	AAAGTATTCG
  1151	GCGAAGCCGA TATTGTGGAT AAGTACGACA ACACCATCCA
  	CAAAACCGAC
  1201	CAAATGATTC AAACCGTATT CGAGCAGCTG CAAAAGCAGC
  	CTGACGGCAA
  1251	CTGGCTGTTT GCCTATACCT CCGATCATGG CCAGTATGTT
  	CGCCAAGATA
  1301	TCTACAATCA AGGCACGGTG CAGCCCGACA GCTATCTCGT
  	GCCGCTAGTG
  1351	TTGTACAGCC CGGATAAGGC CGTGCAACAG GCTGCCAACC
  	AGGCTTTTGC
  1401	GCCTTGCGAG ATTGCCTTCC ATCAGCAGCT TTCAACGTTC
  	CTGATTCACA
  1451	CGTTGGGCTA CGATATGCCG GTTTCAGGTT GTCGCGAAGG
  	CTCGGTAACG
  1501	GGCAACCTGA TTACGGGTGA TGCAGGCAGC TTGAACATTC
  	GCGACGGCAA
  1551	GGCGGAATAT GTTTATCCGC AATGA

• This corresponds to the amino acid sequence <SEQ ID 306; ORF81-1>:

• 1	MKKSFLTLVL YSSLLTASEI AYRFVFGIET LPAAKIAETF
  	ALTFVIAALY
  51	LFARYKVTRL LIAVFFAFSI IANNVHYAVY QSWMTGINYW
  	LMLKEVTEVG
  101	SAGASMLDKL WLPVLWGVLE VMLFCSLAKF RRKTHFSADI
  	LFAFLMLMIF
  151	VRSFDTKQEH GISPKPTYSR IKANYFSFGY FVGRVLPYQL
  	FDLSRIPAFK
  201	QPAPSKIGQG SVQNIVLIMG ESESAAHLKL FGYGRETSPF
  	LTRLSQADFK
  251	PIVKQSYSAG FMTAVSLPSF FNAIPHANGL EQISGGDTNM
  	FRLAKEQGYE
  301	TYFYSAQAEN EMAILNLIGK KWIDHLIQPT QLGYGNGDNM
  	PDEKLLPLFD
  351	KINLQQGKHF IVLHQRGSHA PYGALLQPQD KVFGEADIVD
  	KYDNTIHKTD
  401	QMIQTVFEQL QKQPDGNWLF AYTSDHGQYV RQDIYNQGTV
  	QPDSYLVPLV
  451	LYSPDKAVQQ AANQAFAPCE IAFHQQLSTF LIHTLGYDMP
  	VSGCREGSVT
  501	GNLITGDAGS LNIRDGKAEY VYPQ*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF81 shows 84.7% identity over a 85aa overlap and 99.2% identity over a 121aa overlap with an ORF (ORF81a) from strain A of N. meningitidis:
• 
• The complete length ORF81a nucleotide sequence <SEQ ID 307> is:

• 1	ATGAAAAAAT CCCTTTTCGT TCTCTTTCTG TATTCGTCCC
  	TACTTACTGC
  51	CAGCGAAATT GCTTATCGCT TTGTATTCGG AATTGAAACC
  	TTACCGGCTG
  101	CAAAAATGGC AGAAACGTTT GCGCTGACAT TTGTGATTGC
  	TGCGCTGTAT
  151	CTGTTTGCGC GTTATAAGGC AACGCGTTTG TTGATTGCGG
  	TGTTTTTCGC
  201	GTTCAGCATT ATTGCCAACA ATGTGCATTA CGCGGTTTAT
  	CAAAGCTGGA
  251	TAACGGGCAT TAATTATTGG CTGATGCTGA AAGAGATTAC
  	CGAAGTTGGC
  301	GGCGCAGGGG CGTCGATGTT GGATAAGTTG TGGCTGCCTG
  	CGTTGTGGGG
  351	CGTGTTGGAA GTCATGTTGT TTTGCAGCCT TGCCAAGTTC
  	CGCCGTAAGA
  401	CGCATTTTTC TGCCGATATA CTGTTTGCCT TCCTAATGCT
  	GATGATTTTC
  451	GTGCGTTCGT TCGACACGAA ACAAGAACAC GGTATTTCGC
  	CCAAACCGAC
  501	ATACAGCCGC ATCAAAGCCA ATTATTTCAG CTTCGGTTAT
  	TTTGTCGGAC
  551	GCGTGTTGCC GTATCAGTTG TTTGATTTAA GCAAGATTCC
  	TGTGTTCAAA
  601	CAGCCTGCTC CAAGCAGAAT CGGGCAAGGC AGTATTCAAA
  	ATATCGTCCT
  651	GATTATGGGC GAAAGCGAAA GCGCGGCGCA TTTGAAATTG
  	TTTGGCTACG
  701	GGCGCGAAAC TTCGCCGTTT TTGACCCAGC TTTCGCAAGC
  	CGATTTTAAG
  751	CCGATTGTGA AACAAAGTTA TTCCGCAGGC TTTATGACGG
  	CAGTATCCCT
  801	GCCCAGTTTC TTTAACGTCA TACCGCATGC CAACGGCTTG
  	GAACAAATCA
  851	GCGGCGGCGA TATTGTGGAT AAGTACGACA ACACCATCCA
  	CAAAACCGAC
  901	CAAATGATTC AAACCGTATT CGAGCAGCTG CAAAAGCAGC
  	CTGACGGCAA
  951	CTGGCTGTTT GCCTATACCT CCGATCATGG CCAGTATGTT
  	CGCCAAGATA
  1001	TCTACAATCA AGGCACGGTG CAGCCCGACA GCTATCTCGT
  	GCCGCTGGTG
  1051	TTGTACAGCC CGGATAAGGC CGTGCAACAG GCTGCCAACC
  	AGGCTTTTGC
  1101	GCCTTGCGAG ATTGCCTTCC ATCAGCAGCT TTCAACGTTC
  	CTGATTCACA
  1151	CGTTGGGCTA CGATATGCCG GTTTCAGGTT GTCGCGAAGG
  	CTCGGTAACG
  1201	GGCAACCTGA TTACGGGTGA TGCAGGCAGC TTGAACATTC
  	GCGACGGCAA
  1251	GGCGGAATAT GTTTATCCGC AATGA

• This encodes a protein having amino acid sequence <SEQ ID 308>:

• 1	MKKSLFVLFL YSSLLTASEI AYRFVFGIET LPAAKMAETF
  	ALTFVIAALY
  51	LFARYKATRL LIAVFFAFSI IANNVH YAVY QSWITGINYW
  	LMLKEITEVG
  101	GAGASMLDKL WLPALWGVLE VMLFCSLAKF RRKTHFSADI
  	LFAFLMLMIF
  151	VRSFDTKQEH GISPKPTYSR IKANYFSFGY FVGRVLPYQL
  	FDLSKIPVFK
  201	QPAPSRIGQG SIQNIVLIMG ESESAAHLKL FGYGRETSPF
  	LTQLSQADFK
  251	PIVKQSYSAG FMTAVSLPSF FNVIPHANGL EQISGGDIVD
  	KYDNTIHKTD
  301	QMIQTVFEQL QKQPDGNWLF AYTSDHGQYV RQDIYNQGTV
  	QPDSYLVPLV
  351	LYSPDKAVQQ AANQAFAPCE IAFHQQLSTF LIHTLGYDMP
  	VSGCREGSVT
  401	GNLITGDAGS LNIRDGKAEY VYPQ*

• ORF81a and ORF81-1 show 77.9% identity in 524 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• The aligned aa sequences of ORF81 and a predicted ORF (ORF81.ng) from N. gonorrhoeae of the N- and C-termini show 82.4% and 97.5% identity in 85 and 121 overlap, respectively:
• 
• The complete length ORF81ng nucleotide sequence <SEQ ID 309> is:

• 1	ATGAAAAAAT CCCTTTTCGT TCTCTTTCTG TATTCATCCC
  	TACTTACCGC
  51	CAGCGAAATC GCCTATCGCT TTGTATTCGG AATTGAAACC
  	TTACCGGCTG
  101	CAAAAATGGC GGAAACGTTT GCGCTGACAT TTATGATTGC
  	TGCGCTGTAT
  151	CTGTTTGCGC GTTATAAGGC TTCGCGGCTG CTGATTGCGG
  	TGTTTTTCGC
  201	GTTCAGCATG ATTGCCAACA ATGTGCATTA CGCGGTTTAT
  	CAAAGCTGGA
  251	TGACGGGTAT TAACTATTGG CTGATGCTGA AAGAGGTTAC
  	CGAAGTCGGC
  301	AGCGCGGGCG CGTCGATGTT GGATAAGTTG TGGCTGCCTG
  	CTTTGTGGGG
  351	CGTGGCGGAA GTCATGTTGT TTTGCAGCCT TGCCAAGTTC
  	CGCCGTAAGA
  401	CGCATTTTTC TGCCGATATA CTGTTTGCCT TCCTAATGCT
  	GATGATTTTC
  451	GTGCGTTCGT TCGACACGAA ACAAGAGCAC GGTATTTCGC
  	CCAAACCGAC
  501	ATACAGCCGC ATCAAAGCCA ATTATTTCAG CTTCGGTTAT
  	TTTGTCGGGC
  551	GCGTGTTGCC GTATCAGTTG TTTGATTTAA GCAAGATCCC
  	TGTGTTCAAA
  601	CAGCCTGCTC CAAGCAAAAT CGGGCAAGGC AGTATTCAAA
  	ATATCGTCCT
  651	GATTATGGGC GAAAGCGAAA GCGCGGCGCA TTTGAAATTG
  	TTTGGTTACG
  701	GGCGCGAAAC TTCGCCGTTT TTAACCCGGC TGTCGCAAGC
  	CGATTTTAAG
  751	CCGATTGTGA AACAAAGTTA TTCCGCAGGC TTTATGACGG
  	CAGTATCCCT
  801	GCCCAGTTTC TTTAACGTCA TACCGCACGC CAACGGCTTG
  	GAACAAATCA
  851	GCGGCGGCGA TACCAATATG TTCCGCCTCG CCAAAGAGCA
  	GGGCTATGAA
  901	ACGTATTTTT ACAGTGCCCA GGCTGAAAAC CAAATGGCAA
  	TTTTGAACTT
  951	AATCGGTAAG AAATGGATAG ACCATCTGAT TCAGCCGACG
  	CAACTTGGCT
  1001	ACGGCAACGG CGACAATATG CCCGATGAGA AGCTGCTGCC
  	GTTGTTCGAC
  1051	AAAATCAATT TGCAGCAGGG CAGGCATTTT ATCGTGTTGC
  	ACCAACGCGG
  1101	TTCGCACGCC CCATACGGCG CATTGTTGCA GCCTCAAGAT
  	AAAGTATTCG
  1151	GCGAAGCCGA TATTGTGGAT AAGTACGACA ACACCATCCA
  	CAAAACCGAC
  1201	CAAATGATTC AAACCGTATT CGAGCAGCTG CAAAAGCAGC
  	CTGACGGCAA
  1251	CTGGCTGTTT GCCTATACCT CCGATCATGG CCAGTATGTG
  	CGCCAAGATA
  1301	TCTACAATCA AGGCACGGTG CAGCCCGACA GCTATATTGT
  	GCCTCTGGTT
  1351	TTGTACAGCC CGGATAAGGC CGTGCAACAG GCTGCCAACC
  	AGGCTTTTGC
  1401	GCCTTGCGAG ATTGCCTTCC ATCAGCAGCT TTCAACGTTC
  	CTGATTCACA
  1451	CGTTGGGCTA CGATATGCCG GTTTCAGGTT GTCGCGAAGG
  	CTCGGTAACA
  1501	GGCAACCTGA TTACGGGCGA TGCAGGCAGC TTGAACATTC
  	GCAACGGCAA
  1551	GGCGGAATAT GTTTATCCGC AATAA

• This encodes a protein having amino acid sequence <SEQ ID 310>:

• 1	MKKSLFVLFL YSSLLTASEI AYRFVFGIET LPAAKMAETF
  	ALTFMIAALY
  51	LFARYKASRL LIAVFFAFSM IANNVH YAVY QSWMTGINYW
  	LMLKEVTEVG
  101	SAGASMLDKL WLPALWGVAE VMLFCSLAKF RRKTHFSADI
  	LFAFLMLMIF
  151	VRSFDTKQEH GISPKPTYSR IKANYFSFGY FVGRVLPYQL
  	FDLSKIPVFK
  201	QPAPSKIGQG SIQNIVLIMG ESESAAHLKL FGYGRETSPF
  	LTRLSQADFK
  251	PIVKQSYSAG FMTAVSLPSF FNVIPHANGL EQISGGDTNM
  	FRLAKEQGYE
  301	TYFYSAQAEN QMAILNLIGK KWIDHLIQPT QLGYGNGDNM
  	PDEKLLPLFD
  351	KINLQQGRHF IVLHQRGSHA PYGALLQPQD KVFGEADIVD
  	KYDNTIHKTD
  401	QMIQTVFEQL QKQPDGNWLF AYTSDHGQYV RQDIYNQGTV
  	QPDSYIVPLV
  451	LYSPDKAVQQ AANQAFAPCE IAFHQQLSTF LIHTLGYDMP
  	VSGCREGSVT
  501	GNLITGDAGS LNIRNGKAEY VYPQ*

• ORF81ng and ORF81-1 show 96.4% identity in 524 aa overlap:
• 
• Furthermore, ORF81ng shows significant homology to an E. coli OMP:

• gi|1256380 (U50906) outer membrane adherence protein-associated
  protein [E. coli] Length = 547
  Score = 87.4 bits (213), Expect = 2e−16
  Identities = 122/468 (26%), Positives = 198/468 (42%),
  Gaps = 70/468 (14%)

  Query:	25	VFGIETLPAAKMAETFA-LTFMIAALYLFARYKAS--RLLIAVFFAFSMIANNVHYAVYQ	81
  		VFGI  L A+  A     L F +  + +  R  +   RLL+A  F   + A ++  ++Y
  Sbjct:	29	VFGITNLVASSGAHMVQRLLFFVLTILVVKRISSLPLRLLVAAPFVL-LTAADMSISLY-	86
  Query:	82	SWMT-------GINYWLMLKEVTEVGSAGASMLDKLWLPALWGVAEVMLFCSLAKFRRKT	134
  		SW T       G    ++  +  EV    A ML  ++ P L   A + L       +
  Sbjct:	87	SWCTFGTTFNDGFAISVLQSDPDEV----AKMLG-MYSPYLCAFAFLSLLFLAVIIKYDV	141
  Query:	135	HFSADILFAFLMLMIFVRSF---------DTKQEHGISPKPTYSRIKAN--YFSFGYFVG	183
  		+   L+L++   S          D K ++  SP    SR      +F+  YF
  Sbjct:	142	SLPTKKVTGILLLIVISGSLFSACQFAYKDAKNKNAFSPYILASRFATYTPFFNLNYFAL	201
  Query:	184	RVLPYQ--LFDLSKIPVFKQPAPSKIGQGSIQNIVLIMGESESAAHLKLFGYGRETSPFL	241
  		+Q  L   + +P F+      +    I   VLI+GES    ++ L+GY R T+P +
  Sbjct:	202	AAKEHQRLLSIANTVPYFQL----SVRDTGIDTYVLIVGESVRVDNMSLYGYTRSTTPQV	257
  Query:	242	TRLSQADFKPIVKQSYSAGFMTAVSLP---SFFNVIPHANGLEQISGGDTNMFRLAKEQG	298
  		+Q     +  Q+ S    TA+S+P   +  +V+ H      I     N+  +A + G
  Sbjct:	258	E--AQRKQIKLFNQAISGAPYTALSVPLSLTADSVLSH-----DIHNYPDNIINMANQAG	310
  Query:	299	YETYFYSAQA---ENQMAILNLIGKKWIDHLIQPTQLGYGNGDNMPDEKLLPLFDKINLQ	355
  		++T++ S+Q+   +N  A+ ++         ++  +  Y  G    DE LLP   +   Q
  Sbjct:	311	FQTFWLSSQSAFRQNGTAVTSI--------AMRAMETVYVRGF---DELLLPHLSQALQQ	359
  Query:	356	--QGRHFIVLHQRGSHAPYGALLQPQDKVFGEADIVDK-YDNTIHKTDQMIQTVFEQLQK	412
  		Q +  IVLH  GSH P  +       VF   D  D  YDN+IH TD ++  VFE L+
  Sbjct:	360	NTQQKKLIVLHLNGSHEPACSAYPQSSAVFQPQDDQDACYDNSIHYTDSLLGQVFELLK-	418
  Query:	413	QPDGNWLFAYTSDHG---QYVRQDIYNQG--TVQPDSYIVPL-VLYSP	454
  		D      Y +DHG      ++++Y  G       +Y VP+ + YSP
  Sbjct:	419	--DRRASVMYFADHGLERDPTKKNVYFHGGREASQQAYHVPMFIWYSP	464

• Based on this analysis, including the presence of a putative leader sequence (double-underlined) and several putative transmembrane domains (single-underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 37
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 311>:

• 1	...ACCCTGCTCC TCTTCATCCC CCTCGTCCTC ACAC.GTGCG
  	GCACACTGAC
  51	CGGCATACTC GCCCaCGGCG GCGGCAAACG CTTTGCCGTC
  	GAACAAGAAC
  101	TCGTCGCCGC ATCGTCCCGC GCCGCCGTCA AAGAAATGGA
  	TTTGTCCGCC
  151	yTAAAAGGAC GCAAAGCCGC CyTTTACGTC TCCGTTATGG
  	GCGACCAAGG
  201	TTCGGGCAAC ATAAGCGGCG GACGCTACTC TATCGACGCA
  	CTGATACGCG
  251	GCGGCTACCA CAACAACCCC GAAAGTGCCA CCCAATACAG
  	CTACCCCGCC
  301	TACGACACTA CCGCCACCAC CAAATCCGAC GCGCTCTCCA
  	GCGTAACCAC
  351	TTCCACATCG CTTTTGAACG CCCCCGCCGC CGyCyTGACG
  	AAAAACAGCG
  401	GACGCAAAGG CGAACGcTCC GCCGGACTGT CCGTCAACGG
  	CACGGGCGAC
  451	TACCGCAACG AAACCCTGCT CGCCAACCCC CGCGACGTTT
  	CCTTCCTGAC
  501	CAACCTCATC CAAACCGTCT TCTACCTGCG CGGCATCGAA
  	GTCgTACCGC
  551	CCGrATACGC CGACACCGAC GTATTCGTAA CCGTCGACGT
  	A...

• This corresponds to the amino acid sequence <SEQ ID 312; ORF83>:

• 1	..TLLLFIPLVL TXCGTLTGIL AHGGGKRFAV EQELVAASSR
  	AAVKEMDLSA
  51	LKGRKAAXYV SVMGDQGSGN ISGGRYSIDA LIRGGYHNNP
  	ESATQYSYPA
  101	YDTTATTKSD ALSSVTTSTS LLNAPAAXLT KNSGRKGERS
  	AGLSVNGTGD
  151	YRNETLLANP RDVSFLTNLI QTVFYLRGIE VVPPXYADTD
  	VFVTVDV..

• Further work revealed the complete nucleotide sequence <SEQ ID 313>:

• 1	ATGAAAACCC TGCTCCTCCT CATCCCCCTC GTCCTCACAG
  	CCTGCGGCAC
  51	ACTGACCGGC ATACCCGCCC ACGGCGGCGG CAAACGCTTT
  	GCCGTCGAAC
  101	AAGAACTCGT CGCCGCATCG TCCCGCGCCG CCGTCAAAGA
  	AATGGATTTG
  151	TCCGCCCTAA AAGGACGCAA AGCCGCCCTT TACGTCTCCG
  	TTATGGGCGA
  201	CCAAGGTTCG GGCAACATAA GCGGCGGACG CTACTCTATC
  	GACGCACTGA
  251	TACGCGGCGG CTACCACAAC AACCCCGAAA GTGCCACCCA
  	ATACAGCTAC
  301	CCCGCCTACG ACACTACCGC CACCACCAAA TCCGACGCGC
  	TCTCCAGCGT
  351	AACCACTTCC ACATCGCTTT TGAACGCCCC CGCCGCCGCC
  	CTGACGAAAA
  401	ACAGCGGACG CAAAGGCGAA CGCTCCGCCG GACTGTCCGT
  	CAACGGCACG
  451	GGCGACTACC GCAACGAAAC CCTGCTCGCC AACCCCCGCG
  	ACGTTTCCTT
  501	CCTGACCAAC CTCATCCAAA CCGTCTTCTA CCTGCGCGGC
  	ATCGAAGTCG
  551	TACCGCCCGA ATACGCCGAC ACCGACGTAT TCGTAACCGT
  	CGACGTATTC
  601	GGCACCGTCC GCAGCCGTAC CGAACTGCAC CTCTACAACG
  	CCGAAACCCT
  651	TAAAGCCCAA ACCAAGCTCG AATATTTCGC CGTTGACCGC
  	GACAGCCGGA
  701	AACTGCTGAT TACCCCTAAA ACCGCCGCCT ACGAATCCCA
  	ATACCAAGAA
  751	CAATACGCCC TTTGGACCGG CCCTTACAAA GTCAGCAAAA
  	CCGTCAAAGC
  801	CTCAGACCGC CTGATGGTCG ATTTCTCCGA CATTACCCCC
  	TACGGCGACA
  851	CAACCGCCCA AAACCGTCCC GACTTCAAAC AAAACAACGG
  	TAAAAAACCC
  901	GATGTCGGCA ACGAAGTCAT CCGCCGCCGC AAAGGAGGAT
  	AA

• This corresponds to the amino acid sequence <SEQ ID 314; ORF83-1>:

• 1	MKTLLLLIPL VLTACGTLTG IPAHGGGKRF AVEQELVAAS
  	SRAAVKEMDL
  51	SALKGRKAAL YVSVMGDQGS GNISGGRYSI DALIRGGYHN
  	NPESATQYSY
  101	PAYDTTATTK SDALSSVTTS TSLLNAPAAA LTKNSGRKGE
  	RSAGLSVNGT
  151	GDYRNETLLA NPRDVSFLTN LIQTVFYLRG IEVVPPEYAD
  	TDVFVTVDVF
  201	GTVRSRTELH LYNAETLKAQ TKLEYFAVDR DSRKLLITPK
  	TAAYESQYQE
  251	QYALWTGPYK VSKTVKASDR LMVDFSDITP YGDTTAQNRP
  	DFKQNNGKKP
  301	DVGNEVIRRR KGG*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF83 shows 96.4% identity over a 197aa overlap with an ORF (ORF83a) from strain A of N. meningitidis:
• 
• The complete length ORF83a nucleotide sequence <SEQ ID 315> is:

• 1	ATGAAAACCC TGCTCNTCCT CATCCCCCTC GTCCTCACAG
  	CCTGCGGCAC
  51	ACTGACCGGC ATACCCGCCC ACGGCGGCGG CAAACGCTTT
  	GCCGTCGAAC
  101	AAGAACTCGT CGCCGCATCG TCCCGCGCCG CCGTCAAAGA
  	AATGGACTTG
  151	TCCGCCCTGA AAGGACGCAA AGCCGCCCTT TACGTCTCCG
  	TTATGGGCGA
  201	CCAAGGTTCG GGCAACATAA GCGGCGGACG CTACTCTATC
  	GACGCACTGA
  251	TACGCGGCGG CTACCACAAC AACCCCGAAA GTGCCACCCA
  	ATACAGCTAC
  301	CCCGCCTACG ACACTACCGC CACCACCAAA TCCGACGCGC
  	TCTCCAGCGT
  351	AACCACTTCC ACATCGCTTT TGAACGCCCC CGCCGCCGCC
  	CTGACGAAAA
  401	ACAGCGGACG CAAAGGCGAA CGCTCCGCCG GACTGTCCGT
  	CAACGGCACG
  451	GGCGACTACC GCAACGAAAC CCTGCTCGCC AACCCCCGCG
  	ACGTTTCCTT
  501	CCTGACCAAC CTCATCCAAA CCGTCTTCTA CCTGCGCGGC
  	ATCGAAGTCG
  551	TACCGCCCGA ATACGCCGAC ACCGACGTAT TCGTAACCGT
  	CGACGTATTC
  601	GGCACCGTCC GCAGCCGCAC CGAACTGCAC CTCTACAACG
  	CCGAAACCCT
  651	TAAAGCCCAA ACCAAGCTCG AATATTTCGC CGTTGACCGC
  	GACAGCCGGA
  701	AACTGCTGAT TGCCCCTAAA ACCGCCGCCT ACGAATCCCA
  	ATACCAAGAA
  751	CAATACGCCC TCTGGATGGG ACCTTACAGC GTCGGCAAAA
  	CCGTCAAAGC
  801	CTCAGACCGC CTGATGGTCG ATTTCTCCGA CATCACCCCC
  	TACGGCGACA
  851	CAACCGCCCA AAACCGTCCC GACTTCAAAC AAAACAACGG
  	TAAAAAACCC
  901	GATGTCGGCA ACGAAGTCAT CCGCCGCCGC AAAGGAGGAT
  	AA

• This encodes a protein having amino acid sequence <SEQ ID 316>:

• 1	MKTLLXLIPL VLTACGTLTG IPAHGGGKRF AVEQELVAAS
  	SRAAVKEMDL
  51	SALKGRKAAL YVSVMGDQGS GNISGGRYSI DALIRGGYHN
  	NPESATQYSY
  101	PAYDTTATTK SDALSSVTTS TSLLNAPAAA LTKNSGRKGE
  	RSAGLSVNGT
  151	GDYRNETLLA NPRDVSFLTN LIQTVFYLRG IEVVPPEYAD
  	TDVFVTVDVF
  201	GTVRSRTELH LYNAETLKAQ TKLEYFAVDR DSRKLLIAPK
  	TAAYESQYQE
  251	QYALWMGPYS VGKTVKASDR LMVDFSDITP YGDTTAQNRP
  	DFKQNNGKKP
  301	DVGNEVIRRR KGG*

• ORF83a and ORF83-1 show 98.4% identity in 313 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF83 shows 94.9% identity over a 197aa overlap with a predicted ORF (ORF83.ng) from N. gonorrhoeae:
• 
• The complete length ORF83ng nucleotide sequence <SEQ ID 317> is:

• 1	ATGAAAACCC TGCTCCTCCT CATCCCCCTC GTACTCACCG
  	CCTGCGGCAC
  51	ACTGACCGGC ATACCCGCCC ACGGCGGCGG CAAACGCTTT
  	GCCGTCGAAC
  101	AGGAACTCGT CGCCGCATCG TCCCGCGCCG CCGTCAAAGA
  	AATGGACTTG
  151	TCCGCCCTGA AAGGACGCAA AGCCGCCCTT TACGTCTCCG
  	TTATGGGCGA
  201	CCAAGGTTCG GGCAACATAA GCGGCGGACG CTACTCCATC
  	GACGCACTGA
  251	TACGCGGCGG CTACCACAAC AACCCCGACA GCGCCACCCG
  	ATACAGCTAC
  301	CCCGCCTATG ACACTACCGC CACCACCAAA TCCGACGCGC
  	TCTCCGGCGT
  351	AACCACTTCC ACATCGCTTT TGAACGCCCC CGCCGCCGCC
  	CTGACGAAAA
  401	ACAACGGACG CAAAGGCGAA CGCTCCGCCG GACTGTCCGT
  	CAACGGCACG
  451	GGCGACTACC GCAACGAAAC CCTGCTCGCC AACCCCCGCG
  	ACGTTTCCTT
  501	CCTGACCAAC CTCATCCAAA CCGTCTTCTA CCTGCGCGGC
  	ATCGAAGTCG
  551	TACCGCCCGA ATACGCCGAC ACCGACGTAT TCGTAACCGT
  	CGACGTATTC
  601	GGCACCGTCC GCAGCCGTAC CGAACTGCAC CTCTACAACG
  	CCGAAACCCT
  651	TAAAGCCCAA ACCAAGCTCG AATATTTCGC CGTCGACCGC
  	GACAGCCGGA
  701	AACTGCTGAT TGCCCCTAAA ACCGCCGCCT ACGAATCCCA
  	ATACCAAGAA
  751	CAATACGCCC TCTGGATGGG ACCTTACAGC GTCGGCAAAA
  	CCGTCAAAGC
  801	CTCAGACCGC CTGATGGTCG ATTTCTCCGA CATCACCCCC
  	TACGGCGACA
  851	CAACCGCCCA AAACCGTCCC GACTTCAAAC AAAACAACGG
  	TAAAAACCCC
  901	GATGTCGGCA ACGAAGTCAT CCGCCGCCGC AAAGGAGGAT
  	AA

• This encodes a protein having amino acid sequence <SEQ ID 318>:

• 1	MKTLLLLIPL VLTACGTLTG IPAHGGGKRF AVEQELVAAS
  	SRAAVKEMDL
  51	SALKGRKAAL YVSVMGDQGS GNISGGRYSI DALIRGGYHN
  	NPDSATRYSY
  101	PAYDTTATTK SDALSGVTTS TSLLNAPAAA LTKNNGRKGE
  	RSAGLSVNGT
  151	GDYRNETLLA NPRDVSFLTN LIQTVFYLRG IEVVPPEYAD
  	TDVFVTVDVF
  201	GTVRSRTELH LYNAETLKAQ TKLEYFAVDR DSRKLLIAPK
  	TAAYESQYQE
  251	QYALWMGPYS VGKTVKASDR LMVDFSDITP YGDTTAQNRP
  	DFKQNNGKNP
  301	DVGNEVIRRR KGG*

• ORF83ng and ORF83-1 show 97.1% identity in 313 aa overlap
• 
• Based on this analysis, including the presence of a putative ATP/GTP-binding site motif A (P-loop) in the gonococcal protein (double-underlined) and a putative prokaryotic membrane lipoprotein lipid attachment site (single-underlined), it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 38
• The following DNA sequence, believed to be complete, was identified in N. meningitidis <SEQ ID 319>:

• 1	ATGGCAGAGA TCTGTTTGAT AACCGGCACG CCCGGTTCAG
  	GGAAAACATT
  51	AAAAATGGTT TCCATGATGG CGAATGATGA AATGTTTAAG
  	CCTGATGAAA
  101	AAGCCATACG CCGTAAAGTA TTTACGAACA TAAAAGGCTT
  	GAAAATACCG
  151	CACACCTACA TAGAAACGGA CGCAAAAAAG CTGCCGAAAT
  	CGACAGATGA
  201	GCAGCTTTCG GCGCATGATA TGTACGAATG GATAAAGAAG
  	CCCGAAAATA
  251	TCGGGTCTAT TGTCATTGTA GATGAAGCTC AAGACGTATG
  	GCCGGCACGC
  301	TCGGCAGGTT CAAAAATCCC TGAAAATGTC CAATGGCTGA
  	ATACGCACAG
  351	ACATCAGGGC ATTGATATAT TTGTTTTGAC TCAAGGTCCT
  	AAGCTTCTAG
  401	ATCAAAATCT TAGAACGCTT GTACGGAAAC ATTACCACAT
  	CGCTTCAAAC
  451	AAGATGGGTA TGCGTACGCT TTTAGAATGG AAAATATGCG
  	CGGACGATCC
  501	CGTAAAAATG GCATCAAGCG CATTCTCCAG TATCTATACA
  	CTGGATAAAA
  551	AAGTTTATGA CTTGTAysrr TmmGCGGAAG TTCATACCGT
  	AAATAAGGTC
  601	AAGCGGTCAA AGTGGTTTTA CACTCTGCCa GTAATAGTAT
  	TGCTGATTCC
  651	CGTGTTTGTC GGCCTGTCCT ATAAAATGTT GagCaGTTAC
  	GGAAAAAAAC
  701	aGGAAGAACC CGCAGCACAA GAATCGGCGG CAACAGAACA
  	GCAGGCAGTA
  751	CTTCCGGATA AAACAGAAGG CGAGCCGGTA AATAACGGCA
  	ACCTTACCGC
  801	AGATATGTTT GTTCCGACAT TGTCCGAaAA ACCCGrAAGC
  	AAGCcgaTTT
  851	ATAACGGTGT AAGGCAGGTA AGAACCTTTG AATATATAGC
  	AGGCTGTATA
  901	GAAGGCGGAA GAACCGGATG CGCCTGCTAT TCGCaTCAAG
  	GGACGGCATt
  951	gaAAGAAGTG ACGGaGTTGA TGTGccaAgG aCTATGTaAA
  	AAacGGCTTG
  1001	CCGTTTAACC CaTACAAAGA AGAAAGCCAA GGGCAGGAAG
  	TTCAGCAAAG
  1051	CGCGCAgCAA CATTCGGACA GGGCGcCAAG TTGCCACATT
  	GGGCGGAAAA
  1101	CCGTAGCAGA ACCTAATGTA CGATAATTGG GAAGAACGCG
  	GGAAACCGTT
  1151	TGAAGGAATC GGaCGQGGGC GTGGTCGGAT CGGCAAACTG
  	A

• This corresponds to the amino acid sequence <SEQ ID 320; ORF84>:

• 1	MAEICLITGT PGSGKTLKMV SMMANDEMFK PDEKAIRRKV
  	FTNIKGLKIP
  51	HTYIETDAKK LPKSTDEQLS AHDMYEWIKK PENIGSIVIV
  	DEAQDVWPAR
  101	SAGSKIPENV QWLNTHRHQG IDIFVLTQGP KLLDQNLRTL
  	VRKHYHIASN
  151	KMGMRTLLEW KICADDPVKM ASSAFSSIYT LDKKVYDLYX
  	XAEVHTVNKV
  201	KRSKWFYTLP VIVLLIPVFV GLSYKMLSSY GKKQEEPAAQ
  	ESAATEQQAV
  251	LPDKTEGEPV NNGNLTADMF VPTLSEKPXS KPIYNGVRQV
  	RTFEYIAGCI
  301	EGGRTGCACY SHQGTALKEV TELMCKDYVK NGLPFNPYKE
  	ESQGQEVQQS
  351	AQQHSDRAQV ATLGGKPXQN LMYDNWEERG KPFEGIGGGV
  	VGSAN*

• Further work revealed the complete nucleotide sequence <SEQ ID 321>:

• 1	ATGGCAGAGA TCTGTTTGAT AACCGGCACG CCCGGTTCAG
  	GGAAAACATT
  51	AAAAATGGTT TCCATGATGG CGAATGATGA AATGTTTAAG
  	CCTGATGAAA
  101	ACGGCATACG CCGTAAAGTA TTTACGAACA TAAAAGGCTT
  	GAAAATACCG
  151	CACACCTACA TAGAAACGGA CGCAAAAAAG CTGCCGAAAT
  	CGACAGATGA
  201	GCAGCTTTCG GCGCATGATA TGTACGAATG GATAAAGAAG
  	CCCGAAAATA
  251	TCGGGTCTAT TGTCATTGTA GATGAAGCTC AAGACGTATG
  	GCCGGCACGC
  301	TCGGCAGGTT CAAAAATCCC TGAAAATGTC CAATGGCTGA
  	ATACGCACAG
  351	ACATCAGGGC ATTGATATAT TTGTTTTGAC TCAAGGTCCT
  	AAGCTTCTAG
  401	ATCAAAATCT TAGAACGCTT GTACGGAAAC ATTACCACAT
  	CGCTTCAAAC
  451	AAGATGGGTA TGCGTACGCT TTTAGAATGG AAAATATGCG
  	CGGACGATCC
  501	CGTAAAAATG GCATCAAGCG CATTCTCCAG TATCTATACA
  	CTGGATAAAA
  551	AAGTTTATGA CTTGTACGAA TCAGCGGAAG TTCATACCGT
  	AAATAAGGTC
  601	AAGCGGTCAA AGTGGTTTTA CACTCTGCCA GTAATAGTAT
  	TGCTGATTCC
  651	CGTGTTTGTC GGCCTGTCCT ATAAAATGTT GAGCAGTTAC
  	GGAAAAAAAC
  701	AGGAAGAACC CGCAGCACAA GAATCGGCGG CAACAGAACA
  	GCAGGCAGTA
  751	CTTCCGGATA AAACAGAAGG CGAGCCGGTA AATAACGGCA
  	ACCTTACCGC
  801	AGATATGTTT GTTCCGACAT TGTCCGAAAA ACCCGAAAGC
  	AAGCCGATTT
  851	ATAACGGTGT AAGGCAGGTA AGAACCTTTG AATATATAGC
  	AGGCTGTATA
  901	GAAGGCGGAA GAACCGGATG CGCCTGCTAT TCGCATCAAG
  	GGACGGCATT
  951	GAAAGAAGTG ACGGAGTTGA TGTGCAAGGA CTATGTAAAA
  	AACGGCTTGC
  1001	CGTTTAACCC ATACAAAGAA GAAAGCCAAG GGCAGGAAGT
  	TCAGCAAAGC
  1051	GCGCAGCAAC ATTCGGACAG GGCGCAAGTT GCCACATTGG
  	GCGGAAAACC
  1101	GTAGCAGAAC CTAATGTACG ATAATTGGGA AGAACGCGGG
  	AAACCGTTTG
  1151	AAGGAATCGG CGGGGGCGTG GTCGGATCGG CAAACTGA

• This corresponds to the amino acid sequence <SEQ ID 322; ORF84-1>:

• 1	MAEICLITGT PGSGKTLKMV SMMANDEMFK PDENGIRRKV
  	FTNIKGLKIP
  51	HTYIETDAKK LPKSTDEQLS AHDMYEWIKK PENIGSIVIV
  	DEAQDVWPAR
  101	SAGSKIPENV QWLNTHRHQG IDIFVLTQGP KLLDQNLRTL
  	VRKHYHIASN
  151	KMGMRTLLEW KICADDPVKM ASSAFSSIYT LDKKVYDLYE
  	SAEVHTVNKV
  201	KRSKWFYTLP VIVLLIPVFV GL SYKMLSSY GKKQEEPAAQ
  	ESAATEQQAV
  251	LPDKTEGEPV NNGNLTADMF VPTLSEKPES KPIYNGVRQV
  	RTFEYIAGCI
  301	EGGRTGCACY SHQGTALKEV TELMCKDYVK NGLPFNPYKE
  	ESQGQEVQQS
  351	AQQHSDRAQV ATLGGKP*QN LMYDNWEERG KPFEGIGGGV
  	VGSAN*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF84 shows 93.9% identity over a 395aa overlap with an ORF (ORF84a) from strain A of N. meningitidis:
• 
• The complete length ORF84a nucleotide sequence <SEQ ID 323> is:

• 1	ATGGCAGAGA TCTGTTTGAT AACCGGCACG CCCGGTTCAG
  	GGAAAACATT
  51	AAAAATGGTT TCCATGATGG CAAACGATGA AATGTTTAAG
  	CCGGATGAAA
  101	ACGGCATACG CCGTAAAGTA TTTACGAACA TCAAAGGCTT
  	GAAGATACCG
  151	CACACCTACA TAGAAACGGA CGCGAAAAAG CTGCCGAAAT
  	CGACAGATGA
  201	GCAGCTTTCG GCGCATGATA TGTACGAATG GATAAAGAAG
  	CCCGAAAATA
  251	TCGGGTCTAT TGTCATTGTA GATGAAGCTC AAGACGTATG
  	GCCGGCACGC
  301	TCGGCAGGTT CAAAAATCCC TGAAAATGTC CAATGGCTGA
  	ATACGCACAG
  351	ACATCAGGGC ATTGATATAT TTGTTTTGAC TCAAGGCTCT
  	AAGCTTCTAG
  401	ATCAAAATCT TAGAACGCTT GTACGGAAAC ATTACCACAT
  	CGCTTCAAAC
  451	AAGATGGGTA TGCGTACGCT TTTAGAATGG AAAATATGCG
  	CGGACGATCC
  501	CGTAAAAATG GCATCAAGCG CATTCTCCAG TATCTATACA
  	CTGGATAAAA
  551	AAGTTTATGA CTTGTACGAA TCAGCGGAAG TTCATACCGT
  	AAATAAGGTC
  601	AAGCGGTCAA AATGGTTTTA TACTCTGCCA GTAATAATAT
  	TGCTGATTCC
  651	CGTTTTTGTC GGCCTGTCCT ATAAAATGTT AAGTAGTTAT
  	GGAAAAAAAC
  701	AGGAAGAACC CGCAGCACAA GAATCGGCGG CAACAGAACA
  	TCAGGCAGTA
  751	TTTCAGGATA AAACAGAAGG CGAGCCGGTA AACAACGGTA
  	ACCTTACCGC
  801	AGATATGTTT GTTCCGACAT TGTCCGAAAA ACCCGAAAGC
  	AAGCCGATTT
  851	ATAACGGTGT AAGGCAGGTA AGAACCTTTG AATATATAGC
  	AGGCTGTGTA
  901	GAAGGCGGAA GAACCGGATG CACATGCTAT TCGCATCAAG
  	GGACGGCATT
  951	GAAAGAAATT ACAAAGGAAA TGTGCAAGGA TTACGCAAGA
  	AACGGATTGC
  1001	CGTTTAACCC ATATAAAGAA GAAAGCCAAG GGCGGGATGT
  	CCAGCAAAGT
  1051	GAGCAGCACC ATTCGGACAG ACCGCAAGTT GCCACGTTGG
  	GCGGAAAGCC
  1101	GTGGCAAAAT CTTATGTATG ATAATTGGCA GGAGCGCGGA
  	AAACCGTTTG
  1151	AAGGAATCGG CGGGGGCGTG GTCGGATCGG CAAACTGA

• This encodes a protein having amino acid sequence <SEQ ID 324>:

• 1	MAEICLITGT PGSGKTLKMV SMMANDEMFK PDENGIRRKV
  	FTNIKGLKIP
  51	HTYIETDAKK LPKSTDEQLS AHDMYEWIKK PENIGSIVIV
  	DEAQDVWPAR
  101	SAGSKIPENV QWLNTHRHQG IDIFVLTQGS KLLDQNLRTL
  	VRKHYHIASN
  151	KMGMRTLLEW KICADDPVKM ASSAFSSIYT LDKKVYDLYE
  	SAEVHTVNKV
  201	KRSKWFYTLP VIILLIPVFV GL SYKMLSSY GKKQEEPAAQ
  	ESAATEHQAV
  251	FQDKTEGEPV NNGNLTADMF VPTLSEKPES KPIYNGVRQV
  	RTFEYIAGCV
  301	EGGRTGCTCY SHQGTALKEI TKEMCKDYAR NGLPFNPYKE
  	ESQGRDVQQS
  351	EQHHSDRPQV ATLGGKPWQN LMYDNWQERG KPFEGIGGGV
  	VGSAN*

• ORF84a and ORF84-1 show 95.2% identity in 395 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF84 shows 94.2% identity over a 395aa overlap with a predicted ORF (ORF84.ng) from N. gonorrhoeae:
• 
• The complete length ORF84ng nucleotide sequence <SEQ ID 325> is:

• 1	ATGGCAGAAA TCTGTTTGAT AACCGGCACG CCCGGTTCAG
  	GGAAAACATT
  51	AAAAATGGTT TCCATGATGG CAAACGATGA AATGTTTAAG
  	CCAGATGAAA
  101	ACGGCGTACG CCGTAAAGTA TTTACGAACA TCAAAGGTTT
  	GAAGATACCG
  151	CACACCCACA TAGAAACAGA CGCAAAGAAG CTGCCGAAAT
  	CAACCGATGA
  201	ACAGCTTTCG GCGCATGATA TGTATGAATG GATCAAGAAG
  	CCTGAAAacg
  251	tcggcgCAAT CGTTATTGTC GATGAGGCGC AAGACGTATG
  	GCCCGCACGC
  301	TccgCAGGTT CGAAAATCCC CGAAAACGTC CAATGGCTGA
  	ACACACACAG
  351	GCATCAGGGC ATAGATATAT TTGTATTGAC ACAAGGTCCT
  	AAACTCTTAG
  401	ATCAGAACTT GCGAACATTG GTTAAAAGAC ATTACCACAT
  	TGCGGCCAAC
  451	AAAATGGGTT TGCGTACCCT GCTTGAATGG AAAGTATGCG
  	CGGATGACCC
  501	GGTAAAAATG GCATCAAGTG CATTTTCCAG TATCTACACA
  	CTGGATAAAA
  551	AAGTTTATGA CTTGTACGAA TCCGCAGAAA TTCACACGGT
  	AAACAAAGTC
  601	AAGCGTTCAA AATGGTTTTA TGCATTGCCC GTCATCATAT
  	TATTGATTCC
  651	GCTATTTGTC GGTTTGTCTT ACAAAATGTT GGGCAGTTAC
  	GGAAAAAAAC
  701	AGGAAGAACC CGCAGCACAA GAATCGGCGG CAACAGAACA
  	GCAGGCAGTA
  751	CTTCCGGATA AAACAGAAGG AGAATCGGTG AATAACGGAA
  	ACCTTACGGC
  801	AGATATGTTT GTTCCGACAT TGCCCGAAAA ACCCGAAAGC
  	AAGCCGATTT
  851	ATAACGGTGT AAGGCAGGTA AGGACCTTTG AATATATAGC
  	AGGCTGTATA
  901	GAAGGCGGAA GAACCGGATG CACCTGCTAT TCGCATCAAG
  	GGACGGCATT
  951	GAAAGAAGTG ACGGAGTTGA TGTGCAAGGA CTATGTAAAA
  	AACGGCTTGC
  1001	CGTTTAACCC ATACAAAGAA GAAAGCCAAG GGCAGGAAGT
  	TCAGCAAAGC
  1051	GCGCAGCAAC ATTCGGACAG GGCGCAAGTT GCCACCTTGG
  	GCGGAAAACC
  1101	GCAGCAGAAC CTAATGTACG ACAATTGGGA AGAACGCGGG
  	AAACCGTTTG
  1151	AAGGAATCGG CGGGGGCGTG GTCGGATCGG CAAACTGA

• This encodes a protein having amino acid sequence <SEQ ID 326>:

• 1	MAEICLITGT PGSGKTLKMV SMMANDEMFK PDENGVRRKV
  	FTNIKGLKIP
  51	HTHIETDAKK LPKSTDEQLS AHDMYEWIKK PENVGAIVIV
  	DEAQDVWPAR
  101	SAGSKIPENV QWLNTHRHQG IDIFVLTQGP KLLDQNLRTL
  	VKRHYHIAAN
  151	KMGLRTLLEW KVCADDPVKM ASSAFSSIYT LDKKVYDLYE
  	SAEIHTVNKV
  201	KRSKWFYALP VIILLIPLFV GL SYKMLGSY GKKQEEPAAQ
  	ESAATEQQAV
  251	LPDKTEGESV NNGNLTADMF VPTLPEKPES KPIYNGVRQV
  	RTFEYIAGCI
  301	EGGRTGCTCY SHQGTALKEV TELMCKDYVK NGLPFNPYKE
  	ESQGQEVQQS
  351	AQQHSDRAQV ATLGGKPQQN LMYDNWEERG KPFEGIGGGV
  	VGSAN*

• ORF84ng and ORF84-1 show 95.4% identity in 395 aa overlap:
• 
• Based on this analysis, includng the presence of a putative transmembrane domain (single-underlined) in the gonococcal protein, and a putative ATP/GTP-binding site motif A (P-loop, double-underlined), it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 39
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 327>:

• 1	GTGGTTTTCC TGAATGCCGA CAACGGGATA TTGGTTCAGG
  	ACTTGCCTTT
  51	TGAAGTCAAA CTGAAAAAAT TCCATATCGA TTTTTACAAT
  	ACGGGTATGC
  101	CGCGTGATTT CGCCAGCGAT ATTGAAGTGA CGGACAAGGC
  	AACCGGTGAG
  151	AAACTCGAGC GCACCATCCG CGTGAACCAT CCTTTGACCT
  	TGCACGGCAT
  201	CACGATTTAT CAGGCGAGTT TTGCCGACGG CGGTTCGGAT
  	TTGACATTCA
  251	AGGCGTGGAA TTTGGGTGAT GCTTCGCGCG AGCCTGTCGT
  	GTTGAAGGCA
  301	ACATCCATAC ACCAGTTTCC GTTGGAAATT GGCAAACACA
  	AATATCGTCT
  351	TGAGTTCGAT CAGTTCACTT CTATGAATGT GGAGGACATG
  	AGCGAGGGCG
  401	CGGAACGGGA AAAAAGCCTG AAATCCACGC TGCCCGATGT
  	CCGCGCCGTT
  451	ACTCAGGAAG GTCACAAATA CACCAAT... ..........
  	.....TACCG
  501	TATCCGTGAT GCGCCAGGCC AGGCGGTCGA ATATAAAAAC
  	TATATGCTGC
  551	CGGTTTTGCA GGAACAGGAT TATTTTTGGA TTACCGGCAC
  	GCGCAGCGC.
  601	TTGCAGCAGC AATACCGCTG GCTGCGTATC CCCTTGGACA
  	AGCAGTTGAA
  651	AGCGGACACC TTTATGGCAT TGCGTGAGTT TTTGAAAGAT
  	GGGGAAGGGC
  701	GCAAACGTCT .GTTGCCGAC GCAACCAAAG GCGCACCTGC
  	CGAAATCCGC
  751	GAACAATTCA TGCTGGCTGC GGAAAACACG CTGAACATCT
  	TTGCACAAAA
  801	AGGCTATTTG GGATTGGACG AATTTATTAC GTCCAATATC
  	CCGAAAGAGC
  851	AGCAGGATAA GATGCAGGGC TATTTCTACG AAATGCTTTA
  	CGGCGTGATG
  901	AACGCTGCTT TGGATGAAAC CAT.ACCCGG TACGGCTTGC
  	CCGAATGGCA
  951	GCAGGATGAA GCGCGGAATC GTTTCCTGCT GCACAGTATG
  	GATGCGTACA
  1001	CGGGTTTGAC CGAATATCCC GCGCCTATGC TGCTGCAACT
  	TGATGGGTTT
  1051	TCCGAGGTGC GTTCGTCGGG TTTGCAGATG ACCCGTTCCC
  	C.GGTCCGCT
  1101	TTTGGTCTAT CTC...

• This corresponds to the amino acid sequence <SEQ ID 328; ORF88>:

• 1	MVFLNADNGI LVQDLPFEVK LKKFHIDFYN TGMPRDFASD
  	IEVTDKATGE
  51	KLERTIRVNH PLTLHGITIY QASFADGGSD LTFKAWNLGD
  	ASREPVVLKA
  101	TSIHQFPLEI GKHKYRLEFD QFTSMNVEDM SEGAEREKSL
  	KSTLPDVRAV
  151	TQEGHKYTNX XXXXXYRIRD APGQAVEYKN YMLPVLQEQD
  	YFWITGTRSX
  201	LQQQYRWLRI PLDKQLKADT FMALREFLKD GEGRKRXVAD
  	ATKGAPAEIR
  251	EQFMLAAENT LNIFAQKGYL GLDEFITSNI PKEQQDKMQG
  	YFYEMLYGVM
  301	NAALDETXTR YGLPEWQQDE ARNRFLLHSM DAYTGLTEYP
  	APMLLQLDGF
  351	SEVRSSGLQM TRSXGPLLVY L...

• Further work revealed the complete nucleotide sequence <SEQ ID 329>:

• 1	ATGAGTAAAT CCCGTAGATC TCCCCCACTT CTTTCCCGTC
  	CGTGGTTCGC
  51	TTTTTTCAGC TCCATGCGCT TTGCAGTCGC TTTGCTCAGT
  	CTGCTGGGTA
  101	TTGCATCGGT TATCGGTACG GTGTTGCAGC AAAACCAGCC
  	GCAGACGGAT
  151	TATTTGGTCA AATTCGGATC GTTTTGGGCG CAGATTTTTG
  	GTTTTCTGGG
  201	ACTGTATGAC GTCTATGCTT CGGCATGGTT TGTCGTTATC
  	ATGATGTTTT
  251	TGGTGGTTTC TACCAGTTTG TGCCTGATTC GCAATGTGCC
  	GCCGTTCTGG
  301	CGCGAAATGA AGTCTTTTCG GGAAAAGGTT AAAGAAAAAT
  	CTCTGGCGGC
  351	GATGCGCCAT TCTTCGCTGT TGGATGTAAA AATTGCGCCC
  	GAGGTTGCCA
  401	AACGTTATCT GGAAGTACAA GGTTTTCAGG GAAAAACCAT
  	TAACCGTGAA
  451	GACGGGTCGG TTCTGATTGC CGCCAAAAAA GGCACAATGA
  	ACAAATGGGG
  501	CTATATCTTT GCCCATGTTG CTTTGATTGT CATTTGCCTG
  	GGCGGGTTGA
  551	TAGACAGTAA CCTGCTGTTG AAACTGGGTA TGCTGACCGG
  	TCGGATTGTT
  601	CCGGACAATC AGGCGGTTTA TGCCAAGGAT TTCAAGCCCG
  	AAAGTATTTT
  651	GGGTGCGTCC AATCTCTCAT TTAGGGGCAA CGTCAATATT
  	TCCGAGGGGC
  701	AGAGTGCGGA TGTGGTTTTC CTGAATGCCG ACAACGGGAT
  	ATTGGTTCAG
  751	GACTTGCCTT TTGAAGTCAA ACTGAAAAAA TTCCATATCG
  	ATTTTTACAA
  801	TACGGGTATG CCGCGTGATT TCGCCAGCGA TATTGAAGTG
  	ACGGACAAGG
  851	CAACCGGTGA GAAACTCGAG CGCACCATCC GCGTGAACCA
  	TCCTTTGACC
  901	TTGCACGGCA TCACGATTTA TCAGGCGAGT TTTGCCGACG
  	GCGGTTCGGA
  951	TTTGACATTC AAGGCGTGGA ATTTGGGTGA TGCTTCGCGC
  	GAGCCTGTCG
  1001	TGTTGAAGGC AACATCCATA CACCAGTTTC CGTTGGAAAT
  	TGGCAAACAC
  1051	AAATATCGTC TTGAGTTCGA TCAGTTCACT TCTATGAATG
  	TGGAGGACAT
  1101	GAGCGAGGGC GCGGAACGGG AAAAAAGCCT GAAATCCACG
  	CTGAACGATG
  1151	TCCGCGCCGT TACTCAGGAA GGTAAAAAAT ACACCAATAT
  	CGGCCCTTCC
  1201	ATTGTTTACC GTATCCGTGA TGCGGCAGGG CAGGCGGTCG
  	AATATAAAAA
  1251	CTATATGCTG CCGGTTTTGC AGGAACAGGA TTATTTTTGG
  	ATTACCGGCA
  1301	CGCGCAGCGG CTTGCAGCAG CAATACCGCT GGCTGCGTAT
  	CCCCTTGGAC
  1351	AAGCAGTTGA AAGCGGACAC CTTTATGGCA TTGCGTGAGT
  	TTTTGAAAGA
  1401	TGGGGAAGGG CGCAAACGTC TGGTTGCCGA CGCAACCAAA
  	GGCGCACCTG
  1451	CCGAAATCCG CGAACAATTC ATGCTGGCTG CGGAAAACAC
  	GCTGAACATC
  1501	TTTGCACAAA AAGGCTATTT GGGATTGGAC GAATTTATTA
  	CGTCCAATAT
  1551	CCCGAAAGAG CAGCAGGATA AGATGCAGGG CTATTTCTAC
  	GAAATGCTTT
  1601	ACGGCGTGAT GAACGCTGCT TTGGATGAAA CCATACGCCG
  	GTACGGCTTG
  1651	CCCGAATGGC AGCAGGATGA AGCGCGGAAT CGTTTCCTGC
  	TGCACAGTAT
  1701	GGATGCGTAC ACGGGTTTGA CCGAATATCC CGCGCCTATG
  	CTGCTGCAAC
  1751	TTGATGGGTT TTCCGAGGTG CGTTCGTCGG GTTTGCAGAT
  	GACCCGTTCC
  1801	CCGGGTGCGC TTTTGGTCTA TCTCGGCTCG GTGCTGTTGG
  	TATTGGGTAC
  1851	GGTATTGATG TTTTATGTGC GCGAAAAACG GGCGTGGGTA
  	TTGTTTTCAG
  1901	ACGGCAAAAT CCGTTTTGCC ATGTCTTCGG CCCGCAGCGA
  	ACGGGATTTG
  1951	CAGAAGGAAT TTCCAAAACA CGTCGAGAGT CTGCAACGGC
  	TCGGCAAGGA
  2001	CTTGAATCAT GACTGA

• This corresponds to the amino acid sequence <SEQ ID 330; ORF88-1>:

• 1	MSKSRRSPPL LSRPWFAFFS SMRFAVALLS LLGIASVIGT
  	VLQQNQPQTD
  51	YLVKFGSFWA QIFGFLGLYD VYASAWFVVI MMFLVVSTSL
  	CLIRNVPPFW
  101	REMKSFREKV KEKSLAAMRH SSLLDVKIAP EVAKRYLEVQ
  	GFQGKTINRE
  151	DGSVLIAAKK GTMNKWGYIF AHVALIVICL GGLIDSNLLL
  	KLGMLTGRIV
  201	PDNQAVYAKD FKPESILGAS NLSFRGNVNI SEGQSADVVF
  	LNADNGILVQ
  251	DLPFEVKLKK FHIDFYNTGM PRDFASDIEV TDKATGEKLE
  	RTIRVNHPLT
  301	LHGITIYQAS FADGGSDLTF KAWNLGDASR EPVVLKATSI
  	HQFPLEIGKH
  351	KYRLEFDQFT SMNVEDMSEG AEREKSLKST LNDVRAVTQE
  	GKKYTNIGPS
  401	IVYRIRDAAG QAVEYKNYML PVLQEQDYFW ITGTRSGLQQ
  	QYRWLRIPLD
  451	KQLKADTFMA LREFLKDGEG RKRLVADATK GAPAEIREQF
  	MLAAENTLNI
  501	FAQKGYLGLD EFITSNIPKE QQDKMQGYFY EMLYGVMNAA
  	LDETIRRYGL
  551	PEWQQDEARN RFLLHSMDAY TGLTEYPAPM LLQLDGFSEV
  	RSSGLQMTRS
  601	PGALLVYLGS VLLVLGTVLM FYVREKRAWV LFSDGKIRFA
  	MSSARSERDL
  651	QKEFPKHVES LQRLGKDLNH D*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF88 shows 95.7% identity over a 371aa overlap with an ORF (ORF88a) from strain A of N. meningitidis:
• 
• The complete length ORF88a nucleotide sequence <SEQ ID 331> is:

• 1	ATGAGTAAAT CCCGTAGATC TCCCCCACTT CTTTCCCGTC
  	CGTGGTTCGC
  51	TTTTTTCAGC TCCATGCGCT TTGCGGTCGC TTTGCTCAGT
  	CTGCTGGGTA
  101	TTGCATCGGT TATCGGTACG GTGTTGCAGC AAAACCAGCC
  	GCAGACGGAT
  151	TATTTGGTCA AATTCGGATC GTTTTGGGCG CAGATTTTTG
  	GTTTTCTGGG
  201	ACTGTATGAC GTCTATGCTT CGGCATGGTT TGTCGTTATC
  	ATGATGTTTT
  251	TGGTGGTTTC TACCAGTTTG TGCCTGATTC GCAATGTGCC
  	GCCGTTCTGG
  301	CGCGAAATGA AGTCTTTTCG GGAAAAGGTT AAAGAAAAAT
  	CTCTGGCGGC
  351	GATGCGCCAT TCTTCGCTGT TGGATGTAAA AATTGCGCCC
  	GAGGTTGCCA
  401	AACGTTATCT GGAAGTACAA GGTTTTCAGG GAAAAACCAT
  	TAACCGTGAA
  451	GACGGGTCGG TTCTGATTGC CGCCAAAAAA GGCACAATGA
  	ACAAATGGGG
  501	CTATATCTTT GCCCATGTTG CTTTGATTGT CATTTGCCTG
  	GGCGGGTTGA
  551	TAGACAGTAA CCTGCTGTTG AAACTGGGTA TGCTGACCGG
  	TCGGATTGTT
  601	CCGGACAATC AGGCGGTTTA TGCCAAGGAT TTCAAGCCCG
  	AAAGTATTTT
  651	GGGTGCGTCC AATCTCTCAT TTAGGGGCAA CGTCAATATT
  	TCCGAGGGGC
  701	AGAGTGCGGA TGTGGTTTTC CTGAATGCCG ACAACGGGAT
  	ATTGGTTCAG
  751	GACTTGCCTT TTGAAGTCAA ACTGAAAAAA TTCCATATCG
  	ATTTTTACAA
  801	TACGGGTATG CCGCGCGATT TTGCCAGTGA TATTGAAGTA
  	ACGGATAAGG
  851	CAACCGGTGA GAAACTCGAG CGCACCATCC GCGTGAACCA
  	TCCTTTGACC
  901	TTGCACGGCA TCACGATTTA TCAGGCGAGT TTTGCCGACG
  	GCGGTTCGGA
  951	TTTGACATTC AAGGCGTGGA ATTTGGGTGA TGCTTCGCGC
  	GAGCCTGTCG
  1001	TGTTGAAGGC AACATCCATA CACCAGTTTC CGTTGGAAAT
  	TGGCAAACAC
  1051	AAATATCGTC TTGAGTTCGA TCAGTTTACT TCTATGAATG
  	TGGAGGACAT
  1101	GAGCGAGGGC GCGGAACGGG AAAAAAGCCT GAAATCCACG
  	CTGAACGATG
  1151	TCCGCGCCGT TACTCAGGAA GGTAAAAAAT ACACCAATAT
  	CGGCCCTTCC
  1201	ATTGTTTACC GTATCCGTGA TGCGGCAGGG CAGGCGGTCG
  	AATATAAAAA
  1251	CTATATGCTG CCGGTTTTGC AGGAACAGGA TTATTTTTGG
  	ATTACCGGCA
  1301	CGCGCAGCGG CTTGCAGCAG CAATACCGCT GGCTGCGTAT
  	CCCCTTGGAC
  1351	AAGCAGTTGA AAGCGGACAC CTTTATGGCA TTGCGTGAGT
  	TTTTGAAAGA
  1401	TGGGGAAGGG CGCAAACGTC TGGTTGCCGA CGCAACCAAA
  	GGCGCACCTG
  1451	CCGAAATCCG CGAACAATTC ATGCTGGCTG CGGAAAACAC
  	GCTGAACATC
  1501	TTTGCACAAA AAGGCTATTT GGGATTGGAC GAATTTATTA
  	CGTCCAATAT
  1551	CCCGAAAGAG CAGCAGGATA AGATGCAGGG CTATTTCTAC
  	GAAATGCTTT
  1601	ACGGCGTGAT GAACGCTGCT TTGGATGAAA CCATACGCCG
  	GTACGGCTTG
  1651	CCCGAATGGC AGCAGGATGA AGCGCGGAAT CGTTTCCTGC
  	TGCACAGTAT
  1701	GGATGCGTAC ACGGGTTTGA CCGAATATCC CGCGCCTATG
  	CTGCTGCAAC
  1751	TTGATGGGTT TTCCGAGGTG CGTTCGTCGG GTTTGCAGAT
  	GACCCGTTCC
  1801	CCGGGTGCGC TTTTGGTCTA TCTCGGCTCG GTGCTGTTGG
  	TATTGGGTAC
  1851	GGTATTGATG TTTTATGTGC GCGAAAAACG GGCGTGGGTA
  	TTGTTTTCAG
  1901	ACGGCAAAAT CCGTTTTGCC ATGTCTTCGG CCCGCAGCGA
  	ACGGGATTTG
  1951	CAGAAGGAAT TTCCAAAACA CGTCGAGAGT CTGCAACGGC
  	TCGGCAAGGA
  2001	CTTGAATCAT GACTGA

• This encodes a protein having amino acid sequence <SEQ ID 332>:

• 1	MSKSRRSPPL LSRPWFAFFS SMRFAVALLS LLGIASVIGT
  	VLQQNQPQTD
  51	YLVKFGSFWA QIFGFLGLYD VYASAWFVVI MMFLVVSTSL
  	CLIRNVPPFW
  101	REMKSFREKV KEKSLAAMRH SSLLDVKIAP EVAKRYLEVQ
  	GFQGKTINRE
  151	DGSVLIAAKK GTMNKWGYIF AHVALIVICL GGLIDSNLLL
  	KLGMLTGRIV
  201	PDNQAVYAKD FKPESILGAS NLSFRGNVNI SEGQSADVVF
  	LNADNGILVQ
  251	DLPFEVKLKK FHIDFYNTGM PRDFASDIEV TDKATGEKLE
  	RTIRVNHPLT
  301	LHGITIYQAS FADGGSDLTF KAWNLGDASR EPVVLKATSI
  	HQFPLEIGKH
  351	KYRLEFDQFT SMNVEDMSEG AEREKSLKST LNDVRAVTQE
  	GKKYTNIGPS
  401	IVYRIRDAAG QAVEYKNYML PVLQEQDYFW ITGTRSGLQQ
  	QYRWLRIPLD
  451	KQLKADTFMA LREFLKDGEG RKRLVADATK GAPAEIREQF
  	MLAAENTLNI
  501	FAQKGYLGLD EFITSNIPKE QQDKMQGYFY EMLYGVMNAA
  	LDETIRRYGL
  551	PEWQQDEARN RFLLHSMDAY TGLTEYPAPM LLQLDGFSEV
  	RSSGLQMTRS
  601	PGALLVYLGS VLLVLGTVLM FYVREKRAWV LFSDGKIRFA
  	MSSARSERDL
  651	QKEFPKHVES LQRLGKDLNH D*

• ORF88a and ORF88-1 100.0% identity in 671 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF88 shows 93.8% identity over a 371aa overlap with a predicted ORF (ORF88.ng) from N. gonorrhoeae:
• 
• An ORF88ng nucleotide sequence <SEQ ID 333> was predicted to encode a protein having amino acid sequence <SEQ ID 334>:

• 1	MVFLNADNGM LVQDLPFEVK LKKFHIDFYN TGMPRDFASD
  	IEVTDKATGE
  51	KLERTIRVNH PLTLHGITIY QASFADGGSD LTFKAWNLRD
  	ASREPVVLKA
  101	TSIHQFPLEI GKHKYRLEFD QFTSMNVEDM SEGAEREKSL
  	KSTLNDVRAV
  151	TQEGKKYTNI GPSIVYRIRD AAGQAVEYKN YMLPILQDKD
  	YFWLTGTRSG
  201	LQQQYRWLRI PLDKQLKADT FMALREFLKD GEGRKRLVAD
  	ATKDAPAEIR
  251	EQFMLAAENT LNIFAQKGYL GLDEFITSNI PKGQQDKMQG
  	YFYEMLYGVM
  301	NAALDETIRR YGLPEWQQDE ARNRFLLHSM DAYTGLTEYP
  	APMLLQLDGF
  351	SEVRSSGLQM TRSPGALLVY LGSVLLVLGT VFMFYVPKKR
  	AWVLFSNXKI
  401	RFAMSSARSE RDLQKEFPKH VESLQRLGKD LNHD*

• Further work revealed the complete gonococcal DNA sequence <SEQ ID 335>:

• 1	ATGAGTAAAT CCCGTATATC TCCCACACTT CTTTCCCGTC
  	CGTGGTTCGC
  51	TTTTTTCAGC TCCATGCGCT TTGCGGTCGC TTTGCTCAGT
  	CTGCTGGGTA
  101	TTGCATCGGT TATCGGCACG GTGTTACAGC AAAACCAGCC
  	GCAGACGGAT
  151	TATTTGGTCA AATTCGGACC GTTTTGGACT CGGATTTTTG
  	ATTTTTTGGG
  201	TTTGTATGAT GTCTATGCTT CGGCATGGTT TGTCGTTATC
  	ATGATGTTTC
  251	TGGTGGTTTC TACCAGTTTG TGTTTAATCC GTAACGTTCC
  	GCCGTTTTGG
  301	CGCGAAATGA AGTCTTTCCG GGAAAAGGTT AAAGAAAAAT
  	CTCTGGCGGC
  351	GATGCGCCAT TCTTCGCTGT TGGATGTAAA AATTGCCCCC
  	GAAGTTGCCA
  401	AACGTTATCT GGAGGTGCGG GGTTTTCAGG GAAAAACCGT
  	CAGCCGTGAG
  451	GACGGGTCGG TTCTGATTGC CGCCAAAAAA GGCAcaatga
  	acaaATGGGG
  501	CTATATCTTT GCccaagtag ctTTGATTGT CATTTGCCTG
  	GGCGGGTTGA
  551	TAGACAGTAA CCTGCTGCTG AAGCTGGGTA TGCTGGCCGG
  	TCGGATTGTT
  601	CCGGACAATC AGGCGGTTTA TGCCAAGGAT TTCAAGCCCG
  	AAAGTATTTT
  651	GGGTGCGTCC AATCTCTCAT TTAGGGGCAA CGTCAATATT
  	TCCGAGGGGC
  701	AAAGTGCGGA TGTGGTTTTC CTGAATGCCG ACAACGGGAT
  	GTTGGTTCAG
  751	GACTTGCCTT TTGAAGTCAA ACTGAAAAAA TTCCATATCG
  	ATTTTTACAA
  801	TACGGGTATG CCGCGCGATT TTGCCAGCGA TATTGAAGTA
  	ACGGACAAGG
  851	CAACCGGTGA GAAACTCGAG CGCACCATCC GCGTGAACCA
  	TCCTTTGACC
  901	TTGCACGGCA TCACGATTTA TCAGGCGAGT TTTGCCGACG
  	GCGGTTCGGA
  951	TTTGACATTC AAGGCGTGGA ATTTGAGGGA TGCTTCGCGC
  	GAACCTGTCG
  1001	TGTTGAAGGC AACCTCCATA CACCAGTTTC CGTTGGAAAT
  	CGGCAAACAC
  1051	AAATATCGTC TTGAGTTCGA TCAGTTCACT TCTATGAATG
  	TGGAGGACAT
  1101	GAGCGAGGGT GCGGAACGGG AAAAAAGCCT GAAATCCACT
  	CTGAACGATG
  1151	TCCGCGCCGT TACTCAGGAA GGTAAAAAAT ACACCAATAT
  	CGGCCCTTCC
  1201	ATCGTGTACC GCATCCGTGA TGcggCAGGG CAGGCGGTCG
  	AATATAAAAA
  1251	CTATATGCTG CCGATTTTGC AGGACAAAGA TTATTTTTGG
  	CTGACCGGCA
  1301	CGCGCAGCGG CTTGCAGCAG CAATACCGCT GGCTGCGTAT
  	CCCCTTGGAC
  1351	AAGCAGTTGA AAGCGGACAC CTTTATGGCA TTGCGTGAGT
  	TTTTGAAAGA
  1401	TGGGGAAGGG CGCAAACGTC TGGTTGCCGA CGCAACCAAA
  	GACGCACCTG
  1451	CCGAAATCCG CGAACAATTC ATGCTGGCTG CGGAAAACAC
  	GCTGAATATC
  1501	TTTGCGCAAA AAGGCTATTT GGGATTGGAC GAATTTATTA
  	CGTCCAATAT
  1551	CCCGAAAGGG CAGCAGGATA AGATGCAGGG CTATTTCTAC
  	GAAATGCTTT
  1601	ACGGCGTGAT GAACGCTGCT TTGGATGAAA CCATACGCCG
  	GTACGGCTTG
  1651	CCCGAATGGC AGCAGGATGA AGCGCGGAAC CGTTTCCTGC
  	TGCACAGTAT
  1701	GGATGCCTAT ACGGGGCTGA CGGAATATCC CGCGCCTATG
  	CTGCTCCAGC
  1751	TTGACGGGTT TTCCGAGGTG CGTTCCTCAG GTTTGCAGAT
  	GACCCGTTCG
  1801	CCGGGTGCGC TTTTGGTCTA TCtcggctcg gtattgttgg
  	TTTTGGgtac
  1851	ggtaTttatg tTTTATGTGC GCGAAAAACG GGCGTGGgta
  	tTGTTTTCag
  1901	aCGGCAAAAT CCGTTTTGCT ATGtCTTcgg CCcgcagcga
  	ACGGGATTTG
  1951	cAGAaggaaT TTCCAAAACA CGtcgAGAGC CTGCAACggc
  	tcggcaaggA
  2001	CttgaaTCAT GACTga

• This corresponds to the amino acid sequence <SEQ ID 336; ORF88ng-1>:

• 1	MSKSRISPTL LSRPWFAFFS SMRFAVALLS LLGIASVIGT
  	VLQQNQPQTD
  51	YLVKFGPFWT RIFDFLGLYD VYASAWFVVI MMFLVVSTSL
  	CLIRNVPPFW
  101	REMKSFREKV KEKSLAAMRH SSLLDVKIAP EVAKRYLEVR
  	GFQGKTVSRE
  151	DGSVLIAAKK GTMNKWGYIF AQVALIVICL GGLIDSNLLL
  	KLGMLAGRIV
  201	PDNQAVYAKD FKPESILGAS NLSFRGNVNI SEGQSADVVF
  	LNADNGMLVQ
  251	DLPFEVKLKK FHIDFYNTGM PRDFASDIEV TDKATGEKLE
  	RTIRVNHPLT
  301	LHGITIYQAS FADGGSDLTF KAWNLRDASR EPVVLKATSI
  	HQFPLEIGKH
  351	KYRLEFDQFT SMNVEDMSEG AEREKSLKST LNDVRAVTQE
  	GKKYTNIGPS
  401	IVYRIRDAAG QAVEYKNYML PILQDKDYFW LTGTRSGLQQ
  	QYRWLRIPLD
  451	KQLKADTFMA LREFLKDGEG RKRLVADATK DAPAEIREQF
  	MLAAENTLNI
  501	FAQKGYLGLD EFITSNIPKG QQDKMQGYFY EMLYGVMNAA
  	LDETIRRYGL
  551	PEWQQDEARN RFLLHSMDAY TGLTEYPAPM LLQLDGFSEV
  	RSSGLQMTRS
  601	PGALLVYLGS VLLVLGTVFM FYVREKRAWV LFSDGKIRFA
  	MSSARSERDL
  651	QKEFPKHVES LQRLGKDLNH D*

• ORF88ng-1 and ORF88-1 show 97.0% identity in 671 aa overlap:
• 
• Furthermore, ORG88ng-1 shows homology with a hypothetical protein from Aquifex aeolicus:

• gi|2984296 (AE000771) hypothetical protein [Aquifex aeolicus]
  Length = 537 Score = 94.4 bits (231), Expect = 2e−18.
  Identities = 91/334 (27%), Positives = 159/334 (47%), Gaps = 59/334 (17%)

  Query:	16	FAFFSSMRFAVALLSLLGIASVIG-TVLQQNQPQTDYLVKFGPFWTRIFDFLGLYDVYAS	74
  		+ F +S++ A+ ++ +LGI S++G T ++QNQ    YL +FG         L L DV+ S
  Sbjct:	80	YDFLASLKLAIFIMLVLGILSMLGSTYIKQNQSFEWYLDQFGYDVGIWIWKLWLNDVFHS	139
  Query:	75	AWFVVIMMFLVVSTSLCLIRNVPPFWREMKSFREKVKEKSLAAMRHSSLLDVKIAPEVAK	134
  		++++ ++ L V+   C I+ +P  W++  S +E++ +    A +H   + VKI P+  K
  Sbjct:	140	WYYILFIVLLAVNLIFCSIKRLPRVWKQAFS-KERILKLDEHAEKHLKPITVKI-PDKDK	197
  Query:	135	--RYLEVRGFQGKTVSREDGSVLIAAKKGTMNKWGYIFAQVALIVICLGGLIDSNLLLKL	192
  		++L  +GF+   V  E   + + A+KG  ++ G     +AL+VI  G LID
  Sbjct:	198	VLKFLLKKGFK-VFVEEEGNKLYVFAEKGRFSRLGVYITHIALLVIMAGALID-------	249
  Query:	193	GMLAGRIVPDNQAVYAKDFKPESILGASNLSFRGNVNISEGQSADVVFLNADNGMLVQDL	252
  		+I+G      RG++ ++EG + DV+ + A+       L
  Sbjct:	250	----------------------AIVGV-----RGSLIVAEGDTNDVMLVGAE--QKPYKL	280
  Query:	253	PFEVKLKKFHIDFY---NTGMPRDFA-------SDIEVTDKATGEKLER--TIRVNHPLT	300
  		PF V L  F I  Y   N  + + FA       SDIE+ +   G K+E   T++VN P
  Sbjct:	281	PFAVHLIDFRIKTYAEENPNVDKRFAQAVSSYESDIEIIN---GGKVEAKGTVKVNEPFD	337
  Query:	301	LHGITIYQASFA--DGGSDLTFKAWNLRDASREP	332
  		++QA++   DG S +     + + A  +P
  Sbjct:	338	FGRYRLFQATYGILDGTSGMGVIVVDRKKAHEDP	371

• Based on this analysis, including the putative transmembrane domain in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 40
• The following DNA sequence, believed to be complete, was identified in N. meningitidis <SEQ ID 337>:

• 1	ATGATGAGTA ATAmAATGGm ACAAAAAGGG TTTACATTGA
  	TTGmGmTGAT
  51	GATAGTCGTC GCGATACTCG GCATTATCAG CGTCATTGCC
  	ATACCTTCTT
  101	ATCmAAGTTA TATTGAAAAA GGCTATCAGT CCCAGCTTTA
  	TACGGAGATG
  151	GyCGGTATCA ACAATATTTC CAAACAGTTT ATTTTGAAAA
  	ATCCCCTGGA
  201	CGATAATCAG ACCATCGAGA ACAAACTGGA AATATTTGTC
  	TCAGGCTATA
  251	AGATGAATCC GAAAATTGCC AAAAAaTATA GTGTTTCGGT
  	AAAGTTTGTC
  301	GATAAGGAAA AATCAAGGGC ATACAGGTTG GTCGGCGTTC
  	CGAAGGCGGG
  351	GACGGGTTAT ACTTTGTCGG TATGGATGAA CAGCGTGGGC
  	GACGGATACA
  401	AATGCCGTGA TGCCGCTTCT GCCCAAGCCC ATTTGGAGAC
  	CTTGTCCTCA
  451	GATGTCGGCT GTGAAGCCTT CTCTAATCGT AAAAAATAA

• This corresponds to the amino acid sequence <SEQ ID 338; ORF89>:

• 1	MMSNXMXQKG FTLIXXMIVV AILGIISVIA IPSYXSYIEK
  	GYQSQLYTEM
  51	XGINNISKQF ILKNPLDDNQ TIENKLEIFV SGYKMNPKIA
  	KKYSVSVKFV
  101	DKEKSRAYRL VGVPKAGTGY TLSVWMNSVG DGYKCRDAAS
  	AQAHLETLSS
  151	DVGCEAFSNR KK*

• Further work revealed the complete nucleotide sequence <SEQ ID 339>:

• 1	ATGATGAGTA ATAAAATGGA ACAAAAAGGG TTTACATTGA
  	TTGAGATGAT
  51	GATAGTCGTC GCGATACTCG GCATTATCAG CGTCATTGCC
  	ATACCTTCTT
  101	ATCAAAGTTA TATTGAAAAA GGCTATCAGT CCCAGCTTTA
  	TACGGAGATG
  151	GTCGGTATCA ACAATATTTC CAAACAGTTT ATTTTGAAAA
  	ATCCCCTGGA
  201	CGATAATCAG ACCATCGAGA ACAAACTGGA AATATTTGTC
  	TCAGGCTATA
  251	AGATGAATCC GAAAATTGCC AAAAAATATA GTGTTTCGGT
  	AAAGTTTGTC
  301	GATAAGGAAA AATCAAGGGC ATACAGGTTG GTCGGCGTTC
  	CGAAGGCGGG
  351	GACGGGTTAT ACTTTGTCGG TATGGATGAA CAGCGTGGGC
  	GACGGATACA
  401	AATGCCGTGA TGCCGCTTCT GCCCAAGCCC ATTTGGAGAC
  	CTTGTCCTCA
  451	GATGTCGGCT GTGAAGCCTT CTCTAATCGT AAAAAATAA

• This corresponds to the amino acid sequence <SEQ ID 340; ORF89-1>:

• 1	MMSNKMEQKG FTLIEMMIVV AILGIISVIA IPSYQSYIEK
  	GYQSQLYTEM
  51	VGINNISKQF ILKNPLDDNQ TIENKLEIFV SGYKMNPKIA
  	KKYSVSVKFV
  101	DKEKSRAYRL VGVPKAGTGY TLSVWMNSVG DGYKCRDAAS
  	AQAHLETLSS
  151	DVGCEAFSNR KK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with PilE of N. gonorrhoeae (Accession Number Z69260).
• ORF89 and PilE protein show 30% aa identity in 120a overlap:

• orf89	8	QKGFTLIXXMIVVAILGIISVIAIPSYXSYIEKGYQSQLYTEMXGINNISKQFILKNPL-	66
  		QKGFTLI  MIV+AI+GI++ +A+P+Y  Y  +   S+      G  +   ++ L + +
  PilE	5	QKGFTLIELMIVIAIVGILAAVALPAYQDYTARAQVSEAILLAEGQKSAVTEYYLNHGIW	64
  orf89	67	-DDNQTIENKLEIFVSGYKMNPKIAKKYSVSVKFVDKEKSRAYRLVGVPKAGTGYTLSVW	125
  		DN +         +G   + KI  KY  SV       +      GV K   G  LS+W
  PilE	65	PKDNTS---------AGVASSDKIKGKYVQSVTVAKGVVTAEMASTGVNKEIQGKKLSLW	115

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF89 shows 83.3% identity over a 162aa overlap with an ORF (ORF89a) from strain A of N. meningitidis:
• 
• The complete length ORF89a nucleotide sequence <SEQ ID 341> is:

• 1	ATGATGAGTA ATAAAATGGA ACAAAAAGGG TTTACATTGA
  	TTGNGANGNT
  51	NATNGNCNTC GCGATACNCN GCNTTANCAG CGTCATTNCN
  	ATNNNTNCNT
  101	ATCNNAGTTA TATTGAAAAA GGCTATCAGT CCCAGCTTTA
  	TACGGAGATG
  151	GTCGGTATCA ACAATATTTC CAAACAGTNT ATTTTGAAAA
  	ATCCCCTGGA
  201	CGATAATCAG ACCATCAAGA GCAAACTGGA AATATTTGTC
  	TCAGGCTATA
  251	AGATGAATCC GAAAATTGCC GAAAAATATA ATGTTTCGGT
  	GCATTTTGTC
  301	AATGAGGAAA AACCNAGGGC ATACAGCTTG GTCGGCGTTC
  	CAAAGACGGG
  351	GACGGGTTAT ACTTTGTCGG TATGGATGAA CAGCGTGGGC
  	GACGGATACA
  401	AATGCCGTGA TGCCGCTTCT GCCCGAGCCC ATTTGGAGAC
  	CTTGTCCTCA
  451	GATGTCGGCT GTGAAGCCTT CTCTAATCGT AAAAAATAG

• This encodes a protein having amino acid sequence <SEQ ID 342>:

• 1	MMSNKMEQKG FTLIXXXXXX AIXXXXSVIX XXXYXSYIEK
  	GYQSQLYTEM
  51	VGINNISKQX ILKNPLDDNQ TIKSKLEIFV SGYKMNPKIA
  	EKYNVSVHFV
  101	NEEKPRAYSL VGVPKTGTGY TLSVWMNSVG DGYKCRDAAS
  	ARAHLETLSS
  151	DVGCEAFSNR KK*

• ORF89a and ORF89-1 show 83.3% identity in 162 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF89 shows 84.6% identity over a 162aa overlap with a predicted ORF (ORF89.ng) from N. gonorrhoeae:
• 
• The complete length ORF89ng nucleotide sequence <SEQ ID 343> is:

• 1	aTGATGAGCA ATAAAATGGA ACAAAAAGGG TTTACATTGA
  	TTGAGATGAT
  51	GATAGTTGTC ACGATACTCG GCATCATCAG CGTCATTGCC
  	ATACCTTCTT
  101	ATCAGAGTTA TATTGAAAAA GGCTATCAGT CCCAGCTTTA
  	TACGGAGATG
  151	GTCGGTATCA ACAATGTTCT CAAACAGTTT ATTTTGAAAA
  	ATCCCCAGGA
  201	CGATAATGAT ACCCTCAAGA GCAAACTGAA AATATTTGTC
  	TCAGGCTATA
  251	AGATGAATCC GAAAAttgCC AAAAAATATA GTGTTTCGGt
  	aaggtttGTC
  301	gatGCGGAAA AACCAAGGGC ATACAGGTTG GTCGGCGTTC
  	CGAACGCGGG
  351	GACGGGTTAT ACTTTGTCGG TATGGATGAA CAGCGTGGGC
  	GACGGATACA
  401	AATGCCGTGA TGCCACTTCT GCCCAGGCCT ATTCGGACAC
  	CTTGTCCGCA
  451	GATAGCGGCT GTGAAGCTTT CTCTAATCGT AAAAAATAG

• This encodes a protein having amino acid sequence <SEQ ID 344>:

• 1	MMSNKMEQ KG FTLIEMMIVV TILGIISVIA IPSYQSYIEK
  	GYQSQLYTEM
  51	VGINNVLKQF ILKNPQDDND TLKSKLKIFV SGYKMNPKIA
  	KKYSVSVRFV
  101	DAEKPRAYRL VGVPNAGTGY TLSVWMNSVG DGYKCRDATS
  	AQAYSDTLSA
  151	DSGCEAFSNR KK*

• This gonococcal protein has a putative leader peptide (underlined) and N-terminal methylation site (NMePhe or type-4 pili, double-underlined). In addition, ORF89ng and ORF89-1 show 88.3% identity in 162 aa overlap:
• 
• Based on this analysis, including the gonococcal motifs and the homology with the known PilE protein, it was predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF89-1 (13.6 kDa) was cloned in the pGex vector and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 11A shows the results of affinity purification of the GST-fusion protein. Purified GST-fusion protein was used to immunise mice, whose sera gave a positive result in the ELISA test, confirming that ORF89-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 41
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 345>:

• 1	ATGAAAAAAT CCTCCCTCAT CAGCGCATTG GGCATCGGTA
  	TTTTGAGCAT
  51	CGGCATGGCA TTTGCCGCCC CTGCCGACGC GGTAAGCCAA
  	ATCCGTCAAA
  101	ACGCCACTCA AGTATTGAGC ATCTTAAAAA ACGGCGATGC
  	CAACACCGCT
  151	CGCCAAAAAG CCGAAGCCTA TGCGATTCCC TATTTCGATT
  	TCCAACGTAT
  201	GACCGCATTG GCGGTCGGCA ACCCTTGGsG CACCG.GTCC
  	GACG.GCAAA
  251	AACAAGCGTT GGCCn.AGAA TTTCAACCC...

• This corresponds to the amino acid sequence <SEQ ID 346; ORF91>:

• 1	MKKSSLISAL GIGILSIGMA FAAPADAVSQ IRQNATQVLS
  	ILKNGDANTA
  51	RQKAEAYAIP YFDFQRMTAL AVGNPWXTXS DXQKQALAXE
  	FQP...

• Further work revealed the complete nucleotide sequence <SEQ ID 347>:

• 1	ATGAAAAAAT CCTCCCTCAT CAGCGCATTG GGCATCGGTA
  	TTTTGAGCAT
  51	CGGCATGGCA TTTGCCGCCC CTGCCGACGC GGTAAGCCAA
  	ATCCGTCAAA
  101	ACGCCACTCA AGTATTGAGC ATCTTAAAAA ACGGCGATGC
  	CAACACCGCT
  151	CGCCAAAAAG CCGAAGCCTA TGCGATTCCC TATTTCGATT
  	TCCAACGTAT
  201	GACCGCATTG GCGGTCGGCA ACCCTTGGCG CACCGCGTCC
  	GACGCGCAAA
  251	AACAAGCGTT GGCCAAAGAA TTTCAAACCC TGCTGATCCG
  	CACCTATTCC
  301	GGCACGATGC TGAAATTAAA AAACGCCAAC GTCAACGTCA
  	AAGACAATCC
  351	CATCGTCAAT AAAGGCGGCA AAGAAATCAT CGTCCGCGCC
  	GAAGTCGGCG
  401	TACCCGGGCA AAAACCCGTC AACATGGACT TCACCACCTA
  	CCAAAGCGGC
  451	GGTAAATACC GTACCTACAA CGTCGCCATC GAAGGCGCGA
  	GCCTGGTTAC
  501	CGTGTACCGC AACCAATTCG GCGAAATTAT CAAAGCGAAA
  	GGCGTGGACG
  551	GACTGATTGC CGAGTTGAAA GCCAAAAACG GCGGCAAATA
  	A

• This corresponds to the amino acid sequence <SEQ ID 348; ORF91-1>:

• 1	MKKSSLISAL GIGILSIGMA FAAPADAVSQ IRQNATQVLS
  	ILKNGDANTA
  51	RQKAEAYAIP YFDFQRMTAL AVGNPWRTAS DAQKQALAKE
  	FQTLLIRTYS
  101	GTMLKLKNAN VNVKDNPIVN KGGKEIIVRA EVGVPGQKPV
  	NMDFTTYQSG
  151	GKYRTYNVAI EGASLVTVYR NQFGEIIKAK GVDGLIAELK
  	AKNGGK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF91 shows 92.4% identity over a 92aa overlap with an ORF (ORF91a) from strain A of N. meningitidis:
• 
• The complete length ORF91a nucleotide sequence <SEQ ID 349> is:

• 1	ATGAAAAAAT CCTCCTTCAT CAGCGCATTG GGCATCGGTA
  	TTTTGAGCAT
  51	CGGCATGGCA TTTGCCGCCC CTGCCGACGC GGTAAACCAA
  	ATCCGTCAAA
  101	ACGCCACTCA AGTATTGAGC ATCTTAAAAA GCGGTGATGC
  	CAACACCGCC
  151	CGCCAAAAAG CCGAAGCCTA TGCGATTCCC TATTTCGATT
  	TCCAACGTAT
  201	GACCGCATTG GCGGTCGGCA ACCCTTGGCG CACCGCGTCC
  	GACGCGCAAA
  251	AACAAGCGTT GGCCAAAGAA TTTCAAACCC TGCTGATCCG
  	CACCTATTCC
  301	GGCACGATGC TGAAATTAAA AAACGCCAAC GTCAACGTCA
  	AAGACAATCC
  351	CATCGTCAAT AAAGGCGGCA AAGAAATCAT CGTCCGCGCC
  	GAAGTCGGCG
  401	TACCCGGGCA AAAACCCGTC AACATGGACT TCACCACCTA
  	CCAAAGCGGC
  451	GGTAAATACC GTACCTACAA CGTCGCCATC GAAGGCGCGA
  	GCCTGGTTAC
  501	CGTGTACCGC AACCAATTCG GCGAAATTAT CAAAGCGAAA
  	GGCGTGGACG
  551	GACTGATTGC CGAGTTGAAG GCTAAAAACG GCAGCAAGTA
  	A

• This encodes a protein having amino acid sequence <SEQ ID 350>:

• 1	MKKSSFISAL GIGILSIGMA FAAPADAVNQ IRQNATQVLS
  	ILKSGDANTA
  51	RQKAEAYAIP YFDFQRMTAL AVGNPWRTAS DAQKQALAKE
  	FQTLLIRTYS
  101	GTMLKLKNAN VNVKDNPIVN KGGKEIIVRA EVGVPGQKPV
  	NMDFTTYQSG
  151	GKYRTYNVAI EGASLVTVYR NQFGEIIKAK GVDGLIAELK
  	AKNGSK*

• ORF91a and ORF91-1 show 98.0% identity in 196 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF91 shows 84.8% identity over a 92aa overlap with a predicted ORF (ORF91.ng) from N. gonorrhoeae:
• 
• The complete length ORF91ng nucleotide sequence <SEQ ID 351> is predicted to encode a protein having amino acid sequence <SEQ ID 352>:

• 1	VKKSSFISAL GIGILSIGMA FASPADAVGQ IRQNATQVLT
  	ILKSGDAASA
  51	RPKAEAYAVP YFDFQRMTAL AVGNPWRTAS DAQKQALAKE
  	FQTLLIRTYS
  101	GTMLKFKNAT VNVKDNPIVN KGGKEIVVRA EVGIPGQKPV
  	NMDFTTYQSG
  151	GKYRTYNVAI EGTSLVTVYR NQFGEIIKAK GIDGLIAELK
  	AKNGGK*

• Further work revealed the complete nucleotide sequence <SEQ ID 353>:

• 1	ATGAAAAAAT CCTCCTTCAT CAGCGCATTG GGCATCGGTA
  	TTTTGAGCAT
  51	CGGCATGGCA TTTGCCTCCC CGGCCGACGC AGTGGGACAA
  	ATCCGCCAAA
  101	ACGCCACACA GGTTTTGACC ATCCTCAAAA GCGGCGACGC
  	GGCTTCTGCA
  151	CGCCCAAAAG CCGAAGCCTA TGCGGTTCCC TATTTCGATT
  	TCCAACGTAT
  201	GACCGCATTG GCGGTCGGCA ACCCTTGGCG TACCGCGTCC
  	GACGCGCAAA
  251	AACAAGCGTT GGCCAAAGAA TTTCAAACCC TGCTGATCCG
  	CACCTATTCC
  301	GGCACGATGC TGAAATTCAA AAACGCGACC GTCAACGTCA
  	AAGACAATCC
  351	CATCGTCAAT AAGGGCGGCA AGGAAATCGT CGTCCGTGCC
  	GAAGTCGGCA
  401	TCCCCGGTCA GAAGCCCGTC AATATGGACT TTACCACCTA
  	CCAAAGCGGC
  451	GGCAAATACC GTACCTACAA CGTCGCCATC GAAGGCACGA
  	GCCTGGTTAC
  501	CGTGTACCGC AACCAATTCG GCGAAATCAT CAAAGCCAAA
  	GGCATCGACG
  551	GGCTGATTGC CGAGTTGAAA GCCAAAAACG GCGGCAAATA
  	A

• This corresponds to the amino acid sequence <SEQ ID 354; ORF91ng-1>:

• 1	MKKSSFISAL GIGILSIGMA FASPADAVGQ IRQNATQVLT
  	ILKSGDAASA
  51	RPKAEAYAVP YFDFQRMTAL AVGNPWRTAS DAQKQALAKE
  	FQTLLIRTYS
  101	GTMLKFKNAT VNVKDNPIVN KGGKEIVVRA EVGIPGQKPV
  	NMDFTTYQSG
  151	GKYRTYNVAI EGTSLVTVYR NQFGEIIKAK GIDGLIAELK
  	AKNGGK*

• ORF91ng-1 and ORF91-1 show 92.3% identity in 196 aa overlap:
• 
• In addition, ORF91ng-1 shows homology to a hypothetical E. coli protein:

• sp|P45390|YRBC_ECOLI HYPOTHETICAL 24.0 KD PROTEIN IN MURA-RPON
  INTERGENIC
  REGION PRECURSOR (F211) >gi|606130 (U18997) ORF_f211 [Escherichia coli]
  >gi|1789583 (AE000399) hypothetical 24.0 kD protein in murZ-rpoN
  intergenic region [Escherichia coli]Length = 211
  Score = 70.6 bits (170), Expect = 6e−12
  Identities = 42/137 (30%), Positives = 76/137 (54%), Gaps = 6/137 (4%)

  Query:	59	VPYFDFQRMTALAVGNPWRTASDAQKQALAKEFQTLLIRTYSGTMLKFKNATVNVKDNPI	118
  		+PY   +   AL +G  +++A+ AQ++A    F+  L + Y   +  +   T  +   P
  Sbjct:	65	LPYVQVKYAGALVLGQYYKSATPAQREAYFAAFREYLKQAYGQALAMYHGQTYQIA--PE	122
  Query:	119	VNKGGKEIV-VRAEVGIP-GQKPVNMDFTTYQSG--GKYRTYNVAIEGTSLVTVYRNQFG	174
  		G K IV +R  +  P G+ PV +DF   ++   G ++ Y++  EG S++T  +N++G
  Sbjct:	123	QPLGDKTIVPIRVTIIDPNGRPPVRLDFQWRKNSQTGNWQAYDMIAEGVSMITTKQNEWG	182
  Query:	175	EIIKAKGIDGLIAELKA	191
  		+++ KGIDGL A+LK+
  Sbjct:	183	TLLRTKGIDGLTAQLKS	199

• Based on this analysis, including the presence of a putative leader sequence in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 42
• The following DNA sequence was identified in N. meningitidis <SEQ ID 355>:

• 1	ATGAAACACA TACTCCCCCT GATTGCCGCA TCCGCACTCT
  	GCATTTCAAC
  51	CGCTTCGGCA CATCCTGCCA GCGAACCGTC CACTCAAAAC
  	GAAACCGCTA
  101	TGATCACGCA TACCCTCATC TCAAAATACA GTTTTGGnnn
  	nnnnnnnnnn
  151	nnnnnnnnnn nnGCCATAAA AAGCAAAGGG ATGGACATTT
  	TTGCCGTCAT
  201	CGACCATCAG GAAGCCGCAC GCCGAAACGG CTTAACGATG
  	CAGCCGGCAA
  251	AAGTCATCGT CTTCGGCACG CCCAAAGCCG GCACGCCGCT
  	GATGGTCAAA
  301	GACCCCGCCT TCGCCCTGCA ACTGCCCCTA CGCGTCCTCG
  	TTACCGAAAC
  351	GGACGGCAAA GTACGCGCCG CCTATACCGA TACGCGCGCC
  	CTCATCGCCG
  401	GCAGCCGCAT CGGTTTCGAC GAAGTGGCAA ACACTTTGGC
  	AAACGCCGAA
  451	AAACTGATAC AAAAAACCGT AGGCGAATAA

• This corresponds to the amino acid sequence <SEQ ID 356; ORF97>:

• 1	MKHILPLIAA SALCISTASA HPASEPSTQN ETAMITHTLI
  	SKYSFGXXXX
  51	XXXXAIKSKG MDIFAVIDHQ EAARRNGLTM QPAKVIVFGT
  	PKAGTPLMVK
  101	DPAFALQLPL RVLVTETDGK VRAAYTDTRA LIAGSRIGFD
  	EVANTLANAE
  151	KLIQKTVGE*

• Further work revealed the complete nucleotide sequence <SEQ ID 357>:

• 1	ATGAAACACA TACTCCCCCT GATTGCCGCA TCCGCACTCT
  	GCATTTCAAC
  51	CGCTTCGGCA CATCCTGCCA GCGAACCGTC CACCCAAAAC
  	GAAACCGCTA
  101	TGACCACGCA TACCCTCACC TCAAAATACA GTTTTGACGA
  	AACCGTCAGC
  151	CGCCTTGAAA CCGCCATAAA AAGCAAAGGG ATGGACATTT
  	TTGCCGTCAT
  201	CGACCATCAG GAAGCCGCCC GCCGAAACGG CTTAACGATG
  	CAGCCGGCAA
  251	AAGTCATCGT CTTCGGCACG CCCAAAGCCG GCACGCCGCT
  	GATGGTCAAA
  301	GACCCCGCCT TCGCCCTGCA ACTGCCCCTA CGCGTCCTCG
  	TTACCGAAAC
  351	GGACGGCAAA GTACGCGCCG CCTATACCGA TACGCGCGCC
  	CTCATCGCCG
  401	GCAGCCGCAT CGGTTTCGAC GAAGTGGCAA ACACTTTGGC
  	AAACGCCGAA
  451	AAACTGATAC AAAAAACCGT AGGCGAATAA

• This corresponds to the amino acid sequence <SEQ ID 358; ORF97-1>:

• 1	MKHILPLIAA SALCISTASA HPASEPSTQN ETAMTTHTLT
  	SKYSFDETVS
  51	RLETAIKSKG MDIFAVIDHQ EAARRNGLTM QPAKVIVFGT
  	PKAGTPLMVK
  101	DPAFALQLPL RVLVTETDGK VRAAYTDTRA LIAGSRIGFD
  	EVANTLANAE
  151	KLIQKTVGE*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF97 shows 88.7% identity over a 159aa overlap with an ORF (ORF97a) from strain A of N. meningitidis:
• 
• The complete length ORF97a nucleotide sequence <SEQ ID 359> is:

• 1	ATGANACACA TACTCCCCCT GANTGNCGCA TCCGCACTCT
  	GCATTTCAAC
  51	CGCTTCGGNN CATCCTGCCA GCGAACCGCA AACCCAAAAC
  	GAAACCGCTA
  101	TGACCACGCA TACCCTCACC TCAAAATACA GTTTTGACGA
  	AACCGTCAGC
  151	CGCCTTGAAA CCGCCATAAA AAGCAAAGGG ATGGACATTT
  	TTGCCGTCAT
  201	CGACCATCAG GAAGCCGCCC GCCGAAACGG CTTAACGATG
  	CAGCCGGCAA
  251	AAGTCATCGT CTTCGGCACG CCCAAAGCCG GTACGCCGCT
  	GATGGTCAAA
  301	GACCCCGCCT TCGCCCTGCA ACTGCCCCTG CGCGTCNTCG
  	TTACCGAAAC
  351	GGACGGCAAA GTACGCGCCG CCTATACCGA TACGCGCGCC
  	CTCATCGCCG
  401	GCAGCCGCAT CGGTTTCGAC GAAGTGGCAA ACACTTTGGC
  	AAACGCCGAA
  451	AAACTGATAC AAAAAACCAT AGGCGAATAA

• This encodes a protein having amino acid sequence <SEQ ID 360>:

• 1	MXHILPLXXA SALCISTASX HPASEPQTQN ETAMTTHTLT
  	SKYSFDETVS
  51	RLETAIKSKG MDIFAVIDHQ EAARRNGLTM QPAKVIVFGT
  	PKAGTPLMVK
  101	DPAFALQLPL RVXVTETDGK VRAAYTDTRA LIAGSRIGFD
  	EVANTLANAE
  151	KLIQKTIGE*

• ORF97a and ORF97-1 show 95.6% identity in 159 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF97 shows 88.1% identity over a 159aa overlap with a predicted ORF (ORF97.ng) from N. gonorrhoeae:
• 
• The complete length ORF97ng nucleotide sequence <SEQ ID 361> is predicted to encode a protein having amino acid sequence <SEQ ID 362>:

• 1	MKHILPPIAA SAFCISTASA HPAGKPPTQN ETAMTTHTLT
  	SKYSFDETVS
  51	RLETAIKSKG MDIFAVIDHQ EAARRNGLTM QPAKVIVFGT
  	PKAGTPLMVK
  101	DPAFALQLPL RVLVTETDGK VRTAYTDTRA LIVGSRISFD
  	EVANTLANAE
  151	KLIQKTVGE*

• Further work revealed the complete nucleotide sequence <SEQ ID 363>:

• 1	ATGAAACACA TACTCCCcct gatcgccgca TccgcactCT
  	GCATTTCAAC
  51	CGCTTCGGCA CACCCTGCCG GCAAACCGCC CACCCAAAAC
  	GAAACCGCTA
  101	TGACCACGCA CACCCTCACC TCGAAATACA GTTTTGACGA
  	AACCGTCAGC
  151	CGCCTTGAAA CCGCCATAAA AAGCAAAGGG ATGGACATTT
  	TTGCCGTCAT
  201	CGACCATCAG GAAGCGGCAC GCCGAAACGG CCTGACCATG
  	CAGCCGGCAA
  251	AAGTCATCGT CTTCGGCACG CCCAAGGCCG GTACGCCgct
  	GATGGTCAAA
  301	GACCCCGCCT TCGCCCTGCA ACTGCCCCTG CGCGTCCTCG
  	TTACCGAAAC
  351	GGACGGCAAA GTACGCACCG CCTATACCGA TACGCGCGCC
  	CTCATCGTCG
  401	GCAGCCGCAT CAGTTTCGAC GAAGTGGCAA ACACTTTGGC
  	AAACGCCGAA
  451	AAACTGATAC AAAAAACCGT AGGCGAATAA

• This corresponds to the amino acid sequence <SEQ ID 364; ORF97ng-1>:

• 1	MKHILPLIAA SALCISTASA HPAGKPPTQN ETAMTTHTLT
  	SKYSFDETVS
  51	RLETAIKSKG MDIFAVIDHQ EAARRNGLTM QPAKVIVFGT
  	PKAGTPLMVK
  101	DPAFALQLPL RVLVTETDGK VRTAYTDTRA LIVGSRISFD
  	EVANTLANAE
  151	KLIQKTVGE*

• ORF97ng-1 and ORF97-1 show 96.2% identity in 159 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence in the gonococcal protein, it was predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF97-1 (15.3 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIGS. 12A & 12B show, respectively, the results of affinity purification of the GST-fusion and His-fusion proteins. Purified GST-fusion protein was used to immunise mice, whose sera were used for Western Blot (FIG. 12C), ELISA (positive result), and FACS analysis (FIG. 12D). These experiments confirm that ORF97-1 is a surface-exposed protein, and that it is a useful immunogen. FIG. 12E shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF97-1.
Example 43
• The following DNA, believed to be complete, sequence was identified in N. meningitidis <SEQ ID 365>:

• 1	ATGGCTTTTA TTACGCGCTT ATTCAAAAGC AGTAAATGGC
  	TGATTGTGCC
  51	GCTGATGCTC CCCGCCTTTC AGAATGTGGC GGCGGAGGGG
  	ATAGATGTGA
  101	GCCGTGCCGA AGCGAGGATA ACCGACGGCG GGCAGCTTTC
  	CATCAGCAGC
  151	CGCTTCCAAA CCGAGCTGCC CGACCAGCTC CAACAGGCGT
  	TGCGCCGGGg
  201	CGTGCCGCTC AACTTTACCT TAAGCTGGCA GCTTTCCGCC
  	CCGATAATCG
  251	CTTCTTATCG GTTTAAATTG GGGCAACTGA TTGGCGATGA
  	CGACaATATT
  301	GACTACAAAC TGAGTTTCCA TCCGCTGACc AaACGCTACC
  	GCGTTACCgT
  351	CGgCGCGTTT TCGACAGACT ACGACACCTT GGATGCGGCA
  	TTGCGCGCGA
  401	CCGGCGCGGT TGCCAACTGG AAAGTCCTGA ACAAAGGCGC
  	GCTGTCCGGT
  451	GCGGAAGCAG GGGAAACCAA GGCGGAAATC CGCCTGACGC
  	TGTCCACTTC
  501	AAAACTGCCC AAGCCTTTTC AAATCAATGC ATTGACTTCT
  	CAAAACTGGC
  551	ATTTGGATTC GGGTTGGAAA CCTCTAAACA TCATCGGGAA
  	CAAATAA

• This corresponds to the amino acid sequence <SEQ ID 366; ORF106>:

• 1	MAFITRLFKS SKWLIVPLML PAFQNVAAEG IDVSRAEARI
  	TDGGQLSISS
  51	RFQTELPDQL QQALRRGVPL NFTLSWQLSA PIIASYRFKL
  	GQLIGDDDNI
  101	DYKLSFHPLT KRYRVTVGAF STDYDTLDAA LRATGAVANW
  	KVLNKGALSG
  151	AEAGETKAEI RLTLSTSKLP KPFQINALTS QNWHLDSGWK
  	PLNIIGNK*

• Further work revealed the following DNA sequence <SEQ ID 367>:

• 1	ATGGCTTTTA TTACGCGCTT ATTCAAAAGC AGTAAATGGC
  	TGATTGTGCC
  51	GCTGATGCTC CCCGCCTTTC AGAATGTGGC GGCGGAGGGG
  	ATAGATGTGA
  101	GCCGTGCCGA AGCGAGGATA ACCGACGGCG GGCAGCTTTC
  	CATCAGCAGC
  151	CGCTTCCAAA CCGAGCTGCC CGACCAGCTC CAACAGGCGT
  	TGCGCCGGGG
  201	CGTGCCGCTC AACTTTACCT TAAGCTGGCA GCTTTCCGCC
  	CCGATAATCG
  251	CTTCTTATCG GTTTAAATTG GGGCAACTGA TTGGCGATGA
  	CGACAATATT
  301	GACTACAAAC TGAGTTTCCA TCCGCTGACC AACCGCTACC
  	GCGTTACCGT
  351	CGGCGCGTTT TCGACAGACT ACGACACCTT GGATGCGGCA
  	TTGCGCGCGA
  401	CCGGCGCGGT TGCCAACTGG AAAGTCCTGA ACAAAGGCGC
  	GCTGTCCGGT
  451	GCGGAAGCAG GGGAAACCAA GGCGGAAATC CGCCTGACGC
  	TGTCCACTTC
  501	AAAACTGCCC AAGCCTTTTC AAATCAATGC ATTGACTTCT
  	CAAAACTGGC
  551	ATTTGGATTC GGGTTGGAAA CCTCTAAACA TCATCGGGAA
  	CAAATAA

• This corresponds to the amino acid sequence <SEQ ID 368; ORF106-1>:

• 1	MAFITRLFKS SKWLIVPLML PAFQNVAAEG IDVSRAEARI
  	TDGGQLSISS
  51	RFQTELPDQL QQALRRGVPL NFTLSWQLSA PIIASYRFKL
  	GQLIGDDDNI
  101	DYKLSFHPLT NRYRVTVGAF STDYDTLDAA LRATGAVANW
  	KVLNKGALSG
  151	AEAGETKAEI RLTLSTSKLP KPFQINALTS QNWHLDSGWK
  	PLNIIGNK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF106 shows 87.4% identity over a 199aa overlap with an ORF (ORF106a) from strain A of N. meningitidis:
• 
• Due to the K→N substitution at residue 111, the homology between ORF106a and ORF106-1 is 87.9% over the same 199 aa overlap.
• The complete length ORF106a nucleotide sequence <SEQ ID 369> is:

• 1	ATGGCTTTTA TTACGCGCTT ATTCAAAAGC ATTAAACAAT
  	GGCTTGTGCT
  51	GCTGCCGATG CTTTCCGTTT TGCCGGACGC GGCGGCGGAG
  	GGGATAGATG
  101	TGAGCCGCGC CGAAGCGAGG ATAANCGACG GCGGGCAGCT
  	TTCCATNAGN
  151	AGCCGCTTCC AAACCGAGCT GCCCGACCAG CTCCAANNNG
  	CGNNGNGCCG
  201	GGGCGTGNCG CTCAACTNTA CCTTAAGNTG GCAGCTTTCC
  	GCCCCGATAA
  251	TCGCTTCTTA TCGGTTTNAA TTGGGGCAAC TGATTGGCGA
  	TGACGACNAT
  301	ATTGACTACA AACTGAGTTT CCATCCGCTG ACCAACCGCT
  	ACCGCGTTAC
  351	CGTCGGCGCG TTTTCGACAG ANTACGACAC CTTGGATGCG
  	GCATTGCGCG
  401	CGACCGGCGC GGTTGCCAAC TGGAAAGTCC TGAACAAAGG
  	CGCGCTGTCC
  451	GGTGCGGAAG CAGGGGAAAC CAAGGCGGAA ATCCGCCTGA
  	CGCTGTCCAC
  501	TTCAAAACTG CCCAAGCCTT TTCAAATCAA TGCATTGACT
  	TCTCAAAACT
  551	GGCATTTGGA TTCGGGTTGG AAACCTCTAA ACATCATCGG
  	GAACAAATAA

• This encodes a protein having amino acid sequence <SEQ ID 370>:

• 1	MAFITRLFKS IKQWLVLLPM LSVLPDAAAE GIDVSRAEAR
  	IXDGGQLSXX
  51	SRFQTELPDQ LQXAXXRGVX LNXTLXWQLS APIIASYRFX
  	LGQLIGDDDX
  101	IDYKLSFHPL TNRYRVTVGA FSTXYDTLDA ALRATGAVAN
  	WKVLNKGALS
  151	GAEAGETKAE IRLTLSTSKL PKPFQINALT SQNWHLDSGW
  	KPLNIIGNK*

  
  Homology with a Predicted ORF from N. gonorrhoeae
• ORF106 shows 90.5% identity over a 199aa overlap with a predicted ORF (ORF106.ng) from N. gonorrhoeae:
• 
• Due to the K→N substitution at residue 111, the homology between ORF106ng and ORF106-1 is 91.0% over the same 199 aa overlap.
• The complete length ORF106ng nucleotide sequence <SEQ ID 371> is:

• 1	ATGGCTTTTA TTACGCGCTT ATTCAAAAGC ATTAAACAAT
  	GGCTTGTGCT
  51	GTTGCCGATA CTCTCCGTTT TGCCGGACGC GGCGGCGGAG
  	GGCATTGCCG
  101	CGACCCGCGC CGAAGCGAGG ATAACCGACG GCGGGCGGCT
  	TTCCATCAGC
  151	AGCCGCTTCC AAACCGAGCT GCCCGACCAG CTCCAACAGG
  	CGTTGCGCCG
  201	GGGCGTACCG CTCAACTTTA CCTTAAGCTG GCAGCTTTCC
  	GCCCCGACAA
  251	TCGCTTCTTA TCGGTTTAAA TTGGGGCAAC TGATTGGCGA
  	TGACGACAAT
  301	ATTGACTACA AACTAAGTTT CCATCCGCTG ACCAACCGCT
  	ACCGCGTTAC
  351	CGTCGGCGCA TTTTCCACCG ATTACGACAC TTTGGATGCG
  	GCATTGCGCG
  401	CGACCGGCGC GGTTGCCAAC TGGAAAGTCC TGAACAAAGG
  	CGCGTTGTCC
  451	GGTGCGGAAG CAGGGGAAAC CAAGGCGGAA ATCCGCCTGA
  	CGCTGTCCAC
  501	TTCAAAACTG CCCAAGCCTT TCCAAATCAA CGCATTGACT
  	TCTCAAAACT
  551	GGCATTTGGA TTCGGGTTGG AAACCTCTAA ACATCATCGG
  	GAACAAATAA

• This encodes a protein having amino acid sequence <SEQ ID 372>:

• 1	MAFITRLFKS IKQWLVLLPI LSVLPDAAAE GIAATRAEAR
  	ITDGGRLSIS
  51	SRFQTELPDQ LQQALRRGVP LNFTLSWQLS APTIASYRFK
  	LGQLIGDDDN
  101	IDYKLSFHPL TNRYRVTVGA FSTDYDTLDA ALRATGAVAN
  	WKVLNKGALS
  151	GAEAGETKAE IRLTLSTSKL PKPFQINALT SQNWHLDSGW
  	KPLNIIGNK*

• Based on this analysis, including the presence of a putative leader sequence in the gonococcal protein, it was predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF106-1 (18 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 13A shows the results of affinity purification of the His-fusion protein, and FIG. 13B shows the results of expression of the GST-fusion in E. coli. Purified His-fusion protein was used to immunise mice, whose sera were used for FACS analysis (FIG. 13C) These experiments confirm that ORF106-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 44
• The following DNA sequence, believed to be complete, was identified in N. meningitidis <SEQ ID 373>:

• 1	ATGGACACAA AAGAAATCCT CGG.TACGCG GcAGGcTCGA
  	TCGGCAGCGC
  51	GGTTTTAGCC GTCATCATCc TGCCGCTGCT GTCGTGGTAT
  	TTCCCCGCCG
  101	ACGACATCGG GCGCATCGTG CTGATGCAGA CGGCGGCGGG
  	GCTgACGGTG
  151	TCGGTGTTGT GCCTCGGGCT GGATCAGGCA TACGTCCGCG
  	AATACTATGC
  201	CACCGCCGAC AAAGACAcCT TGTTCAAAAC CCTGTTCCTG
  	CCGCCGCTGC
  251	TGTCTGCCGC CGCGATAGCC GCCCTGCTGC TTTCCCGCCC
  	GTCCCTGCCG
  301	TCTGAAATCC TGTTTTCACT CGACGATGCC gCCGCCGGCa
  	TCGGGCTGGT
  351	GCTGTTTGAA CtGAGCTTCC TGCCCATCCG cTTTCTCTTA
  	CTGGTTTTGC
  401	GTATGGAAGG ACGCGCCcTT GCCTTTTCGT CCGCGCAACT
  	CGTGCcCAAG
  451	CTCGCCATCC TGCTGCTG.T GCCGCTGACG GTCGGGCTGC
  	TGCACTTTCC
  501	AGCGAACACC GCCGTCCTGA CCGCCGTTTA CGCGCTGGCA
  	AACCTTGCCG
  551	CCGCCGCCTT TTTGCTGTTT CAAAACCGAT GCCGTCTGAA
  	GGCCGTCCGG
  601	CACGCACCGT TTTCGCCCGC CGTCCTGCAC CGGGGG.TGC
  	GCTACGGCAT
  651	ACCGATCGCA CTGAGCAGCA TCGCCTATTG GGGGCTGGCA
  	TCCGCCGACC
  701	GTTTGTTCCT GAAAAAATAT GCCGGCCTGG AACAGCTCGG
  	CGTTTATTCG
  751	ATGGGTATTT CGTTCGGCGG GGCGGCATTA TTGTTCCAAA
  	GCATCTTTTC
  801	AACGGTCTGG ACACCGTATA TTTTCCGCGC AATCGAAGAA
  	AACGCCCCGC
  851	CCGCTCGCCT CTCGGCAACG GCAGAATCCG CCGCCGCCCT
  	GCTTGCCTCC
  901	GCCCTCTGC. TGACCGGCAT TTTCTCGCCC CTTGCCTCCC
  	TCCTGCTGCC
  951	GGAAAACTAC GCCGCCGTCC GGTTTATCGT CGTATCGTGT
  	ATG.TGCCGC
  1001	CGCTGTTTTG CACGCTGGCG GAAATCAGCG GCATCGGTTT
  	GAACGTCGTT
  1051	CGCAAAACGC GCCCGATCGC GCTCGCCACC TTGGGCGCGC
  	TGGCGGCAAA
  1101	CCTGCTGCTG CTGGGGCTTG ACCGTGCCGT ACCGGCGAGG
  	CCGCC.GGCG
  1151	CGGCGGTTGC CTGTGCCGCC TCATTCTGGC TGTTTTTTGC
  	CTTCAAGACC
  1201	GAAAGCTCyT GCCGCCTGTG GCAGCCGCTC AAACGCCTGC
  	CGCTTTATCT
  1251	GCACACATTG TTCTGCCTGA CCTCCTCGGC GGCCTACACC
  	TGCTTCGGCA
  1301	CGCCGGCAAA CTATCCCCTG TTTGCCGGCG TATGGGCGGC
  	ATATCTGGCA
  1351	GGCTGCATCC TGCGCCACCG GAAAGATTTG CACAAACTGT
  	TTCATTATTT
  1401	GAAAAAACAA GGTTTCCCAT TATGA

• This corresponds to the amino acid sequence <SEQ ID 374; ORF10>:

• 1	MDTKEILXYA AGSIGSAVLA VIILPLLSWY FPADDIGRIV
  	LMQTAAGLTV
  51	SVLCLGLDQA YVREYYATAD KDTLFKTLFL PPLLSAAAIA
  	ALLLSRPSLP
  101	SEILFSLDDA AAGIGLVLFE LSFLPIRFLL LVLRMEGRAL
  	AFSSAQLVPK
  151	LAILLLXPLT VGLLHFPANT AVLTAVYALA NLAAAAFLLF
  	QNRCRLKAVR
  201	HAPFSPAVLH RGXRYGIPIA LSSIAYWGLA SADRLFLKKY
  	AGLEQLGVYS
  251	MGISFGGAAL LFQSIFSTVW TPYIFRAIEE NAPPARLSAT
  	AESAAALLAS
  301	ALCXTGIFSP LASLLLPENY AAVRFIVVSC MXPPLFCTLA
  	EISGIGLNVV
  351	RKTRPIALAT LGALAANLLL LGLDRAVPAR PXGAAVACAA
  	SFWLFFAFKT
  401	ESSCRLWQPL KRLPLYLHTL FCLTSSAAYT CFGTPANYPL
  	FAGVWAAYLA
  451	GCILRHRKDL HKLFHYLKKQ GFPL*

• Further sequence analysis revealed the complete DNA sequence <SEQ ID 375> to be:

• 1	ATGGACACAA AAGAAATCCT CGGCTACGCG GCAGGCTCGA
  	TCGGCAGCGC
  51	GGTTTTAGCC GTCATCATCC TGCCGCTGCT GTCGTGGTAT
  	TTCCCCGCCG
  101	ACGACATCGG GCGCATCGTG CTGATGCAGA CGGCGGCGGG
  	GCTGACGGTG
  151	TCGGTGTTGT GCCTCGGGCT GGATCAGGCA TACGTCCGCG
  	AATACTATGC
  201	CACCGCCGAC AAAGACACCT TGTTCAAAAC CCTGTTCCTG
  	CCGCCGCTGC
  251	TGTCTGCCGC CGCGATAGCC GCCCTGCTGC TTTCCCGCCC
  	GTCCCTGCCG
  301	TCTGAAATCC TGTTTTCACT CGACGATGCC GCCGCCGGCA
  	TCGGGCTGGT
  351	GCTGTTTGAA CTGAGCTTCC TGCCCATCCG CTTTCTCTTA
  	CTGGTTTTGC
  401	GTATGGAAGG ACGCGCCCTT GCCTTTTCGT CCGCGCAACT
  	CGTGCCCAAG
  451	CTCGCCATCC TGCTGCTGCT GCCGCTGACG GTCGGGCTGC
  	TGCACTTTCC
  501	AGCGAACACC GCCGTCCTGA CCGCCGTTTA CGCGCTGGCA
  	AACCTTGCCG
  551	CCGCCGCCTT TTTGCTGTTT CAAAACCGAT GCCGTCTGAA
  	GGCCGTCCGG
  601	CACGCACCGT TTTCGCCCGC CGTCCTGCAC CGGGGGCTGC
  	GCTACGGCAT
  651	ACCGATCGCA CTGAGCAGCA TCGCCTATTG GGGGCTGGCA
  	TCCGCCGACC
  701	GTTTGTTCCT GAAAAAATAT GCCGGCCTGG AACAGCTCGG
  	CGTTTATTCG
  751	ATGGGTATTT CGTTCGGCGG GGCGGCATTA TTGTTCCAAA
  	GCATCTTTTC
  801	AACGGTCTGG ACACCGTATA TTTTCCGCGC AATCGAAGAA
  	AACGCCCCGC
  851	CCGCCCGCCT CTCGGCAACG GCAGAATCCG CCGCCGCCCT
  	GCTTGCCTCC
  901	GCCCTCTGCC TGACCGGCAT TTTCTCGCCC CTTGCCTCCC
  	TCCTGCTGCC
  951	GGAAAACTAC GCCGCCGTCC GGTTTATCGT CGTATCGTGT
  	ATGCTGCCGC
  1001	CGCTGTTTTG CACGCTGGCG GAAATCAGCG GCATCGGTTT
  	GAACGTCGTC
  1051	CGCAAAACGC GCCCGATCGC GCTCGCCACC TTGGGCGCGC
  	TGGCGGCAAA
  1101	CCTGCTGCTG CTGGGGCTTG CCGTGCCGTC CGGCGGCGCG
  	CGCGGCGCGG
  1151	CGGTTGCCTG TGCCGCCTCA TTCTGGCTGT TTTTTGCCTT
  	CAAGACCGAA
  1201	AGCTCCTGCC GCCTGTGGCA GCCGCTCAAA CGCCTGCCGC
  	TTTATCTGCA
  1251	CACATTGTTC TGCCTGACCT CCTCGGCGGC CTACACCTGC
  	TTCGGCACGC
  1301	CGGCAAACTA TCCCCTGTTT GCCGGCGTAT GGGCGGCATA
  	TCTGGCAGGC
  1351	TGCATCCTGC GCCACCGGAA AGATTTGCAC AAACTGTTTC
  	ATTATTTGAA
  1401	AAAACAAGGT TTCCCATTAT GA

• This corresponds to the amino acid sequence <SEQ ID 376; ORF10-1>:

• 1	MDTKEILGYA AGSIGSAVLA VIILPLLSWY FPADDIGRIV
  	LMQTAAGLTV
  51	SVLCLGLDQA YVREYYATAD KDTLFKTLFL PPLLSAAAIA
  	ALLLSRPSLP
  101	SEILFSLDDA AAGIGLVLFE LSFLPIRFLL LVLRMEGRAL
  	AFSSAQLVPK
  151	LAILLLLPLT VGLLHFPANT AVLTAVYALA NLAAAAFLLF
  	QNRCRLKAVR
  201	HAPFSPAVLH RGLRYGIPIA LSSIAYWGLA SADRLFLKKY
  	AGLEQLGVYS
  251	MGISFGGAAL LFQSIFSTVW TPYIFRAIEE NAPPARLSAT
  	AESAAALLAS
  301	ALCLTGIFSP LASLLLPENY AAVRFIVVSC MLPPLFCTLA
  	EISGIGLNVV
  351	RKTRPIALAT LGALAANLLL LGLAVPSGGA RGAAVACAAS
  	FWLFFAFKTE
  401	SSCRLWQPLK RLPLYLHTLF CLTSSAAYTC FGTPANYPLF
  	AGVWAAYLAG
  451	CILRHRKDLH KLFHYLKKQG FPL*

• Computer analysis of this amino acid sequence gave the following results:
Prediction
• ORF10-1 is predicted to be the precursor of an integral membrane protein, since it comprises several (12-13) potential transmembrane segments, and a probable cleavable signal peptide
• Homology with EpsM from Streptococcus thermophilus (Accession Number U40830).
• ORF10 shows homology with the epsM gene of S. thermophilus, which encodes a protein of a size similar to ORF10 and is involved in expolysaccharide synthesis. Other homologies are with prokaryotic membrane proteins:

• Identities = (25%)

  Query:	213	LRYGIPLALSSLAYWGLASADRLFLKKYAGLEQLGVYSMGISFGGAALLLQSIFSTVW	270
  		L Y +PL  SS+ +W L ++ R F+  + G    G+ ++         +  +IF+  W
  Sbjct:	210	LYYALPLIPSSILWWLLNASSRYFVLFFLGAGANGLLAVATKIPSIISIFNTIFTQAW	267

  Identities = 15/57 (26%), Positives = 31/57 (54%)

  Query:	7	LGYAAGSIGSAVLAVIILPLLSWYFPADDIGRIVLMQTAAGLTVSVLCLGLDQAYVR	63
  		L +  G++GS +L  +++PL ++     + G   L QT A L + ++ + +  A +R
  Sbjct:	12	LVFTIGNLGSKLLVFLLVPLYTYAMTPQEYGMADLYQTTANLLLPLITMNVFDATLR	68

  Identities = 16/96 (16%), Positives = 36/96 (37%)

  Query:	307	IFSPLASLLLPENYAAVRFTVVSCMLPPLFYTLTEISGIGLNVVRKTRPIXXXXXXXXXX	366
  		+  P+   ++  +YA+    V   ML  LF + ++  G      ++T+ +
  Sbjct:	305	VLKPIVEKVVSSDYASSWQYVPFFMLSMLFSSFSDFFGTNYIAAKQTKGVFMTSIYGTIV	364

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF10 shows 95.4% identity over a 475aa overlap with an ORF (ORF10a) from strain A of N. meningitidis:
• 
• The complete length ORF10a nucleotide sequence <SEQ ID 377> is:

• 1	ATGGACACAA AAGAAATCCT CGGCTACGCG GCAGGCTCGA
  	TCGGCAGCGC
  51	GGTTTTAGCC GTCATCATCC TGCCGCTGCT GTCGTGGTAT
  	TTCCCTGCCG
  101	ACGACATCGG ACGCATCGTG CTGATGCAGA CGGCGGCGGG
  	GCTGACGGTG
  151	TCGGTGTTGT GCCTCGGGCT GGATCAGGCA TACGTCCGCG
  	AATACTATGC
  201	CGCCGCCGAC AAAGACACTT TGTTCAAAAC CCTGTTCCTG
  	CCGCCGCTGC
  251	TGTCTGCCGC CGCGATAGCC GCCCTGCTGC TTTCCCGCCC
  	ATCCCTGCCG
  301	TCTGAAATCC TGTTTTCGCT CGACGATGCC GCCGCCGGCA
  	TCGGGCTGGT
  351	GCTGTTTGAA CTGAGCTTCC TGCCCATCCG CTTTCTCTTA
  	CTGGTTTTGC
  401	GTATGGAAGG ACGCGCCCTT GCCTTTTCGT CCGCGCAACT
  	CGTGTCCAAG
  451	CTCGCCATCC TGCTGCTGCT GCCGCTGACG GTCGGGCTGC
  	TGCACTTTCC
  501	GGCGAACACC GCCGTCCTGA CCGCCGTTTA CGCGCTGGCA
  	AACCTTGCCG
  551	CCGCCGCCTT TTTGCTGTTT CAAAACCGAT GCCGTCTGAA
  	GGCCGTCCGG
  601	CGCGCACCGT TTTCATCCGC CGTCCTGCAT CGCGGCCTGC
  	GCTACGGCAT
  651	ACCGATCGCA CTAAGCAGCA TCGCCTATTG GGGGCTGGCA
  	TCCGCCGACC
  701	GTTTGTTCCT GAAAAAATAT GCCGGCCTAG AACAGCTCGG
  	CGTTTATTCG
  751	ATGGGTATTT CGTTCGGCGG AGCGGCATTA TTGTTCCAAA
  	GCATCTTTTC
  801	AACGGTCTGG ACACCGTATA TTTTCCGCGC AATCGAAGCA
  	AACGCCCCGC
  851	CCGCCCGCCT CTCGGCAACG GCAGAATCCG CCGCCGCCCT
  	GCTTGCCTCC
  901	GCCCTCTGCC TGACCGGCAT TTTCTCGCCC CTCGCCTCCC
  	TCCTGCTGCC
  951	GGAAAACTAC GCCGCCGTCC GGTTTATCGT CGTATCGTGT
  	ATGCTGCCTC
  1001	CGCTGTTTTG CACGCTGGTA GAAATCAGCG GCATCGGTTT
  	GAACGTCGTC
  1051	CGAAAAACAC GCCCGATCGC GCTCGCCACC TTGGGCGCGC
  	TGGCGGCAAA
  1101	CCTGCTGCTG CTGGGGCTTG CCGTACCGTC CGGCGGCGCG
  	CGCGGCGCGG
  1151	CGGTTGCCTG TGCCGCCTCA TTTTGGCTGT TTTTTGTTTT
  	CAAGACCGAA
  1201	AGCTCCTGCC GCCTGTGGCA GCCGCTCAAA CGCCTGCCGC
  	TTTATATGCA
  1251	CACATTGTTC TGCCTGGCCT CCTCGGCGGC CTACACCTGC
  	TTCGGCACTC
  1301	CGGCAAACTA CCCCCTGTTT GCCGGCGTAT GGGCGGTATA
  	TCTGGCAGGC
  1351	TGCATCCTGC GCCACCGGAA AGATTTGCAC AAACTGTTTC
  	ATTATTTGAA
  1401	AAAACAAGGT TTCCCATTAT GA

• This encodes a protein having amino acid sequence <SEQ ID 378>:

• 1	MDTKEILGYA AGSIGSAVLA VIILPLLSWY FPADDIGRIV
  	LMQTAAGLTV
  51	SVLCLGLDQA YVREYYAAAD KDTLFKTLFL PPLLSAAAIA
  	ALLLSRPSLP
  101	SEILFSLDDA AAGIGLVLFE LSFLPIRFLL LVLRMEGRAL
  	AFSSAQLVSK
  151	LAILLLLPLT VGLLHFPANT AVLTAVYALA NLAAAAFLLF
  	QNRCRLKAVR
  201	RAPFSSAVLH RGLRYGIPIA LSSIAYWGLA SADRLFLKKY
  	AGLEQLGVYS
  251	MGISFGGAAL LFQSIFSTVW TPYIFRAIEA NAPPARLSAT
  	AESAAALLAS
  301	ALCLTGIFSP LASLLLPENY AAVRFIVVSC MLPPLFCTLV
  	EISGIGLNVV
  351	RKTRPIALAT LGALAANLLL LGLAVPSGGA RGAAVACAAS
  	FWLFFVFKTE
  401	SSCRLWQPLK RLPLYMHTLF CLASSAAYTC FGTPANYPLF
  	AGVWAVYLAG
  451	CILRHRKDLH KLFHYLKKQG FPL*

• ORF10a and ORF10-1 show 95.4% identity in 475 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF10 shows 94.1% identity over a 475aa overlap with a predicted ORF (ORF10.ng) from N. gonorrhoeae:
• 
• The complete length ORF10ng nucleotide sequence <SEQ ID 379> is:

• 1	ATGGACACAA AAGAAATCCT CGGCTACGCG GCAGGCTCGA
  	TCGGCAGCGC
  51	GGTTTTAGCC GTCATCATCC TGCCGCTGCT GTCGTGGTAT
  	TTCcccgCCG
  101	ACGACATCGG GCGCATCGTG CTGATGCAGA CGGCGGCGGG
  	ACTGACGGTG
  151	TCGGTATTGT GCCTCGGGCT GGATCAGGCA TACGTCCGCG
  	AATACTATGC
  201	CGCCGCCGAC AAAGACACTT TGTTCAAAAC CCTGTTCCTG
  	CCGCCGCTGC
  251	TGTTTTCCGC CGCGATAGCC GCCCTGCTGC TTTCCCGCCC
  	GTCCCTGCCG
  301	TCTGAAATCC TGTTTTCGCT CGACGATGCC GCCGCCGGCA
  	TCGGGCTGGT
  351	GCTGTTTGAA CTGAGCTTCC TGCCCATCCG CTTTCTCTTA
  	CTGGTTTTGC
  401	GTATGGAAGG GCGCGCCCTT GCCTTTTCGT CCGCGCAACT
  	CGTGCCCAAA
  451	CTCGCCATTC TGCTGCTGTT GCCGCTGACG GTCGGGCTGC
  	TGCACTTTCC
  501	GGCGAACACC TCCGTCCTGA CCGCCGTTTA CGCGCTGGCA
  	AACCTTGCCG
  551	CCGCCGCCTT TTTGCTGTTT CAAAACCGAT GCCGTCTGAA
  	GGCCGTCCGG
  601	CGCGCGCCGT TTTCGCCCGC CGTCCTGCAC CGGGGGCTGC
  	GCTACGGCAT
  651	ACCGCTCGCA CTGAGCAGCC TTGCCTATTG GGGGCTGGCA
  	TCCGCCGACC
  701	GTTTGTTCCT GAAAAAATAT GCGGGCCTGG AACAGCTCGG
  	CGTTTATTCG
  751	ATGGGTATTT CGTTCGGCGG GGCGGCATTA TTGCTCCAAA
  	GCATCTTTTC
  801	AACGGTCTGG ACACCGTATA TTTTCCGTGC AATCGAAGAA
  	AACGCCACGC
  851	CCGCCCGCCT CTCGGCAACG GCAGAATCCG CCGCCGCCCT
  	GCTTGCCTCC
  901	GCCCTCTGCC TGACCGGAAT TTTCTCGCCC CTCGCCTCCC
  	TCCTGCTGCC
  951	GGAAAACTAC GCCGCCGTCC GGTTTACCGT CGTATCGTGT
  	ATGCTGccgc
  1001	cgctGTTTTA CACGCTGACC GAAATCAGCG GCATCGGTTT
  	GAACGTCGTC
  1051	CGCAAAACGC GTCCGATCGC GCTTGCCACC TTGGGCGCGC
  	TGGCGGCAAA
  1101	CCTGCTGCTG CTGGGGCTTG CCGTACCGTC CGGCGGCACG
  	CGCGGCGCGG
  1151	CGGTTGCCTG TGCCGCCTCA TTCTGGTTGT TTTTTGTTTT
  	CAAGACAGAA
  1201	AGCTCCTGCC GCCTGTGGCA GCCGCTCAAA CGCCTGCCGC
  	TTTATATGCA
  1251	CACATTGTTC TGCCTgGCCT CCTCGGCGGC CTACACCTGC
  	TTCGGCACAC
  1301	CGGCAAACTA CCCcctgttt gccggcgtAT GGGCGGCATA
  	TCTGGCAGGC
  1351	TGCATCCTGC GCCACCGGAA AAATTTGCAC AAACTGTTTC
  	ATTATTTGAA
  1401	AAAACAAGGT TTCCCATTAT GA

• This encodes a protein having amino acid sequence <SEQ ID 380>:

• 1	MDTKEILGYA AGSIGSAVLA VIILPLLSWY FPADDIGRIV
  	LMQTAAGLTV
  51	SVLCLGLDQA YVREYYAAAD KDTLFKTLFL PPLLFSAAIA
  	ALLLSRPSLP
  101	SEILFSLDDA AAGIGLVLFE LSFLPIRFLL LVLRMEGRAL
  	AFSSAQLVPK
  151	LAIL LLLPLT VGLLHFPANT SVLTAVYALA NLAAAAFLLF
  	QNRCRLKAVR
  201	RAPFSPAVLH RGLRYGIPLA LSSLAYWGLA SADRLFLKKY
  	AGLEQLGVYS
  251	MGISFGGAAL LLQSIFSTVW TPYIFRAIEE NATPARLSAT
  	AESAAALLAS
  301	ALCLTGIFSP LASLLLPENY AAVRFTVVSC MLPPLFYTLT
  	EISGIGLNVV
  351	RKTRPIALAT LGALAANLLL LGLAVPSGGT RGAAVACAAS
  	FWLFFVFKTE
  401	SSCRLWQPLK RLPLYMHTLF CLASSAAYTC FGTPANYPLF
  	AGVWAAYLAG
  451	CILRHRKNLH KLFHYLKKQG FPL*

• ORF10ng and ORF10-1 show 96.4% identity in 473 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader peptide and several transmembrane segments and the presence of a leucine-zipper motif(4 Leu residues spaced by 6 aa, shown in bold), it is predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 45
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 381>:

• 1	..ATCCTGAAAC CGCATAACCA GCTTAAGGAA GACATCCAAC
  	CTGATCCGGC
  51	CGATCAAAAC GCCTTGTCCG AACCGGATGC TGCGACAGAG
  	GCAGAGCAGT
  101	CGGATGCGGA AAATGCTGCC GACAAGCAGC CCGTTGCCGA
  	TAAAGCCGAC
  151	GAGGTTGAAG AAAAGGCGGG CGAGCCGGAA CGGGAAGAGC
  	CGGACGGACA
  201	GGCAGTGCGT AAGAAAGCGC TGACGGAAGA GCGTGAACAA
  	ACCGTCAGGG
  251	AAAAAGCGCA GAAGAAAGAT GCCGAAACGG TTAAAATACA
  	AGCGGTAAAA
  301	CCGTCTAAAG AAACAGAGAA AAAAGCTTCA AAAGAAGAGA
  	AAAAGGCGGC
  351	GAAGGAAAAA GTTGCACCCA AACCAACCCC GGAACAAATC
  	CTCAACAGCG
  401	GCAgCATCGA AAAmGCGCGC AgTGCCGCCG CCAAAGAAGT
  	GCAGAAAATG
  451	AA.AACGTCC GACAAGGCGG AAGC.AACGC ATTATCTGCA
  	AATGGGCGCG
  501	TATGCCGACC GTCAGAGCGC GGAAGGGCAG CGTGCCAAAC
  	TGGCAATCTT
  551	GGGCATATCT TCCAAGGTGG TCGGTTATCA GGCGGGACAT
  	AAAACGCTTT
  601	ACCGGGTGCA AAGCGGCAAT ATGTCTGCCG ATGCGGTGA

• This corresponds to the amino acid sequence <SEQ ID 382; ORF65>:

• 1	..ILKPHNQLKE DIQPDPADQN ALSEPDAATE AEQSDAENAA
  	DKQPVADKAD
  51	EVEEKAGEPE REEPDGQAVR KKALTEEREQ TVREKAQKKD
  	AETVKIQAVK
  101	PSKETEKKAS KEEKKAAKEK VAPKPTPEQI LNSGSIEXAR
  	SAAAKEVQKM
  151	XNVRQGGSXR IICKWARMPT VRARKGSVPN WQSWAYLPRW
  	SVIRRDIKRF
  201	TGCKAAICLP MR*

• Further work revealed the complete nucleotide sequence <SEQ ID 383>:

• 1	ATGTTTATGA ACAAATTTTC CCAATCCGGA AAAGGTCTGT
  	CCGGTTTTTT
  51	CTTCGGTTTG ATACTGGCGA CGGTCATTAT TGCCGGTATT
  	TTGTTTTATC
  101	TGAACCAGAG CGGTCAAAAT GCGTTCAAAA TCCCGGCTTC
  	GTCGAAGCAG
  151	CCTGCAGAAA CGGAAATCCT GAAACCGAAA AACCAGCCTA
  	AGGAAGACAT
  201	CCAACCTGAA CCGGCCGATC AAAACGCCTT GTCCGAACCG
  	GATGCTGCGA
  251	CAGAGGCAGA GCAGTCGGAT GCGGAAAAAG CTGCCGACAA
  	GCAGCCCGTT
  301	GCCGATAAAG CCGACGAGGT TGAAGAAAAG GCGGGCGAGC
  	CGGAACGGGA
  351	AGAGCCGGAC GGACAGGCAG TGCGTAAGAA AGCGCTGACG
  	GAAGAGCGTG
  401	AACAAACCGT CAGGGAAAAA GCGCAGAAGA AAGATGCCGA
  	AACGGTTAAA
  451	AAACAAGCGG TAAAACCGTC TAAAGAAACA GAGAAAAAAG
  	CTTCAAAAGA
  501	AGAGAAAAAG GCGGCGAAGG AAAAAGTTGC ACCCAAACCA
  	ACCCCGGAAC
  551	AAATCCTCAA CAGCGGCAGC ATCGAAAAAG CGCGCAGTGC
  	CGCCGCCAAA
  601	GAAGTGCAGA AAATGAAAAC GTCCGACAAG GCGGAAGCAA
  	CGCATTATCT
  651	GCAAATGGGC GCGTATGCCG ACCGTCAGAG CGCGGAAGGG
  	CAGCGTGCCA
  701	AACTGGCAAT CTTGGGCATA TCTTCCAAGG TGGTCGGTTA
  	TCAGGCGGGA
  751	CATAAAACGC TTTACCGGGT GCAAAGCGGC AATATGTCTG
  	CCGATGCGGT
  801	GAAAAAAATG CAGGACGAGT TGAAAAAACA TGAAGTCGCC
  	AGCCTGATCC
  851	GTTCTATCGA AAGCAAATAA

• This corresponds to the amino acid sequence <SEQ ID 384; ORF65-1>:

• 1	MFMNKFSQSG KGLSGFFFGL ILATVIIAGI LFYLNQSGQN
  	AFKIPASSKQ
  51	PAETEILKPK NQPKEDIQPE PADQNALSEP DAATEAEQSD
  	AEKAADKQPV
  101	ADKADEVEEK AGEPEREEPD GQAVRKKALT EEREQTVREK
  	AQKKDAETVK
  151	KQAVKPSKET EKKASKEEKK AAKEKVAPKP TPEQILNSGS
  	IEKARSAAAK
  201	EVQKMKTSDK AEATHYLQMG AYADRQSAEG QRAKLAILGI
  	SSKVVGYQAG
  251	HKTLYRVQSG NMSADAVKKM QDELKKHEVA SLIRSIESK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF65 shows 92.0% identity over a 150aa overlap with an ORF (ORF65a) from strain A of N. meningitidis:
• 
• The complete length ORF65a nucleotide sequence <SEQ ID 385> is:

• 1	ATGTTTATGA ACAAATTTTC CCAATCCGGA AAAGGTCTGT
  	CCGGTTTTTT
  51	CTTCGGTTTG ATACTGGCGA CGGTCATTAT TGCCGGTATT
  	TTGTTTTATC
  101	TGAACCAGAG CGGTCAAAAT GCGTTCAAAA TCCCGGTTCC
  	GTCGAAGCAG
  151	CCTGCAGAAA CGGAAATCCT GAAACCGAAA AACCAGCCTA
  	AGGAAGACAT
  201	CCAACCTGAA CCGGCCGATC AAAACGCCTT GTCCGAACCG
  	GATGCTGCGA
  251	AAGAGGCAGA GCAGTCGGAT GCGGAAAAAG CTGCCGACAA
  	GCAGCCCGTT
  301	GCCGACAAAG CCGACGAGGT TGAGGAAAAG GCGGACGAGC
  	CGGAGCGGGA
  351	AAAGTCGGAC GGACAGGCAG TGCGCAAGAA AGCACTGACG
  	GAAGAGCGTG
  401	AACAAACCGT CGGGGAAAAA GCGCAGAAGA AAGATGCCGA
  	AACGGTTAAA
  451	AAACAAGCGG TAAAACCATC TAAAGAAACA GAGAAAAAAG
  	CTTCAAAAGA
  501	AGAGAAAAAG GCGGAGAAGG AAAAAGTTGC ACCCAAACCG
  	ACCCCGGAAC
  551	AAATCCTCAA CAGCGGCAGC ATCGAAAAAG CGCGCAGTGC
  	CGCTGCCAAA
  601	GAAGTGCAGA AAATGAAAAC GCCCGACAAG GCGGAAGCAA
  	CGCATTATCT
  651	GCAAATGGGC GCGTATGCCG ACCGCCGGAG CGCGGAAGGG
  	CAGCGTGCCA
  701	AACTGGCAAT CTTGGGCATA TCTTCCAAGG TGGTCGGTTA
  	TCAGGCGGGA
  751	CATAAAACGC TTTACCGGGT GCAAAGCGGC AATATGTCTG
  	CCGATGCGGT
  801	GAAAAAAATG CAGGACGAGT TGAAAAAACA TGAAGTCGCC
  	AGCCTGATCC
  851	GTTCTATCGA AAGCAAATAA

• This encodes a protein having amino acid sequence <SEQ ID 386>:

• 1	MFMNKFSQSG KGLSGFFFGL ILATVIIAGI LFYLNQSGQN
  	AFKIPVPSKQ
  51	PAETEILKPK NQPKEDIQPE PADQNALSEP DAAKEAEQSD
  	AEKAADKQPV
  101	ADKADEVEEK ADEPEREKSD GQAVRKKALT EEREQTVGEK
  	AQKKDAETVK
  151	KQAVKPSKET EKKASKEEKK AEKEKVAPKP TPEQILNSGS
  	IEKARSAAAK
  201	EVQKMKTPDK AEATHYLQMG AYADRRSAEG QRAKLAILGI
  	SSKVVGYQAG
  251	HKTLYRVQSG NMSADAVKKM QDELKKHEVA SLIRSIESK*

• ORF65a and ORF65-1 show 96.5% identity in 289 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF65 shows 89.6% identity over a 212aa overlap with a predicted ORF (ORF65.ng) from N. gonorrhoeae:
• 
• An ORF65ng nucleotide sequence <SEQ ID 387> was predicted to encode a protein having amino acid sequence <SEQ ID 388>:

• 1	MFMNKFSQSG KGLSGFFFGL ILATVIIAGI LLYLNQGGQN
  	AFKIPAPSKQ
  51	PAETEILKLK NQPKEDIQPE PADQNALSEP DVAKEAEQSD
  	AEKAADKQPV
  101	ADKADEVEEK AGEPEREEPD GQAVRKKALT EEREQTVREK
  	AQKKDAETVK
  151	KKAVKPSKET EKKASKEEKK AAKEKVAPKP TPEQILNSRS
  	IEKARSAAAK
  201	EVQKMKNFGQ GGSQRIICKW ARMPNPGARK GSVPNWQSWA
  	YLPKWSAIRR
  251	DIKRFTACKA AICPPMR*

• After further analysis, the complete gonococcal DNA sequence <SEQ ID 389> was found to be:

• 1	ATGTTTATGA ACAAATTTTC CCAATCCGGA AAAGGTCTGT
  	CCGGTTTCTT
  51	CTTCGGTTTG ATACTGGCAA CGGTCATTAT TGCCGGTATT
  	TTGCTTTATC
  101	TGAACCAGGG CGGTCAAAAT GCGTTCAAAA TCCCGGCTCC
  	GTCGAAGCAG
  151	CCTGCAGAAA CGGAAATCCT GAAACTGAAA AACCAGCCTA
  	AGGAAGACAT
  201	CCAACCTGAA CCGGCCGATC AAAACGCCTT GTCCGAACCG
  	GATGTTGCGA
  251	AAGAGGCAGA GCAGTCGGAT GCGGAAAAAG CTGCCGACAA
  	GCAGCCCGTT
  301	GCCGACAAag ccgacgAGGT TGAAGAAAag GcGGgcgAgc
  	cggaACGGga
  351	aGAGCCGGAC ggACAGGCAG TGCGCAAGAA AGCACTGAcg
  	gAAGAgcGTG
  401	AACAAACcgt cagggAAAAA GCGCagaaga AAGATGCCGA
  	AACGgTTAAA
  451	AAacaaGCgg tAaaaccgtc tAAAGAAACa gagaaaaaag
  	cTtcaaaaga
  501	agagaaaaag gcggcgaaag aaaAAGttgc acccaaaccg
  	accccggaaC
  551	aaatcctcaa cagccgCagc atcgaaaaag cgcgtagtgc
  	cgctgccaaa
  601	gaAgtgcaGA AAatgaaaaa ctTtgggcaa ggcgGaagcc
  	aacgcattaT
  651	CTGcaaatgg gcgcgtatgc cgaccgtccg gagcgcggaA
  	gggcagcgtg
  701	ccaaACtggc aAtcttgGgc atatctTccg aagtggtcgG
  	CTATCAGGCG
  751	GGACATAAAA CGCTTTACCG CGTGCAAagc GGCAatatgt
  	ccgccgatgc
  801	gGTGAAAAAA ATGCAGGACG AGTTGAAAAA GCATGGGGtt
  	gcCAGCCTGA
  851	TCCGTGcgAT TGAAGGCAAA TAA

• This encodes the following amino acid sequence <SEQ ID 390>:

• 1	MFMNKFSQSG KGLSGFFFGL ILATVIIAGI LLYLNQGGQN
  	AFKIPAPSKQ
  51	PAETEILKLK NQPKEDIQPE PADQNALSEP DVAKEAEQSD
  	AEKAADKQPV
  101	ADKADEVEEK AGEPEREEPD GQAVRKKALT EEREQTVREK
  	AQKKDAETVK
  151	KQAVKPSKET EKKASKEEKK AAKEKVAPKP TPEQILNSRS
  	IEKARSAAAK
  201	EVQKMKNFGQ GGSQRIICKW ARMPTVRSAE GQRAKLAILG
  	ISSEVVGYQA
  251	GHKTLYRVQS GNMSADAVKK MQDELKKHGV ASLIRAIEGK
  	*

• ORF65ng-1 and ORF65-1 show 89.0% identity in 290 aa overlap:
• 
• On this basis, including the presence of a putative transmembrane domain in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 46
• The following DNA sequence, believed to be complete, was identified in N. meningitidis <SEQ ID 391>:

• 1	ATGAACCACG ACATCACTTT CCTCACCCTG TTCCTACTCG
  	GTkTCTTCGG
  51	CGGAAcGCAC TGCATCGGTA TGTGCGGCGG ATTAAGCAGC
  	GcGTTTGs.s
  101	TCCAACTCCC CCCGCATATC AACCGCTTTT GGCTGATCCT
  	GCTGCTTAAC
  151	ACAGGACGGG TAAGCAGCTA TACGGCAAtC GGCCTGATAC
  	TCGGATTAAT
  201	CGGACAGGTC GGCGTTTCAC TCGAcCAaAC CCGCGTCCTG
  	CAGAATATTT
  251	TATACACGGC CGCCAACCTC CTGCTGCTCT TTTTAGGCTT
  	ATACTTGAGC
  301	GGTATTTCTT CCTTGGCGGC AAAAATCGAG AAaATCGGCA
  	AACCGATATG
  351	GCGGAACCTG AACCCGATAC TCAACCGGCT GTTACCCATA
  	AAATCCATAC
  401	CCGCCTGCCT tGCGgTCGGA ATATTATGGG GCTGGCTGCC
  	GTGCGGACTG
  451	GTTTACAGCG CGTCGCTTTA CGCGCTGGGA AgCGGTAGTG
  	CGGCAACGGG
  501	CGGGTTATAT ATGCTTGCCT TTGCACTGGG TACGCTGCCC
  	AATCTTtTAG
  551	CAATCGGCAT TTTtTCCCTG CAACTGAAwA AAATCATGCA
  	AAACCGATAT
  601	ATCCGCCTGT GTACGGGATT ATCCGTATCA TTATGGGCAT
  	TATGGAAACT
  651	TGCCGTCCTG TGGCTGTAA

• This corresponds to the amino acid sequence <SEQ ID 392; ORF103>:

• 1	MNHDITFLTL FLLGXFGGTH CIGMCGGLSS AFXXQLPPHI
  	NRFWLILLLN
  51	TGRVSSYTAI GLILGLIGQV GVSLDQTRVL QNILYTAANL
  	LLLFLGLYLS
  101	GISSLAAKIE KIGKPIWRNL NPILNRLLPI KSIPACLAVG
  	ILWGWLPCGL
  151	VYSASLYALG SGSAATGGLY MLAFALGTLP NLLAIGIFSL
  	QLXKIMQNRY
  201	IRLCTGLSVS LWALWKLAVL WL*

• Further work elaborated the DNA sequence <SEQ ID 393> as:

• 1	ATGAACCACG ACATCACTTT CCTCACCCTG TTCCTACTCG
  	GTTTCTTCGG
  51	CGGAACGCAC TGCATCGGTA TGTGCGGCGG ATTAAGCAGC
  	GCGTTTGCGC
  101	TCCAACTCCC CCCGCATATC AACCGCTTTT GGCTGATCCT
  	GCTGCTTAAC
  151	ACAGGACGGG TAAGCAGCTA TACGGCAATC GGCCTGATAC
  	TCGGATTAAT
  201	CGGACAGGTC GGCGTTTCAC TCGACCAAAC CCGCGTCCTG
  	CAGAATATTT
  251	TATACACGGC CGCCAACCTC CTGCTGCTCT TTTTAGGCTT
  	ATACTTGAGC
  301	GGTATTTCTT CCTTGGCGGC AAAAATCGAG AAAATCGGCA
  	AACCGATATG
  351	GCGGAACCTG AACCCGATAC TCAACCGGCT GTTACCCATA
  	AAATCCATAC
  401	CCGCCTGCCT TGCGGTCGGA ATATTATGGG GCTGGCTGCC
  	GTGCGGACTG
  451	GTTTACAGCG CGTCGCTTTA CGCGCTGGGA AGCGGTAGTG
  	CGGCAACGGG
  501	CGGGTTATAT ATGCTTGCCT TTGCACTGGG TACGCTGCCC
  	AATCTTTTAG
  551	CAATCGGCAT TTTTTCCCTG CAACTGAAAA AAATCATGCA
  	AAACCGATAT
  601	ATCCGCCTGT GTACGGGATT ATCCGTATCA TTATGGGCAT
  	TATGGAAACT
  651	TGCCGTCCTG TGGCTGTAA

• This corresponds to the amino acid sequence <SEQ ID 394; ORF103-1>:

• 1	MNHDITFLTL FLLGFFGGTH CIGMCGGLSS AFALQLPPHI
  	NRFWLILLLN
  51	TGRVSSYTAI GLILGLIGQV GVSL DQTRVL QNILYTAANL
  	LLLFLGLYLS
  101	GISSLAAKIE KIGKPIWRNL NPILNRLLPI KSIPACLAVG
  	ILWGWLPCGL
  151	VYSASLYALG SGSAATGGLY MLAFALGTLP NLLAIGIFSL
  	QLKKIMQNRY
  201	IRLCTGLSVS LWALWKLAVL WL*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF103 shows 93.8% identity over. a 222aa overlap with an ORF (ORF103a) from strain A of N. meningitidis:
• 
• The complete length ORF103a nucleotide sequence <SEQ ID 395> is:

• 1	ATGAACCANG ACATCACTTT CCTCACCCTG TTCCTACTCG
  	GTTTCTTCGG
  51	CGGAACGCAC TGCATCGGTA TGTGCGGCGG ATTAAGCAGC
  	GCGTTTGCGC
  101	TCCAACTCCC CCCGCATATC AACCGCTTNT GGCTGATCCT
  	GCTGCTTAAC
  151	ACAGGACGGG TAAGCAGCTA TACGGCAATC GGCCTGATAC
  	TCGGATTAAT
  201	CGGACAGGTC GGCGTTTCAC TCGACCAAAC CCGCGTCNTG
  	CAGAATATTT
  251	TATACACGGC CGCCAACCTC CTGCTGCTCT TTTTAGGCTT
  	ATACTTGAGC
  301	GGTATTTCTT CCTTGGCGGC AAAAATCGAG AAAATCGGCA
  	AACCGATATG
  351	GCGGAACCTG AACCCGATAC TCAACCGGCT GTTACCCATA
  	AAATCCATAC
  401	CCGCCTGCCT TGCGGTCGGA ATATTATGGG GCTGGCTGCC
  	GTGCGGACTA
  451	GTTTACAGCG CGTCGCTTTA CGCGCTGGGA AGCGGTAGTG
  	CGGCAACGGG
  501	CGGGTTATAT ATGCTTGCCT TTGCACTGGG TACGCTGCCC
  	AATCTTTNGG
  551	CAATCGGCAT TTTTTCCCTG CAACTGNAAA AAATCATGCA
  	AAACCGATAT
  601	ATCCGCCTGT GTACGGGATT ATCCGTATCA TTATGGGCAT
  	TATGGAAACT
  651	TGCCGTCCTG TGGCTGTAA

• This encodes a protein having amino acid sequence <SEQ ID 396>:

• 1	MNXDITFLTL FLLGFFGGTH CIGMCGGLSS AFALQLPPHI
  	NRXWLILLLN
  51	TGRVSSYTAI GLILGLIGQV GVSL DQTRVX QNILYTAANL
  	LLLFLGLYLS
  101	GISSLAAKIE KIGKPIWRNL NPILNRLLPI KSIPACLAVG
  	ILWGWLPCGL
  151	VYSASLYALG SGSAATGGLY MLAFALGTLP NLXAIGIFSL
  	QLXKIMQNRY
  201	IRLCTGLSVS LWALWKLAVL WL*

• ORF103a and ORF103-1 show 97.7% identity in 222 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF103 shows 95.5% identity over a 222aa overlap with a predicted ORF (ORF103.ng) from N. gonorrhoeae:
• 
• The complete length ORF103ng nucleotide sequence <SEQ ID 397> is:

• 1	ATGAACCACG ACATCACTTT CCTCACCCTG TTCCTGCTCG
  	GTTTCTTCGG
  51	CGGAACTCAC TGCATCGGTA TGTGCGGCGG ATTAAGCAGC
  	GCGTTTGCGC
  101	TCCAACTCCC CCCGCATATC AACCGCTTTT GGCTGATTCT
  	GCTGCTTAAC
  151	ACAGGACGGA TAAGCAGCTA TACGGCAATC GGCCTGATGC
  	TCGGATTAAT
  201	CGGACAACTC GGCATTTCAC TCGACCAAAc ccgcgTCCTG
  	CAAAATATTT
  251	tatacacagc ctccaaCCTC CTGCTGCTCT TTTTAGGCTT
  	ATACTTGAGC
  301	GGTATTTCTT CCTTGGCGGC AAAAATCGAG AAAATCGGCA
  	AACCGATATG
  351	GCGCAACCTG AACCCGATAC TCAACCGGCT GCTGCCCATA
  	AAATCCATAC
  401	CCGCCTGCCT TGCTGTCGGA ATATTATGGG GCTGGCTGCC
  	GTGCGGACTG
  451	GTTTACAGCG CATCACTTTA CGCGCTGGGA AGCGGTAGTG
  	CGACAACCGG
  501	CGGACTGTAT ATGCTTGCCT TTGCACTGGG TACGCTGCCC
  	AATCTTTTGG
  551	CAATCGGCAT TTTTTCCCTG CAACTGAAAA AAATCATGCA
  	AAACCGATAT
  601	ATCCGCCTGT GTACAGGATT ATCCGTATCA TTATGGGCAT
  	TATGGAAGCT
  651	TGCCGTCCTG TGGCTGTAA

• This encodes a protein having amino acid sequence <SEQ ID 398>:

• 1	MNHDITFLTL FLLGFFGGTH CIGMCGGLSS AFALQLPPHI
  	NRFWLILLLN
  51	TGRISSYTAI GLMLGLIGQL GISL DQTRVL QNILYTASNL
  	LLLFLGLYLS
  101	GISSLAAKIE KIGKPIWRNL NPILNRLLPI KSIPACLAVG
  	ILWGWLPCGL
  151	VYSASLYALG SGSATTGGLY MLAFALGTLP NLLAIGIFSL
  	QLKKIMQNRY
  201	IRLCTGLSVS LWALWKLAVL WL*

• In addition, ORF103ng and ORF103-1 show 97.3% identity in 222 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence (double-underlined) and several putative transmembrane domains (single-underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 47
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 399>:

• 1	ATGGAAAACC AAAGGCCGCT CCTAGGCTTT CGCTTGGCAC
  	TTTTGGCGGC
  51	GATGACGTGG GGAACGCTGC CGAT.TCCGT GCGGCAGGTA
  	TTGAAGTTTG
  101	TCGATGCGCC GACGCTGGTG TGGGTGCGTT TTACCGTGGC
  	GGCGGCGGTA
  151	TTGTTTGTTT TGCTGGCACT GGGCGGGCGG CTGCcGAAGC
  	GGCGaGGATT
  201	TTTCTTGGTG CTCATTCAGG CTGCTGCTGC TCGGCGTGGC
  	GGGCATTTCG
  251	GCAAACTTTG TGCTGATTGC CCAAGGGCTG CATTATATTT
  	CGCCGACCAC
  301	GACGCAGGTT TTGTGGCAGA TTTCGCCGTT TACGATGATT
  	GTwGTCGGTG
  351	TGTTGGTGTT TAAAGACCGG ATGACTGCCG CTCAGAAAAT
  	CGGCTTGGTT
  401	TTGCTGCTTG CCGGTTTGCT TATGTATTTT AACGATAAAT
  	TCGGCGAGTT
  451	GTCGGGTTTG GGCGCGTATG C.AAGGGCGT GTTGCTGTGT
  	GCGGCAGGCA
  501	GTATGGCATG GGTGTGTAAT GCCGTGGCGC AAAAGCTGCT
  	GTCGGCGCAA
  551	TTCGGGCCGC AACAGATTCT GCTGTTGATT TATGCGGCAA
  	GTGCCGCCGT
  601	GTTCCTGCCG TTTGCCGAAC CGGCACACAT CGGAAGTATG
  	GACGGTACGT
  651	TGGCGTGGGT ATGTATTGCG TATTGCTGCT TGAATACGTT
  	AATCGGTTAC
  701	GGCTCGTTCG GCGAGGCGTT GAAACATTGG GAGGCTTCCA
  	AAGTCAGCGC
  751	GGTAACAACC TTGCTCCCCG TGTTTACCGT AATAAATACT
  	TTGCTCGGGC
  801	ATTATGTGAT GCCTGAAACT TTTGCCGCGC CGGA..

• This corresponds to the amino acid sequence <SEQ ID 400; ORF104>:

• 1	MENQRPLLGF RLALLAAMTW GTLPXSVRQV LKFVDAPTLV
  	WVRFTVAAAV
  51	LFVLLALGGR LPKRRDFSWC SFRLLLLGVA GISANFVLIA
  	QGLHYISPTT
  101	TQVLWQISPF TMIVVGVLVF KDRMTAAQKI GLVLLLAGLL
  	MYFNDKFGEL
  151	SGLGAYXKGV LLCAAGSMAW VCNAVAQKLL SAQFGPQQIL
  	LLIYAASAAV
  201	FLPFAEPAHI GSMDGTLAWV CIAYCCLNTL IGYGSFGEAL
  	KHWEASKVSA
  251	VTTLLPVFTV INTLLGHYVM PETFAAP...

• Further work revealed further, partial DNA sequence <SEQ ID 401>:

• 1	ATGGAAAACC AAAGGCCGCT CCTAGGCTTC GCGTTGGCAC
  	TTTTGGCGGC
  51	GATGACGTGG GGAACGCTGC CGATTGCCGT GCGGCAGGTA
  	TTGAAGTTTG
  101	TCGATGCGCC GACGCTGGTG TGGGTGCGTT TTACCGTGGC
  	GGCGGCGGTA
  151	TTGTTTGTTT TGCTGGCACT GGGCGGGCGG CTGCCGAAGC
  	GGCGGGATTT
  201	TTCTTGGTGC TCATTCAGGC TGCTGCTGCT CGGCGTGGCG
  	GGCATTTCGG
  251	CAAACTTTGT GCTGATTGCC CAAGGGCTGC ATTATATTTC
  	GCCGACCACG
  301	ACGCAGGTTT TGTGGCAGAT TTCGCCGTTT ACGATGATTG
  	TTGTCGGTGT
  351	GTTGGTGTTT AAAGACCGGA TGACTGCCGC TCAGAAAATC
  	GGCTTGGTTT
  401	TGCTGCTTGC CGGTTTGCTT ATGTTTTTTA ACGATAAATT
  	CGGCGAGTTG
  451	TCGGGTTTGG GCGCGTATGC GAAGGGCGTG TTGCTGTGTG
  	CGGCAGGCAG
  501	TATGGCATGG GTGTGTTATG CCGTGGCGCA AAAGCTGCTG
  	TCGGCGCAAT
  551	TCGGGCCGCA ACAGATTCTG CTGTTGATTT ATGCGGCAAG
  	TGCCGCCGTG
  601	TTCCTGCCGT TTGCCGAACC GGCACACATC GGAAGTTTGG
  	ACGGTACGTT
  651	GGCGTGGGTT TGTTTTGCGT ATTGCTGCTT GAATACGTTA
  	ATCGGTTACG
  701	GCTCGTTCGG CGAGGCGTTG AAACATTGGG AGGCTTCCAA
  	AGTCAGCGCG
  751	GTAACAACCT TGCTCCCCGT GTTTACCGTA ATAwTwwCTT
  	TGCTCGGGCA
  801	TTATGTGATG CCTGAAACTT TTGCCGCGCC GGA...

• This corresponds to the amino acid sequence <SEQ ID 402; ORF104-1>:

• 1	MENQRPLLGF ALALLAAMTW GTLPIAVRQV LKFVDAPTLV
  	WVRFTVAAAV
  51	LFVLLALGGR LPKRRDFSWC SFRLLLLGVA GISANFVLIA
  	QGLHYISPTT
  101	TQVLWQISPF TMIVVGVLVF KDRMTAAQKI GLVLLLAGLL
  	MFFNDKFGEL
  151	SGLGAYAKGV LLCAAGSMAW VCYAVAQKLL SAQFGPQQIL
  	LLIYAASAAV
  201	FLPFAEPAHI GSLDGTLAWV CFAYCCLNTL I GYGSFGEAL
  	KHWEASKVSA
  251	VTTLLPVFTV IXXLLGHYVM PETFAAP...

• Computer analysis of this amino acid sequence gave the following results:
• Homology with Hypothetical HI0878 Protein of H. influenzae (Accession Number U32769)
• ORF104 and HI0878 show 40% aa identity in 277aa overlap:

• orf104	4	QRPLLGFRLALLAAMTWGTLPXSVRQVLKFVDAPTLVWXXXXXXXXXXXXXXXXXXXXP-	62
  		Q+PLLGF  AL+ AM WG+LP +++QVL  ++A T+VW                    P
  HI0878	3	QQPLLGFTFALITAMAWGSLPIALKQVLSVMNAQTIVWYRFIIAAVSLLALLAYKKQLPE	62
  orf104	63	--KRRDFSWCSFRLLLLGVAGISANFVLIAQGLHYISPTTTQVLWQISPFTMIVVGVLVF	120
  		K R ++W    ++L+GV G+++NF+L +  L+YI P+  Q+   +S F M++ GVL+F
  HI0878	63	LMKVRQYAW----IMLIGVIGLTSNFLLFSSSLNYIEPSVAQIFIHLSSFGMLICGVLIF	118
  orf104	121	KDRMTAAQKIXXXXXXXXXXMYFNDKFGELSGLGAYXKGVLLCAAGSMAWVCNAVAQKLL	180
  		K+++   QKI          ++FND+F   +GL  Y  GV+L   G++ WV   +AQKL+
  HI0878	119	KEKLGLHQKIGLFLLLIGLGLFFNDRFDAFAGLNQYSTGVILGVGGALIWVAYGMAQKLM	178
  orf104	181	SAQFGPQQILLLIYAASAAVFLPFAEPAHIGSMDGTLAWVCIAYCCLNTLIGYGSFGEAL	240
  		+F  QQILL++Y   A  F+P A+ + +  +   LA +C  YCCLNTLIGYGS+ EAL
  HI0878	179	LRKFNSQQILLMMYLGCAIAFMPMADFSQVQELT-PLALICFIYCCLNTLIGYGSYAEAL	237
  orf104	241	KHWEASKVSAVTTLLPVFTVINTLLGHYVMPETFAAP	277
  		W+ SKVS V TL+P+FT++ + + HY  P  FAAP
  HI0878	238	NRWDVSKVSVVITLVPLFTILFSHIAHYFSPADFAAP	274

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF104 shows 95.3% identity over a 277aa overlap with an ORF (ORF104a) from strain A of N. meningitidis:
• 
• The complete length ORF104a nucleotide sequence <SEQ ID 403> is:

• 1	ATGGAAAACC AAAGGCCGCT CCTAGGCTTC GCGTTGGCAC
  	TTTTGGCGGC
  51	GATGACGTGG GGAACGCTGC CGATTGCCGT GCGGCAGGTA
  	TTGAAGTTTG
  101	TCGATGCGCC GACGCTGGTG TGGGTGCGTT TTACCGTGGC
  	GGCGGCGGTA
  151	TTGTTTGTTT TGCTGGCATT GGGCGGGCGG CTGCCGAAGT
  	GGCGGGATTT
  201	TTCTTGGTGC TCATTCAGGC TGCTGCTGCT CGGCGTGGCG
  	GGCATTTCGG
  251	CAAACTTTGT GCTGATTGCC CAAGGGCTGC ATTATATTTC
  	GCCGACCACG
  301	ACGCAGGTTT TGTGGCAGAT TTCGCCGTTT ACGATGATTG
  	TTGTCGGTGT
  351	GTTGGTGTTT AAAGACCGGA TGACTGCCGC TCAGAAAATC
  	GGCTTGGTTT
  401	TGCTGCTTGC CGGTTTGCTT ATGTTTTTTA ACGATAAATT
  	CGGCGAGTTG
  451	TCGGGTTTGG GCGCGTATGC GAAGGGCGTG TTGCTGTGTG
  	CGGCAGGCAG
  501	TATGGCATGG GTGTGTTATG CCGTGGCGCA AAAGCTGCTG
  	TCGGCGCAAT
  551	TCGGGCCGCA ACAGATTCTG CTGTTGATTT ATGCGGCAAG
  	TGCCGCCGTG
  601	TTCCTGCCGT TTGCCGAACT GGCACACATC GGAAGTTTGG
  	ACGGTACGTT
  651	GGCGTGGGTT TGTTTTGCGT ATTGCTGCTT GAATACGTTA
  	ATCGGTTACG
  701	GCTCGTTCGG CGAGGCGTTG AAACATTGGG AGGCTTCCAA
  	AGTCAGCGCG
  751	GTAACAACCT TGCTCCCCGT GTTTACCGTA ATATTTTCTT
  	TGCTCGGGCA
  801	TTATGTGATG CCTGATACTT TTGCCGCGCC GGATATGAAC
  	GGTTTGGGTT
  851	ATGCCGGCGC ACTGGTCGTG GTCGGGGGTG CGGTTACGGC
  	GGCGGTGGGG
  901	GACAGGCTGT TCAAACGCCG CTAG

• This encodes a protein having amino acid sequence <SEQ ID 404>:

• 1	MENQRPLLGF ALALLAAMTW GTLPIAVRQV LKFVDAPTLV
  	WVRFTVAAAV
  51	LFVLLALGGR LPKWRDFSWC SFRLLLLGVA GISANFVLIA
  	QGLHYISPTT
  101	TQVLWQISPF TMIVVGVLVF KDRMTAAQKI GLVLLLAGLL
  	MFFNDKFGEL
  151	SGLGAYAKGV LLCAAGSMAW VCYAVAQKLL SAQFGPQQIL
  	LLIYAASAAV
  201	FLPFAELAHI GSLDGTLAWV CFAYCCLNTL I GYGSFGEAL
  	KHWEASKVSA
  251	VTTLLPVFTV IFSLLGHYVM PDTFAAPDMN GLGYAGALVV
  	VGGAVTAAVG
  301	DRLFKRR*

• ORF104a and ORF104-1 show 98.2% identity in 277 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF104 shows 93.9% identity over a 277aa overlap with a predicted ORF (ORF104.ng) from N. gonorrhoeae:
• 
• The complete length ORF104ng nucleotide sequence <SEQ ID 405> is predicted to encode a protein having amino acid sequence <SEQ ID 406>:

• 1	MENQRPLLGF ALALLAAMTW GTLPIAVRQV LKFVDAPTLV
  	WVRFTVAAAV
  51	LFVLLALGGR LPKRRDFSWH SFRLLLLGVT GISANFVLIA
  	QGLHYISPTT
  101	TQVLWQISPF TMIVVGVLVF KDRMTAAQKI GLVLLLVGLL
  	MFFNDKFGEL
  151	SGLGAYAKGV LLCAAGSMAW VCYAVAQKLL SAQFGPQQIL
  	LLIYAASAAV
  201	FLLXAEPAHI GSLDGTLAWV CFVYCCLNTL  IGYGSFGEAL
  	KHWEASKVSA
  251	VTTLLPVFTV IFSLLGHYVM PDTFAAPDMN GLGYVGALVV
  	VGGAVTAAVG
  301	DRPFKRR*

• Further work revealed the complete gonococcal nucleotide sequence <SEQ ID 407>:

• 1	ATGGAAAACC AAAGGCCGCT CCTAGGCTTC GCGTTGGCAC
  	TTTTGGCGGC
  51	GATGACGTGG GGGACGCTGC CGATTGCCGT GCGGCAGGTA
  	TTGAAGTTTG
  101	TCGATGCGCC GACGCTGGTG TGGGTGCGTT TTACCGTGGC
  	GGCGGCGGTA
  151	TTGTTTGTTT TGCTGGCATT GGGCGGGCGG CTGCCGAAGC
  	GGCGGGATTT
  201	TTCTTGGCAT TCATTCAGGC TGCTGCTGCT CGGCGTGACG
  	GGCATTTCGG
  251	CAAACTTTGT GCTGATTGCC CAAGGGCTGC ATTATATTTC
  	GCCGACCACG
  301	ACGCAGGTTT TGTGGCAGAT TTCGCCGTTT ACGATGATTG
  	TTGTCGGCGT
  351	GTTGGTGTTT AAAGACCGGA tgaCTGCCGC GCAGAAAATC
  	GGTTTGGTTT
  401	TGCTGCttgT CGGTttgCTT ATGTTTTtta ACGACAAATT
  	CGGCGAGTTG
  451	TCGGGTTTGG GCGCGTATGC GAAGGGCGTG TTGCTGTGTG
  	CGGCAGGCAG
  501	TATGGCCTGG GTGTGTTATG CCGTGGCGCA AAAGCTGCTG
  	TCGGCGCAAT
  551	TCGGGCCGCA ACAGATTCTG CTGTTGATTT ATGCGGcaag
  	tgccgccGTG
  601	TTCCtgccgT TTGccgaaCC GGCACACATC GGAAGTTTgg
  	aCGGTACGtt
  651	GGCGTGGGTT TGTTTTGTGT ATTGCTGCTT GAATACGTTA
  	ATCGGTTACG
  701	GCTCGTTCGG CGAGGCGTTG AAACATTGGG AGGCTTCCAA
  	AGTCAGCGCG
  751	GTAACAACCT TGCTCCCCGT GTTTACCGTA ATATTTTCTT
  	TGCTCGGGCA
  801	TTATGTGATG CCTGATACTT TTGCCGCGCC GGATATGAAC
  	GGTTTGGGTT
  851	ATGTCGGCGC ACTGGTCGTG GTCGGGGGTG CGGTTACGGC
  	GGCGGTGGGG
  901	GACAGGCCGT TCAAACGCCG CTAG

• This corresponds to the amino acid sequence <SEQ ID 408; ORF104ng-1>:

• 1	MENQRPLLGF ALALLAAMTW GTLPIAVRQV LKFVDAPTLV
  	WVRFTVAAAV
  51	LFVLLALGGR LPKRRDFSWH SFRLLLLGVT GISANFVLIA
  	QGLHYISPTT
  101	TQVLWQISPF TMIVVGVLVF KDRMTAAQKI GLVLLLVGLL
  	MFFNDKFGEL
  151	SGLGAYAKGV LLCAAGSMAW VCYAVAQKLL SAQFGPQQIL
  	LLIYAASAAV
  201	FLPFAEPAHI GSLDGTLAWV CFVYCCLNTL I GYGSFGEAL
  	KHWEASKVSA
  251	VTTLLPVFTV IFSLLGHYVM PDTFAAPDMN GLGYVGALVV
  	VGGAVTAAVG
  301	DRPFKRR*

• ORF104ng-1 and ORF104-1 show 97.5% identity in 277 aa overlap:
• 
• In addition, ORF104ng-1 shows significant homology with a hypothetical H. influenzae protein:

• gi|1573895 (U32769) hypothetical [Haemophilus influenzae] Length = 306
  Score = 237 bits (598), Expect = 8e−62
  Identities = 114/280 (40%), Positives = 168/280 (59%), Gaps = 8/280 (2%)

  Query:	30	QRPXXXXXXXXXXXMTWGTLPIAVRQVLKFVDAPTLVWXXXXXXXXXXXXXXXXXXXXP-	88
  		Q+P           M WG+LPIA++QVL  ++A T+VW                    P
  Sbjct:	3	QQPLLGFTFALITAMAWGSLPIALKQVLSVMNAQTIVWYRFIIAAVSLLALLAYKKQLPE	62
  Query:	89	--KRRDFSWHSFRLLLLGVTGISANFVLIAQGLHYISPTTTQVLWQISPFTMIVVGVLVF	146
  		K R ++W    ++L+GV G+++NF+L +  L+YI P+  Q+   +S F M++ GVL+F
  Sbjct:	63	LMKVRQYAW----IMLIGVIGLTSNFLLFSSSLNYIEPSVAQIFIHLSSFGMLICGVLIF	118
  Query:	147	KDRMTAAQKIXXXXXXXXXXMFFNDKFGELSGLGAYAKGVLLCAAGSMAWVCYAVAQKLL	206
  		K+++   QKI          +FFND+F   +GL  Y+ GV+L   G++ WV Y +AQKL+
  Sbjct:	119	KEKLGLHQKIGLFLLLIGLGLFFNDRFDAFAGLNQYSTGVILGVGGALIWVAYGMAQKLM	178
  Query:	207	SAQFGPQQILLLIYAASAAVFLPFAEPAHIGSLDGTLAWVCFVYCCLNTLIGYGSFGEAL	266
  		+F  QQILL++Y   A  F+P A+ + +  L   LA +CF+YCCLNTLIGYGS+ EAL
  Sbjct:	179	LRKFNSQQILLMMYLGCAIAFMPMADFSQVQELT-PLALICFIYCCLNTLIGYGSYAEAL	237
  Query:	267	KHWEASKVSAVTTLLPVFTVIFSLLGHYVMPDTFAAPDMN	306
  		W+ SKVS V TL+P+FT++FS + HY  P  FAAP++N
  Sbjct:	238	NRWDVSKVSVVITLVPLFTILFSHIAHYFSPADFAAPELN	277

• Based on this analysis, including the presence of a putative leader sequence and several putative transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 48
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 409>:

• 1	ATGGTAGCTC GTCGGGCTCA TAACCCGAAG GTCGTAGGTT
  	CGAATCCTGT
  51	.CCCGCAACC TAATTTCAAA CCCCTCGGTT CAATGCCGAG
  	GG.GTTTTGT
  101	T.TTGCCTGT TTCCTGTTTC CTGTTTCCTG CCGCCTCCGT
  	TTTTTGCCGG
  151	ATTTTCCTTC CGGCCGCAAT ATCGGAACGG CAGACCGCCG
  	TCTGTTTGCG
  201	GTTGCAAATT CAGGCAGTTT GGCTACAATC TTCCGCATTG
  	TCTTCAAGAA
  251	AGCCAACCAT GCCGACCGTC CGTTTTACCG AATCCGTCAG
  	CAAACAAGAC
  301	CTTGATGCTC TGTTCGAGTG GGCAAAAGCA AGTTACGGTG
  	CAGAAAGTTG
  351	CTGGAAAACG CTGTATCTGA ACGGTCysCC TTTGGGCAAC
  	CTGTCGCCGG
  401	AATGGGTGGA ACGCGTsmmA AAAGACTGGG AGGCAGGCTG
  	CyCGGAGTCT
  451	TCAGACGGCA TTTTTCTGAA TgCGGACGGc TGgCctGATA
  	TGGgCGGAcg
  501	cTTACAGCAC CTCGCCCTCG GTTGGCACTG TGCGGGGCTG
  	TTGGACGgsT
  551	GGCGCAACGA GTGTTTCGAC CTGACCGACG GCGGCGGCAA
  	CCCCTTGTTC
  601	ACGCTCGaAc GCGCCGyTTT mCGTCCTkTC GGACTGCTCA
  	GCCGCGCCGT
  651	CCATCTCAAC GGTCTGACCG AATCGGACGG CCGATGGCAT
  	TTCTGGATAG
  701	GCAGGCGCAG TCCGCACAAA GCAGTCGATC CCAACAAACT
  	CGACAATACT
  751	rCCGCCGGCG GTGTTTCCGG CGGCGAAATG CCGTCTGAAG
  	CCGTGTGTCG
  801	CGAAAGCAGC GAAGAAGCCG GTTTGGATAA AACGCTGcTT
  	CCGCTCATCC
  851	GCCCGGTATC GCAGCTGCAC AGCCTGCGCT CCGTCAGCCG
  	GGGTGTACAC
  901	AATGAAATCC TGTATGTATT CGATGCCGTC CTGCCG...

• This corresponds to the amino acid sequence <SEQ ID 410; ORF105>:

• 1	MVARRAHNPK VVGSNPXPAT XFQTPRFNAE XVLXLPVSCF
  	LFPAASVFCR
  51	IFLPAAISER QTAVCLRLQI QAVWLQSSAL SSRKPTMPTV
  	RFTESVSKQD
  101	LDALFEWAKA SYGAESCWKT LYLNGXPLGN LSPEWVERVX
  	KDWEAGCXES
  151	SDGIFLNADG WPDMGGRLQH LALGWHCAGL LDGWRNECFD
  	LTDGGGNPLF
  201	TLERAXXRPX GLLSRAVHLN GLTESDGRWH FWIGRRSPHK
  	AVDPNKLDNT
  251	XAGGVSGGEM PSEAVCRESS EEAGLDKTLL PLIRPVSQLH
  	SLRSVSRGVH
  301	NEILYVFDAV LP...

• Further work revealed the complete nucleotide sequence <SEQ ID 411>:

• 1	ATGCCGACCG TCCGTTTTAC CGAATCCGTC AGCAAACAAG
  	ACCTTGATGC
  51	TCTGTTCGAG TGGGCAAAAG CAAGTTACGG TGCAGAAAGT
  	TGCTGGAAAA
  101	CGCTGTATCT GAACGGTCTG CCTTTGGGCA ACCTGTCGCC
  	GGAATGGGTG
  151	GAACGCGTCA AAAAAGACTG GGAGGCAGGC TGCTCGGAGT
  	CTTCAGACGG
  201	CATTTTTCTG AATGCGGACG GCTGGCCTGA TATGGGCGGA
  	CGCTTACAGC
  251	ACCTCGCCCT CGGTTGGCAC TGTGCGGGGC TGTTGGACGG
  	CTGGCGCAAC
  301	GAGTGTTTCG ACCTGACCGA CGGCGGCGGC AACCCCTTGT
  	TCACGCTCGA
  351	ACGCGCCGCT TTCCGTCCTT TCGGACTGCT CAGCCGCGCC
  	GTCCATCTCA
  401	ACGGTCTGAC CGAATCGGAC GGCCGATGGC ATTTCTGGAT
  	AGGCAGGCGC
  451	AGTCCGCACA AAGCAGTCGA TCCCAACAAA CTCGACAATA
  	CTGCCGCCGG
  501	CGGTGTTTCC GGCGGCGAAA TGCCGTCTGA AGCCGTGTGT
  	CGCGAAAGCA
  551	GCGAAGAAGC CGGTTTGGAT AAAACGCTGC TTCCGCTCAT
  	CCGCCCGGTA
  601	TCGCAGCTGC ACAGCCTGCG CTCCGTCAGC CGGGGTGTAC
  	ACAATGAAAT
  651	CCTGTATGTA TTCGATGCCG TCCTGCCCGA AACCTTCCTG
  	CCTGAAAATC
  701	AGGATGGCGA AGTGGCGGGT TTTGAGAAAA TGGACATCGG
  	CGGTCTGTTG
  751	GATGCCATGT TGTCGGGAAA CATGATGCAC GACGCGCAAC
  	TGGTTACGCT
  801	GGACGCGTTT TGCCGTTACG GTCTGATTGA TGCCGCCCAT
  	CCGCTGTCCG
  851	AGTGGCTGGA CGGCATACGT TTATAG

• This corresponds to the amino acid sequence <SEQ ID 412; ORF105-1>:

• 1	MPTVRFTESV SKQDLDALFE WAKASYGAES CWKTLYLNGL
  	PLGNLSPEWV
  51	ERVKKDWEAG CSESSDGIFL NADGWPDMGG RLQHLALGWH
  	CAGLLDGWRN
  101	ECFDLTDGGG NPLFTLERAA FRPFGLLSRA VHLNGLTESD
  	GRWHFWIGRR
  151	SPHKAVDPNK LDNTAAGGVS GGEMPSEAVC RESSEEAGLD
  	KTLLPLIRPV
  201	SQLHSLRSVS RGVHNEILYV FDAVLPETFL PENQDGEVAG
  	FEKMDIGGLL
  251	DAMLSGNMMH DAQLVTLDAF CRYGLIDAAH PLSEWLDGIR
  	L*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF105 shows 89.4% identity over a 226aa overlap with an ORF (ORF105a) from strain A of N. meningitidis:
• 
• The complete length ORF105a nucleotide sequence <SEQ ID 413> is:

• 1	ATGCCGACCG TCCGTTTTAC CGAATCCGTC AGCAAACACG
  	ACCTTGATGC
  51	CCTATTCGAG TGGGCAAAGG CAAGTTACGG TGCGGAAAGT
  	TGCTGGAAAA
  101	CGCTGTATCT GAACGGTCTG CCTTTGGGCA ATCTGTCGCC
  	GGAATGGGCG
  151	GAGCGCGTCA AAAAAGACTG GGAGGCAGGC TGCTCGGAGT
  	CTTCAGACGG
  201	CATTTTCCTG AATGCGGACG GCTGGCCAGA TATGGGCAGA
  	CGCTTGCAGC
  251	ACCTCGCCCG AATATGGAAA GAAGCGGGAC TGCTTCACGG
  	CTGGCGCGAC
  301	GAGTGTTTCG ACCTGACCGA CGGCGGCAGC AATCCCTTGT
  	TCGCGCTCGA
  351	ACGCGCCGCT TTCCGTCCGT TCGGACTGCT CAGCCGCGCC
  	GTCCATCTCA
  401	ACGGTTTGGT CGAATCGGAC GGCCGATGGC ATTTCTGGAT
  	AGGCAGGCGC
  451	AGTCCGCACA AAGCAGTCGA TCCCGACAAA CTCGACAATA
  	CTGCCGCCGG
  501	CGGTGTTTCC AGCGGTGAAT TGCCGTCTGA AACCGTGTGT
  	CGCGAAAGCA
  551	GCGAAGAAGC CGGTTTGGAT AAAACGCTGC TTCCGCTCAT
  	CCGCCCGGTA
  601	TCGCAGCTGC ACAGCCTGCG CCCCGTCAGC CGGGGTGTGC
  	ACAATGAAAT
  651	CCTGTATGTA TTCGATGCCG TCCTGCCCGA AACCTTCCTG
  	CCTGAAAATC
  701	AGGATGGCGA AGTGGCGGGT TTTGAGAAAA TGGACATCGG
  	CGGTCTGTTG
  751	GCTGCCATGT TGTCGGGAAA CATGATGCAC GACGCGCAAC
  	TGGTTACGCT
  801	GGACGCGTTT TGCCGTTACG GTCTGATTGA TGCCGCCCAT
  	CCGCTGTCCG
  851	AGTGGCTGGA CGGCATACGT TTATAG

• This encodes a protein having amino acid sequence <SEQ ID 414>:

• 1	MPTVRFTESV SKHDLDALFE WAKASYGAES CWKTLYLNGL
  	PLGNLSPEWA
  51	ERVKKDWEAG CSESSDGIFL NADGWPDMGR RLQHLARIWK
  	EAGLLHGWRD
  101	ECFDLTDGGS NPLFALERAA FRPFGLLSRA VHLNGLVESD
  	GRWHFWIGRR
  151	SPHKAVDPDK LDNTAAGGVS SGELPSETVC RESSEEAGLD
  	KTLLPLIRPV
  201	SQLHSLRPVS RGVHNEILYV FDAVLPETFL PENQDGEVAG
  	FEKMDIGGLL
  251	AAMLSGNMMH DAQLVTLDAF CRYGLIDAAH PLSEWLDGIR
  	L*

• ORF105a and ORF105-1 show 93.8% identity in 291 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF105 shows 87.5% identity over a 312aa overlap with a predicted ORF (ORF105.ng) from N. gonorrhoeae:
• 
• A complete length ORF105ng nucleotide sequence <SEQ ID 415> was predicted to encode a protein having amino acid sequence <SEQ ID 416>:

• 1	MVARRAHNPK VVGSNPAPAT KYQTPRFNAE GVLFFLFPAA
  	SVFCRIFLPA
  51	AISERQAAVC LRLQIQAVWL QSSALCSRKP AMPTVRFTES
  	VSKQDLDALF
  101	ERAKASYGAE SCWKTLYLNR LPLGNLSPEW AERIKKDWEA
  	GCSESSNGIF
  151	LNADGWPDMG GRLQHLARTW NKAGLLHGWR NECFDLTDGG
  	GNPLFTLERA
  201	AFRPFGLLIR AVHLNGLVES NGRWHFWIGR RSPHKAVDPG
  	KLDNIAGGGV
  251	SGGEMPSEAV CRESSEEAGL DKTLFPLIRP VSRLHSLRPV
  	SRGVHNEILY
  301	VFDAVLPETF LPENQDGEVA GFEKMDIGGL LDAMLSKNMM
  	HDAQLVTLDA
  351	FYRYGLIDAA HPLSEWLDGI RL*

• Further work revealed the complete nucleotide sequence <SEQ ID 417>:

• 1	ATGCCGACCG TCCGTTTTAC CGAATCCGTC AGCAAACAAG
  	ACCTTGATGC
  51	CCTGTTCGAG CGGGCAAAAG CAAGTTACGG TGCCGAAAGT
  	TGCTGGAAAA
  101	CGCTGTATCT GAACCGTCTT CCTTTGGGCA ATCTGTCGCC
  	GGAATGGGCT
  151	GAGCGCATCA AAAAAGACTG GGAGGCAGGC TGCTCCGAGT
  	CTTCAGACGG
  201	CATTTTTCTG AATGCGGACG GCTGGCCGGA TATGGGCGGA
  	CGCTTGCAGC
  251	ACCTCGCCCG CACATGGAAC AAGGCGGGGC TGCTTCACGG
  	ATGGCGCAAC
  301	GAGTGTTTCG ACCTGACCGA CGGCGGCGGC AACCCCTTGT
  	TCACGCTCGA
  351	ACGCGCCGCT TTCCGTCCGT TCGGACTACT CAGCCGCGCC
  	GTCCATCTCA
  401	ACGGTTTGGT CGAATCGAAC GGCAGATGGC ATTTTTGGAT
  	AGGCAGGCGC
  451	AGTCCGCACA AAGCAGTCGa tcCCGGCAAG CTCGACAATA
  	TTGCCGGCGG
  501	CGGTGTTTCC GGCGGCGAAA TGCCGTCTGA AGCCGTGTGC
  	CGCGAAAGCA
  551	GCGAAGAAGC CGGTTTGGAT AAAACGCTGT TTCCGCTCAT
  	CCGCCCAGTA
  601	TCGCGGCTGC ACAGCCTTCG CCCCGTCAGC CGAGGTGTGC
  	ACAATGAAAT
  651	CCTGTATGTG TTCGATGCCG TCCTGCCCGA AACCTTCCTG
  	CCTGAAAATC
  701	AGGATGGCGA GGTAGCGGGT TTTGAAAAGA TGGACATTGG
  	CGGCCTATTG
  751	GATGCCATGT TGTCGAAAAA CATGATGCAC GACGCGCAAC
  	TGGTTACGCT
  801	GGACGCGTTT TACCGTTACG GTCTGATTGA TGCCGCCCAT
  	CCGCTGTCCG
  851	AGTGGCTGGA CGGCATACGT TTATAG

• This corresponds to the amino acid sequence <SEQ ID 418; ORF105ng-1>:

• 1	MPTVRFTESV SKQDLDALFE RAKASYGAES CWKTLYLNRL
  	PLGNLSPEWA
  51	ERIKKDWEAG CSESSDGIFL NADGWPDMGG RLQHLARTWN
  	KAGLLHGWRN
  101	ECFDLTDGGG NPLFTLERAA FRPFGLLSRA VHLNGLVESN
  	GRWHFWIGRR
  151	SPHKAVDPGK LDNIAGGGVS GGEMPSEAVC RESSEEAGLD
  	KTLFPLIRPV
  201	SRLHSLRPVS RGVHNEILYV FDAVLPETFL PENQDGEVAG
  	FEKMDIGGLL
  251	DAMLSKNMMH DAQLVTLDAF YRYGLIDAAH PLSEWLDGIR
  	L*

• ORG105ng-1 and ORF105-1 show 93.5% identity in 291 aa overlap:
• 
• Furthermore, ORF105ng-1 shows homology with a yeast enzyme:

• sp|P41888|TNR3_SCHPO THIAMIN PYROPHOSPHOKINASE (TPK) (THIAMIN KINASE)
  >gi|1076928|pir||S52350 thiamin pyrophosphokinase (EC 2.7.6.2) - fission
  yeast (Schizosaccharomyces pombe) >gi|666111 (X84417) thiamin
  pyrophosphokinase [Schizosaccharomyces pombe]
  >gi|2330852|gnl|PID|e334056 (Z98533) thiamin
  pyrophosphokinase [Schizosaccharomyces pombe] Length = 569
  Score = 105 bits (259), Expect = 4e−22
  Identities = 64/192 (33%), Positives = 94/192 (48%), Gaps = 3/192 (1%)

  Query:	268	NKAGLLHGWRNECFDLTDGGGNPLFTLERAAFRPFGLLSRAVHLNGLVESNGRW--HFWI	441
  		N  G+   WRNE + +      P+  +ER  F  FG LS  VH    + +        W+
  Sbjct:	96	NTFGIADQWRNELYTVYGKSKKPVLAVERGGFWLFGFLSTGVHCTMYIPATKEHPLRIWV	155
  Query:	442	GRRSPHKAVDPGKLDNIAGGGVSGGEMPSEAVCRESSEEAGLDKTLFPLIRPVSRLHSLR	621
  		RRSP K   P  LDN   GG++ G+     + +E SEEA LD +   LI P   +  ++
  Sbjct:	156	PRRSPTKQTWPNYLDNSVAGGIAHGDSVIGTMIKEFSEEANLDVSSMNLI-PCGTVSYIK	214
  Query:	622	PVSRG-VHNEILYVFDAVLPETFLPENQDGEVAGFEKMDIGGLLDAMLSKNMMHDAQLVT	798
  		R  +  E+ YVFD  + +  +P   DGEVAGF  + +  +L  +  K+   +  LV
  Sbjct:	215	MEKRHWIQPELQYVFDLPVDDLVIPRINDGEVAGFSLLPLNQVLHELELKSFKPNCALVL	274
  Query:	799	LDAFYRYGLIDAAHP	843
  		LD   R+G+I   HP
  Sbjct:	275	LDFLIRHGIITPQHP	289

• Based on this analysis, including the presence of a putative transmembrane domain in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 49
• The following DNA sequence, believed to be complete, was identified in N. meningitidis <SEQ ID 419>:

• 1	ATGAATAGAC CCAAGCAACC CTTCTTCCGT CCCGAAGTCG
  	CCGTTGCCCG
  51	CCAAACCAGC CTGACGGGTA AAGTGATTCT GACACGACCG
  	TTGTCATTTT
  101	CCCTATGGAC GACATTTGCA TCGATATCTG CGTTATTGAT
  	TATCCTGTTT
  151	TTGATATTTG GTAACTATAC GCGAAAGACA ACAGTGGAGG
  	GACAAATTTT
  201	ACCTGCATCG GGCGTAATCA GGGTGTATGC ACCGgATACG
  	rGkACAATTA
  251	CAGCGAAATT CGTGGAAGAT GGmsAAAAGG TTAAGGCTGG
  	CGACAAGCTA
  301	TTTGCGCTTT CGACCTCACG TTTCGGCGCA GGAGGTAGCG
  	TGCAGCAGCA
  351	GTTGAAAACG GAGGCAGTTT TGAAGAAAAC GTTGGCAGAA
  	CAGGAACTGG
  401	GTCGTCTGAA GCTGATACAC GGGAATGAAA CGCGCAgCcT
  	TAAAGCAACT
  451	GTCGAACGTT TGGAAAACCA GGAACTCCAT ATTTCGCAAC
  	AGATAGACGG
  501	TCAGAAAAGG CGCATTAGAC TTGCGGAAGA AATGTTGCAG
  	AAATATCGTT
  551	TCCTATCCGC .CAATGA

• This corresponds to the amino acid sequence <SEQ ID 420; ORF107>:

• 1	MNRPKQPFFR PEVAVARQTS LTGKVILTRP LSFSLWTTFA
  	SISALLIILF
  51	LIFGNYTRKT TVEGQILPAS GVIRVYAPDT XTITAKFVED
  	GXKVKAGDKL
  101	FALSTSRFGA GGSVQQQLKT EAVLKKTLAE QELGRLKLIH
  	GNETRSLKAT
  151	VERLENQELH ISQQIDGQKR RIRLAEEMLQ KYRFLSXQ*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF107 shows 97.8% identity over a 186aa overlap with an ORF (ORF107a) from strain A of N. meningitidis:
• 
• The complete length ORF107a nucleotide sequence <SEQ ID 421> is:

• 1	ATGAATAGAC CCAAGCAACC NTTCTTCCGT CCCGAAGTCG
  	CCGTTGCCCG
  51	CCAAACCAGC CTGACGGGTA AAGTGATTCT GACACGACCG
  	TTGTCATTTT
  101	CCCTATGGAC GACATTTGCA TCGATATCTG CGTTATTGAT
  	TATCCTGTTT
  151	TTGATATTTG GTAACTATAC GCGAAAGACA ACAGTGGAGG
  	GACAAATTTT
  201	ACCTGCATCG GGCGTAATCA GGGTGTATGC ACCGGATACG
  	GGGACAATTA
  251	CNGCGAAATT CNTGGAAGAT GGAGAAAAGG TTAAGGCTGG
  	CGACAAGCTA
  301	TTTGCGCTTT CGACCTCACG TTTCGGCGCA GGAGATAGCG
  	TGCAGCAGCA
  351	GTTGAAAACG GAGGCAGTTT TGAAGAAAAC GTTGGCAGAA
  	CAGGAACTGG
  401	GTCGTCTGAA GCTGATACAC GGGAATGAAA CGCGCAGCCT
  	TAAAGCAACT
  451	GTCGAACGTT TGGAAAACCA GGAACTCCAT ATTTCGCAAC
  	AGATAGACGG
  501	TCAGAAAAGG CGCATTAGAC TTGCGGAAGA AATGTTGCAG
  	AAATATCGTT
  551	TCCTATCCGC CAATGATGCA GTGCCAAAAC AAGAAATGAT
  	GAATGTCAAG
  601	GCAGAGCTTT TAGAGCAGAA AGCCAAACTT GATGCCTACC
  	GCCGAGAAGA
  651	AGTCGGGCTG CTTCAGGAAA TCCGCACGCA GAATCTGACA
  	TTGGNNAGCC
  701	TCCCCCAAGC GGCATGA

• This encodes a protein having amino acid sequence <SEQ ID 422>:

• 1	MNRPKQPFFR PEVAVARQTS LTGKVILTRP LSFSLWTTFA
  	SISALLIILF
  51	LIFGNYTRKT TVEGQILPAS GVIRVYAPDT GTITAKFXED
  	GEKVKAGDKL
  101	FALSTSRFGA GDSVQQQLKT EAVLKKTLAE QELGRLKLIH
  	GNETRSLKAT
  151	VERLENQELH ISQQIDGQKR RIRLAEEMLQ KYRFLSANDA
  	VPKQEMMNVK
  201	AELLEQKAKL DAYRREEVGL LQEIRTQNLT LXSLPQAA*

  
  Homology with a Predicted ORF from N. gonorrhoeae
• ORF107 shows 95.7% identity over a 188aa overlap with a predicted ORF (ORF107.ng) from N. gonorrhoeae:
• 
• The complete length ORF107ng nucleotide sequence <SEQ ID 423> is predicted to encode a protein having amino acid sequence <SEQ ID 424>:

• 1	MNRPKQPFFR PEVAIARQTS LTGKVILTRP LSFSLWTTFA
  	SISALLIILF
  51	LIFGNYTRKT TMEGQILPAS GVIRVYAPDT GTITAKFVED
  	GEKVKAGDKL
  101	FALSTSRFGA GGSVQQQLKT EAVLKKTLAE QELGRLKLIH
  	ENETRSLKAT
  151	VERLENQKLH ISQQIDGQKR RIRLAEEMLR KYRFLSAQ*

• Based on the presence of a putative transmembrane domain in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 50
• The following DNA sequence, believed to be complete, was identified in N. meningitidis <SEQ ID 425>:

• 1	ATGCTGAATA CTTTTTTTGC CGTATTGGGC GGCTGCCTGC
  	TGCT.TTGCC
  51	GTGCGGCAAA TCCGTAAATA CGGCGGTACA GCCGCAAAAC
  	GCGGTACAAA
  101	GCGCGCCGAA ACCGGTTTTC AAAGTCATAT ATATCGACAA
  	TACGGCGATT
  151	GCCGGTTTGG ATTTGGGACA AAGCAGCGAA GGCAAAACCA
  	ACGACGGCAA
  201	AAAACAAATC AGTTATCCGA TTAAAGGCTT GCCGGAACAA
  	AATGTTATCC
  251	GACTGATCGG CAAGCATCCC GGCGACTTGG AAGCCGTCAG
  	CGGCAAATGT
  301	ATGGAAACCG ATGATAAGGA CAGTCCGGCA GGTTGGGCAG
  	AAAACGGCGT
  351	GTGCCATACC TTGTTTGCCA AACTGGTGGG CAATATCGCC
  	GAAGACGGCG
  401	GCAAACTGAC GGATTACCTA GTTTCGCATG CCGCCCTGCA
  	ACCCTATCAG
  451	GCAGGCAAAA GCGGCTATGC CGCCGTGCAG AACGGACGCT
  	ATGTGCTGGA
  501	AATCGACAGC GAAGGGGCGT TTTATTTCCG CCGCCGCCAT
  	TATTGA

• This corresponds to the amino acid sequence <SEQ ID 426; ORF108>:

• 1	MLNTFFAVLG GCLLXLPCGK SVNTAVQPQN AVQSAPKPVF
  	KVIYIDNTAI
  51	AGLDLGQSSE GKTNDGKKQI SYPIKGLPEQ NVIRLIGKHP
  	GDLEAVSGKC
  101	METDDKDSPA GWAENGVCHT LFAKLVGNIA EDGGKLTDYL
  	VSHAALQPYQ
  151	AGKSGYAAVQ NGRYVLEIDS EGAFYFRRRH Y*

• Further work revealed the following DNA sequence <SEQ ID 427>:

• 1	ATGCTGAAAA CATCTTTTGC CGTATTGGGC GGCTGCCTGC
  	TGCTTGCCGC
  51	CTGCGGCAAA TCCGAAAATA CGGCGGAACA GCCGCAAAAC
  	GCGGTACAAA
  101	GCGCGCCGAA ACCGGTTTTC AAAGTCAAAT ATATCGACAA
  	TACGGCGATT
  151	GCCGGTTTGG ATTTGGGACA AAGCAGCGAA GGCAAAACCA
  	ACGACGGCAA
  201	AAAACAAATC AGTTATCCGA TTAAAGGCTT GCCGGAACAA
  	AATGTTATCC
  251	GACTGATCGG CAAGCATCCC GGCGACTTGG AAGCCGTCAG
  	CGGCAAATGT
  301	ATGGAAACCG ATGATAAGGA CAGTCCGGCA GGTTGGGCAG
  	AAAACGGCGT
  351	GTGCCATACC TTGTTTGCCA AACTGGTGGG CAATATCGCC
  	GAAGACGGCG
  401	GCAAACTGAC GGATTACCTA GTTTCGCATG CCGCCCTGCA
  	ACCCTATCAG
  451	GCAGGCAAAA GCGGCTATGC CGCCGTGCAG AACGGACGCT
  	ATGTGCTGGA
  501	AATCGACAGC GAAGGGGCGT TTTATTTCCG CCGCCGCCAT
  	TATTGA

• This corresponds to the amino acid sequence <SEQ ID 428; ORF108-1>:

• 1	MLKTSFAVLG GCLLLAACGK SENTAEQPQN AVQSAPKPVF
  	KVKYIDNTAI
  51	AGLDLGQSSE GKTNDGKKQI SYPIKGLPEQ NVIRLIGKHP
  	GDLEAVSGKC
  101	METDDKDSPA GWAENGVCHT LFAKLVGNIA EDGGKLTDYL
  	VSHAALQPYQ
  151	AGKSGYAAVQ NGRYVLEIDS EGAFYFRRRH Y*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF108 shows 88.4% identity over a 181 aa overlap with a predicted ORF (ORF 108.ng) from N. gonorrhoeae:
• 
• ORF108-1 shows 92.3% identity with ORF108ng over the same 181 aa overlap:
• 
• The complete length ORF108ng nucleotide sequence <SEQ ID 429> is:

• 1	ATGCTGAAAa tacctTTTGC CGTGTtgggc ggCtgcctGC
  	TGCTTGCCGC
  51	CTGCGGCAAA TCCGAAAATa cggcggaACA GCCGCAAAAT
  	gcggCACAAA
  101	GCGCGCCGAA ACCGGTTTTC AAAGTCAAAT ACATCGACAA
  	TACGGCGATT
  151	GCCGGTTTGG CTTTGGGACA AAGTAGCGAA GGCAAAACCA
  	acgacgGCAA
  201	AAAACAAATC AGTTATccgA TTAAAGGCTT GCCGGAACAA
  	Aacgccgtcc
  251	gGCTGACCGG AAAGCATCCC AACGACTTGG AagccgtcgT
  	CGGCAAATGT
  301	ATGGAAACCG ACGGAAAGGA CGCGCCTTCG GGCTGGGCGG
  	AAAACGGCGT
  351	GTGCCATACC TTGTTTGCCA AACTGGTGGG CAATATCGCC
  	GAAGACGGCG
  401	GCAAACTGAC TGATTACCTG ATTTCGCATT CCGCCCTGCA
  	ACCCTATCAG
  451	GCAGGCAAAA GCGGCTATGC CGCCGTGCAG AACGGACGCT
  	ATGTGCTGGA
  501	AATCGACAGC GagggGGCGT TTTATttccg ccgccgccat
  	tattgA

• This encodes a protein having amino acid sequence <SEQ ID 430>:

• 1	MLKIPFAVLG GCLLLAACGK SENTAEQPQN AAQSAPKPVF
  	KVKYIDNTAI
  51	AGLALGQSSE GKTNDGKKQI SYPIKGLPEQ NAVRLTGKHP
  	NDLEAVVGKC
  101	METDGKDAPS GWAENGVCHT LFAKLVGNIA EDGGKLTDYL
  	ISHSALQPYQ
  151	AGKSGYAAVQ NGRYVLEIDS EGAFYFRRRH Y*

• Based on this analysis, including the presence of a predicted prokaryotic membrane lipoprotein lipid attachment site (underlined) and a putative ATP/GTP-binding site motif A (P-loop, double-underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 51
• The following DNA sequence was identified in N. meningitidis <SEQ ID 431>:

• 1	ATGGAAGATT TATATATAAT ACTCGCTTTG GGTTTGGTTG
  	CGATGATTGC
  51	CGgATTTATC GATgcgatTg cGggCGGGGG TGGTTTGATT
  	ACGCTGCCCG
  101	CACTCTTGTT GGCAGGTATT CCTCCCGTGT CGGCAATTGC
  	CACCAACAAG
  151	CTGCAAgCAG CCGCTGCTAC GTTTTCAGCT ACGGTTTCTT
  	TTGCACGCAA
  201	AGGTTTGATT GATTGGAAGA AAGGTCTCCC GATTGCCGCA
  	GCATCGTTTG
  251	TAGGCGGCGT GGcCGGTGCA TTATCGGTCA GCTTGGTTTC
  	CAAAGATATT
  301	CTgCTgGCGG TCGTGCCGGT TTTGTTGATA TTTGTCGCAC
  	TGTATTTTGT
  351	GTTTTCGCCC AAGCTCGACG GCAGTAAGGA AGGCAAAGCC
  	AGAATGTCTT
  401	TTTTTCTGTT cGGGCTGACG GTCGC.ACCG CTTTTGGGTT
  	TTTACGACGG
  451	TGTGTTCGGA CCGGGTGTCG GCTCGTTTTT TCTGATTGCC
  	TTTATTGTTT
  501	TGCTCGGCTG CAAgCTGTTG AACGCGATGT CTTACACCAA
  	ATTGGCGAAC
  551	GTTGCCTGCA ATCTTGGTTC GCTATCGGTA TTCCTGCTGC
  	ACGGTTCGAT
  601	TATTTTCCCG ATTGCGGCAA CGaTGGCGGT CGGTGCGTTT
  	GTCGGtGCGA
  651	ATTTAgGTGC GAGATTTGCC GTaCgctTCG GTTCGAAGCT
  	GATTAA

• This corresponds to the amino acid sequence <SEQ ID 432; ORF109>:

• 1	MEDLYIILAL GLVAMIAGFI DAIAGGGGLI TLPALLLAGI
  	PPVSAIATNK
  51	LQAAAATFSA TVSFARKGLI DWKKGLPIAA ASFVGGVAGA
  	LSVSLVSKDI
  101	LLAVVPVLLI FVALYFVFSP KLDGSKEGKA RMSFFLFGLT
  	VXTAFGFLRR
  151	CVRTGCRLVF SDCLYCFARL QAVERDVLHQ IGERCLQSWF
  	AIGIPAARFD
  201	YFPDCGNDGG RCVCRCEFRC EICRTLRFEA D*

• Further work revealed the following DNA sequence <SEQ ID 433>:

• 1	ATGGAAGATT TATATATAAT ACTCGCTTTG GGTTTGGTTG
  	CGATGATTGC
  51	CGGATTTATC GATGCGATTG CGGGCGGGGG TGGTTTGATT
  	ACGCTGCCCG
  101	CACTCTTGTT GGCAGGTATT CCTCCCGTGT CGGCAATTGC
  	CACCAACAAG
  151	CTGCAAGCAG CCGCTGCTAC GTTTTCAGCT ACGGTTTCTT
  	TTGCACGCAA
  201	AGGTTTGATT GATTGGAAGA AAGGTCTCCC GATTGCCGCA
  	GCATCGTTTG
  251	TAGGCGGCGT GGCCGGTGCA TTATCGGTCA GCTTGGTTTC
  	CAAAGATATT
  301	CTGCTGGCGG TCGTGCCGGT TTTGTTGATA TTTGTCGCAC
  	TGTATTTTGT
  351	GTTTTCGCCC AAGCTCGACG GCAGTAAGGA AGGCAAAGCC
  	AGAATGTCTT
  401	TTTTTCTGTT CGGGCTGACG GTCGCACCGC TTTTGGGTTT
  	TTACGACGGT
  451	GTGTTCGGAC CGGGTGTCGG CTCGTTTTTT CTGATTGCCT
  	TTATTGTTTT
  501	GCTCGGCTGC AAGCTGTTGA ACGCGATGTC TTACACCAAA
  	TTGGCGAACG
  551	TTGCCTGCAA TCTTGGTTCG CTATCGGTAT TCCTGCTGCA
  	CGGTTCGATT
  601	ATTTTCCCGA TTGCGGCAAC GATGGCGGTC GGTGCGTTTG
  	TCGGTGCGAA
  651	TTTAGGTGCG AGATTTGCCG TCCGCTTCGG TTCGAAGCTG
  	ATTAAGCCGC
  701	TGCTGATTGT CATCAGCATT TCGATGGCTG TGAAATTGTT
  	GATAGACGAG
  751	AGAAATCCGC TGTATCAGAT GATTGTTTCG ATGTTTTAA

• This corresponds to the amino acid sequence <SEQ ID 434; ORF109-1>:

• 1	MEDLYIILAL GLVAMIAGFI DAIAGGGGLI TLPALLLAGI
  	PPVSAIATNK
  51	LQAAAATFSA TVSFARKGLI DWKKGLPIAA ASFVGGVAGA
  	LSVSLVSKDI
  101	LLAVVPVLLI FVALYFVFSP KLDGSKEGKA RMSFFLFGLT
  	VAPLLGFYDG
  151	VFGPGVGSFF LIAFIVLLGC KLLNAMSYTK LANVACNLGS
  	LSVFLLHGSI
  201	IFPIAATMAV GAFVGANLGA RFAVRFGSKL IKPLLIVISI
  	SMAVKLLIDE
  251	RNPLYQMIVS MF*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF109 shows 95.9% identity over a 147aa overlap with an ORF (ORF109a) from strain A of N. meningitidis:
• 
• The complete length ORF109a nucleotide sequence <SEQ ID 435> is:

• 1	ATGGAAGATT TATACATAAT ACTCGCTTTG GGTTTGGTTG
  	CGATGATTGC
  51	CGGATTTATC GATGCGATTG CGGGTGGGGG TGGTTTGATT
  	ACGCTGCCTG
  101	CACTCTTGTT GGCAGGTATT CCTCCCGTGT CGGCAATTGC
  	CACCAACAAG
  151	CTGCAAGCAG CCGCTGCTAC GTTTTCGGCT ACGGTTTCTT
  	TTGCACGCAA
  201	AGGTTTGATT GATTGGAAGA AAGGTCTCCC GATTGCGGCA
  	GCATCGTTTG
  251	CAGGCGGCGT GGTCGGTGCA TTATCGGTCA GCTTGGTTTC
  	CAAAGATATT
  301	CTGCTGGCGG TCGTGCCGGT TTTGTTGATA TTTGTCGCGC
  	TGTATTTTGT
  351	GTTTTCGCCC AAGCTCGACG GCAGTAAGGA AGGCAAAGCC
  	AGAATGTCTT
  401	TTTTTCTGTT CGGTCTGACG GTTGCACCAC TTTTGGGTTT
  	TTACGACGGT
  451	GTGTTCGGAC CGGGTGTCGG CTCGTTTTTT CTGATTGCCT
  	TTATTGTTTT
  501	GCTCGGCTGC AAGCTGTTGA ACGCGATGTC TTACACCAAA
  	TTGGCGAACG
  551	TTGCCTGCAA TCTTGGTTCG CTATCGGTAT TCCTGCTGCA
  	CGGTTCGATT
  601	ATTTTCCCGA TTGCGGCAAC GATGGCGGTC GGTGCGTTTG
  	TCGGTGCGAA
  651	TTTAGGTGCG AGATTTGCCG TCCGCTTCGG TTCGAAGCTG
  	ATTAAGCCGC
  701	TGCTGATTGT CATCAGCATT TCGATGGCTG TGAAATTGTT
  	GATAGACGAG
  751	AGAAATCCGC TGTATCAGAT GATTGTTTCG ATGTTTTAA

• This encodes a protein having amino acid sequence <SEQ ID 436>:

• 1	MEDLYIILAL GLVAMIAGFI DAIAGGGGLI TLPALLLAGI
  	PPVSAIATNK
  51	LQAAAATFSA TVSFARKGLI DWKKGLPIAA ASFAGGVVGA
  	LSVSLVSKDI
  101	LLAVVPVLLI FVALYFVFSP KLDGSKEGKA RMSFFLFGLT
  	VAPLLGFYDG
  151	VFGPGVGSFF LIAFIVLLGC KLNAMSYTK LANVACNLGS
  	LSVFLLHGSI
  201	IFPIAATMAV GAFVGANLGA RFAVRFGSKL IKPLLIVISI
  	SMAVKLLIDE
  251	RNPLYQMIVS MF*

• ORF109a and ORF109-1 show 99.2% identity in 262 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF109 shows 98.3% identity over a 231aa overlap with a predicted ORF (ORF109.ng) from N. gonorrhoeae:
• 
• An ORF109ng nucleotide sequence <SEQ ID 437> was predicted to encode a protein having amino acid sequence <SEQ ID 438>:

• 1	MEDLYIILAL GLVAMIAGFI DAIAGGGGLI TLPALLLAGI
  	PPVSAIATNK
  51	LQAAAATFSA TVSFARKGLI DWKKGLPIAA ASFAGGVVGA
  	LSVSLVSKDI
  101	LLAVVPVLLI FVALYFVFSP KLDGSKEGKA RMSFFLFGLT
  	VATAFGFLRR
  151	CVRTGCRLVF SDCLYCFARL QAVERDVLHQ IGERCLQSWF
  	AIGIPAARFD
  201	YFPDCGNDGG RCVCRCEFRC EICRPLRFEA D*

• Further work revealed the following gonococcal DNA sequence <SEQ ID 439>:

• 1	ATGGAAGATT TATACATAAT ACTCGCTTTG GGTTTGGTTG
  	CGATGATCGC
  51	CGGATTTATC GATGCGATTG CGGGCGGGGG TGGTTTGATT
  	ACGCTGCCTG
  101	CACTCTTGTT GGCAGGTATT CCTCCCGTGT CGGCAATTGC
  	CACCAACAAG
  151	CTGCAAGCAG CCGCTGCTAC GTTTTCGGCT ACGGTTTCTT
  	TTGCACGCAA
  201	AGGTTTGATT GATTGGAAGA AAGGTCTCCC GATTGCCGCA
  	GCATCGTTTG
  251	CAGGCGGCGT GGTCGGTGCA TTATCGGTCA GCTTGGTTTC
  	CAAAGATATT
  301	TTGCTGGCGG TCGTGCCGGT TTTGTTGATA TTTGTCGCGC
  	TGTATTTTGT
  351	GTTTTCGCCC AAGCTCGACG GCAGTAAGGA AGGCAAAGCC
  	AGAATGTCTT
  401	TTTTTCTATT CGGGCTGACG GTTGCACCGC TTTTGGGTTT
  	TTACGACGGT
  451	GTGTTCGGAC CGGGTGTCGG CTCGTTTTTT CTGATTGCCT
  	TTATTGTTTT
  501	GCTCGGCTGC AAGCTGTTGA ACGCGATGTC TTACACCAAA
  	TTGGCGAACG
  551	TTGCTTGCAA TCTTGGTTCG CTATCGGTAT TCCTGCTGCA
  	CGGTTCGATT
  601	ATTTTCCCGA TTGTGGCAAC GATGGCGGTC GGTGCGTTTG
  	TCGGTGCGAA
  651	TTTAGGTGCG AGATTTGCCG TCCGCTTCGG TTCGAAGCTG
  	ATTAAGCCGC
  701	TGCTGATTGT CATCAGCATT TCGATGGCTG TGAAATTGTT
  	GATAGACGAG
  751	AGAAATCCGC TGTATCAGAT GATTGTTTCG ATGTTTTAA

• This corresponds to the amino acid sequence <SEQ ID 440; ORF109ng-1>:

• 1	MEDLYIILAL GLVAMIAGFI DAIAGGGGLI TLPALLLAGI
  	PPVSAIATNK
  51	LQAAAATFSA TVSFARKGLI DWKKGLPIAA ASFAGGVVGA
  	LSVSLVSKDI
  101	LLAVVPVLLI FVALYFVFSP KLDGSKEGKA RMSFFLFGLT
  	VAPLLGFYDG
  151	VFGPGVGSFF LIAFIVLLGC KLLNAMSYTK LANVACNLGS
  	LSVFLLHGSI
  201	IFPIVATMAV GAFVGANLGA RFAVRFGSKL IKPLLIVISI
  	SMAVKLLIDE
  251	RNPLYQMIVS MF*

• ORF109ng-1 and ORF109-1 show 98.9% identity in 262 aa overlap:
• 
• In addition, ORF109ng-1 shows homology to a hypothetical Pseudomonas protein:

• sp|P29942|YCB9_PSEDE HYPOTHETICAL 27.4 KD PROTEIN IN COBO 3′REGION (ORF9)
  >gi|94984|pir||I38164 hypothetical protein 9 - Pseudomonas sp >gi|551929
  (M62866) ORF9 [Pseudomonas denitrificans] Length = 261
  Score = 175 bits (439), Expect = 3e−43
  Identities = 83/214 (38%), Positives = 131/214 (60%), Gaps = 1/214 (0%)

  Query:	41	PPVSAIATNKLQXXXXXXXXXXXXXRKGLIDWKKGLPIXXXXXXXXXXXXXXXXXXXKDI	100
  		PP+  + TNKLQ             R+G ++ K+ LP+                    D+
  Sbjct:	43	PPLQTLGTNKLQGLFGSGSATLSYARRGHVNLKEQLPMALMSAAGAVLGALLATIVPGDV	102
  Query:	101	LLAVVPVLLIFVALYFVFSPKLDGSKEGKARMSFFLFGLTVAPLLGFYDGVFGPGVGSFF	160
  		L A++P LLI +ALYF   P + G  +  +R++ F+F LT+ PL+GFYDGVFGPG GSFF
  Sbjct:	103	LKAILPFLLIAIALYFGLKPNM-GDVDQHSRVTPFVFTLTLVPLIGFYDGVFGPGTGSFF	161
  Query:	161	LIAFIVLLGCKLLNAMSYTKLANVACNLGSLSVFLLHGSIIFPIVATMAVGAFVGANLGA	220
  		++ F+ L G  +L A ++TK  N   N+G+  VFL  G++++ +   M +G F+GA +G+
  Sbjct:	162	MLGFVTLAGFGVLKATAHTKFLNFGSNVGAFGVFLFFGAVLWKVGLLMGLGQFLGAQVGS	221
  Query:	221	RFAVRFGSKLIKPLLIVISISMAVKLLIDERNPL	254
  		R+A+  G+K+IKPLL+++SI++A++LL D  +PL
  Sbjct:	222	RYAMAKGAKIIKPLLVIVSIALAIRLLADPTHPL	255

• Based on this analysis, including the presence of a putative leader sequence (double-underlined) and several putative transmembrane domains (single-underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 52
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 441>:

• 1	..CTGCTAGGGT ATTGCATCGG TTATCGGTAC GGCTGTTGCA
  	GCAAAACCAG
  51	CCGCAGACGG ATTATTTGGT CAAATTCGGA TCGTTTTGGG
  	CGAG.ATTTT
  101	TGGTTTTCTG GGACTGTATG ACGTCTATGC TTCGGCATGG
  	TTTGTCGTTA
  151	TCATGATGTT TTTGGTGGTT TCTACCAGTT TGTGCCTGAT
  	TCGCAATGTG
  201	CCGCCGTTCT GGCGCGAAAT GAAGTCTTTT CGGGAAAAGG
  	TTAAAGAAAA
  251	ATCTCTGGCG GCGATGCGCC ATTCTTCGCT GTTGGATGTA
  	AAAATTGCGC
  301	CCGAGGTTGC CAAACGTTAT CTGGAAGTAC AAGGTTTTCA
  	GGGGAAAACC
  351	ATTAACCGTG AAGACGGGTC GGTTCTGATT GCCGCCAAAA
  	AAGGCACAAT
  401	GAACAAATGG GGCTATATCT TTGCCCATGT TGCTTTGATT
  	GTCATTTGCC
  451	TGGGCGGGTT GATAGACAGT AACCTGCTGT TGAAACTGGG
  	TATGCTGACC
  501	GGTCGGATTG TTCCGGACAA TCAGGCGGTT TATGCCAAGG
  	ATTTC.AAGC
  551	CCGAAAGTAT .TTTGGGTGC gTCCAATCTC TCATTTAGGG
  	GCAACGTCAA
  601	TATTTCCG.A GGGGCAGAgT GCGGATGTGG TTTTCCTGA

• This corresponds to the amino acid sequence <SEQ ID 442; ORF110>:

• 1	..LLGIASVIGT LLQQNQPQTD YLVKFGSFWA XIFGFLGLYD
  	VYASAWFVVI
  51	MMFLVVSTSL CLIRNVPPFW REMKSFREKV KEKSLAAMRH
  	SSLLDVKIAP
  101	EVAKRYLEVQ GFQGKTINRE DGSVLIAAKK GTMNKWGYIF
  	AHVALIVICL
  151	GGLIDSNLLL KLGMLTGRIF RTIRRFMPRI XKPESXFGCV
  	QSLI*GQRQY
  201	FXRGRVRMWF S*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with ORF88a from N. meningitidis (Strain A)
• ORF110 shows 91.5% identity over a 188aa overlap with ORF88a from strain A of N. meningitidis:
• 
• However, ORF88 and ORF110 do not align, because they represent two different fragments of the same protein.
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF110 shows 88.6% identity over a 211 as overlap with a predicted ORF (ORF110.ng) from N. gonorrhoeae:
• 
• The complete length ORF110ng nucleotide sequence <SEQ ID 443> is predicted to encode a protein having amino acid sequence <SEQ ID 444>:

• 1	MSKSRISPTL LSRPWFAFFS SMRFAVALLS LLGIASVIGT
  	VLQQNQPQTD
  51	YLVKFGPFWT RIFDFLGLYD VYASAWFVVI MMFLVVSTSL
  	CLIRNVPPFW
  101	REMKSFREKV KEKSLAAMRH SSLLDVKIAP EVAKRYLEVR
  	GFQGKTVSRE
  151	DGSVLIAAKK GTMNKWGYIX AHVALIVICL GRLINXNLLL
  	KLGMLAGSIF
  201	RNNRRVMPRI SKPESIWGGV QSLIKGQRQY FQRGKVRMWF
  	S*

• Based on the putative transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 53
• The following DNA sequence was identified in N. meningitidis <SEQ ID 445>:

• 1	ATGCCGTCTG AAACACGCCT GCCGAACTTT ATCCGCGTCT
  	TGATATTTGC
  51	CCTGGGTTTC ATCTTCCTGA ACGCCTGTTC GGAACAAACC
  	GCGCAAACCG
  101	TTACCCTGCA AGGCGAAACG ATGGGCACGA CCTATACCGT
  	CAAATACCTT
  151	TCAAATAATC GGGACAAACT CCCCTCACCT GCCGAAATAC
  	AAAAACGCAT
  201	CGATGACGCG CTTAAAGAAG TCAACCGGCA GATGTCCACC
  	TATCAGCCCG
  251	ACTCCGAAAT CAGCCGGTTC AACCAACACA CAGCCGGCAA
  	GCCCCTCCGC
  301	ATTTCAAGCG ACTTCGCACA CGTTACTGCC GAAGCCGTCC
  	GCCTGAACCG
  351	CCTGACACAC GGCGCGCTGG ACGTAACCGT CGGCCCCTTG
  	GTCAACCTTT
  401	GGGGATTCGG CCCCGACAAA TCCGTTACCC GTGAACCGTC
  	GCCGGAACAA
  451	ATCAAACAGG CGGCATCTTA TACGGGCATA GACAAAATCA
  	TTTTGAAACA
  501	AGGCAAAGAT TACGCTTCCT TGAGCAAAAC CCACCCCAAG
  	GCCTATTTGG
  551	ATTTATCTTC GATTGCCAAA GGCTTCGGCG TTGATAAAGT
  	TGCGGGCGAA
  601	CTGGAAAAAT ACGGCATTCA AAATTATCTG GTCGAAATCG
  	GCGGCGAGTT
  651	GCACGGCAAA GGCAAAAACG CGCGCGGCGA ACCGTGGCGC
  	ATCGGTATCG
  701	AGCAGCCCAA TATCGTCCAA GGCGGCAATA CGCAGATTAT
  	CGTCCCGCTG
  751	AACAACCGTT CGCTTGCCAC TTCCGGCGAT TACCGTATTT
  	TCCACGTCGA
  801	TAAAAACGGC AAACGCCTCT CCCATATCAT CAACCCGAAC
  	AACAAACGAC
  851	CCATCAGCCA CAACCTCGCC TCCATCAGCG TGGTCGCAGA
  	CAGTGCGATG
  901	ACGGCGGACG GCTTGTCCAC AGGATTATTC GTATTGGGCG
  	AAACCGAAGC
  951	CTTAAAGCTG GCAGAGCGCG AAAAACTCGC TGTTTTCCTG
  	ATTGTCAGGG
  1001	ATAAAGGCGG CTACCGCACC GCCATGTCTT CCGAATTTGA
  	AAAACTGCTC
  1051	CGCTAA

• This corresponds to the amino acid sequence <SEQ ID 446; ORF111>:

• 1	MPSETRLPNF IRVLIFALGF IFLNACSEQT AQTVTLQGET
  	MGTTYTVKYL
  51	SNNRDKLPSP AEIQKRIDDA LKEVNRQMST YQPDSEISRF
  	NQHTAGKPLR
  101	ISSDFAHVTA EAVRLNRLTH GALDVTVGPL VNLWGFGPDK
  	SVTREPSPEQ
  151	IKQAASYTGI DKIILKQGKD YASLSKTHPK AYLDLSSIAK
  	GFGVDKVAGE
  201	LEKYGIQNYL VEIGGELHGK GKNARGEPWR IGIEQPNIVQ
  	GGNTQIIVPL
  251	NNRSLATSGD YRIFHVDKNG KRLSHIINPN NKRPISHNLA
  	SISVVADSAM
  301	TADGLSTGLF VLGETEALKL AEREKLAVFL IVRDKGGYRT
  	AMSSEFEKLL
  351	R*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF111 shows 96.9% identity over a 351aa overlap with an ORF (ORF111a) from strain A of N. meningitidis:
• 
• The complete length ORF111a nucleotide sequence <SEQ ID 447> is:

• 1	ATGCCGTCTG AAACACGCCT GCCGAACTTT ATCCGCACCT
  	TGATATTTGC
  51	CCTGAGTTTT ATCTTCCTGA ACGCCTGTTC GGAACAAACC
  	GCGCAAACCG
  101	TTACCCTGCA AGGTGAAACG ATGGGCACGA CCTATACCGT
  	CAAATACCTT
  151	TCAAATAATC GGGACNAACT CCCNTCACCT GCCGAAATAC
  	AAAANCGCAT
  201	CGATGACGCG CTTAAAGAAG TCAACCGGCA GATGTCCACC
  	TATCAGCCCG
  251	ACTCCGAAAT CAGCCGGTTC AACCAACACA CAGCCGGCAA
  	GCCCCTCCGC
  301	ATTTCAAGCG ACTTCGCACA CGTTACTGCC GAAGCCGTCC
  	ACCTGAACCG
  351	CCTGACACAC GGCGCGCTGG ACGTAACCGT CGGCCCCTTG
  	GTCAACCTTT
  401	GGGGATTCGG CCCCGACAAA TCCGTTACCC GTGAACCGTC
  	GCCGGAACAA
  451	ATCAAACAAG CAGCATCTTA TACGGGCATA GACAAAATCA
  	TTTTGAAACA
  501	AGGCAAAGAT TACGCTTCCT TGAGCAAAAC CCACCCCAAG
  	GCCTATTTGG
  551	ATTTATCTTC GATTGCCAAA GGCTTCGGCG TTGATNANGT
  	TGCGGGCGAA
  601	CTGGAAAAAT ACGGCATTCA AAATTATCTG GTCGAAATCG
  	GCGGNGAGTT
  651	GCACGGCAAA GNCAAAAACG CGCGCGGCGA ACCTTGGCGC
  	ATCGGCATCG
  701	AACAGCCCAA CATCGTCCAA GGCGGCAATA CGCAGATTAT
  	CGTCCCGCTG
  751	AACAACCGTT CGNTTGCCAC TTCCGGCGAT TACCGTATTT
  	TCCACGTCGA
  801	TAAAAGCGGC AAACGCCTCT CCCATATCAT TAATCCGAAC
  	AACAAACGAC
  851	CCATCAGCCA CAACCTCGCC TCCATCAGCG TGNTCGCAGA
  	CAGTGCGATG
  901	ACGGCGGACG GCTTNTCCAC AGGATTATTC GTATTGGGCG
  	AAACCGAAGC
  951	CTTAAAGCTG GCAGAGCGCG AAAAACTCGC TGTTTTCCTG
  	ATTGTCAGGG
  1001	ATAAAGGCGG CTACCGCACC GCCATGTCTT CCGAATTTGA
  	AAAACTGCTC
  1051	CGCTAA

• This encodes a protein having amino acid sequence <SEQ ID 448>:

• 1	MPSETRLPNF IRTLIFALSF IFLNACSEQT AQTVTLQGET
  	MGTTYTVKYL
  51	SNNRDXLPSP AEIQXRIDDA LKEVNRQMST YQPDSEISRF
  	NQHTAGKPLR
  101	ISSDFAHVTA EAVHLNRLTH GALDVTVGPL VNLWGFGPDK
  	SVTREPSPEQ
  151	IKQAASYTGI DKIILKQGKD YASLSKTHPK AYLDLSSIAK
  	GFGVDXVAGE
  201	LEKYGIQNYL VEIGGELHGK XKNARGEPWR IGIEQPNIVQ
  	GGNTQIIVPL
  251	NNRSXATSGD YRIFHVDKSG KRLSHIINPN NKRPISHNLA
  	SISVXADSAM
  301	TADGXSTGLF VLGETEALKL AEREKLAVFL IVRDKGGYRT
  	AMSSEFEKLL
  351	R*

  
  Homology with a Predicted ORF from N. gonorrhoeae
• ORF111 shows 96.6% identity over a 351aa overlap with a predicted ORF (ORF111.ng) from N. gonorrhoeae:
• 
• The complete length ORF111ng nucleotide sequence <SEQ ID 449> is:

• 1	ATGCCGTCTG AAACACGCCT GCCGAACCTT ATCCGCGCCT
  	TGATATTTGC
  51	CCTGGGTTTC ATCTTCCTGA ACGCCTGTTC GGaacaaacC
  	GCGCAaaccg
  101	TTACCCTGCA AGGCGAAAcg aTGGGTACGA CCTATACCGT
  	CAAATACCTT
  151	TCAAATAATC GGGACAAACT CCCCTCCCCT GCCAAAATAC
  	AAAAGCGCAT
  201	TGATGATGCG CTTAAAGAAG TCAACCGGCA GATGTCCACC
  	TACCAGACCG
  251	ATTCCGAAAT CAGCCGGTTC AACCAACACA CAGCCGGCAA
  	GCCCCTCCGC
  301	ATTTCAAGCG ATTTCGCACA CGTTACCGCC GAAGCCGTCC
  	GCCTGAACCG
  351	CCTGACTCAC GGCGCACTGG ACGTAACCGT CGGCCCTTTG
  	GTCAACCTTT
  401	GGGGGTTCGG CCCCGACAAA TCCGTTACCC GTGAACCGTC
  	GCCGGAACAA
  451	ATCAAACAGG CGGCATCTTA TACGGGCATA GACAAAATCA
  	TTTTGCAACA
  501	AGGCAAAGAT TACGCTTCCT TGAGCAAAAC CCACCCCAAA
  	GCCTATTTGG
  551	ATTTATCTTC GATTGCCAAA GGCTTCGGCG TTGATAAAGT
  	TGCGGGCGAA
  601	CTGGAAAAAT ACGGCATTCA AAATTATCTG GTCGAAAtcg
  	gcggcGAGTT
  651	GCACGGCAAA GGCAAAAATG CGCACGGCGA ACCGTGGCGC
  	ATCGGTATAG
  701	AGCAACCCAA TATCATCCAA GgcgGCAata CGCAGATTAt
  	cgtcccgctg
  751	aaCaaccgtt cgctTGCCAC TTCCGGCGAT TAccgtaTTT
  	tccacgtcgA
  801	TAAAAAcggc aaacgccttt cccacaTCAT CAATCCCaAC
  	aacAAACgac
  851	ccATCAGcca caacctcgcc tccatcagcg tggtctcAGA
  	CAGTGCAATG
  901	ACGGCGGACG GTTtatCCAC AGGATTATTT GTTTTAGGCG
  	AAACCGAAGC
  951	CTTAAGGCTG GCAGAACAAG AAAAACTCGC TGTTTTCCTA
  	ATTGTCCGGG
  1001	ATAAGGACGG CTACCGCACC GCCATGTCTT CCGAATTTGC
  	CAAGCTGCTC
  1051	CGCTAA

• This encodes a protein having amino acid sequence <SEQ ID 450>:

• 1	MPSETRLPNL IRALIFALGF IFLNACSEQT AQTVTLQGET
  	MGTTYTVKYL
  51	SNNRDKLPSP AKIQKRIDDA LKEVNRQMST YQTDSEISRF
  	NQHTAGKPLR
  101	ISSDFAHVTA EAVRLNRLTH GALDVTVGPL VNLWGFGPDK
  	SVTREPSPEQ
  151	IKQAASYTGI DKIILQQGKD YASLSKTHPK AYLDLSSIAK
  	GFGVDKVAGE
  201	LEKYGIQNYL VEIGGELHGK GKNAHGEPWR IGIEQPNIIQ
  	GGNTQIIVPL
  251	NNRSLATSGD YRIFHVDKNG KRLSHIINPN NKRPISHNLA
  	SISVVSDSAM
  301	TADGLSTGLF VLGETEALRL AEQEKLAVFL IVRDKDGYRT
  	AMSSEFAKLL
  351	R*

• This protein shows homology with a hypothetical lipoprotein precursor from H. influenzae:

• sp|P44550|YOJL_HAEIN HYPOTHETICAL LIPOPROTEIN HI0172 PRECURSOR >gi|1074292|pir|4
  hypothetical protein HI0172 - Haemophilus influenzae (strain Rd KW20)
  >gi|1573128 (U32702) hypothetical [Haemophilus influenzae] Length = 346
  Score = 353 bits (896), Expect = 9e−97
  Identities = 181/344 (52%), Positives = 247/344 (71%), Gaps = 4/344 (1%)

  Query:	7	LPNLIRALIFALGFIFLNACSEQTAQTVTLQGETMGTTYTVKYLSNNRDKLPSPAKIQKR	66
  		+  LI  +I     + L AC ++T + ++L G+TMGTTY VKYL +      S  K  +
  Sbjct:	1	MKKLISGIIAVAMALSLAACQKET-KVISLSGKTMGTTYHVKYLDDGSITATSE-KTHEE	58
  Query:	67	IDDALKEVNRQMSTYQTDSEISRFNQHT-AGKPLRISSDFAHVTAEAVRLNRLTHGALDV	125
  		I+  LK+VN +MSTY+ DSE+SRFNQ+T    P+ IS+DFA V AEA+RLN++T GALDV
  Sbjct:	59	IEAILKDVNAKMSTYKKDSELSRFNQNTQVNTPIEISADFAKVLAEAIRLNKVTEGALDV	118
  Query:	126	TVGPLVNLWGFGPDKSVTREPSPEQIKQAASYTGIDKIILQQGKDYASLSKTHPKAYLDL	185
  		TVGP+VNLWGFGP+K   ++P+PEQ+ +  ++ GIDKI L   K+ A+LSK  P+ Y+DL
  Sbjct:	119	TVGPVVNLWGFGPEKRPEKQPTPEQLAERQAWVGIDKITLDTNKEKATLSKALPQVYVDL	178
  Query:	186	SSIAKGFGVDKVAGELEKYGIQNYLVEIGGELHGKGKNAHGEPWRIGIEQPNIIQGGNTQ	245
  		SSIAKGFGVD+VA +LE+   QNY+VEIGGE+  KGKN  G+PW+I IE+P        +
  Sbjct:	179	SSIAKGFGVDQVAEKLEQLNAQNYMVEIGGEIRAKGKNIEGKPWQIAIEKPTTTGERAVE	238
  Query:	246	IIVPLNNRSLATSGDYRIFHVDKNGKRLSHIINPNNKRPISHNLASISVVSDSAMTADGL	305
  		++ LNN  +A+SGDYRI+  ++NGKR +H I+P    PI H+LASI+V++ ++MTADGL
  Sbjct:	239	AVIGLNNMGMASSGDYRIY-FEENGKRFAHEIDPKTGYPIQHHLASITVLAPTSMTADGL	297
  Query:	306	STGLFVLGETEALRLAEQEKLAVFLIVRDKDGYRTAMSSEFAKL	349
  		STGLFVLGE +AL +AE+  LAV+LI+R  +G+ T  SS F KL
  Sbjct:	298	STGLFVLGEDKALEVAEKNNLAVYLIIRTDNGFVTKSSSAFKKL	341

• Based on this analysis, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 54
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 451>:

• 1	..CCGTGCCGCC GACAGGGCGA CGACGTGTAT GCGGCGCACG
  	CGTCCCGTCA
  51	AAAATTGTGG CTGCGCTTCA TCGGCGGCCG GTCGCATCAA
  	AATATACGGG
  101	GCGGCGCGGC TGCGGACGGG TGGCGCAAAG GCGTGCAAAT
  	CGGCGGCGAG
  151	GTGTTTGTAC GGCAAAATGA AGGCAGCCkA yTGGCAATCG
  	GCGTGATGGG
  201	CGGCAGGGCC GGCCAGCACG CwTCAGTCAA CGGCAAAGGC
  	GGTGCGGCAG
  251	gCAGTGATTT GTATGGTTAT GgCGGGGgTG TTTATGCTgC
  	GTGGCATCAG
  301	TTGCGCGATA AACAAACGGG TgCGTATTTG GACGGCTGGT
  	TGCAATACCA
  351	ACGTTTCAAA CACCGCATCA ATGATGAAAA CCGTGCGGAA
  	CgCTACAAAA
  401	CCAAAGGTTG GACGGCTTCT GTCGAAGGCG GCTACAACGC
  	GCTTGTGGCG
  451	GAAGGCATTG TCGGAAAAGG CAATAATGTG CGGTTTTACC
  	TACAACCGCA
  501	GgCGCAGTTT ACCTACTTGG GCGTAAACGG CGGCTTTACC
  	GACAGCGAGG
  551	GGACGGCGGT CGGACTGCTC GGCAGCGGTC AGTGGCAAAG
  	CCGCGCCGGC
  601	AtTCGGGCAA AAACCCGTTT TGCTTTGCGT AACGGTGTCA
  	ATCTTCAGCC
  651	TTTTGCCGCT TTTAATGTtt TGCACAGGTC AAAATCTTTC
  	GGCGTGGAAA
  701	TGGACGGCGA AAAACAGACG CTGGCAGGCA GGACGGCACT
  	CGAAGGGCGG
  751	TTCGGTATTG AAGCCGGTTG GAAAGGCCAT ATGTCCGCA..

• This corresponds to the amino acid sequence <SEQ ID 452; ORF35>:

• 1	..PCRRQGDDVY AAHASRQKLW LRFIGGRSHQ NIRGGAAADG
  	WRKGVQIGGE
  51	VFVRQNEGSX LAIGVMGGRA GQHASVNGKG GAAGSDLYGY
  	GGGVYAAWHQ
  101	LRDKQTGAYL DGWLQYQRFK HRINDENRAE RYKTKGWTAS
  	VEGGYNALVA
  151	EGIVGKGNNV RFYLQPQAQF TYLGVNGGFT DSEGTAVGLL
  	GSGQWQSRAG
  201	IRAKTRFALR NGVNLQPFAA FNVLHRSKSF GVEMDGEKQT
  	LAGRTALEGR
  251	FGIEAGWKGH MSA..

• Computer analysis of this amino acid sequence gave the following results:
• Homology with Putative Secreted VirG-Homolgue of N. meningitidis (Accession Number A32247)
• ORF and virg-h protein show 51% aa identity in 261aa overlap:

• Orf35	5	QGDDVYAAHASRQKLWLRFIGGRSHQNIRGGAA-ADGWRKGVQIGGEVFVRQNEGSXLAI	63
  		+  D++     R+ LWLR I G S+Q ++G  A  +G+RKGVQ+GGEVF  QNE + L+I
  virg-h	396	KNSDIFDRTLPRKGLWLRVIDGHSNQWVQGKTAPVEGYRKGVQLGGEVFTWQNESNQLSI	455
  Orf35	64	GVMGGRAGQHASVNGKG--GAAGSDLYGYGGGVYAAWHQLRDKQTGAYLDGWLQYQRFKH	121
  		G+MGG+A Q ++ +          ++ G+G GVYA WHQL+DKQTGAY D W+QYQRF+H
  virg-h	456	GLMGGQAEQRSTFHNPDTDNLTTGNVKGFGAGVYATWHQLQDKQTGAYADSWMQYQRFRH	515
  Orf35	122	RINDENRAERYKTKGWTASVEGGYNALVAEGIVGKGNNVRFYLQPQAQFTYLGVNGGFTD	181
  		RIN E+  ER+ +KG TAS+E GYNAL+AE    KGN++R YLQPQAQ TYLGVNG F+D
  virg-h	516	RINTEDGTERFTSKGITASIEAGYNALLAEHFTKKGNSLRVYLQPQAQLTYLGVNGKFSD	575
  Orf35	182	SEGTAVGLLGSGQWQSRAGIRAKTRFALRNGVNLQPFAAFNVLHRSKSFGVEMDGEKQTL	241
  		SE   V LLGS Q Q+R G++AK +F+L   + ++PFAA N L+ +K FGVEMDGE++ +
  virg-h	576	SENAHVNLLGSRQLQTRVGVQAKAQFSLYKNIAIEPFAAVNALYHNKPFGVEMDGERRVI	635
  Orf35	242	AGRTALEGRFGIEAGWKGHMS	262
  		+TA+E + G+    K H++
  virg-h	636	NNKTAIESQLGVAVKIKSHLT	656

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF35 shows 96.9% identity over a 259aa overlap with an ORF (ORF35a) from strain A of N. meningitidis:
• 
• The complete length ORF35a nucleotide sequence <SEQ ID 453> is:

• 1	ATGTTCAGAG CTCAGCTTGG TTCAAATACT CGTTCTACCA
  	AAATCGGCGA
  51	CGATGCCGAT TTTTCATTTT CAGACAAGCC GAAACCCGGC
  	ACTTCCCATT
  101	ATTTTTCCAG CGGTAAAACC GATCAAAATT CATCCGAATA
  	TGGGTATGAC
  151	GAAATCAATA TCCAAGGTAA AAACTACAAT AGCGGCATAC
  	TCGCCGTCGA
  201	TAATATGCCC GTTGTTAAGA AATATATTAC AGATACTTAC
  	GGGGATAATT
  251	TAAAGGATGC GGTTAAGAAG CAATTACAGG ATTTATACAA
  	AACAAGACCC
  301	GAAGCTTGGG AAGAAAATAA AAAACGGACT GAGGAGGCGT
  	ATATAGAACA
  351	GCTTGGACCA AAATTTAGTA TACTCAAACA GAAAAACCCC
  	GATTTAATTA
  401	ATAAATTGGT AGAAGATTCC GTACTCACTC CTCATAGTAA
  	TACATCACAG
  451	ACTAGTCTCA ACAACATCTT CAATAAAAAA TTACACGTCA
  	AAATCGAAAA
  501	CAAATCCCAC GTCGCCGGAC AGGTGTTGGA ACTGACCAAG
  	ATGACGCTGA
  551	AAGATTCCCT TTGGGAACCG CGCCGCCATT CCGACATCCA
  	TATGCTGGAA
  601	ACTTCCGATA ATGCCCGCAT CCGCCTGAAC ACGAAAGATG
  	AAAAACTGAC
  651	CGTCCATAAA GCGTATCAGG GCGGTGCGGA TTTCCTGTTC
  	GGCTACGACG
  701	TGCGGGAGTC GGACAAACCC GCCCTGACCT TTGAAGAAAA
  	AGTCAGCGGA
  751	CAATCCGGCG TGGTTTTGGA ACGCCGGCCG GAAAATCTGA
  	AAACGCTCGA
  801	CGGGCGCAAA CTGATTGCGG CGGAAAAGGC AGACTCTAAT
  	TCGTTTGCGT
  851	TTAAACAAAA TTACCGGCAG GGACTGTACG AATTATTGCT
  	CAAGCAATGC
  901	GAAGGCGGAT TTTGCTTGGG CGTGCAGCGT TTGGCTATCC
  	CCGAGGCGGA
  951	AGCGGTTTTA TATGCCCAAC AGGCTTATGC GGCAAATACT
  	TTGTTCGGGC
  1001	TGCGTGCCGC CGACAGGGGC GACGACGTGT ATGCCGCCGA
  	TCCGTCCCGT
  1051	CAAAAATTGT GGCTGCGCTT CATCGGCGGC CGGTCGCATC
  	AAAATATACG
  1101	GGGCGGCGCG GCTGCGGACG GGCGGCGCAA AGGCGTGCAA
  	ATCGGCGGCG
  1151	AGGTGTTTGT ACGGCAAAAT GAAGGCAGCC GGCTGGCAAT
  	CGGCGTGATG
  1201	GGCGGCAGGG CTGGCCAGCA CGCATCAGTC AACGGCAAAG
  	GCGGTGCGGC
  1251	AGGCAGTTAT TTGCATGGTT ATGGCGGGGG TGTTTATGCT
  	GCGTGGCATC
  1301	AGTTGCGCGA TAAACAAACG GGTGCGTATT TGGACGGCTG
  	GTTGCAATAC
  1351	CAACGTTTCA AACACCGCAT CAATGATGAA AACCGTGCGG
  	AACGCTACAA
  1401	AACCAAAGGT TGGACGGCTT CTGTCGAAGG CGGCTACAAC
  	GCGCTTGTGG
  1451	CGGAAGGCGT TGTCGGAAAA GGCAATAATG TGCGGTTTTA
  	CCTGCAACCG
  1501	CAGGCGCAGT TTACCTACTT GGGCGTAAAC GGCGGCTTTA
  	CCGACAGCGA
  1551	GGGGACGGCG GTCGGACTGC TCGGCAGCGG TCAGTGGCAA
  	AGCCGCGCCG
  1601	GCATTCGGGC AAAAACCCGT TTTGCTTTGC GTAACGGTGT
  	CAATCTTCAG
  1651	CCTTTTGCCG CTTTTAATGT TTTGCACAGG TCAAAATCTT
  	TCGGCGTGGA
  1701	AATGGACGGC GAAAAACAGA CGCTGGCAGG CAGGACGGCG
  	CTCGAAGGGC
  1751	GGTTCGGCAT TGAAGCCGGT TGGAAAGGCC ATATGTCCGC
  	ACGCATCGGA
  1801	TACGGCAAAA GGACGGACGG CGACAAAGAA GCCGCATTGT
  	CGCTCAAATG
  1851	GCTGTTTTGA

• This encodes a protein having amino acid sequence <SEQ ID 454>:

• 1	MFRAQLGSNT RSTKIGDDAD FSFSDKPKPG TSHYFSSGKT
  	DQNSSEYGYD
  51	EINIQGKNYN SGILAVDNMP VVKKYITDTY GDNLKDAVKK
  	QLQDLYKTRP
  101	EAWEENKKRT EEAYIEQLGP KFSILKQKNP DLINKLVEDS
  	VLTPHSNTSQ
  151	TSLNNIFNKK LHVKIENKSH VAGQVLELTK MTLKDSLWEP
  	RRHSDIHMLE
  201	TSDNARIRLN TKDEKLTVHK AYQGGADFLF GYDVRESDKP
  	ALTFEEKVSG
  251	QSGVVLERRP ENLKTLDGRK LIAAEKADSN SFAFKQNYRQ
  	GLYELLLKQC
  301	EGGFCLGVQR LAIPEAEAVL YAQQAYAANT LFGLRAADRG
  	DDVYAADPSR
  351	QKLWLRFIGG RSHQNIRGGA AADGRRKGVQ IGGEVFVRQN
  	EGSRLAIGVM
  401	GGRAGQHASV NGKGGAAGSY LHGYGGGVYA AWHQLRDKQT
  	GAYLDGWLQY
  451	QRFKHRINDE NRAERYKTKG WTASVEGGYN ALVAEGVVGK
  	GNNVRFYLQP
  501	QAQFTYLGVN GGFTDSEGTA VGLLGSGQWQ SRAGIRAKTR
  	FALRNGVNLQ
  551	PFAAFNVLHR SKSFGVEMDG EKQTLAGRTA LEGRFGIEAG
  	WKGHMSARIG
  601	YGKRTDGDKE AALSLKWLF*

  
  Homology with a Predicted ORF from N. gonorrhoeae
• ORF35 shows 51.7% identity over a 261aa overlap with a predicted ORF (ORF35ngh) from N. gonorrhoeae:
• 
• A partial ORF35ngh nucleotide sequence <SEQ ID 455> is predicted to encode a protein having partial amino acid sequence <SEQ ID 456>:

• 1	..KKLRDRNSEY WKEETYHIKS NGRTYPNIPA LFPKHPFDPF
  	ENINNSKKIS
  51	FYDKEYTEDY LVGFARGFGV EKRNGEEEKP LRQYFKDCVN
  	TENSNNDNCK
  101	ISSFGNYGPI LIKSDIFALA SQIKNSHINS EILSVGNYIE
  	WLRPTLNKLT
  151	GWQEHLYAGL DPFHYIEVTD NSHVIGQTID LGALELTNSL
  	WKPRWNSNID
  201	YLITKNAEIR FNTKNESLLV KEDYAGGARF RFAYDLKDKV
  	PEIPVLTFEK
  251	NITGTSDIIF EGKALDNLKH LDGHQIVKVN DTADKDAFRL
  	SSKYRKGIYT
  301	LSLQQRPEGF FTKVQERDDI AIYAQQAQAA NTLFALRLND
  	KNSDIFDRTL
  351	PRKGLWLRVI DGHSNQWVQG KTAPVEGYRK GVQLGGEVFT
  	WQNESNQLSI
  401	GLMGGQAEQR STFRNPDTDN LTTGNVKGFG AGVYATWHQL
  	QDKQTGAYVD
  451	SWMQYQRFRH RINTEYATER FTSKGITASI EAGYNALLAE
  	HFTKKGNSLR
  501	VYLQPQAQLT YLGVNGKFSD SENAQVNLLG SRQLQSRVGV
  	QAKAQFAFTN
  551	GVTFQPFVAV NSIYQQKPFG VEIDGDRRVI NNKTVIETQL
  	GVAAKIKSHL
  601	TLQASFNRQT SKHHHAKQGA LNLQWTF*

• Based on this prediction, these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 55
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 457>:

• 1	..GCGGAATATG TTCAGTTCTC TATAGATTTG TTCAGTGTGG
  	GTAAATCGGG
  51	GGGCGGTATA CCTAAGGCTA AGCCTGTGTT TGATGCGAAA
  	CCGAGATGGG
  101	AGGTTGATAG GAAGCTTAAT AAATTGACAA CTCGTGAGCA
  	GGTGGAGAAA
  151	AATGTTCAGG AAACGAGAAG AAGGAGTCAG AGTAGTCAGT
  	TTAAAGCCCA
  201	TGCGCAACGA GAATGGGAAA ATAAAACAGG GTTAGATTTT
  	AATCATTTTA
  251	TAGGTGGTGA TATCAATAAA AAAGGCACAG TAACAGGAGG
  	GCATAGTCTA
  301	ACCCGTGGTG ATGTACGGGT GATACAACAA ACCTCGGCAC
  	CTGATAAACA
  351	TGGGGT.TTA TCAAGCGACA GTGGAAATTN A

• This corresponds to the amino acid sequence <SEQ ID 458; ORF46>:

• 1	..AEYVQFSIDL FSVGKSGGGI PKAKPVFDAK PRWEVDRKLN
  	KLTTREQVEK
  51	NVQETRRRSQ SSQFKAHAQR EWENKTGLDF NHFIGGDINK
  	KGTVTGGHSL
  101	TRGDVRVIQQ TSAPDKHGXL SSDSGNX

• Further work revealed further partial nucleotide sequence <SEQ ID 459>:

• 1	..GCAGTGTGCC TnCCGATGCA TGCACACGCC TCAnATTTGG
  	CAAACGATTC
  51	TTTTATCCGG CAGGTTCTCG ACCGTCAGCA TTTCGAACCC
  	GACGGGAAAT
  101	ACCACCTATT CGGCAGCAGG GGGGAACTTG CCGAGCGCCA
  	GTCTCATATC
  151	GGATTGGGAA AAATACAAAG CCATCAGTTG GGCAACCTGA
  	TGATTCAACA
  201	GGCGGCCATT AAAGGAAATA TCGGCTACAT TGTCCGCTTT
  	TCCGATCACG
  251	GGCACGAAGT CCATTCCCCs TTCGACAACC ATGCCTCACA
  	TTCCGATTCT
  301	GATGAAGCCG GTAGTCCCGT TGACGGATTT AGCCTTTACC
  	GCATCCATTG
  351	GGACGGATAC GAACACCATC CCGCCGACGG CTATGACGGG
  	CCACAGGGCG
  401	GCGGCTATCC CGCTCCCAAA GGCGCGAGGG ATATATACAG
  	TTACGACATA
  451	AAAGGCGTTG CCCAAAATAT CCGCCTCAAC CTGACCGACA
  	ACCGCAGCAC
  501	CGGACAACGG CTTGCCGACC GTTTCCACAA TGCCGGTAGT
  	ATGCTGACGC
  551	AAGGAGTAGG CGACGGATTC AAACGCGCCA CCCGATACAG
  	CCCCGAGCTG
  601	GACAGATCGG GCAATGCCGC CGAAGCCTTC AACGGCACTG
  	CAGATATCGT
  651	TAAAAACATC ATCGGCGCTG CAGGAGAAAT TGT

• This corresponds to the amino acid sequence <SEQ ID 460; ORF46-1>:

• 1	..AVCLPMHAHA SXLANDSFIR QVLDRQHFEP DGKYHLFGSR
  	GELAERQSHI
  51	GLGKIQSHQL GNLMIQQAAI KGNIGYIVRF SDHGHEVHSP
  	FDNHASHSDS
  101	DEAGSPVDGF SLYRIHWDGY EHHPADGYDG PQGGGYPAPK
  	GARDIYSYDI
  151	KGVAQNIRLN LTDNRSTGQR LADRFHNAGS MLTQGVGDGF
  	KRATRYSPEL
  201	DRSGNAAEAF NGTADIVKNI IGAAGEI

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF46 shows 98.2% identity over, a 111 aa overlap with a predicted ORF (ORF46ng) from N. gonorrhoeae:
• 
• A partial ORF46ng nucleotide sequence <SEQ ID 461> is predicted to encode a protein having partial amino acid sequence <SEQ ID 462>:

• 1	..RRLKHCCHAR LGSAFHRKQD GAHQRFGRYG ATQRLCRSSH
  	PRLGSPKPQC
  51	RTRHRSRQQY LYGSHPHQRD WSCPGKIQLG RHHGTSCRAV
  	ADXRDRICER
  101	EIRRQRQXCR CRLGKIPSLS IPKYPLKLEQ RYGKENITSS
  	TVPPSNGKNV
  151	KLADQRHPKT GVPFDGKGFP NFEKHVKYDT KLDIQELSGG
  	GIPKAKPVFD
  201	AKPRWEVDRK LNKLTTREQV EKNVQETRRR SQSSQFKAHA
  	QREWENKTGL
  251	DFNHFIGGDI NKKGAVTGGH SLTRGDVRVI QQTSAPDKHG
  	VLSSDSGN*

• Further work revealed the complete gonococcal DNA sequence <SEQ ID 463>:

• 1	TTGGGCATTT CCCGCAAAAT ATCCCTTATT CTGTCCATAC
  	TGGCAGTGTG
  51	CCTGCCGATG CATGCACACG CCTCAGATTT GGcaAACGAT
  	CCCTTTATCC
  101	GgCaggttcT CGaccGTCAG CATTTCGaac ccgacggGAa
  	ATACCaCCTA
  151	TTcggCaGCA GGGGGGAGCT TgccnagcGC aacggccATa
  	tcggattggG
  201	aaacaTAcaa Agccatcagt tGggccacct gatgattcaa
  	caggcggccg
  251	ttgaaggaaA TAtcgGctac attgtccgct tttccgatca
  	cgggcacaaa
  301	ttccattcgc ccttcGAcaa ccaTGCCTCA CATTCCGATT
  	CTGACGAAGC
  351	CGGTAGTCCC GTTGACGGAT TCAGCCTTTA CCGCATCCAT
  	TGGGACGGAT
  401	ACGAACACCA TCCCGCCGAC GGCTATGACG GGCCACAGGG
  	CGGCGGCTAT
  451	CCCGCTCCCA AAGGCGCGAG GGATATATAC AGCTACGACA
  	TAAAAGGCGT
  501	TGCCCAAAAT ATCCGCCTCA ACCTGACCGA CAACCGCAGC
  	ACCGGACAAC
  551	GGCTTGCCGA CCGTTTCCAC AATGCCGGCG CTATGCTGAC
  	GCAAGGAGTA
  601	GGCGACGGAT TCAAACGCGC CACCCGATAC AGCCCCGAGC
  	TGGACAGATC
  651	GGGCAATGCc gccGAAGCCT TCAACGGCAC TGCAGATATC
  	GTCAAAAACA
  701	TCATCGGCGC GGCAGGAGAA ATTGTCGGCG CAGGCGATGC
  	CGTGCagGGT
  751	ATAAGCGAAG GCTCAAACAT TGCTGTCATG CACGGCTTGG
  	GTCTGCTTTC
  801	CACCGAAAAC AAGATGGCGC GCATCAACGA TTTGGCAGAT
  	ATGGCGCAAC
  851	TCAAAGACTA TGCCGCAGCA GCCATCCGCG ATTGGGCAGT
  	CCAAAACCCC
  901	AATGCCGCAC AAGGCATAGA AGCCGTCAGC AATATCTTTA
  	TGGCAGCCAT
  951	CCCCATCAAA GGGATTGGAG CTGTCCGGGG AAAATACGGC
  	TTGGGCGGCA
  1001	TCACGGCACA TCCTGTCAAG CGGTCGCAGA TGGGCGCGAT
  	CGCATTGCCG
  1051	AAAGGGAAAT CCGCCGTCAG CGACAATTTT GCCGATGCGG
  	CATACGCCAA
  1101	ATACCCGTCC CCTTACCATT CCCGAAATAT CCGTTCAAAC
  	TTGGAGCAGC
  1151	GTTACGGCAA AGAAAACATC ACCTCCTCAA CCGTGCCGCC
  	GTCAAACGGC
  1201	AAAAATGTCA AACTGGCAGA CCAACGCCAC CCGAAGACAG
  	GCGTACCGTT
  1251	TGACGGTAAA GGGTTTCCGA ATTTTGAGAA GCACGTGAAA
  	TATGATACGA
  1301	AGCTCGATAT TCAAGAATTA TCGGGGGGCG GTATACCTAA
  	GGCTAAGCCT
  1351	GTGTTTGATG CGAAACCGAG ATGGGAGGTT GATAGGAAGC
  	TTAATAAATT
  1401	GACAACTCGT GAGCAGGTGG AGAAAAATGT TCAGGAAACG
  	AGAAGAAGGA
  1451	GTCAGAGTAG TCAGTTTAAA GCCCATGCGC AACGAGAATG
  	GGAAAATAAA
  1501	ACAGGGTTAG ATTTTAATCA TTTTATAGGT GGTGATATCA
  	ATAAGAAAGG
  1551	CACAGTAACA GGAGGGCATA GTCTAACCCG TGGTGATGTA
  	CGGGTGATAC
  1601	AACAAACCTC GGCACCTGAT AAACATGGGG TTTATCAAGC
  	GACAGTGGAA
  1651	ATTAAAAAGC CTGATGGAAG TTGGGAGGTG AAAACGAAAA
  	AAGGTGGGAA
  1701	AGTGATGACC AAGCACACCA TGTTCCCAAA AGATTGGGAT
  	GAGGCTAGAA
  1751	TTAGGGCTGA AGTTACTTCG GCTTGGGAAA GTAGAATAAT
  	GCTTAAGGAT
  1801	AATAAATGGC AGGGTACAAG TAAATCGGGT ATTAAAATAG
  	AAGGATTTAC
  1851	CGAACCTAAT AGAACAGCAT ATCCCATTTA TGAATAG

• This corresponds to the amino acid sequence <SEQ ID 464; ORF46ng-1>:

• 1	LGISRKISLI LSILAVCLPM HAHASDLAND PFIRQVLDRQ
  	HFEPDGKYHL
  51	FGSRGELAXR NGHIGLGNIQ SHQLGHLMIQ QAAVEGNIGY
  	IVRFSDHGHK
  101	FHSPFDNHAS HSDSDEAGSP VDGFSLYRIH WDGYEHHPAD
  	GYDGPQGGGY
  151	PAPKGARDIY SYDIKGVAQN IRLNLTDNRS TGQRLADRFH
  	NAGAMLTQGV
  201	GDGFKRATRY SPELDRSGNA AEAFNGTADI VKNIIGAAGE
  	IVGAGDAVQG
  251	ISEGSNIAVM HGLGLLSTEN KMARINDLAD MAQLKDYAAA
  	AIRDWAVQNP
  301	NAAQGIEAVS NIFMAAIPIK GIGAVRGKYG LGGITAHPVK
  	RSQMGAIALP
  351	KGKSAVSDNF ADAAYAKYPS PYHSRNIRSN LEQRYGKENI
  	TSSTVPPSNG
  401	KNVKLADQRH PKTGVPFDGK GFPNFEKHVK YDTKLDIQEL
  	SGGGIPKAKP
  451	VFDAKPRWEV DRKLNKLTTR EQVEKNVQET RRRSQSSQFK
  	AHAQREWENK
  501	TGLDFNHFIG GDINKKGTVT GGHSLTRGDV RVIQQTSAPD
  	KHGVYQATVE
  551	IKKPDGSWEV KTKKGGKVMT KHTMFPKDWD EARIRAEVTS
  	AWESRIMLKD
  601	NKWQGTSKSG IKIEGFTEPN RTAYPIYE*

• ORF46ng-1 and ORF46-1 show 94.7% identity in 227 aa overlap:
• 
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF46ng-1 shows 87.4% identity over a 486aa overlap with an ORF (ORF46a) from strain A of N. meningitidis:
• 
• The complete length ORF46a DNA sequence <SEQ ID 465> is:

• 1	TTGGGCATTT CCCGCAAAAT ATCCCTTATT CTGTCCATAC
  	TGGCAGTGTG
  51	CCTGCCGATG CATGCACACG CCTCAGATTT GGCAAACGAT
  	TCTTTTATCC
  101	GGCAGGTTCT CGACCGTCAG CATTTCGAAC CCGACGGGAA
  	ATACCACCTA
  151	TTCGGCAGCA GGGGGGAACT TGCCGAGCGC AGCGGTCATA
  	TCGGATTGGG
  201	AAACATACAA AGCCATCAGT TGGGCAACCT GTTCATCCAG
  	CAGGCGGCCA
  251	TTAAAGGAAA TATCGGCTAC ATTGTCCGCT TTTCCGATCA
  	CGGGCACGAA
  301	GTCCATTCCC CCTTCGACAA CCATGCCTCA CATTCCGATT
  	CTGATGAAGC
  351	CGGTAGTCCC GTTGACGGAT TCAGCCTTTA CCGCATCCAT
  	TGGGACGGAT
  401	ACGAACACCA TCCCGCCGAC GGCTATGACG GGCCACAGGG
  	CGGCGGCTAT
  451	CCCGCTCCCA AAGGCGCGAG GGATATATAC AGCTACGACA
  	TAAAAGGCGT
  501	TGCCCAAAAT ATCCGCCTCA ACCTGACCGA CAACCGCAGC
  	ACCGGACAAC
  551	GGCTTGTCGA CCGTTTCCAC AATACCGGTA GTATGCTGAC
  	GCAAGGAGTA
  601	GGCGACGGAT TCAAACGCGC CACCCGATAC AGCCCCGAGC
  	TGGACAGATC
  651	GGGCAATGCC GCCGAAGCTT TCAACGGCAC TGCAGATATC
  	GTCAAAAACA
  701	TCATCGGCGC GGCAGGAGAA ATTGTCGGCG CAGGCGATGC
  	CGTGCAGGGT
  751	ATAAGCGAAG GCTCAAACAT TGCTGTTATG CACGGCTTGG
  	GTCTGCTTTC
  801	CACCGAAAAC AAGATGGCGC GCATCAACGA TTTGGCAGAT
  	ATGGCGCAAC
  851	TCAAAGACTA TGCCGCAGCA GCCATCCGCG ATTGGGCAGT
  	CCAAAACCCC
  901	AATGCCGCAC AAGGCATAGA AGCCGTCAGC AATATCTTTA
  	CGGCAGTCAT
  951	CCCCGTCAAA GGGATTGGAG CTGTTCGGGG AAAATACGGC
  	TTGGGCGGCA
  1001	TCACGGCACA TCCTGTCAAG CGGTCGCAGA TGGGCGAGAT
  	CGCATTGCCG
  1051	AAAGGGAAAT CCGCCGTCAG CGACAATTTT GCCGATGCGG
  	CATACGCCAA
  1101	ATACCCGTCC CCTTACCATT CCCGAAATAT CCGTTCAAAC
  	TTGGAGCAGC
  1151	GTTACGGCAA AGAAAACATC ACCTCCTCAA CCGTGCCGCC
  	GTCAAACGGA
  1201	AAGAATGTGA AACTGGCAAA CAAACGCCAC CCGAAGACCA
  	AAGTGCCGTT
  1251	TGACGGTAAA GGGTTTCCGA ATTTTGAAAA AGACGTAAAA
  	TACGATACGA
  1301	GAATTAATAC CGCTGTACCA CAAGTGAATC CTATAGATGA
  	ACCCGTCTTT
  1351	AATCCTAAAG GTTCTGTCGG ATCGGCTCAT TCTTGGTCTA
  	TAACTGCCAG
  1401	AATTCAATAC GCAAAATTAC CAAGGCAAGG TAGAATCAGA
  	TATATCCCAC
  1451	CTAAAAATTA CTCTCCTTCA GCACCGCTAC CAAAAGGACC
  	TAATAATGGA
  1501	TATTTGGATA AATTTGGTAA TGAATGGACT AAAGGTCCAT
  	CAAGAACTAA
  1551	AGGTCAAGAA TTTGAATGGG ATGTTCAATT GTCTAAAACA
  	GGAAGAGAGC
  1601	AACTTGGATG GGCTAGTAGG GATGGTAAGC ATTTAAATAT
  	ATCAATTGAT
  1651	GGAAAGATTA CACACAAATG A

• This corresponds to the amino acid sequence <SEQ ID 466>:

• 1	LGISRKISLI LSILAVCLPM HAHASDLAND SFIRQVLDRQ
  	HFEPDGKYHL
  51	FGSRGELAER SGHIGLGNIQ SHQLGNLFIQ QAAIKGNIGY
  	IVRFSDHGHE
  101	VHSPFDNHAS HSDSDEAGSP VDGFSLYRIH WDGYEHHPAD
  	GYDGPQGGGY
  151	PAPKGARDIY SYDIKGVAQN IRLNLTDNRS TGQRLVDRFH
  	NTGSMLTQGV
  201	GDGFKRATRY SPELDRSGNA AEAFNGTADI VKNIIGAAGE
  	IVGAGDAVQG
  251	ISEGSNIAVM HGLGLLSTEN KMARINDLAD MAQLKDYAAA
  	AIRDWAVQNP
  301	NAAQGIEAVS NIFTAVIPVK GIGAVRGKYG LGGITAHPVK
  	RSQMGEIALP
  351	KGKSAVSDNF ADAAYAKYPS PYHSRNIRSN LEQRYGKENI
  	TSSTVPPSNG
  401	KNVKLANKRH PKTKVPFDGK GFPNFEKDVK YDTRINTAVP
  	QVNPIDEPVF
  451	NPKGSVGSAH SWSITARIQY AKLPRQGRIR YIPPKNYSPS
  	APLPKGPNNG
  501	YLDKFGNEWT KGPSRTKGQE FEWDVQLSKT GREQLGWASR
  	DGKHLNISID
  551	GKITHK*

• Based on this analysis, including the presence of a RGD sequence in the gonococcal protein, typical of adhesins, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 56
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 467>:

• 1	ATGAATATTC ACACCCTGCT CTCCAAACAA TGGACGCTGC
  	CGCCATTCCT
  51	GCCGAAACGG CTGCTGCTGT CCCTGCTGAT ACTGCTTGCC
  	CCCAATGCGG
  101	TGTTTTGGGT TTTGGCACTG CTGACCGCCA CCGCCCGCCC
  	GATTGTCAAT
  151	TTGGACTATC TTCCCGCCGC GCTGCTGATC GCCCTGCCTT
  	GGCGTTTCGT
  201	CAAAATTGCC GGCGTATTGG CGTTTTGGCT GGCGGTTTTG
  	TTTGACGGGC
  251	TGATGATGGT GATCCAACTC TTCCCTTTTA TGGATCTCAT
  	CGGCGCCATC
  301	AACCTCGTCC CCTTCATCCT GACCGCCCCC GCCCCTTATC
  	AGATAATGAC
  351	CGGGCTG...

• This corresponds to the amino acid sequence <SEQ ID 468; ORF48>:

• 1	MNIHTLLSKQ WTLPPFLPKR LLLSLLILLA PNAVFWVLAL
  	LTATARPIVN
  51	LDYLPAALLI ALPWRFVKIA GVLAFWLAVL FDGLMMVIQL
  	FPFMDLIGAI
  101	NLVPFILTAP APYQIMTGL...

• Further work revealed the complete nucleotide sequence <SEQ ID 469>:

• 1	ATGAATATTC ACACCCTGCT CTCCAAACAA TGGACGCTGC
  	CGCCATTCCT
  51	GCCGAAACGG CTGCTGCTGT CCCTGCTGAT ACTGCTTGCC
  	CCCAATGCGG
  101	TGTTTTGGGT TTTGGCACTG CTGACCGCCA CCGCCCGCCC
  	GATTGTCAAT
  151	TTGGACTATC TTCCCGCCGC GCTGCTGATC GCCCTGCCTT
  	GGCGTTTCGT
  201	CAAAATTGCC GGCGTATTGG CGTTTTGGCT GGCGGTTTTG
  	TTTGACGGGC
  251	TGATGATGGT GATCCAACTC TTCCCTTTTA TGGATCTCAT
  	CGGCGCCATC
  301	AACCTCGTCC CCTTCATCCT GACCGCCCCC GCCCCTTATC
  	AGATAATGAC
  351	CGGGCTGTTG CTGCTGTATA TGCTGGCGAT GCCGTTTGTG
  	TTGCAGAAAG
  401	CCGCCGCCAA AACCGACTTC CGGCACATTG CCGTCTGCGC
  	CGCCGTTGTG
  451	GCGGCAGCCG GCTATTTCAC CGGCCATTTG AGTTACTACG
  	ACCGGGGTCG
  501	GATGGCCAAT ATCTTCGGCG CAAACAACTT CTACTACGCC
  	AAAAGTCAGG
  551	CGATGCTCTA CACCGTCAGC CAGAATGCCG ACTTTATTAC
  	CGCCGGCCTG
  601	GTCGATCCCG TCTTCCTCCC CTTGGGCAAT CAACAGCGTG
  	CCGCCACGCA
  651	TCTGAACGAG CCGAAATCTC AAAAAATCCT CTTTATCGTC
  	GCCGAATCTT
  701	GGGGGCTGCC GGCCAATCCC GAACTTCAAA ACGCCACTTT
  	TGCCAAACTG
  751	CTGGCGCAAA AAGACCGTTT TTCGGTTTGG GAAAGCGGCA
  	GTTTTCCCTT
  801	CATCGGCGCG ACGGTCGAAG GCGAAATGCG CGAACTGTGT
  	GCCTACGGCG
  851	GTTTGCGCGG GTTCGCACTG CGCCGCGCGC CCGACGAAAA
  	ATTTGCCCGC
  901	TGCCTCCCCA ACCGTTTGAA ACAAGAAGGT TACGCCACCT
  	TTGCGATGCA
  951	CGGCGCGGGC AGTTCGCTTT ACGACCGCTT CAGCTGGTAT
  	CCGAGGGCGG
  1001	GCTTTCAAGA AATCAAAACC GCCGAAAACC TGATCGGTAA
  	AAAAACCTGC
  1051	GCCATTTTCG GCGGCGTGTG CGACAGCGAG CTGTTCGGCG
  	AAGTGTCGGC
  1101	ATTTTTCAAA AAACACGACA AGGGACTGTT TTACTGGATG
  	ACGCTGACCA
  1151	GCCACGCCGA CTATCCCGAA TCCGACATTT TCAACCACAG
  	GCTCAAATGC
  1201	ACCGAATATG GCCTGCCCGC CGAAACCGAC CTCTGCCGCA
  	ATTTCAGCCT
  1251	GCACACCCAA TTCTTCGACC AACTGGCGGA TTTGATCCAA
  	CGCCCCGAAA
  1301	TGAAAGGCAC GGAAGTCATC ATCGTCGGCG ACCATCCGCC
  	GCCCGTCGGC
  1351	AACCTCAATG AAACCTTCCG CTACCTCAAA CAGGGGCACG
  	TCGCCTGGCT
  1401	GAACTTCAAA ATCAAATAA

• This corresponds to the amino acid sequence <SEQ ID 470; ORF48-1>:

• 1	MNIHTLLSKQ WTLPPFLPKR LLLSLLILLA PNAVFWVLAL
  	LTATARPIVN
  51	LDYLPAALLI ALPWRFVKIA GVLAFWLAVL FDGLMMVIQL
  	FPFMDLIGAI
  101	NLVPFILTAP APYQIMTGLL LLYMLAMPFV LQKAAAKTDF
  	RHIAVCAAVV
  151	AAAGYFTGHL SYYDRGRMAN IFGANNFYYA KSQAMLYTVS
  	QNADFITAGL
  201	VDPVFLPLGN QQRAATHLNE PKSQKILFIV AESWGLPANP
  	ELQNATFAKL
  251	LAQKDRFSVW ESGSFPFIGA TVEGEMRELC AYGGLRGFAL
  	RRAPDEKFAR
  301	CLPNRLKQEG YATFAMHGAG SSLYDRFSWY PRAGFQEIKT
  	AENLIGKKTC
  351	AIFGGVCDSE LFGEVSAFFK KHDKGLFYWM TLTSHADYPE
  	SDIFNHRLKC
  401	TEYGLPAETD LCRNFSLHTQ FFDQLADLIQ RPEMKGTEVI
  	IVGDHPPPVG
  451	NLNETFRYLK QGHVAWLNFK IK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF48 shows 94.1% identity over a 119aa overlap with an ORF (ORF48a) from strain A of N. meningitidis:
• 
• The complete length ORF48a nucleotide sequence <SEQ ID 471> is:

• 1	ATGAATATTC ACACCCTGCT CTCCAAACAA TGGACGCTGC
  	CGCCATTCCT
  51	GCCGAAACGG CTGCTGCTGT CCCTGCTGAT ACTGCTNNCC
  	CCCAATGCGG
  101	TGTTTTGGGT TTTGGCACTG CTGACCGCCA CCGCCCGCCC
  	GATTGTCAAT
  151	TTGGANTACC TTCCCGCCGC GCTGCTGATC GCCCTGCCTT
  	GGCGTNTCGT
  201	CAAAATTGNC GGCGTATTGG CGTNTTGGCT GGCGGTTTTG
  	TTTGACGGGC
  251	TGATGATGGT GATCCAACTC TTCCCTTTTA TGGATCTCAT
  	CGGCGCCATC
  301	AACCTCGTCC CCTTCATCNT GACCGCCCCC GCCCTTTATC
  	AGATAATGAC
  351	CGGGCTGTTA CTGCTGTATA TGCTGGCGAT GCCGTTTGTG
  	TTGCAGAAAG
  401	CCGCCGCCAA AACCGACTTC CGACACATTG CCGCCTGTGC
  	CGCCGTTGTG
  451	GTGGCAGCCG GCTATTTTAC CGGCCATTTG AGTTANTACG
  	ACCGGGGGCG
  501	GATGGCCAAT ATCTTCGGCG CAAACAACTT CTATTACGCC
  	AAAAGTCAGG
  551	CGATGCTCTA CACCGTCAGC CAGAATGCCG ACTTTATTAC
  	CGCCGGCCTG
  601	GTCGATCCCG TCTTCCTCCC CTTGGGCAAT CAACAGCGTG
  	CCGCCACGCA
  651	TCTGAACGAG CCGAAATCTC AAAAAATCCT CTTTATCGTC
  	GCCGAATCTT
  701	GGGGGCTGCC GGCCAATCCC GAACTTCAAA ACGCCACTTT
  	TGCCAAACTG
  751	CTGGCGCAAA AAGANCGTTT TTCGGTTTGG GAAAGCGGCA
  	GTTTTCCCTT
  801	CATCGGCGCG ACGATCGAAG GCGAAATGCG CGAACTGTGT
  	GCCTACGGCG
  851	GTTTGCGCGG GTTCGCACTG CGCCGCGCGC CCGACGAAAA
  	ATTTGCCCGC
  901	TGCCTCCCCA ACCGTTTGAA ACAAGAAGGT TACGCCACCT
  	TTGCGATGCA
  951	CGGCGCGGGC AGTTCGCTTT ACGACCGCTT CAGCTGGTAT
  	CCGAGGGCGG
  1001	GCTTTCAAGA AATCAAAACC GCCGAAAACC TGATCGGTAA
  	AAAAACCTGC
  1051	GCCATTTTCG GCGGCGTGTG CGACAGCGAG CTGTTCGGCG
  	AAGTGTCGGC
  1101	ANTTTTCAAA AAACACGACA AGGGACTGTT TTACTGGATG
  	ACGCTGACCA
  1151	GCCACGCCGA CTATCCCGAA TCNGACATTT TCAACCACAG
  	GCTCAAATGC
  1201	ACCGAATATG GCCTGCCCGC CGAAACCGAC NTCTGCCGCA
  	ATTTCAGCCT
  1251	GCACACCCAA TTCTTCGACC AACTGGCGGA TTTGATCCAA
  	CGCCCCGAAA
  1301	TGAAAGGCAC GGAAGTCATC ATCGTCGGCG ACCATCCGCC
  	GCCCGTCGGC
  1351	AACCTCAATG AAACCTTCCG CTACCTCAAA CAGGGGCACG
  	TCGNCTGGCT
  1401	GAACTTCAAA ATCAAATAA

• This encodes a protein having amino acid sequence <SEQ ID 472>:

• 1	MNIHTLLSKQ WTLPPFLPKR LLLSLLILLX PNAVFWVLAL
  	LTATARPIVN
  51	LXYLPAALLI ALPWRXVKIX GVLAXWLAVL FDGLMMVIQL
  	FPFMDLIGAI
  101	NLVPFIXTAP ALYQIMTGLL LLYMLAMPFV LQKAAAKTDF
  	RHIAACAAVV
  151	VAAGYFTGHL SXYDRGRMAN IFGANNFYYA KSQAMLYTVS
  	QNADFITAGL
  201	VDPVFLPLGN QQRAATHLNE PKSQKILFIV AESWGLPANP
  	ELQNATFAKL
  251	LAQKXRFSVW ESGSFPFIGA TIEGEMRELC AYGGLRGFAL
  	RRAPDEKFAR
  301	CLPNRLKQEG YATFAMHGAG SSLYDRFSWY PRAGFQEIKT
  	AENLIGKKTC
  351	AIFGGVCDSE LFGEVSAXFK KHDKGLFYWM TLTSHADYPE
  	SDIFNHRLKC
  401	TEYGLPAETD XCRNFSLHTQ FFDQLADLIQ RPEMKGTEVI
  	IVGDHPPPVG
  451	NLNETFRYLK QGHVXWLNFK IK*

• ORF48a and ORF48-1 show 96.8% identity in 472 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF48 shows 97.5% identity over a 119aa overlap with a predicted ORF (ORF48ng) from N. gonorrhoeae:
• 
• The ORF48ng nucleotide sequence <SEQ ID 473> was predicted to encode a protein having amino acid sequence <SEQ ID 474>:

• 1	MNIHALLSEQ WTLPPFLPKR LLLSLLILLA PNAVFWVLAL
  	LTATARPIVN
  51	LDYLPAALLI ALPWRFVKIA GVLAFWPAVL FDGLMMVIQL
  	FPFMDLIGAI
  101	NLVPFILTAP APYQIMTGLL LLYMLAMPFV LQKAAVKTDF
  	RHIAVCAAVV
  151	AAARYFTGPF ELLRTGGRWQ YVQHRRLLLS GSRASFRRRQ
  	KADVLRRLGN
  201	PYASMGNGG..

• Further work identified the complete gonococcal DNA sequence <SEQ ID 475>:

• 1	ATGAATATTC ACGCCCTGCT CTCCGAACAA TGGACGCTGC
  	CGCCATTCCT
  51	GCCGAAACGG CTGCTGCTGT CCCTGCTGAT ACTGCTGGCC
  	CCCAATGCGG
  101	TGTTTTGGGT TTTGGCACTG CTGACCGCCA CCGCCCGCCC
  	GATTGTCAAT
  151	TTGGACTACC TTCCCGCCGC GCTGCTGATC GCCCTGCCTT
  	GGCGTTTCGT
  201	CAAAATTGCC GGCGTATTGG CGTTTTGGCC GGCGGTTTTG
  	TTTGACGGGC
  251	TGATGATGGT GATCCAACTC TTCCCTTTTA TGGACCTCAT
  	CGGCGCCATC
  301	AACCTCGTCC CCTTCATCCT GACCGCCCCC GCCCCTTATC
  	AGATAATGAC
  351	CGGGCTGTTG CTGCTGTATA TGCTGGCGAT GCCGTTTGTG
  	TTGCAAAAAG
  401	CCGCCGTCAA AACCGACTTC CGACACATTG CCGTCTGTGC
  	CGCCGTTGTG
  451	GCGGCAGCCG GCTATTTCAC CGGCCATTTG AGTTACTACG
  	ACCGGGGGCG
  501	GATGGCCAAT ATCTTCGGCG CAAACAACTT CTATTACGCc
  	aAAAGTCAGG
  551	CGATGCTCTA CACCGTCAGC CAGAATGCCG ACTTTATTAC
  	CGCCGgcctG
  601	GTCGACCCCG TCTTCCTCCC CTTGGGCAAT CAGCAGCGTG
  	CCGCCACGCG
  651	GCTGAGTGAG CCGAAATCTC AAAAAATCCT CTTTATCGTC
  	GCCGAATCTT
  701	GGGGGCTGCC GGGCAATCCC GAGCTTCAAA ACGCCACTTT
  	TGCCAAACTG
  751	CTGGCGCAAA AAGACCGTTT TTCGGTTTGG GAAAGCGGCA
  	GTTTTCCCTT
  801	CATCGGCGCG ACGGTCGAAG GCGAAATGCG CGAATTGTGC
  	GCCTACGGCG
  851	GTTTGCGCGG GTTCGCACTG CGCCGCGCGC CCGACGAAAA
  	ATTTGCCCGC
  901	TGCCTCCCCA ACCGTTTGAA ACAAGAAGGT TACGCCACCT
  	TTGCGATGCA
  951	CGGCGCGGGT AGTTCGCTTT ACGACCGCTT CAGCTGGTAT
  	CCGAGGGCGG
  1001	GCTTTCAAAA AATCAAAACC GCCGAAAACC TGATCGGTAA
  	AAAAACCTGC
  1051	GCCATTTTCG GCGGCGTGTG CGACAGCGAG CTGTTCGGCG
  	AAGTGTCGGC
  1101	ATTTTTCAAA AAACACGACA AGGGACTGTT TTACTGGATG
  	ACGCTGACCA
  1151	GCCACGCCGA CTATCCCGAA TCCGACATTT TCAACCACAG
  	GCTCAAATGC
  1201	ACCGAATACG GCCTGCCCGC CGAAACCGAC CTCTGCCGCA
  	ATTTCAGCCT
  1251	GCACACCCAA TtcttcgACC AACTGGCGGA TTTGATCCGA
  	CGCCCCGAAA
  1301	TGAAAGGCAC GGAAGTCATC ATCGTCGGCG ACCATCCGCC
  	GCCCGTCGGC
  1351	AACCTCAATG AAACCTTCCG CTACCTCAAA CAGGGACACG
  	TCGCCTGGCT
  1401	GCACTTCAAA ATCAAATAA

• This encodes a protein having amino acid sequence <SEQ ID 476; ORF48ng-1>:

• 1	MNIHALLSEQ WTLPPFLPKR LLLSLLILLA PNAVFWVLAL
  	LTATARPIVN
  51	LDYLPAALLI ALPWRFVKIA GVLAFWPAVL FDGLMMVIQL
  	FPFMDLIGAI
  101	NLVPFILTAP APYQIMTGLL LLYMLAMPFV LQKAAVKTDF
  	RHIAVCAAVV
  151	AAAGYFTGHL SYYDRGRMAN IFGANNFYYA KSQAMLYTVS
  	QNADFITAGL
  201	VDPVFLPLGN QQRAATRLSE PKSQKILFIV AESWGLPGNP
  	ELQNATFAKL
  251	LAQKDRFSVW ESGSFPFIGA TVEGEMRELC AYGGLRGFAL
  	RRAPDEKFAR
  301	CLPNRLKQEG YATFAMHGAG SSLYDRFSWY PRAGFQKIKT
  	AENLIGKKTC
  351	AIFGGVCDSE LFGEVSAFFK KHDKGLFYWM TLTSHADYPE
  	SDIFNHRLKC
  401	TEYGLPAETD LCRNFSLHTQ FFDQLADLIR RPEMKGTEVI
  	IVGDHPPPVG
  451	NLNETFRYLK QGHVAWLHFK IK*

• ORG48ng-1 and ORF48-1 show 97.9% identity in 472 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence (double-underlined) and two putative transmembrane domains (single-underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 57
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 477>:

• 1	..GTGAGCGGAC GTTACCGCGC TTTGGATCGC GTTTCCAAAA
  	TCATCATCGT
  51	TACTTTGAGT ATCGCCACGC TTGCCGCCGC CGGCATCGCT
  	ATGTCGCGCG
  101	GTATGCAGAT GCAGTCCGAT TTTATCGAGC CGACACCGTG
  	GACGCTTGCC
  151	GGTTTGGGCT TCCTGATCGC GCTGATGGGC TGGATGCCCG
  	CGCCGATTGA
  201	AATTTCCGCC ATCAATTCTT TGTGGGTAAC CGAAAAACAA
  	CGCATCAATC
  251	CTTCCGAATA CCGCGACGGG ATTTTTGAAT TCAACGTCGG
  	TTATATCGCC
  301	AGTGCGGTTT TGGCTTTGGT TTTCCTTGCA CTGGGCGC.G
  	TAGCGCCGAA
  351	CGGCAACGGC GA.ACAGTGC AGATGGCGGG CGGCAAATAT
  	AACGGGCAAT
  401	TGATCAATAT GTACGCC..

• This corresponds to the amino acid sequence <SEQ ID 478; ORF53>:

• 1	..VSGRYRALDR VSKIIIVTLS IATLAAAGIA MSRGMQMQSD
  	FIEPTPWTLA
  51	GLGFLIALMG WMPAPIEISA INSLWVTEKQ RINPSEYRDG
  	IFEFNVGYIA
  101	SAVLALVFLA LGXVAPNGNG XTVQMAGGKY NGQLINMYA..

• Further work revealed the complete nucleotide sequence <SEQ ID 479>:

• 1	ATGTCCGAAC AACATATTTC GACTTGGAAA AGTAAAATCA
  	ACGCATTGGG
  51	TCCGGGGATC ATGATGGCTT CGGCGGCGGT CGGCGGTTCG
  	CACCTGATTG
  101	CCTCGACGCA GGCGGGCGCG CTTTACGGCT GGCAGATCGC
  	GCTCATCATC
  151	ATCCTGACCA ACCTCTTCAA ATACCCGTTT TTCCGCTTCA
  	GCGCGCATTA
  201	CACGCTGGAC ACGGGCAAGA GCCTGATTGA AGGTTATGCC
  	GAGAAAAGCC
  251	GCGTTTATTT GTGGGTATTC CTGATTTTGT GCATCCTCTC
  	CGCCACGATT
  301	AACGCGGGCG CGGTCGCCAT TGTAACCGCC GCCATCGTCA
  	AAATGGCGAT
  351	TCCCTCGCTG ATGTTTGATG CCGGCACGGT TGCCGCCTTG
  	ATTATGGCAT
  401	CCTGCCTGAT TATTTTGGTG AGCGGACGTT ACCGCGCTTT
  	GGATCGCGTT
  451	TCCAAAATCA TCATCGTTAC TTTGAGTATC GCCACGCTTG
  	CCGCCGCCGG
  501	CATCGCTATG TCGCGCGGTA TGCAGATGCA GTCCGATTTT
  	ATCGAGCCGA
  551	CACCGTGGAC GCTTGCCGGT TTGGGCTTCC TGATCGCGCT
  	GATGGGCTGG
  601	ATGCCCGCGC CGATTGAAAT TTCCGCCATC AATTCTTTGT
  	GGGTAACCGA
  651	AAAACAACGC ATCAATCCTT CCGAATACCG CGACGGGATT
  	TTTGATTTCA
  701	ACGTCGGTTA TATCGCCAGT GCGGTTTTGG CTTTGGTTTT
  	CCTTGCACTG
  751	GGCGCGTTTG TGCAATACGG CAACGGCGAA GCAGTGCAGA
  	TGGCGGGCGG
  801	CAAATATATC GGGCAATTGA TCAATATGTA CGCCGTTACC
  	ATCGGCGGCT
  851	GGTCGCGCCC GCTGGTGGCG TTTATCGCGT TTGCCTGTAT
  	GTACGGCACG
  901	ACGATTACCG TCGTGGACGG CTATGCCCGT GCCATTGCCG
  	AACCCGTGCG
  951	CCTGCTGCGC GGAAAAGACA AAACGGGCAA CGCCGAATTC
  	TTTGCCTGGA
  1001	ATATTTGGGT GGCGGGCAGC GGTTTGGCGG TGATTTTCTG
  	GTTTGACGGC
  1051	GTAATGGCGA ATCTGCTCAA ATTTGCGATG ATTGCCGCTT
  	TTGTGTCCGC
  1101	CCCTGTGTTT GCCTGGCTGA ATTACCGTTT GGTTAAAGGT
  	GATGAAAAAC
  1151	ACAAACTCAC ATCAGGTATG AATGCCCTTG CATTGGCAGG
  	CTTGATTTAT
  1201	CTGACCGGTT TTACCGTTTT GTTCTTATTG AATTTGGCGG
  	GAATGTTCAA
  1251	ATGA

• This corresponds to the amino acid sequence <SEQ ID 480; ORF53-1>:

• 1	MSEQHISTWK SKINALGPGI MMASAAVGGS HLIASTQAGA
  	LYGWQIALII
  51	ILTNLFKYPF FRFSAHYTLD TGKSLIEGYA EKSRVYLWVF
  	LILCILSATI
  101	NAGAVAIVTA AIVKMAIPSL MFDAGTVAAL IMASCLIILV
  	SGRYRALDRV
  151	SKIIIVTLSI ATLAAAGIAM SRGMQMQSDF IEPTPWTLAG
  	LGFLIALMGW
  201	MPAPIEISAI NSLWVTEKQR INPSEYRDGI FDFNVGYIAS
  	AVLALVFLAL
  251	GAFVQYGNGE AVQMAGGKYI GQLINMYAVT IGGWSRPLVA
  	FIAFACMYGT
  301	TITVVDGYAR AIAEPVRLLR GKDKTGNAEF FAWNIWVAGS
  	GLAVIFWFDG
  351	VMANLLKFAM IAAFVSAPVF AWLNYRLVKG DEKHKLTSGM
  	NALALAGLIY
  401	LTGFTVLFLL NLAGMFK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF53 shows 93.5% identity over a 139aa overlap with an ORF (ORF53a) from strain A of N. meningitidis:
• 
• The complete length ORF53a nucleotide sequence <SEQ ID 481> is:

• 1	ATGTCCGAAC AACATATTTC GACTTGGAAA AGTAAAATCA
  	ACGCATTGGG
  51	ACCGGGGATT ATGATGGCTT CGGCGGCGGT CGGCGGTTCG
  	CACCTGATTG
  101	CCTCGACGCA GGCGGGCGCG CTTTACGGCT GGCAGATCGC
  	GCTCATCATC
  151	ATCCTGACCA ACCTCTTCAA ATACCCGTTT TTCCGCTTCA
  	GCGCGCATTA
  201	CACGCTGGAC ACGGGCAAGA GCCTGATTGA AGGTTATGCC
  	GAGAAAAGCC
  251	GCGTTTATTT GTGGGTATTC CTGATTTTGT GCATCCTCTC
  	CGCCACGATT
  301	AACGCGGGCG CGGTCGCCAT TGTAACCGCC GCCATCGTCA
  	AAATGGCGAT
  351	TCCCTCGCTG ATGTTTGATG CCGGCACGGT TGCCGCCTTG
  	ATTATGGCAT
  401	CCTGCCTGAT TATTTTGGTG AGCGGACGTT ACCGCGCTTT
  	GGATCGCGTT
  451	TCCAAAATCA TCATCGTTAC TTTGAGTATC GCCACGCTTG
  	CCGCCGCCGG
  501	CATCGCTATG TCGCGCGGTA TGCAGATGCA GTCCGATTTT
  	ATCGAGCCGA
  551	CACCGTGGAC GCTTGCCGGT TTGGGCTTCC TGATCGCGCT
  	GATGGGCTGG
  601	ATGCCCGCGC CGATTGAAAT TTCCGCCATC AATTCTTTGT
  	GGGTAACCGA
  651	AAAACAACGC ATCAATCCTT CCGAATACCG CGACGGGATT
  	TTTGATTTCA
  701	ACGTCGGTTA TATCGCCAGT GCGGTTTTGG CTTTGGTTTT
  	CCTTGCACTG
  751	GGCGCGTTTG TGCAATACGG CAACGGCGAA GCAGTGCAGA
  	TGGCGGGCGG
  801	CAAATATATC GGGCAATTGA TCAATATGTA CGCCGTTACC
  	ATCGGCGGCT
  851	GGTCGCGCCC GCTGGTGGCG TTTATCGCGT TTGCCTGTAT
  	GTACGGCACG
  901	ACGATTACCG TTGTGGACGG CTATGCCCGT GCCATTGCCG
  	AACCCGTGCG
  951	CCTGCTGCGC GGAAAAGACA AAACGGGCAA CGCCGAATTC
  	TTTGCCTGGA
  1001	ATATTTGGGT GGCGGGCAGC GGTTTGGCGG TGATTTTCTG
  	GTTTGACGGC
  1051	GTAATGGCGA ATCTGCTCAA ATTTGCGATG ATTGCCGCTT
  	TTGTGTCCGC
  1101	CCCTGTGTTT GCCTGGCTGA ATTACCGTTT GGTCAAAGGT
  	GATGAAAAAC
  1151	ACAAACTCAC ATCAGGTATG AATGCCCTTG CATTGGCAGG
  	CTTGATTTAT
  1201	CTGACCGGTT TTACCGTTTT GTTCTTATTG AATTTGGCGG
  	GAATGTTCAA
  1251	ATGA

• This encodes a protein having amino acid sequence <SEQ ID 482>:

• 1	MSEQHISTWK SKINALGPGI MMASAAVGGS HLIASTQAGA
  	LYGWQIALII
  51	ILTNLFKYPF FRFSAHYTLD TGKSLIEGYA EKSRVYLWVF
  	LILCILSATI
  101	NAGAVAIVTA AIVKMAIPSL MFDAGTVAAL IMASCLIILV
  	SGRYRALDRV
  151	SKIIIVTLSI ATLAAAGIAM SRGMQMQSDF IEPTPWTLAG
  	LGFLIALMGW
  201	MPAPIEISAI NSLWVTEKQR INPSEYRDGI FDFNVGYIAS
  	AVLALVFLAL
  251	GAFVQYGNGE AVQMAGGKYI GQLINMYAVT IGGWSRPLVA
  	FIAFACMYGT
  301	TITVVDGYAR AIAEPVRLLR GKDKTGNAEF FAWNIWVAGS
  	GLAVIFWFDG
  351	VMANLLKFAM IAAFVSAPVF AWLNYRLVKG DEKHKLTSGM
  	NALALAGLIY
  401	LTGFTVLFLL NLAGMFK*

• ORF 53a shows 100.0% identity in 417 aa overlap with ORF53-1:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF53 shows 92.1% identity over a 139aa overlap with a predicted ORF (ORF53ng) from N. gonorrhoeae:
• 
• An ORF53ng nucleotide sequence <SEQ ID 483> was predicted to encode a protein having amino acid sequence <SEQ ID 484>:

• 1	MPKKSCVYLW VFLILCIASA TINAGAVAIV TAAIVKMAIP
  	SLMFDAGTVA
  51	ALIMASCLII LVSGRYRALD RVSKIIIVTL SIATLAAAGI
  	AMSRGMQMQP
  101	DFIEPTPWTL AGLGFLIALM GWMPAPIEIS AINSLWVTEK
  	QRINPSEYRD
  151	GIFDFNVGYI ASAVLALVFL ALGAFVQYGN GEAVQMGGGK
  	YIGQLINMYA
  201	VTIGGGSRPL VAFIAFACMY GAASTVV DGY ARAIAEPVRL
  	LRGKDKTARP
  251	IVLLEKLGGR HRFGRDFLV*

• Further analysis revealed further partial DNA gonococcal sequence <SEQ ID 485>:

• 1	..aagaAAAGCT GCGTTTATTT GTGGGTTTTT TTGATTTTGT
  	GTATCGCCTC
  51	CGCCACGATT AACGCGGGCG CGGTCGCCAT TGTAACCGCC
  	GCCATCGTCA
  101	AAATGGCGAT TCCCTCGCTG ATGTTTGATG CCGGCACGGT
  	TGCCGCCTTG
  151	ATTATGGCAT CCTGCCTGAT TATTTTGGTG AGCGGACGTT
  	ACCGCGCTTT
  201	GGATCGTGTT TCCAAAATCA TCATTGTTAC TTTGAGCATC
  	GCCACGCTTG
  251	CCGCCGCCGG CATCGCTATG TCGCGCGGTA TGCAGATGCA
  	GCCCGATTTT
  301	ATCGAGCCGA CACCGTGGAC GCTTGCCGGT TTGGGCTTCC
  	TGATCGCGCT
  351	GATGGGCTGG ATGCCCGCGC CGATCGAAAT TTCCGCCATC
  	AATTCTTTGT
  401	GGGTAACCGA AAAACAACGC ATCAATCCTT CTGAATACCG
  	CGACGGGATT
  451	TTCGATTTCA ACGTCGGTTA TATCGCcagT GCGGTTTTGG
  	CTTTGGTTTT
  501	CCTTGCACTG GGCGCGTTTG TGCAATACGG CAACGGCGAA
  	GCAGTGCAGA
  551	TGGCGGGCGG CAAATATATC GGGCAATTGA TTAATATGTA
  	TGCCGTAACC
  601	ATCGGCGGCT GGTCTCGTCC GCTGGTGGCG TTTATCGCGT
  	TTGCCTGTAT
  651	GTACGGCACG ACGATTACCG TTGTGGACGG TTATGCGCGT
  	GCCATTGCCG
  701	AACCCGTGCG CCTGCTGCGC GGCAGGGATA AAACCGGCAA
  	CGCCGAGTTG
  751	TTtgccTGGA ATATTTGGGT GGCGGGCAGC GGTTTGGCGG
  	TGATTTTCTG
  801	GTTTGACggc gcaaTGGCgG AACtgcTCAA ATTTGCGATG
  	ATtgccgcCT
  851	TTGTGTCCGC CCCTGTGTTC GCCTGGCTCA ACTACCGCCT
  	CGTCAAAGGG
  901	GACAAACGCC ACAGGCTTAC CGCCGGTATG AACGCCCTTG
  	CCATTGTCGG
  951	CCTGCTCTAC CTGGCCGGGT TTGCCGTTTT GTTCCTGTTG
  	AACCTTACCG
  1001	GACTTTTGGC ATAG

• This corresponds to the amino acid sequence <SEQ ID 486; ORF53ng-1>:

• 1	..KKSCVYLWVF LILCIASATI NAGAVAIVTA AIVKMAIPSL
  	MFDAGTVAAL
  51	IMASCLIILV SGRYRALDRV SKIIIVTLSI ATLAAAGIAM
  	SRGMQMQPDF
  101	IEPTPWTLAG LGFLIALMGW MPAPIEISAI NSLWVTEKQR
  	INPSEYRDGI
  151	FDFNVGYIAS AVLALVFLAL GAFVQYGNGE AVQMAGGKYI
  	GQLINMYAVT
  201	IGGWSRPLVA FIAFACMYGT TITVV DGYAR AIAEPVRLLR
  	GRDKTGNAEL
  251	FAWNIWVAGS GLAVIFWFDG AMAELLKFAM IAAFVSAPVF
  	AWLNYRLVKG
  301	DKRHRLTAGM NALAIVGLLY LAGFAVLFLL NLTGLLA*

• ORF53ng-1 and ORF53-1 show 94.0% identity in 336 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence (double-underlined) and several putative transmembrane domains (single-underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 58
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 487>:

• 1	..TTGCGGGAAA CGGCATATGT TTTGGATAGT TTTGATCGTT
  	ATTTTGTTGT
  51	TGCGCTTGCC GGCTTGTTTT TTGTCCGCGC ACAATCCGAA
  	CGCGAGTGGA
  101	TGCGCGAGGT TTCTGCGTGG CAGGAAAAGA AAGGGGAAAA
  	ACAGGCGGAG
  151	CTGCCTGAAA TCAAAGACGG TATGCCCGAT TTTCCCGAAC
  	TTGCCCTGAT
  201	GCTTTTCCAC GCCGTCAAAA CGGCAGTGTA TTGGCTGTTT
  	GTCGGTGTCG
  251	TCCGTTTCTG CCGAAACTAT CTGGCGCACG AATCCGAACC
  	GGACAGGCCC
  301	GTTCCGCCT..

• This corresponds to the amino acid sequence <SEQ ID 488; ORF58>:

• 1	..LRETAYVLDS FDRYFVVALA GLFFVRAQSE REWMREVSAW
  	QEKKGEKQAE
  51	LPEIKDGMPD FPELALMLFH AVKTAVYWLF VGVV RFCRNY
  	LAHESEPDRP
  101	VPP..

• Further work revealed the complete nucleotide sequence <SEQ ID 489>:

• 1	ATGTTTTGGA TAGTTTTGAT CGTTATTTTG TTGCTTGCGC
  	TTGCCGGCTT
  51	GTTTTTTGTC CGCGCACAAT CCGAACGCGA GTGGATGCGC
  	GAGGTTTCTG
  101	CGTGGCAGGA AAAGAAAGGG GAAAAACAGG CGGAGCTGCC
  	TGAAATCAAA
  151	GACGGTATGC CCGATTTTCC CGAACTTGCC CTGATGCTTT
  	TCCATGCCGT
  201	CAAAACGGCA GTGTATTGGC TGTTTGTCGG TGTCGTCCGT
  	TTCTGCCGAA
  251	ACTATCTGGC GCACGAATCC GAACCGGACA GGCCCGTTCC
  	GCCTGCTTCT
  301	GCAAACCGTG CGGATGTTCC GACCGCATCC GACGGATATT
  	CAGACAGTGG
  351	AAACGGGACG GAAGAAGCGG AAACGGAAGA AGCAGAAGCT
  	GCGGAGGAAG
  401	AGGCTGCCGA TACGGAAGAC ATTGCAACTG CCGTAATCGA
  	CAACCGCCGC
  451	ATCCCATTCG ACCGGAGTAT TGCTGAAGGG TTGATGCCGT
  	CTGAAAGCGA
  501	AATTTCGCCC GTCCGTCCGG TTTTTAAAGA AATCACTTTG
  	GAAGAAGCAA
  551	CGCGTGCTTT AAACAGCGCG GCTTTAAGGG AAACGAAAAA
  	ACGCTATATC
  601	GATGCATTTG AGAAAAACGA AACAGCGGTC CCCAAAGTCC
  	GCGTGTCCGA
  651	TACCCCGATG GAAGGGCTGC AGATTATCGG TTTGGACGAC
  	CCTGTGCTTC
  701	AACGCACGTA TTCCCATATG TTCGATGCGG ACAAAGAAGC
  	GTTTTCCGAG
  751	TCTGCGGATT ACGGATTTGA GCCGTATTTT GAGAAGCAGC
  	ATCCGTCTGC
  801	CTTTTCTGCA GTCAAAGCCG AAAATGCACG GAATGCGCCG
  	TTCCACCGTC
  851	ATGCAGGGCA GGGGAAAGGG CAGGCGGAGG CAAAATCCCC
  	GGATGTTTCC
  901	CAAGGGCAGT CCGTTTCAGA CGGCACGGCC GTCCGCGATG
  	CCCGCCGCCG
  951	CGTTTCCGTC AATTTGAAAG AACCGAACAA GGCAACGGTT
  	TCTGCGGAGG
  1001	CGCGAATTTC TCGCCTGATT CCGGAAAGTC AGACGGTTGT
  	CGGGAAACGG
  1051	GATGTCGAAA TGCCGTCTGA AACCGAAAAT GTTTTCACGG
  	AAACCGTTTC
  1101	GTCTGTGGGA TACGGCGGTC CGGTTTATGA TGAAACTGCC
  	GATATCCATA
  1151	TTGAAGAACC TGCCGCGCCC GATGCTTGGG TGGTCGAACC
  	ACCCGAAGTG
  1201	CCGAAAGTTC CCATGACCGC AATCGATATT CAGCCGCCGC
  	CTCCCGTATC
  1251	GGAAATCTAC AACCGTACCT ATGAACCGCC GTCAGGATTC
  	GAGCAGGTGC
  1301	AACGCAGCCG CATTGCCGAG ACCGACCATC TTGCCGATGA
  	TGTTTTGAAT
  1351	GGAGGTTGGC AGGAGGAAAC CGCCGCTATT GCGGATGACG
  	GCAGTGAAGG
  1401	TGCGGCAGAG CGGTCAAGCG GGCAATATCT GTCGGAAACC
  	GAAGCGTTCG
  1451	GGCATGACAG TCAGGCGGTT TGTCCGTTTG AAAATGTGCC
  	GTCTGAACGC
  1501	CCGTCCTGCC GGGTATCGGA TACGGAAGCG GATGAAGGGG
  	CGTTCCCATC
  1551	TGAAGAAACC GGTGCGGTAT CCGAACACCT GCCGACAACC
  	GACCTGCTTC
  1601	TGCCTCCGCT GTTCAATCCC GAGGCGACGC AAACCGAAGA
  	AGAACTGTTG
  1651	GAAAACAGCA TCACCATCGA AGAAAAATTG GCGGAGTTCA
  	AAGTCAAGGT
  1701	CAAGGTTGTC GATTCTTATT CCGGCCCCGT AATTACGCGT
  	TATGAAATCG
  1751	AACCCGATGT CGGCGTGCGC GGCAATTCCG TTCTGAATCT
  	GGAAAAAGAT
  1801	TTGGCGCGTT CGCTCGGCGT GGCTTCCATC CGCGTTGTCG
  	AAACCATCCC
  1851	CGGCAAAACC TGCATGGGTT TGGAACTTCC GAACCCGAAA
  	CGCCAAATGA
  1901	TACGCCTGAG CGAAATCTTC AATTCGCCCG AGTTTGCCGA
  	ATCCAAATCC
  1951	AAGCTGACGC TCGCGCTCGG TCAGGACATC ACCGGACAGC
  	CCGTCGTAAC
  2001	CGACTTGGGA AAAGCACCGC ATTTGTTGGT TGCCGGCACG
  	ACCGGTTCGG
  2051	GCAAATCGGT GGGTGTCAAC GCGATGATTC TGTCTATGCT
  	TTTCAAAGCC
  2101	GCGCCGGAAG ACGTGCGTAT GATTATGATC GATCCGAAAA
  	TGCTGGAATT
  2151	GAGCATTTAC GAAGGCATCC CGCACCTGCT CGCCCCTGTC
  	GTTACCGATA
  2201	TGAAGCTGGC GGCAAACGCG CTGAACTGGT GTGTTAACGA
  	AATGGAAAAA
  2251	CGCTACCGCC TGATGAGCTT TATGGGCGTG CGTAATCTTG
  	CGGGCTTCAA
  2301	TCAAAAAATC GCCGAAGCCG CAGCAAGGGG AGAAAAAATC
  	GGCAATCCGT
  2351	TCAGCCTCAC GCCCGACGAT CCCGAACCTT TGGAAAAACT
  	GCCGTTTATC
  2401	GTGGTCGTGG TCGATGAGTT TGCCGACCTG ATGATGACGG
  	CAGGCAAGAA
  2451	AATCGAAGAA CTGATTGCCC GCCTCGCCCA AAAAGCCCGC
  	GCGGCAGGCA
  2501	TCCATTTGAT TCTTGCCACA CAACGCCCCA GCGTCGATGT
  	CATCACGGGT
  2551	CTGATTAAGG CGAACATCCC GACGCGTATC GCGTTCCAAG
  	TGTCCAGCAA
  2601	AATCGACAGC CGCACGATTC TCGACCAAAT GGGCGCGGAA
  	AACCTGCTCG
  2651	GTCAGGGCGA TATGCTGTTC CTGCTGCCGG GTACTGCCTA
  	TCCGCAGCGC
  2701	GTTCACGGCG CGTTTGCCTC GGATGAAGAG GTGCACCGCG
  	TGGTCGAATA
  2751	TTTGAAACAG TTTGGCGAAC CGGACTATGT TGACGATATT
  	TTGAGCGGCG
  2801	GCGGCAGCGA AGAGCTGCCC GGCATCGGGC GCAGCGGCGA
  	CGACGAAACC
  2851	GATCCGATGT ACGACGAGGC CGTATCCGTT GTCCTGAAAA
  	CGCGCAAAGC
  2901	CAGCATTTCG GGCGTACAGC GCGCCTTGCG TATCGGCTAC
  	AACCGCGCCG
  2951	CGCGTCTGAT TGACCAGATG GAGGCGGAAG GCATTGTGTC
  	CGCACCGGAA
  3001	CACAACGGCA ACCGTACGAT TCTCGTCCCC TTGGACAATG
  	CTTGA

• This corresponds to the amino acid sequence <SEQ ID 490; ORF58-1>:

• 1	MFWIVLIVIL LLALAGL FFV RAQSEREWMR EVSAWQEKKG
  	EKQAELPEIK
  51	DGMPDFPELA LMLFHAVKTA VYWLFVGVV R FCRNYLAHES
  	EPDRPVPPAS
  101	ANRADVPTAS DGYSDSGNGT EEAETEEAEA AEEEAADTED
  	IATAVIDNRR
  151	IPFDRSIAEG LMPSESEISP VRPVFKEITL EEATRALNSA
  	ALRETKKRYI
  201	DAFEKNETAV PKVRVSDTPM EGLQIIGLDD PVLQRTYSHM
  	FDADKEAFSE
  251	SADYGFEPYF EKQHPSAFSA VKAENARNAP FHRHAGQGKG
  	QAEAKSPDVS
  301	QGQSVSDGTA VRDARRRVSV NLKEPNKATV SAEARISRLI
  	PESQTVVGKR
  351	DVEMPSETEN VFTETVSSVG YGGPVYDETA DIHIEEPAAP
  	DAWVVEPPEV
  401	PKVPMTAIDI QPPPPVSEIY NRTYEPPSGF EQVQRSRIAE
  	TDHLADDVLN
  451	GGWQEETAAI ADDGSEGAAE RSSGQYLSET EAFGHDSQAV
  	CPFENVPSER
  501	PSCRVSDTEA DEGAFPSEET GAVSEHLPTT DLLLPPLFNP
  	EATQTEEELL
  551	ENSITIEEKL AEFKVKVKVV DSYSGPVITR YEIEPDVGVR
  	GNSVLNLEKD
  601	LARSLGVASI RVVETIPGKT CMGLELPNPK RQMIRLSEIF
  	NSPEFAESKS
  651	KLTLALGQDI TGQPVVTDLG KAPHLLVAGT TGSGKSVGVN
  	AMILSMLFKA
  701	APEDVRMIMI DPKMLELSIY EGIPHLLAPV VTDMKLAANA
  	LNWCVNEMEK
  751	RYRLMSFMGV RNLAGFNQKI AEAAARGEKI GNPFSLTPDD
  	PEPLEKLPFI
  801	VVVVDEFADL MMTAGKKIEE LIARLAQKAR AAGIHLILAT
  	QRPSVDVITG
  851	LIKANIPTRI AFQVSSKIDS RTILDQMGAE NLLGQGDMLF
  	LLPGTAYPQR
  901	VHGAFASDEE VHRVVEYLKQ FGEPDYVDDI LSGGGSEELP
  	GIGRSGDDET
  951	DPMYDEAVSV VLKTRKASIS GVQRALRIGY NRAARLIDQM
  	EAEGIVSAPE
  1001	HNGNRTILVP LDNA*

• Computer analysis of this amino acid sequence predicts the indicated transmembrane region, and also gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF58 shows 96.6% identity over a 89aa overlap with an ORF (ORF58a) from strain A of N. meningitidis:
• 
• The complete length ORF58a nucleotide sequence <SEQ ID 491> is:

• 1	ATGTTTTGGA TAGTTTTGAT CGTTATTTTG TTGCTTGCGC
  	TTGCCGGCTT
  51	GTTTTTTGTC CGCGCACAAT CCGAACGCGA GTGGATGCGC
  	GAGGTTTCTG
  101	CGTGGCAGGA AAAGAAAGGG GAAAAACAGG CGGAGCTGCC
  	TGAAATCAAA
  151	GACGGTATGC CCGATTTTCC CGAACTTGCC CTGATGCTTT
  	TCCATGCCGT
  201	CAAAACGGCA GTGTATTGGC TGTTTGTCGG TGTCGTCCGT
  	TTCTGCCGAA
  251	ACTATCTGGC GCACGAATCC GAACCGGACA GGCCCGTTCC
  	GCCTGCTTCT
  301	GCAAATCGTG CGGATGTTCC GACCGCATCC GACGGATATT
  	CAGACAGTGG
  351	AAACGGGACG GAAGAAGCGG AAACGGAAGA AGCAGAAGCT
  	GCGGAGGAAG
  401	AGGCTGCCGA TACGGAAGAC ATTGCAACTG CCGTAATCGA
  	CAACCGCCGC
  451	ATCCCATTCG ACCGGAGTAT TGCTGAAGGG TTGATGCCGT
  	CTGAAAGCGA
  501	AATTTCGCCC GTCCGTCCGG TTTTTAAGGA AATCACTTTG
  	GAAGAAGCAA
  551	CGCGTGCTTT AAACAGCGCG GCTTTAAGGG AAACGAAAAA
  	ACGCTATATC
  601	GATGCATTTG AGAAAAACGA AACAGCGGTC CCCAAAGTCC
  	GCGTGTCCGA
  651	TACCCCGATG GAAGGGCTGC AGATTATCGG TTTGGACGAC
  	CCTGTGCTTC
  701	AACGCACGTA TTCCCGTATG TTCGATGCGG ACAAAGAAGC
  	GTTTTCCGAG
  751	TCTGCGGATT ACGGATTTGA GCCGTATTTT GAGAAGCAGC
  	ATCCGTCTGC
  801	CTTTTCTGCA GTCAAAGCCG AAAATGCACG GAATGCGCCG
  	TTCCGCCGTC
  851	ATGCAGGGCA GGGNAAAGGG CAGGCGGAGG CNAAATCCCC
  	GGATGTTTCC
  901	CAAGGGCAGT CCGTTTCAGA CGGCACAGCC GTCCGCGATG
  	CCNGCCGCCG
  951	CGTTTCCGTC AATTTGAAAG AACCGAACAA GGCAACGGTT
  	TCTGCGGAGG
  1001	CGCGGATTTC GCGCCTGATT CCGGAAAGTC GGACGGTTGT
  	CGGGAAACGG
  1051	GATGTCGAAA TGCCGTCTGA AACCGAAAAT GTTTTCACGG
  	AAANTGTTTC
  1101	GTCTGTGGGA TACGGCGNTC CGGTTTATGA TGAAACTGCC
  	GATATCCATA
  1151	TTGAAGAACC TGCCGCGCCC GATGCTTGGG TGGTCGAACC
  	ACCCGAAGTG
  1201	CCGAAAGTTC CCATGCCCGC AATNGATATT CCGCCGCCGC
  	CTCCCGTATC
  1251	GGAAATCTAC AACCGTACCT ATGAACCGCC GGCAGGATTC
  	GAGCAGGTGC
  1301	AACGCAGCCG CATTGCCGAA ACCGATCATC TTGCCGATGA
  	TGTTTTGAAT
  1351	GGAGGTTGGC AGGAGGAAAC CGCCGCTATT GCGAATGACG
  	GCAGTGAGGG
  1401	TGTGGCAGAG CGGTCAAGCG GGCAATATTT GTCGGAAACC
  	GAAGCGTTCG
  1451	GGCATGACAG TCAGGCGGTT TGTCCGTTTG AAAATGTGCC
  	GTCTGAACGC
  1501	CCGTCCCGCC GGGCATNGGA TACGGAAGCG GATGAAGGGG
  	CGTTCCAATC
  1551	TGAAGAAACC GGTGCGGTAT CCGAACACCT GCCGACAACC
  	GACCTGCTTC
  1601	TGCCGCCGCT GTTCAATCCC GGGGCGACGC AAACCGAAGA
  	AGANCTGTTG
  1651	GANAACAGCA TCACCATCGA AGAAAAATNG GCGGAGTTCA
  	AAGTCAAGGT
  1701	CAAGGTTGTC GATTCTTATT CCGGCCCCGT GATTACGCGT
  	TATGAAATCG
  1751	AACCCGATGT CGGCGTGCGC GGCAATTCCG TTCTAAATCT
  	GGAAAAAGAN
  1801	TTGGCGCGTT CGCTCGGCGT GGCTTCCATC CGCGTTGTCG
  	AAACCATCCT
  1851	CGGCAAAACC TGTATGGGTT TGGAACTTCC GAACCCGAAA
  	CGCCAAATGA
  1901	TACGCCTGAG CGAAATCTTC AATTCGCCCG AGTTTGCCGA
  	ATCCAAATCC
  1951	AAGCTGACGC TCGCGCTCGG TCAGGACATC ACCGGACAGC
  	CCGTCGTAAC
  2001	CGACTTGGGC AAAGCACCGC ATTTGTTGGT TGCCGGCACG
  	ACCGGTTCGG
  2051	GCAAATCGGT GGGTGTCAAC GCGATGATTC TGTCTATGCT
  	TTTCAAAGCC
  2101	GCGCCGGAAG ACGTGCGTAT GATTATGATC GATCCGAAAA
  	TGCTGGAATT
  2151	GAGCATTTAC GAAGGCATCC CGCACCTGCT CGCCCCTGTC
  	GTTACCGATA
  2201	TGAAGCTGGC GGCAAACGCG CTGAACTGGT GTGTTAACGA
  	AATGGAAAAA
  2251	CGCTACCGCC TGATGAGCTT TATGGGCGTG CGCAATCTTG
  	CGGGTNTCAA
  2301	TCAAAAAATC GCCGAAGCCG CAGCAAGGGG GGAGAAAATC
  	GGCAACCCGT
  2351	TCAGCCTCAC GCCCGACAAT CCCGAACCTT TGGANAAATT
  	GCCGTTTATC
  2401	GTGGTCGTGG TTGATGAGTT TGCCGACCTG ATGATGACGG
  	CAGGCAAGAA
  2451	AATCGAAGAA CTGATTGCCC GCCTCGCCCA AAAAGCCCGC
  	GCGGCAGGCA
  2501	TCCATCTTAT CCTTGCCACA CAACGCCCCA GTGTCGATGT
  	CATCACGGGT
  2551	CTGATTAAGG CGAACATCCC GACGCGTATC GCGTTCCAAG
  	TGTCCAGCAA
  2601	AATCGACAGC CGCACGATTC TTGACCAAAT GGGTGCGGAA
  	AACCTGCTCG
  2651	GGCAGGGCGA TATGCTGTTC CTGCCGCCGG GTACGGCCTA
  	TCCGCAGCGC
  2701	GTTCACGGCG CGTTTGCCTC GGATGAAGAG GTGCACCGCG
  	TGGTCGAATA
  2751	TCTGAAACAG TTTGGCGAAC CGGACTATGT TGACGATATN
  	TTGAGCGGCG
  2801	GTATGTCCGA CGATTTGCTG GGAATCAGCC GGAGCGGCGA
  	CGGCGAAACC
  2851	GATCCGATGT ACGACGAGGC CGTGTCNGTT GTTTTGAAAA
  	CGCGCAAAGC
  2901	CAGCATTTCT GGCGTGCAGC GCGCATTGCG TATCGGCTAT
  	AATCGCGCCG
  2951	CGCGTCTGAT TGACCAGATG GAGGCGGAAG GCATTGTGTC
  	CGCACCGGAA
  3001	CACAACGGCA ACCGTACGAT TCTCGTCCCC TTNGACAATG
  	CTTGA

• This encodes a protein having amino acid sequence <SEQ ID 492>:

• 1	MFWIVLIVIL LLALAGLFFV RAQSEREWMR EVSAWQEKKG
  	EKQAELPEIK
  51	DGMPDFPELA LMLFHAVKTA VYWLFVGVV R FCRNYLAHES
  	EPDRPVPPAS
  101	ANRADVPTAS DGYSDSGNGT EEAETEEAEA AEEEAADTED
  	IATAVIDNRR
  151	IPFDRSIAEG LMPSESEISP VRPVFKEITL EEATRALNSA
  	ALRETKKRYI
  201	DAFEKNETAV PKVRVSDTPM EGLQIIGLDD PVLQRTYSRM
  	FDADKEAFSE
  251	SADYGFEPYF EKQHPSAFSA VKAENARNAP FRRHAGQGKG
  	QAEAKSPDVS
  301	QGQSVSDGTA VRDAXRRVSV NLKEPNKATV SAEARISRLI
  	PESRTVVGKR
  351	DVEMPSETEN VFTEXVSSVG YGXPVYDETA DIHIEEPAAP
  	wDAWVVEPPEV
  401	PKVPMPAXDI PPPPPVSEIY NRTYEPPAGF EQVQRSRIAE
  	TDHLADDVLN
  451	GGWQEETAAI ANDGSEGVAE RSSGQYLSET EAFGHDSQAV
  	CPFENVPSER
  501	PSRRAXDTEA DEGAFQSEET GAVSEHLPTT DLLLPPLFNP
  	GATQTEEXLL
  551	XNSITIEEKX AEFKVKVKVV DSYSGPVITR YEIEPDVGVR
  	GNSVLNLEKX
  601	LARSLGVASI RVVETILGKT CMGLELPNPK RQMIRLSEIF
  	NSPEFAESKS
  651	KLTLALGQDI TGQPVVTDLG KAPHLLVAGT TGSGKSVGVN
  	AMILSMLFKA
  701	APEDVRMIMI DPKMLELSIY EGIPHLLAPV VTDMKLAANA
  	LNWCVNEMEK
  751	RYRLMSFMGV RNLAGXNQKI AEAAARGEKI GNPFSLTPDN
  	PEPLXKLPFI
  801	VVVVDEFADL MMTAGKKIEE LIARLAQKAR AAGIHLILAT
  	QRPSVDVITG
  851	LIKANIPTRI AFQVSSKIDS RTILDQMGAE NLLGQGDMLF
  	LPPGTAYPQR
  901	VHGAFASDEE VHRVVEYLKQ FGEPDYVDDX LSGGMSDDLL
  	GISRSGDGET
  951	DPMYDEAVSV VLKTRKASIS GVQRALRIGY NRAARLIDQM
  	EAEGIVSAPE
  1001	HNGNRTILVP XDNA*

• ORF58a and ORF58-1 show 96.6% identity in 1014 aa overlap:
• 

  
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF58 shows complete identity over a 9aa overlap with a predicted ORF (ORF58ng) from N. gonorrhoeae:
• 
• The ORF58ng nucleotide sequence <SEQ ID 493> is predicted to encode a protein having partial amino acid sequence <SEQ ID 494>:

• 1	..SEPDRPVPPA SANRADVPTA SDGYSDSGNG TEEAETEAAE
  	AAEEEAADTE
  51	DIATAVIDNR RIPFDRSIAE GLMQSESKTS PVRPVFKEIT
  	LEEATRALSS
  101	AALRETKKRY IDAFEKNGTA VPKVRVSDTP MEGLQIIGLD
  	DPVLQRTYSR
  151	MFDADKEAFS ESADYGFEPY FEKQHPSAFS AVKAENARNA
  	PFRRHAGQEK
  201	GQAEAKSPDV SQGQSVSDGT AVRDARRRVS VNLKEPNKAT
  	VSAEARISRL
  251	IPESRTVVGK RDVEMPSETE NVFTETVSSV GYGGPVYDEA
  	ADIHIEEPAA
  301	PDAWVVEPPE VPEVAVPEID ILPPPPVSEI YNRTYEPPAG
  	FEQAQRSRIA
  351	ETDHLAADVL NGGWQEETAA IADDGSEGAA ERSSGQYLSE
  	TEAFGHDSQA
  401	VCPFEDVPSE RPSCRVSDTE ADEGAFQSEE TGAVSEHLPT
  	TDLLLPPLFN
  451	PEATQTEEEL LENSITIEEK LAEFKVKVKV VDSYSGPVIT
  	RYEIEPDVGV
  501	RGNSVLNLEK DLARSLGVAS IRVVETIPGK TCMGLELPNP
  	KRQMIRLSEI
  551	FNSPEFAESK SKLTLALGQD ITGQPVVTDL GKAPHLLVAG
  	TTGSGKSVGV
  601	NAMILSMLFK AAPEDVRMIM IDPKMLELSI YEGITHLLAP
  	VVTDMKLAAN
  651	ALNWCVNEME KRYRLMSFMG VRNLAGFNQK IAEAAARGEK
  	IGNPFSLTPD
  701	DPEPLEKLPF IVVVVDEFAD LMMTAGKKIE ELIARLAQKA
  	RAAGIHLILA
  751	TQRPSVDVIT GLIKANIPTR IAFQVSSKID SRTILDQMGA
  	ENLLGQGDML
  801	FLPPGTAYPQ RVHGAFASDE EVHRVVEYLK QFGEPDYVDD
  	ILSGGGSEEL
  851	PGIGRSGDGE TDPMYDEAVS VVLKTRKASI SGVQRALRIG
  	YNRAARLIDQ
  901	MEAEGIVSAP EHNGNRTILV PLDNA*

• This partial gonococcal sequence contains a predicted transmembrane region and a predicted ATP/GTP-binding site motif A (P-loop; double underlined). Furthermore, it has a domain homologous to the FTSK cell division protein of E. coli. Alignment of ORF58ng and FtsK (accession number p46889) show a 65% amino acid identity in 459 overlap:

• ORF58ng:  467 IEEKLAEFKVKVKVVDSYSGPVITRYEIEPDVGVRGNSVLNLEKDLARSLGVASIRVVET  526
  +E +LA+F++K  VV+   GPVITR+E+    GV+   + NL +DLARSL    ++RVVE
  FtsK:     868 VEARLADFRIKADVVNYSPGPVITRFELNLAPGVKAARISNLSRDLARSLSTVAVRVVEV  927
  ORF58ng:  527 IPGKTCMGLELPNPKRQMIRLSEIFNSPEFAESKSKLTLALGQDITGQPVVTDLGKAPHL  586
  IPGK  +GLELPN KRQ + L E+ ++ +F ++ S LT+ LG+DI G+PVV DL K  PHL
  FtSK:     928 IPGKPYVGLELPNKKRQTVYLREVLDNAKFRDNPSPLTVVLGKDIAGEPVVADLAKMPHL  987
  ORF58ng:  587 LVAGTTGSGKSVGVNAMILSMLFKAAPEDVRMIMIDPKMLELSIYEGITHLLAPVVTDMK  646
  LVAGTTGSGKSVGVNAMILSML+KA PEDVR IMIDPKMLELS+YEGI HLL  VVTDMK
  FtsK:     988 LVAGTTGSGKSVGVNAMILSMLYKAQPEDVRFIMIDPKMLELSVYEGIPHLLTEVVTDMK 1047
  ORF58ng:  647 LAANALNWCVNEMEKRYRLMSFMGVANLAGFNQKIAEAAARGEKIGNPFSLTPDDPEP--  704
  AANAL WCVNEME+RY+LMS +GVRNLAG+N+KIAEA      I +P+     D  +
  FtsK:    1048 DAANALRWCVNEMERRYKLMSALGVRNLAGYNEKIAEADRMMRPIPDPYWKPGDSMDAQH 1107
  ORF58ng:  705 --LEKLPFIVVVVDEFADLMMTAGKKIEELIARLAQKARAAGIHLILATQRPSVDVITGL  762
  L+K P+IVV+VDEFADLMMT GKK+EELIARLAQKARAAGIHL+LATQRPSVDVITGL
  FtsK:    1108 PVLKKEPYIVVLVDEFADLMMTVGKKVEELIARLAQKARAAGIHLVLATQRPSVDVITGL 1167
  ORF58ng:  763 IKANIPTRIAFQVSSKIDSRTILDQMGAENLLGQGDMLFLPPGTAYPQRVHGAFASDEEV  822
  IKANIPTRIAF VSSKIDSRTILDQ GAE+LLG GDML+  P +  P RVHGAF  D+EV
  FtsK:    1168 IKANIPTRIAFTVSSKIDSRTILDQAGAESLLGMGDMLYSGPNSTLPVRVHGAFVRDQEV 1227
  ORF58ng:  823 HRVVEYLKQFGEPDYVDDILSGGGSEELPGIGRSGDGETDPMYDEAVSVVLKTRKASISG   882
  H VV+  K  G P YVD I S   SE   G G  G  E DP++D+AV  V + RKASISG
  FtsK:    1228 HAVVQDWKARGRPQYVDGITSDSESEGGAG-GFDGAEELDPLFDQAVQFVTEKRKASISG  1286
  ORF58ng:  883 VQRALRIGYNRAARLIDQMEAEGIVSAPEHNGNRTILVP                       921
  VQR  RIGYNRAAR+I+QMEA+GIVS   HNGNR +L P
  FtsK:    1287 VQRQFRIGYNRAARIIEQMEAQGIVSEQGHNGNREVLAP                      1325

• Further work on ORF58ng revealed the complete gonococcal DNA sequence to be <SEQ ID 495>:

• 1	ATGTTTTGGA TAGTTTTGAT CGTTATtgtg TTGCTTGCGC
  	TTGCCGGCCT
  51	GTTTTTTGTC CGCGCACAAT CCGAACGCGA GTGGATGCGC
  	GAGGTTTCTG
  101	CGTGGCAGGA AAAGAAAGGG GAAAAACAGG CGGAGCTGCC
  	TGAAATCAAA
  151	GACGGTATGC CCGATTTTCC CGAGTTTTCC CTGATGCTTT
  	TCCATGCCGT
  201	CAAAACGGCA GTGTATTGGC TGTTTGTCGG TGTCGTCCGT
  	TTCTGCCGAA
  251	ACTATCTGGC GCACGAATCC GAACCGGACA GGCCCGTTCC
  	GCCTGCTTCT
  301	GCAAACCGTG CGGATGTTCC GACCGCATCC GACGGGTATT
  	CAGACAGTGG
  351	AAACGGGACG GAAGAAGCGG AAACGGAAGC AGCAGAAGCT
  	GCGGAGGAAG
  401	AGGCTGCCgA TACgGAAGAC ATTGCAACTG CCGTAATCGA
  	CAACCGCCGC
  451	ATCCcatTCG ACCGGAGTAT TGCTGAAGGG TTGATGCAGT
  	CTGAAAGCAA
  501	AACTTCGCCC GTCCGTCCGG TTTTTAAGGA AATCACTTTG
  	GAAGAAGCAA
  551	CGCGTGCTTT AAGCAGCGCG GCTTTAAGGG AAACGAAAAA
  	ACGCTATATC
  601	GATGCATTTG AGAAAAACGG AACAGCCGTC CCCAAAGTAC
  	GCGTGTCCGA
  651	TACCCCGATG GAAGGGCTGC AGATTATCGG TTTGGACGAC
  	CCTGTGCTTC
  701	AACGCACGTA TTCCCGTATG TTTGATGCGG ACAAAGAAGC
  	GTTTTCCGAG
  751	TCTGCGGATT ACGGATTTGA GCCGTATTTT GAGAAGCAGC
  	ATCCGTCTGC
  801	CTTTTCTGCA GTCAAAGCCG AAAATGCACG GAATGCGCCG
  	TTCCGCCGTC
  851	ATGCAGGGCA GGAGAAAGGG CAGGCGGAGG CAAAATCCCC
  	GGATGTTTCC
  901	CAAGGGCAGT CCGTTTCAGA CGGCACAGCC GTCCGCGATG
  	CCCGCCGCCG
  951	CGTTTCCGTC AATTTGAAAG AACCGAACAA GGCAACGGTT
  	TCTGCGGAGG
  1001	CGCGGATTTC GCGCCTGATT CCGGAAAGTC GGACGGTTGT
  	CGGGAAACGG
  1051	GATGTCGAAA TGCCGTCTGA AACCGAAAAT GTTTTCACGG
  	AAACCGTTTC
  1101	GTCTGTGGGA TACGGCGGTC CGGTTTATGA TGAAGCTGCC
  	GATATCCATA
  1151	TTGAAGAGCC TGCCGCGCCC GATGCTTGGG TGGTCGAACC
  	ACCCGAAGTG
  1201	CCGGAGGTAG CCGTACCCGA AATCGATATT CTGCCGCCGC
  	CTCCCGTATC
  1251	GGAAATCTAC AACCGTACCT ATGAGCCGCC GGCAGGATTC
  	GAGCAGGCGC
  1301	AACGCAGCCG CATTGCCGAA ACCGACCATC TTGCCGCTGA
  	TGTTTTGAAT
  1351	GGAGGTTGGC AGGAGGAAAC CGCCGCTATT GCAGATGACG
  	GCAGTGAGGG
  1401	TGCGGCAGAG CGGTCAAGCG GGCAATATCT GTCGGAAACC
  	GAAGCGTTCG
  1451	GGCATGACAG TCAGGCGGTT TGTCCGTTTG AAGATGTGCC
  	GTCTGAACGC
  1501	CCGTCCTGCC GGGTATCGGA TACGGAAGCG GATGAAGGGG
  	CGTTCCAATC
  1551	GGAAGAGACC GGTGCGGTAT CCGAACACCT GCCGACAACC
  	GACCTGCTTC
  1601	TGCCTCCGCT GTTCAATCCC GAGGCGACGC AAACCGAAGA
  	AGAACTGTTG
  1651	GAAAACAGCA TCACCATCGA AGAAAAATTG GCGGAGTTCA
  	AAGTCAAGGT
  1701	CAAGGTTGTC GATTCTTATT CCGGCCCCGT GATTACGCGT
  	TATGAAATCG
  1751	AACCCGATGT CGGCGTGCGC GGCAATTCCG TTCTGAATTT
  	GGAAAAAGAC
  1801	TTGGCGCGTT CGCTCGGCGT GGCTTCCATC CGCGTTGTCG
  	AAACCATCCC
  1851	CGGCAAAACC TGCATGGGTT TGGAACTTCC GAACCCGAAA
  	CGCCAAATGA
  1901	TACGCCTGAG CGAAATTTTC AATTCGCCCG AGTTTGCCGA
  	ATCCAAATCC
  1951	AAGCTGACGC TCGCGCTCGG TCAGGACATT ACCGGACAGC
  	CCGTCGTAAC
  2001	CGACTTGGGC AAAGCACCGC ATTTGCTGGT TGCCGGCACG
  	ACCGGTTCGG
  2051	GCAAATCGGT GGGTGTCAAC GCGATGATTC TGTCTATGCT
  	TTTCAAAGCC
  2101	GCGCCGGAAG ACGTGCGTAT GATTATGATC GATCCGAAAA
  	TGCTGGAATT
  2151	GAGCATTTAC GAAGGCATCA CGCACCTGCT CGCCCCTGTC
  	GTTACCGATA
  2201	TGAAGCTGGC GGCAAACGCG CTGAACTGGT GTGTTAACGA
  	AATGGAAAAA
  2251	CGCTACCGCC TGATGAGCTT TATGGGCGTG CGCAATCTTG
  	CGGGCTTCAA
  2301	CCAAAAAATC GCCGAAGCCG CAGCAAGGGG AGAAAAAATC
  	GGCAATCCGT
  2351	TCAGCCTCAC GCCCGACGAT CCCGAACCTT TGGAAAAACT
  	GCCGTTTATC
  2401	GTGGTCGTGG TCGATGAGTT TGCCGATTTG ATGATGACGG
  	CAGGCAAGAA
  2451	AATCGAAGAA CTGATTGCGC GCCTCGCCCA AAAAGCCCGC
  	GCGGCAGGCA
  2501	TCCACCTTAT CCTTGCCACA CAACGCCCCA GCGTCGATGT
  	CATCACGGGT
  2551	CTGATTAAGG CGAACATCCC GACGCGTATC GCGTTCCAAG
  	TGTCCAGCAA
  2601	AATCGACAGC CGCACGATTC TCGACCAAAT GGGCGCGGAA
  	AACCTGCTCG
  2651	GTCAGGGCGA TATGCTGTTC CTGCCGCCGG GTACTGCCTA
  	TCCGCAGCGC
  2701	GTTCACGGCG CGTTTGCCTC GGATGAAGAG GTGCACCGCG
  	TGGTCGAATA
  2751	TCTGAAGCAG TTTGGCGAGC CGGACTATGT TGACGATATT
  	TTGAGCGGCG
  2801	GCGGCAGCGA AGAGCTGCCC GGCATCGGGC GCAGCGGCGA
  	CGGCGAAACC
  2851	GATCCGATGT ACGACGAGGC CGTATCCGTT GTCCTGAAAA
  	CGCGCAAAGC
  2901	CAGCATTTCG GGCGTACAGC GCGCCTTGCG CATCGGCTAC
  	AACCGCGCCG
  2951	CGCGTCTGAT TGACCAAATG GAAGCGGAAG GCATTGTGTC
  	CGCACCGGAA
  3001	CACAACGGCA ACCGTACGAT TCTCGTCCCC TTGGACAATG
  	CTTGA

• This corresponds to the amino acid sequence <SEQ ID 496; ORF58ng-1>:

• 1	MFWIVLIVIV LLALAGLFFV RAQSEREWMR EVSAWQEKKG
  	EKQAELPEIK
  51	DGMPDFPEFS LMLFHAVKTA VYWLFVGVV R FCRNYLAHES
  	EPDRPVPPAS
  101	ANRADVPTAS DGYSDSGNGT EEAETEAAEA AEEEAADTED
  	IATAVIDNRR
  151	IPFDRSIAEG LMQSESKTSP VRPVFKEITL EEATRALSSA
  	ALRETKKRYI
  201	DAFEKNGTAV PKVRVSDTPM EGLQIIGLDD PVLQRTYSRM
  	FDADKEAFSE
  251	SADYGFEPYF EKQHPSAFSA VKAENARNAP FRRHAGQEKG
  	QAEAKSPDVS
  301	QGQSVSDGTA VRDARRRVSV NLKEPNKATV SAEARISRLI
  	PESRTVVGKR
  351	DVEMPSETEN VFTETVSSVG YGGPVYDEAA DIHIEEPAAP
  	DAWVVEPPEV
  401	PEVAVPEIDI LPPPPVSEIY NRTYEPPAGF EQAQRSRIAE
  	TDHLAADVLN
  451	GGWQEETAAI ADDGSEGAAE RSSGQYLSET EAFGHDSQAV
  	CPFEDVPSER
  501	PSCRVSDTEA DEGAFQSEET GAVSEHLPTT DLLLPPLFNP
  	EATQTEEELL
  551	ENSITIEEKL AEFKVKVKVV DSYSGPVITR YEIEPDVGVR
  	GNSVLNLEKD
  601	LARSLGVASI RVVETIPGKT CMGLELPNPK RQMIRLSEIF
  	NSPEFAESKS
  651	KLTLALGQDI TGQPVVTDLG KAPHLLVAGT TGSGKSVGVN
  	AMILSMLFKA
  701	APEDVRMIMI DPKMLELSIY EGITHLLAPV VTDMKLAANA
  	LNWCVNEMEK
  751	RYRLMSFMGV RNLAGFNQKI AEAAARGEKI GNPFSLTPDD
  	PEPLEKLPFI
  801	VVVVDEFADL MMTAGKKIEE LIARLAQKAR AAGIHLILAT
  	QRPSVDVITG
  851	LIKANIPTRI AFQVSSKIDS RTILDQMGAE NLLGQGDMLF
  	LPPGTAYPQR
  901	VHGAFASDEE VHRVVEYLKQ FGEPDYVDDI LSGGGSEELP
  	GIGRSGDGET
  951	DPMYDEAVSV VLKTRKASIS GVQRALRIGY NRAARLIDQM
  	EAEGIVSAPE
  1001	HNGNRTILVP LDNA*

• ORF58ng-1 and ORF58-1 show 97.2% identity in 1014 aa overlap:
• 

  
• Furthermore, ORF58ng-1 shows significant homology to the E. coli protein FtsK:

• sp|P46889|FTSK_ECOLI CELL DIVISION PROTEIN FTSK >gi|1651412|gnl|PID|d1015290
  (Dl division protein FtsK [Escherichia coli] >gi|1651418|gnl|PID|d1015296
  (D90727) Cell division protein FtsK [Escherichia coli] >gi|1787117 (AE000191)
  cell division protein FtsK [Escherichia coli] Length = 1329
  Score = 576 bits (1469), Expect = e−163
  Identities = 301/459 (65%), Positives = 353/459 (76%), Gaps = 5/459 (1%)

  Query:	556	IEEKLAEFKVKVKVVDSYSGPVITRYEIEPDVGVRGNSVLNLEKDLARSLGVASIRVVET	615
  		+E +LA+F++K  VV+   GPVITR+E+    GV+   + NL +DLARSL   ++RVVE
  Sbjct:	868	VEARLADFRIKADVVNYSPGPVITRFELNLAPGVKAARISNLSRDLARSLSTVAVRVVEV	927
  Query:	616	IPGKTCMGLELPNPKRQMIRLSEIFNSPEFAESKSKLTLALGQDITGQPVVTDLGKAPHL	675
  		IPGK  +GLELPN KRQ + L E+ ++ +F ++ S LT+ LG+DI G+PVV DL K PHL
  Sbjct:	928	IPGKPYVGLELPNKKRQTVYLREVLDNAKFRDNPSPLTVVLGKDIAGEPVVADLAKMPHL	987
  Query:	676	LVAGTTGSGKSVGVNAMILSMLFKAAPEDVRMIMIDPKMLELSIYEGITHLLAPVVTDMK	735
  		LVAGTTGSGKSVGVNAMILSML+KA  PEDVR IMIDPKMLELS+YEGI HLL  VVTDMK
  Sbjct:	988	LVAGTTGSGKSVGVNAMILSMLYKAQPEDVRFIMIDPKMLELSVYEGIPHLLTEVVTDMK	1047
  Query:	736	LAANALNWCVNEMEKRYRLMSFMGVRNLAGFNQKIAEAAARGEKIGNPFSLTPDDPEP--	793
  		AANAL WCVNEME+RY+LMS +GVRNLAG+N+KIAEA      I +P+    D  +
  Sbjct:	1048	DAANALRWCVNEMERRYKLMSALGVRNLAGYNEKIAEADRMMRPIPDPYWKPGDSMDAQH	1107
  Query:	794	--LEKLPFIVVVVDEFADLMMTAGKKIEELIARLAQKARAAGIHLILATQRPSVDVITGL	851
  		L+K P+IVV+VDEFADLMMT GKK+EELIARLAQKARAAGIHL+LATQRPSVDVITGL
  Sbjct:	1108	PVLKKEPYIVVLVDEFADLMMTVGKKVEELIARLAQKARAAGIHLVLATQRPSVDVITGL	1167
  Query:	852	IKANIPTRIAFQVSSKIDSRTILDQMGAENLLGQGDMLFLPPGTAYPQRVHGAFASDEEV	911
  		IKANIPTRIAF VSSKIDSRTILDQ GAE+LLG GDML+  P +  P RVHGAF  D+EV
  Sbjct:	1168	IKANIPTRIAFTVSSKIDSRTILDQAGAESLLGMGDMLYSGPNSTLPVRVHGAFVRDQEV	1227
  Query:	912	HRVVEYLKQFGEPDYVDDILSGGGSEELPGIGRSGDGETDPMYDEAVSVVLKTRKASISG	971
  		H VV+  K  G P YVD I S   SE   G G  G  E DP++D+AV  V + RKASISG
  Sbjct:	1228	HAVVQDWKARGRPQYVDGITSDSESEGGAG-GFDGAEELDPLFDQAVQFVTEKRKASISG	1286
  Query:	972	VQRALRIGYNRAARLIDQMEAEGIVSAPEHNGNRTILVP	1010
  		VQR  RIGYNRAAR+I+QMEA+GIVS   HNGNR +L P
  Sbjct:	1287	VQRQFRIGYNRAARIIEQMEAQGIVSEQGHNGNREVLAP	1325

• Based on this analysis, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 59
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 497>:
• 
• This corresponds to the amino acid sequence <SEQ ID 498; ORF101>:
• 
• Further work revealed the complete nucleotide sequence <SEQ ID 499>:

• 1	ATGATTTATC AAAGAAACCT CATCAAAGAA CTCTCTTTTA
  	CCGCCGTCGG
  51	CATTTTCGTC GTCCTCTTGG CGGTATTGGT CTCCACGCAG
  	GCAATCAACC
  101	TGCTCGGCCG TGCCGCCGAC GGGCGTGTCG CCATCGATGC
  	CGTGTTGGCA
  151	TTGGTCGGCT TCTGGGTCAT CGGTATGACG CCGCTTTTGC
  	TGGTGTTGAC
  201	CGCATTTATC AGTACGTTGA CCGTGTTGAC CCGCTACTGG
  	CGCGACAGCG
  251	AAATGTCGGT CTGGCTATCC TGCGGATTGG CATTGAAACA
  	ATGGATACGC
  301	CCGGTGATGC AGTTTGCCGT GCCGTTTGCC GTTTTGGTTG
  	CCGTCATGCA
  351	GCTTTGGGTG ATACCGTGGG CAGAGCTACG CAGCCGCGAA
  	TACGCTGAAA
  401	TCCTGAAGCA GAAGCAGGAA TTGTCTTTGG TGGAGGCAGG
  	CGAGTTCAAC
  451	AGTTTGGGCA AGCGCAACGG CAGGGTTTAT TTTGTCGAAA
  	CCTTCGATAC
  501	CGAATCCGGC ATCATGAAAA ACCTGTTCCT GCGCGAACAG
  	GACAAAAACG
  551	GCGGCGACAA CATCATCTTC GCCAAAGAAG GTAACTTCTC
  	GCTGAACGAC
  601	AACAAACGCA CGCTCGAATT GCGCCACGGC TACCGTTACA
  	GCGGCACGCC
  651	CGGACGCGCC GACTACAATC AGGTTTCCTT CCAAAAACTC
  	AACCTGATTA
  701	TCAGCACCAC GCCCAAACTC ATCGACCCCG TTTCCCACCG
  	CCGTACCATT
  751	CCGACCGCCC AACTGATTGG CAGCAGCAAC CCGCAACATC
  	AGGCGGAATT
  801	GATGTGGCGC ATCTCGCTGA CCGTCAGCGT CCTCCTACTC
  	TGCCTGCTTG
  851	CCGTGCCGCT TTCCTATTTC AACCCGCGCA GCGGACATAC
  	CTACAATATC
  901	TTGATTGCCA TCGGTTTGTT TTTAATTTAC CAAAACGGGC
  	TGACCCTGCT
  951	TTTTGAAGCC GTGGAAGACG GCAAAATCCA TTTTTGGCTC
  	GGACTGCTGC
  1001	CTATGCACAT TATCATGTTT GCCGTTGCAC TCATCCTGTT
  	GCGCGTCCGC
  1051	AGTATGCCCA GCCAGCCCTT CTGGCAGGCG GTTGGCAAAA
  	GTCTGACATT
  1101	GAAAGGCGGA AAATGA

• This corresponds to the amino acid sequence <SEQ ID 500; ORF101-1>:

• 1	MIYQRNLIKE LSFTAVGIFV VLLAVLVSTQ AINLLGRAAD
  	GRVAIDAVLA
  51	LVGFWVIGMT PLLLVLTAFI STLTVLTRYW RDSEMSVWLS
  	CGLALKQWIR
  101	PVMQFAVPFA VLVAVMQLWV IPWAELRSRE YAEILKQKQE
  	LSLVEAGEFN
  151	SLGKRNGRVY FVETFDTESG IMKNLFLREQ DKNGGDNIIF
  	AKEGNFSLND
  201	NKRTLELRHG YRYSGTPGRA DYNQVSFQKL NLIISTTPKL
  	IDPVSHRRTI
  251	PTAQLIGSSN PQHQAELMWR ISLTVSVLLL CLLAVPLSYF
  	NPRSGHTYNI
  301	LIAIGLFLIY QNGLTLLFEA VEDGKIHFWL GLLPMHIIMF
  	AVALILLRVR
  351	SMPSQPFWQA VGKSLTLKGG K*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF101 shows 91.2% identity over a 57aa overlap and 95.7% identity over a 69aa overlap with an ORF (ORF101a) from strain A of N. meningitidis:
• 
• The complete length ORF101a nucleotide sequence <SEQ ID 501> is:

• 1	ATGATTTATC AAAGAAACCT CATCAAAGAA CTCTCTTTTA
  	CCGCCGTCGG
  51	CATTTTCGTC GTCCTCTTGG CGGTATTGGT CTCCACGCAG
  	GCAATCAACC
  101	TGCTCGGCCN TGCCGCCGAC NGGCGTNTCG CCATCGATGC
  	CGTGTTGGCA
  151	TTGGTCGGCT TCTGGGTCNN NNGNATGACG CCGCTTTTGC
  	TNGTGTTGAC
  201	CGCATTTATC AGTACGTTGA CCGTGTTGAC CCGCTACTGG
  	CGNGACAGCG
  251	AAATGTCGGT CTGGNTATCC TGCGGATTGG CATTGAAACA
  	ATGGATACGC
  301	CCGGTGATGC AGTTTGCCGT GCCGTTTGCC GTTTTGGTTG
  	CCGTCATGCA
  351	GCTTTGGGTG ATACCGTGGG CAGAGCTACG CAGCCGCGAA
  	TACGCTGAAA
  401	TCCTGAAGCA GAAGCAGGAA TTGTCTTTGG TGGAGGCAGG
  	CGGGTTCAAC
  451	AGTTTGGGCA AGCGCAACGG CAGGGTTTAT TTTGTCGAAA
  	CCTTCGATAC
  501	CGAATCCGGC ATCATGAAAA ACCTGTTCCT GCGCGAACAG
  	GACAAAAACG
  551	GCGGCGACAA CATCATCTTC NCCAAAGAAA GTAACTTCTC
  	GCTGAACGAC
  601	AACAAACGCA CGCTCGAATT GCGCCACGGC TACCGTTACA
  	GCGGCACGCC
  651	CGGACGCGCC GACTACAATC AGGTTTCCTT CCNAAAACTC
  	AACCTGATTA
  701	TCAGCACCAC GCCCAAACTC ATCGACCCCG TTTCCCACCG
  	CCGTACNATN
  751	CCNACNGCCC AACTGATTGG CAGCAGCAAC CCGCAACATC
  	ANGCGGAATT
  801	GATGTGGCGC ATCTCGCTGA CCGTCAGCGT CCTCCTACTC
  	TGCCTGCTTG
  851	CCGTGCCGCT TTCCTATTTC AACCCGCGCA GCGGACATAC
  	CTACAATATC
  901	TTGANTGCCA TCGGTTTGTT TTTAATTTAC CAAAACGGGC
  	TGACCCTGCT
  951	TTTTGAAGCC GTGGAAGACG GCAAAATCCA TTTTTGGCTC
  	GGACTGCTGC
  1001	CTATGCACAT CATCATGTTC GTCATCGCAA TCGTACTTCT
  	GCGCGTCCGC
  1051	AGCATGCCCA GCCAGCCCTT CTGGCAGGCG GTTGGCAAAA
  	GTCTGACATT
  1101	GAAAGGCGGA AAATGA

• This encodes a protein having amino acid sequence <SEQ ID 502>:

• 	1	MIYQRNLIKE LSFTAVGIFV VLLAVLVSTQ AINLLGXAAD
  		XRXAIDAVLA
  	51	LVGFWVXXMT PLLLVLTAFI STLTVLTRYW RDSEMSVWXS
  		CGLALKQWIR
  	101	PVMQFAVPFA VLVAVMQLWV IPWAELRSRE YAEILKQKQE
  		LSLVEAGGFN
  	151	SLGKRNGRVY FVETFDTESG IMKNLFLREQ DKNGGDNIIF
  		XKESNFSLND
  	201	NKRTLELRHG YRYSGTPGRA DYNQVSFXKL NLIISTTPKL
  		IDPVSHRRTX
  	251	PTAQLIGSSN PQHXAELMWR ISLTVSVLLL CLLAVPLSYF
  		NPRSGHTYNI
  	301	LXAIGLFLIY QNGLTLLFEA VEDGKIHFWL GLLPMHIIMF
  		VIAIVLLRVR
  	351	SMPSQPFWQA VGKSLTLKGG K*

• ORF101a and ORF101-1 show 95.4% identity in 371 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF101 shows 96.5% identity in 57aa overlap at the N-terminal domain and 95.1% identity in 61 as overlap at the C-terminal domain, respectively, with a predicted ORF (ORF101ng) from N. gonorrhoeae:
• 
• The ORF101ng nucleotide sequence <SEQ ID 503> is predicted to encode a protein having partial amino acid sequence <SEQ ID 504>:

• 1	MIYQRNLIKE LSFTAVGIFV VLLAVLVSTQ AINLLGRAAD
  	GRVAIDAVLA
  51	LVGFWVIGMT PLLLVLTAFI STLTVLTRYW RDSEMSVWLS
  	CGLALKQWIR
  101	PVMQFAVPFA ILIAVMQLWV IPWAELRSRE YAEILKQKQE
  	LSLVEAGEFN
  151	NLGKRNGRVY FVETFDTESG IMKNLFLREQ DKNGGDNIIF
  	AKEGNFSLKD
  201	NKRTLELRHG YRYSGTPGRA DYNQVSFQKL NLIISTTPKL
  	IDPVSHRRTI
  251	STAQLIGSSN PQHQAELMWR ISLTVSVLLL CLLAVPLSYF
  	NPRSGHTYNI
  301	LIAIGLFLIY QNGLTLLFEA VEDGKIHFWL GLLPMHIIMF
  	VIAIVLLRVR
  351	SMPSQPFWQA VG...

• Further work revealed the complete nucleotide sequence <SEQ ID 505>:

• 1	ATGATTTATC AAAGAAACCT CATCAAAGAA CTCTCTTTTA
  	CCGCCGTCGG
  51	CATTTTCGTC GTCCTCTTGG CGGTGTTGGT GTCCACGCAG
  	GCGATCAACC
  101	TGCTTGGCCG CGCAGCTGAC GGGCGTGTCG CCATCGATGC
  	CGTGTTGGCC
  151	TTAGTCGGCT TCTGGGTCAT CGGTATGACC CCGCTTTTGC
  	TGGTGTTGAC
  201	CGCATTCATC AGCACGCTGA CCGTATTGAC CCGCTACTGG
  	CGCGACAGCG
  251	AAATGTCGGT CTGGCTATCC TGCGGATTGG CGTTGAAACA
  	GTGGATACGC
  301	CCCGTCATGC AGTTTGCCGT GCCGTTTGCC ATCCTGATTG
  	CCGTCATGCA
  351	GCTTTGGGTG ATACCGTGGG CAGAGCTGCG CAGCCGCGAA
  	TATGCCGAAA
  401	TTTTGAAGCA GAAGCAGGAA TTGTCTTTGG TGGAAGCCGG
  	CGAGTTCAAT
  451	AACTTGGGCA AGCGCAACGG CAgggtttaT TtcgtcgaaA
  	CCTTTGACAC
  501	CGaatccgGC ATCATGAAAA ACCTGTtcct GcGCGAACAG
  	GACAAAAACG
  551	gcggcgacaA CATCATCTTC GCcaaaGAag gtaactTctc
  	gctgaaggaC
  601	AACAAAcgca cgctcgaATT GCGCCACGGC TACCGTTACA
  	GCGGcacgcC
  651	CGGacGCGCc gactaCAATC AGGTTtcctt cCAAAAacTc
  	aacctgATta
  701	TCAGCACCAC GCCCAAacTT ATCGaccCCG TTTCCCACCG
  	CCGCACCATT
  751	tcgacCGCCC AAcTGATTGG CAGCAGCAAT CCGCAACATC
  	AGGCAGAATT
  801	GATGTGGCGC ATCTCGCTGA CCGTCAGCGT CCTCCTGCTC
  	TGCCTACTCG
  851	CCGTGCCGCT TTCCTATTTC AACCCGCGCA GCGGACATAC
  	CTACAATATC
  901	TTGATTGCCA TCGGTTTGTT TTTAATTTAC CAAAACGGGC
  	TGACCCTGCT
  951	TTTTGAAGCC GTGGAAGACG GCAAAATCCA TTTTTGGCTC
  	GGACTGCTGC
  1001	CTATGCACAT CATCATGTTC GTCATCGCAA TCGTACTTCT
  	GCGCGTCCGC
  1051	AGTATGCCCA GCCAGCCCTT CTGGCAGGCG GTTGGCAAAA
  	GTCTGACATT
  1101	GAAAGgcgGA AAATGA

• This corresponds to the amino acid sequence <SEQ ID 506; ORF101ng-1>:

• 	1	MIYQRNLIKE LSFTAVGIFV VLLAVLVSTQ AINLLGRAAD
  		GRVAIDAVLA
  	51	LVGFWVIGMT PLLLVLTAFI STLTVLTRYW RDSEMSVWLS
  		CGLALKQWIR
  	101	PVMQFAVPFA ILIAVMQLWV IPWAELRSRE YAEILKQKQE
  		LSLVEAGEFN
  	151	NLGKRNGRVY FVETFDTESG IMKNLFLREQ DKNGGDNIIF
  		AKEGNFSLKD
  	201	NKRTLELRHG YRYSGTPGRA DYNQVSFQKL NLIISTTPKL
  		IDPVSHRRTI
  	251	STAQLIGSSN PQHQAELMWR ISLTVSVLLL CLLAVPLSYF
  		NPRSGHTYNI
  	301	LIAIGLFLIY QNGLTLLFEA VEDGKIHFWL GLLPMHIIMF
  		VIAIVLLRVR
  	351	SMPSQPFWQA VGKSLTLKGG K*

• ORF101ng-1 and ORF101-1 show 97.6% identity in 371 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence (double-underlined) and several putative transmembrane domains (single-underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 60
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 507>:

• 1	..GGTGGTGGTT TTATCAATGC TTCCTGTGCC ACTTTGACGA
  	CAGCCAAACC
  51	GCAATATCAA GCAGGAGACC TTAGCGCTTT TAAGATAAGG
  	CAAGGCAATG
  101	TTGTAATCGC CGGACACGGT TTGGATGCAC GTGATACCGA
  	TTACACACGT
  151	ATTCTCAGTT ATCATTCCAA AATCGATGCA CCCGTATGGG
  	GACAAGATGT
  201	TCGTGTCGTC GCGGGACAAA ACGATGTGGC CGCAACAGGT
  	GATGCACATT
  251	CGCCTATTCT CAATAATGCT GCTGCCAATA CGTCAAACAA
  	TACAGCCAAC
  301	AACGGCACAC ATATCCCTTT ATTTGCGATT GATACAGGCA
  	AATTAGGAGG
  351	TAT.GTATGC CAACAAAATC ACCTTGATCA GTACGGTCGA
  	GCAAGCAGGC
  401	ATTCGTAA

• This corresponds to the amino acid sequence <SEQ ID 508; ORF113>:

• 1	..GGGFINASCA TLTTAKPQYQ AGDLSAFKIR QGNVVIAGHG
  	LDARDTDYTR
  51	ILSYHSKIDA PVWGQDVRVV AGQNDVAATG DAHSPILNNA
  	AANTSNNTAN
  101	NGTHIPLFAI DTGKLGGXVC QQNHLDQYGR ASRHS*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with pspA Putative Secreted Protein of N. meningitidis (Accession AF030941)
• ORF and pspA show 44% aa identity in 179aa overlap:

• orf113	GGGFINASCATLTTAKPQYQAGDLSAFKIRQGNVVIAGHGLDARDTDYTRILSYHSKIDA		60
  	GGG INA+  TLT+  P    G+L+ F +  G VVI G GLD  D DYTRILS  ++I+A
  pspa	GGGLINAASVTLTSGVPVLNNGNLTGFDVSSGKVVIGGKGLDTSDADYTRILSRAAEINA	256
  orf113	PVWGQDVRVVAGQNDVAATGDAHSPILXXXXXXXXXXXXXXGTHIPLFAIDTGKLGGMYA		120
  	VWG+DV+VV+G+N +   G                      +  P  AIDT  LGGMYA
  pspa	GVWGKDVKVVSGKNKLDFDG---------SLAKTASAPSSSDSVTPTVAIDTATLGGMYA	307
  orf113	NKITLISTVEQAGIRNQGQWFASAGNVAVNAEGKLVNTGMIAATGENHAVSLHARNVHN	179
  	+KITLIST   A IRN+G+ FA+ G V ++A+GKL N+G I A      +++ A+ V N
  pspa	DKITLISTDNGAVIRNKGRIFAATGGVTLSADGKLSNSGSIDAA----EITISAQTVDN	362

  
  Homology with a Predicted ORF from N. gonorrhoeae
• ORF113 shows 86.5% identity in 52aa overlap at the N-terminal part and 94.1% identity in 17aa overlap at the C-terminal part with a predicted ORF (ORF113ng) from N. gonorrhoeae:
• 
• The complete length ORF113ng nucleotide sequence <SEQ ID 509> is predicted to encode a protein having amino acid sequence <SEQ ID 510>:

• 1	MNKTLYRVIF NRKRGAVVAV AETTKREGKS CADSGSGSVY
  	VKSVSFIPTH
  51	SKAFCFSALG FSLCLALGTV NIAFADGIIT DKAAPKTQQA
  	TILQTGNGIP
  101	QVNIQTPTSA GVSVNQYAQF DVGNRGAILN NSRSNTQTQL
  	GGWIQGNPWL
  151	TRGEARVVVN QINSSHPSQL NGYIEVGGRR AEVVIANPAG
  	IAVNGGGFIN
  201	ASRATLTTGQ PQYQAGDFSG FKIRQGNAVI AGHGLDARDT
  	DFTRILVCQQ
  251	NHLDQYGRTS RHS*

• Based on this analysis, it is predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 61
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 511>:

• 1	..TCAACGGGAC ATAGCGAACA AAATTACACT TTGCCGCGAG
  	AAATCACACG
  51	CAACATTTCA CTGGGTTCAT TTGCCTATGA ATCGCATCGC
  	AAAGCATTAA
  101	GCCATCATGC GCCCAGCCAA GGCACTGAGT TGCCGCAAAG
  	CAACGGTATT
  151	TCGCTACCCT ATACGTCCAA TTCTTTTACC CCATTACCCA
  	GCAGCAGCTT
  201	ATACATTATC AATCCTGTCA ATAAAGGCTA TCTTGTTGAA
  	ACCGATCCAC
  251	GCTTTGCCAA CTACCGTCAA TGGTTGGGTA GTGACTATAT
  	GCtGGACAGC
  301	CTCAAACTAG ACCCAAACAA TTTACATAAA CGTTTGGGTG
  	ATGGTTATTA
  351	CGAGCAACGT TTAATCAATG AACAAATCGC AGAGCTGACA
  	GGGCATCGTC
  401	GTTTAGAcGG TTATCAAAAC GACGAAGAAC AATTTAAAGC
  	CTTAATGGAT
  451	AATGGCGCGA CTGCGGCACG TTcGATGAAT CTCAGCGTTG
  	GCATTGCATT
  501	AAGTGCCGAG CAAGTAGCGC AACTGACCAG CGATATTGTT
  	TGGTTGGTAC
  551	AAAAAGAAGT TAAGCTTCCT GATGGCGGCA CACAAACCGT
  	ATTGGTGCCA
  601	CAGGTTTATG TACGCGTTAA AAATGGCGAC ATAGACGGTA
  	AAGGTGCATT
  651	GTTGTCAGGC AGCAATACAC AAATCAATGT TTCAGGCAGC
  	CTGAAAAACT
  701	CAGGCACGAT TGCAGGgCGC AATGCGCTTA TTATCAATAC
  	CGATACGCTA
  751	GACAATATCG GTGGGCGTAT TCATGCGCAA AAATCAGCGG
  	TTACGGCCAC
  801	ACAAGACATC AATAATATTG GCGGCATGCT TTCTGCCGAA
  	CAGACATTAT
  851	TGCTCAACGC AGGCAACAAC ATCAACAGCC AAAGCACCAC
  	CGCCAGCAGT
  901	CAAAATACAC AAGGCAGCAG CACCTACCTA GACCGAATGG
  	CAGGTATTTA
  951	TATCACAGGC AAAGAAAAAG GTGTTT..

• This corresponds to the amino acid sequence <SEQ ID 512; ORF115>:

• 1	..STGHSEQNYT LPREITRNIS LGSFAYESHR KALSHHAPSQ
  	GTELPQSNGI
  51	SLPYTSNSFT PLPSSSLYII NPVNKGYLVE TDPRFANYRQ
  	WLGSDYMLDS
  101	LKLDPNNLHK RLGDGYYEQR LINEQIAELT GHRRLDGYQN
  	DEEQFKALMD
  151	NGATAARSMN LSVGIALSAE QVAQLTSDIV WLVQKEVKLP
  	DGGTQTVLVP
  201	QVYVRVKNGD IDGKGALLSG SNTQINVSGS LKNSGTIAGR
  	NALIINTDTL
  251	DNIGGRIHAQ KSAVTATQDI NNIGGMLSAE QTLLLNAGNN
  	INSQSTTASS
  301	QNTQGSSTYL DRMAGIYITG KEKGV..

• Computer analysis of this amino acid sequence gave the following results:
• Homology with the pspA Putative Secreted Protein of N. meningitidis (Accession Number AF030941)
• ORF115 and pspA protein show 50% aa identity in 325aa overlap:

• Orf115:	1	STGHSEQNYTLPREITRNISLGSFAYESHRKALSHHAPSQGTELPQSNGISLPYTSNSFT	60
  		STG+S   Y    E++ +I +G  AY+ +    +   P     +   NGI   +T
  pspA:	778	STGYSRSPYEPAPEVS-SIRMGISAYKGYAPQQASDIPGTVVPVVAENGIHPTFT-----	831
  Orf115:	61	PLPSSSLYIINPVNKGYLVETDPRFANYRQWLGSDYMLDSLKLDPNNLHKRLGDGYYEQR	120
  		LP+SSL+ I P NKGYL+ETDP F +YR+WLGS YML +L+ DPN++HKRLGDGYYEQ+
  pspA:	832	-LPNSSLFAIAPNNKGYLIETDPAFTDYRKWLGSGYMLAALQQDPNHIHKRLGDGYYEQK	890
  Orf115:	121	LINEQIAELTGHRRLDGYQNDEEQFKALMDNGATAARSMNLSVGIALSAEQVAQLTSDIV	180
  		L+NEQIA+LTG+RRLDGY NDEEQFKALMDNG T A+ + L+ GIALSAEQVA+LTSDIV
  pspA:	891	LVNEQIAKLTGYRRLDGYTNDEEQFKALMDNGITIAKELQLTPGIALSAEQVARLTSDIV	950
  Orf115:	181	WLVQKEVKLPDGGTQTVLVPQVYVRVKNGDIDGKGALLSGSNTQINVSGSLKN-SGTIAG	239
  		WL  + V LPDG TQTVL P+VYVR +  D++G+GALLSGS   I  SG+++N  G IAG
  pspA:	951	WLENETVTLPDGTTQTVLKPKVYVRARPKDMNGQGALLSGSVVDIG-SGAIENRGGLIAG	1009
  Orf115:	240	RNALIINTDTLDNIGGRIHAQKSAVTATQDINNIGGMLSAEQTLLLNAGXXXXXXXXXXX	299
  		R ALI+N   + N+ G +  +     A  DI N G  + AE  LLL A
  pspA:	1010	REALILNAQNIKNLQGDLQGKNIFAAAGSDITNTGS-IGAENALLLKASNNIESRSETRS	1068
  Orf115:	300	XXXXXXXXXYLDRMAGIYITGKEKG	324
  		+ R+AGIY+TG++ G
  pspA:	1069	NQNEQGSVRNIGRVAGIYLTGRQNG	1093

  
  Homology with a Predicted ORF from N. gonorrhoeae
• ORF115 shows 91.9% identity over a 334aa overlap with a predicted ORF (ORF115ng) from N. gonorrhoeae:
• 
• An ORF115ng nucleotide sequence <SEQ ID 513> was predicted to encode a protein having amino acid sequence <SEQ ID 514>:

• 1	MLVQTEKDGL HNEQTFGEKK VFSENGKLHN YWRARRKGHD
  	ETGHREQNYT
  51	LPEEITRDIS LGSFAYESHS KALSRHAPSQ GTELPQSNRD
  	NIRTAKSNGI
  101	SLPYTPNSFT PLPGSSLYII NPANKGYLVE TDPRFANYRQ
  	WLGSDYMLGS
  151	LKLDPNNLHK RLGDGYYEQR LINEQIAELT GHRRLDGYQN
  	DEEQFKALMD
  201	NGATAARSMN LSVGIALSAE QAAQLTSDIV WLVQKEVKLP
  	DGGTQTVLMP
  251	QVYVRVKNGG IDGKGALLSG SNTQINVSGS LKNSGTIAGR
  	NALIINTDTL
  301	DNIGGRIHAQ KSAVTATQDI NNIGGILSAE QTLLLNAGNN
  	INNQSTAKSS
  351	QNAQGSSTYL DRMAGIYITG KEKGVLAAQA GKDINIIAGQ
  	ISNQSDQGQT
  401	RLQAGRDINL DTVQTGKYQE IHFDADNHTI RGSTNEVGSS
  	IQTKGDVTLL
  451	SGNNLNAKAA EVGSAKGTLA VYAKNDITIS SGIHAGQVDD
  	ASKHTGRSGG
  501	GNKLVITDKA QSHHETAQSS TFEGKQVVLQ AGNDANILGS
  	NVISDNGTRI
  551	QAGNHVRIGT TQTQSQSETY HQTQKSGLMS AGIGFTIGSK
  	TNTQENQSQS
  601	NEHTGSTVGS LKGDTTIVAS KHYEQTGSNV SSPEGNNLIS
  	TQSMDIGAAQ
  651	NQLNSKTTQT YEQKGLTVAF SSPVTDLAQQ AIAVAHKAAK
  	QFDKAKTTAL
  701	MPWRLPMQVG RLFKQAKAPK K*

• Further work revealed the following partial gonococcal DNA sequence <SEQ ID 515>:

• 1	TTGCTTGTGC AAACAGAAAA AGACGGTTTG CATAACGAGC
  	AAACCTTTGG
  51	CGAGAAGAAA GTCTTCAGCG AAAATGGTAA GTTGCACAAC
  	TACTGGCGTG
  101	CGCGTCGTAA AGGACATGAT GAAACAGGGC ATCGTGAACA
  	AAATTATACT
  151	TTGCCGGAGG AAATCACACG CGACATTTCA CTGGGTTCAT
  	TTGCCTATGA
  201	ATCGCATAGC AAAGCATTAA GCCGTCATGC GCCCAGCCAA
  	GGCACTGAGT
  251	TGCCACAAAG TAACCGGGAT AATATCCGTA CTGCGAAAAG
  	CAACGGTATT
  301	TCGCTACCCT ATACGCCCAA TTCTTTTACC CCATTACCCG
  	GCAGCAGCTT
  351	ATACATTATC AATCCTGCCA ATAAAGGCTA TCTTGTTGAA
  	ACCGATCCAC
  401	GCTTTGCCAA CTACCGTCAA TGGTTGGGTA GTGACTATAT
  	GCTGGGCAGC
  451	CTCAAACTAG ACCCAAACAA TTTACATAAA CGTTTGGGTG
  	ATGGTTATTA
  501	CGAGCAACGT TTAATCAATG AACAAATCGC AGAGCTGACA
  	GGGCATCGTC
  551	GTTTAGACGG TTATCAAAAC GACGAAGAAC AATTTAAAGC
  	CTTAATGGAT
  601	AATGGCGCGA CTGCGGCACG TTCGATGAAT CTCAGCGTTG
  	GCATTGCATT
  651	AAGTGCCGAG CAAGCAGCGC AACTGACCAG CGATATTGTT
  	TGGTTGGTAC
  701	AAAAAGAAGT TAAACTTCCT GATGGCGGCA CACAAACCGT
  	ATTGATGCCA
  751	CAGGTTTATG TACGCGTTAA AAATGGCGGC ATAGACGGTA
  	AAGGTGCATT
  801	GTTGTCAGGC AGCAATACAC AAATCAATGT TTCAGGCAGC
  	CTGAAAAACT
  851	CAGGCACGAT TGCAGGGCGC AATGCGCTTA TTATCAATAC
  	CGATACGCTA
  901	GACAATATCG GTGGGCGTAT TCATGCGCAA AAATCAGCGG
  	TTACGGCCAC
  951	ACAAGACATC AATAATATTG GCGGCATTCT TTCTGCCGAA
  	CAGACATTAT
  1001	TGCTCAATGC GGGTAACAAC ATCAACAACC AAAGCACGGC
  	CAAGAGCAGT
  1051	CAAAATGCAC AAGGTAGCAG CACCTACCTA GACCGAATGG
  	CAGGTATTTA
  1101	TATCACAGGC AAAGAAAAAG GTGTTTTAGC AGCGCAGGCA
  	GGCAAAGACA
  1151	TCAACATCAT TGCCGGTCAA ATCAGCAATC AATCAGATCA
  	AGGGCAAACC
  1201	CGGCTGCAGG CAGGACGCGA CATTAACCTG GATACGGTAC
  	AAACCGGCAA
  1251	ATATCAAGAA ATCCATTTTG ATGCCGATAA CCATACCATC
  	CGAGGTTCAA
  1301	CGAACGAAGT CGGCAGCAGC ATTCAAACAA AAGGCGATGT
  	TACCCtatTG
  1351	TCAGGGAATA ATCTCAATGC CAAAGCTGCC GAAGTCGGCA
  	GCGCAAAAGG
  1401	CACACTTGCC GTGTATGCTA AAAATGACAT TACTATCAGC
  	TCAGGCATCC
  1451	ATGCCGGCCA AGTTGATGAT GCGTCCAAAC ATACAGGCAG
  	AAGCGGCGGC
  1501	GGTAATAAAT TAGTCATTAC CGATAAAGCC CAAAGTCATC
  	ACGAAACTGC
  1551	TCAAAGCAGC ACCTTTGAAG GCAAGCAAGT TGTATTGCAG
  	GCAGGAAACG
  1601	ATGCCAACAT CCTTGGCAGT AATGTTATTT CCGATAATGG
  	CACCCGGATT
  1651	CAAGCAGGCA ATCATGTTCG CATTGGTACA ACCCAAACTC
  	AAAGCCAAAG
  1701	CGAAACCTAT CATCAAACCC AAAAATCAGG ATTGATGAGT
  	GCAGGTATCG
  1751	GCTTCACTAT TGGCAGCAAG ACAAACACAC AAGAAAACCA
  	ATCCCAAAGC
  1801	AACGAACATA CAGGCAGTAC CGTAGGCAGC CTGAAAGGCG
  	ATACCACCAT
  1851	TGTTGCAAGC AAACACTACG AACAAACCGG CAGCAACGTT
  	TCCAGCCCTG
  1901	AGGGCAACAA CCTTATCAGC ACGCAAAGTA TGGATATTGG
  	CGCAGCACAA
  1951	AACCAATTAA ACAGCAAAAC CACCCAAACC TACGAACAAA
  	AAGGCTTAAC
  2001	GGTGGCATTC AGTTCGCCCG TTACCGATTT GGCACAACAA
  	GCGATTGCCG
  2051	TAGCACACAA AGCAGCAAAC AAGTCGGACA AAGCAAAAAC
  	GACCGCGTTA
  2101	ATGCCATGGC GGCTGCCAAT GCAGGTTGGC AGGCCTATCA
  	AACAGGCAAA
  2151	GGCGCACAAA ACTTAG

• This corresponds to the amino acid sequence <SEQ ID 516; ORF115ng-1>:

• 1	LLVQTEKDGL HNEQTFGEKK VFSENGKLHN YWRARRKGHD
  	ETGHREQNYT
  51	LPEEITRDIS LGSFAYESHS KALSRHAPSQ GTELPQSNRD
  	NIRTAKSNGI
  101	SLPYTPNSFT PLPGSSLYII NPANKGYLVE TDPRFANYRQ
  	WLGSDYMLGS
  151	LKLDPNNLHK RLGDGYYEQR LINEQIAELT GHRRLDGYQN
  	DEEQFKALMD
  201	NGATAARSMN LSVGIALSAE QAAQLTSDIV WLVQKEVKLP
  	DGGTQTVLMP
  251	QVYVRVKNGG IDGKGALLSG SNTQINVSGS LKNSGTIAGR
  	NALIINTDTL
  301	DNIGGRIHAQ KSAVTATQDI NNIGGILSAE QTLLLNAGNN
  	INNQSTAKSS
  351	QNAQGSSTYL DRMAGIYITG KEKGVLAAQA GKDINIIAGQ
  	ISNQSDQGQT
  401	RLQAGRDINL DTVQTGKYQE IHFDADNHTI RGSTNEVGSS
  	IQTKGDVTLL
  451	SGNNLNAKAA EVGSAKGTLA VYAKNDITIS SGIHAGQVDD
  	ASKHTGRSGG
  501	GNKLVITDKA QSHHETAQSS TFEGKQVVLQ AGNDANILGS
  	NVISDNGTRI
  551	QAGNHVRIGT TQTQSQSETY HQTQKSGLMS AGIGFTIGSK
  	TNTQENQSQS
  601	NEHTGSTVGS LKGDTTIVAS KHYEQTGSNV SSPEGNNLIS
  	TQSMDIGAAQ
  651	NQLNSKTTQT YEQKGLTVAF SSPVTDLAQQ AIAVAHKAAN
  	KSDKAKTTAL
  701	MPWRLPMQVG RPIKQAKAHK T*

• This gonococcal protein (ORF115ng-1) shows 91.9% identity with ORF115 over 334aa:
• 
• In addition, it shows homology with a secreted N. meningitidis protein in the database:

• gi|2623258 (AF030941) putative secreted protein [Neisseria meningitidis]
  Length = 2273
  Score = 604 bits (1541), Expect = e−172
  Identities = 325/678 (47%), Positives = 449/678 (65%), Gaps = 22/678 (3%)

  Query:	1	LLVQTEKDGLHNEQTFGEKKVFSENGKLHNYWRARRKGHDETGHREQNYTLPEEITRDIS	60
  		L+V T +  L N++T G K + ++ G LH Y R  +KG D TG+    Y    E++  I
  Sbjct:	739	LIVGTPESALDNDETLGTKTI-TDKGDLHRYHRHHKKGRDSTGYSRSPYEPAPEVS-SIR	796
  Query:	61	LGSFAYESHSKALSRHAPSQGTELPQSNRDNIRTAKSNGISLPYTPNSFTPLPGSSLYII	120
  		+G  AY+ +       AP Q +++P +    +     NGI   +T      LP SSL+ I
  Sbjct:	797	MGISAYKGY-------APQQASDIPGTV---VPVVAENGIHPTFT------LPNSSLFAI	840
  Query:	121	NPANKGYLVETDPRFANYRQWLGSDYMLGSLKLDPNNLHKRLGDGYYEQRLINEQIAELT	180
  		P NKGYL+ETDP F +YR+WLGS YML +L+ DPN++HKRLGDGYYEQ+L+NEQIA+LT
  Sbjct:	841	APNNKGYLIETDPAFTDYRKWLGSGYMLAALQQDPNHIHKRLGDGYYEQKLVNEQIAKLT	900
  Query:	181	GHRRLDGYQNDEEQFKALMDNGATAARSMNLSVGIALSAEQAAQLTSDIVWLVQKEVKLP	240
  		G+RRLDGY NDEEQFKALMDNG T A+ + L+ GIALSAEQ A+LTSDIVWL  + V LP
  Sbjct:	901	GYRRLDGYTNDEEQFKALMDNGITIAKELQLTPGIALSAEQVARLTSDIVWLENETVTLP	960
  Query:	241	DGGTQTVLMPQVYVRVKNGGIDGKGALLSGSNTQINVSGSLKN-SGTIAGRNALIINTDT	299
  		DG TQTVL P+VYVR +   ++G+GALLSGS   I  SG+++N  G IAGR ALI+N
  Sbjct:	961	DGTTQTVLKPKVYVRARPKDMNGQGALLSGSVVDIG-SGAIENRGGLIAGREALILNAQN	1019
  Query:	300	LDNIGGRIHAQKSAVTATQDINNIGGILSAEQTLLLNAGNNINNQSTAKSSQNAQGSSTY	359
  		+ N+ G +  +     A  DI N G I  AE  LLL A NNI ++S  +S+QN QGS
  Sbjct:	1020	IKNLQGDLQGKNIFAAAGSDITNTGSI-GAENALLLKASNNIESRSETRSNQNEQGSVRN	1078
  Query:	360	LDRMAGIYITGKEKGVLAAQAGKDINIIAGQISNQSDQGQTRLQAGRDINLDTVQTGKYQ	419
  		+ R+AGIY+TG++ G +   AG +I + A +++NQS+ GQT L AG DI  DT    + Q
  Sbjct:	1079	IGRVAGIYLTGRQNGSVLLDAGNNIVLTASELTNQSEDGQTVLNAGGDIRSDTTGISRNQ	1138
  Query:	420	EIHFDADNHTIRGSTNEVGSSIQTKGDVTLLSGNNLNAKAAEVGSAKGTLAVYAKNDITI	479
  		FD+DN+ IR   NEVGS+I+T+G+++L +  ++  +AAEVGS + G L + A  DI +
  Sbjct:	1139	NTIFDSDNYVIRKEQNEVGSTIRTRGNLSLNAKGDIRIRAAEVGSEQGRLKLAAGRDIKV	1198
  Query:	480	SSGIHAGQVDDASKHTGRSGGGNKLVITDKAQSHHETAQSSTFEGKQVVLQAGNDANILG	539
  		+G    + +DA K+TGRSGGG K  +T   ++ +  A S T +GK+++L +G D  + G
  Sbjct:	1199	EAGKAHTETEDALKYTGRSGGGIKQKMTRHLKNQNGQAVSGTLDGKEIILVSGRDITVTG	1258
  Query:	540	SNVISDNGTRIQAGNHVRIGTTQTQSQSETYHQTQKSGLM-SAGIGFTIGSKTNTQENQS	598
  		SN+I+DN T + A N++ +   +T+S+S   ++ +KSGLM S GIGFT GSK +TQ N+S
  Sbjct:	1259	SNIIADNHTILSAKNNIVLKAAETRSRSAEMNKKEKSGLMGSGGIGFTAGSKKDTQTNRS	1318
  Query:	599	QSNEHTGSTVGSLKGDTTIVASKHYEQTGSNVSSPEGNNLISTQSMDIGAAQNQLNSKTT	658
  		++  HT S VGSL G+T I A KHY QTGS +SSP+G+  IS+  + I AAQN+ + ++
  Sbjct:	1319	ETVSHTESVVGSLNGNTLISAGKHYTQTGSTISSPQGDVGISSGKISIDAAQNRYSQESK	1378
  Query:	659	QTYEQKGLTVAFSSPVTD	676
  		Q YEQKG+TVA S PV +
  Sbjct:	1379	QVYEQKGVTVAISVPVVN	1396

• Based on this analysis, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 62
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 517>:

• 1	..TCAGGGAATA ACCTCAATGC CAAAGCTGCC GAAGTCAGCA
  	GCGCAAACGG
  51	TACACTCGCT GTGTCTGCCA ATAATGACAT CAACATCAGC
  	GCAGGCATCA
  101	ACACGACCCA TGTTGATGAT GCGTCCAAAC ACACAGGCAG
  	AAGCGGTGGT
  151	GGCAATAAAT TAGTCATTAC CGATAAAGCC CAAAGTCATC
  	ACGAAACCGC
  201	CCAAAGCAGC ACCTTTGAAG GCAAGCAAGT TGTATTGCAG
  	GCAGGAAACG
  251	ATGCCAACAT CCTTGGCAGC AATGTTATTT CCGATAATGG
  	CACCCAGATT
  301	CAAGCAGGCA ATCATGTTCG CATTGGTACA ACCCAAACTC
  	AAAGCCAAAG
  351	CGAAACCTAT CATCAAACCC AGAAATCAGG ATTGATGAGT
  	GCAGGTATCG
  401	GCTTCACTAT TGGCAGCAAG ACAAACACAC AAGAAAACCA
  	ATCCCAAAGC
  451	AACGAACATA CAGGCAGTAC CGTAGGCAGC TTGAAAGGCG
  	ATACCACCAT
  501	TGTTGCAGGC AAACACTACG AACAAATCGG CAGTACCGTT
  	TCCAGCCCGG
  551	AAGGCAACAA TACCATCTAT GCCCAAAGCA TAGACATTCA
  	AGCGGCACAC
  601	AACAAATTAA ACAGTAATAC CACCCAAACC TATGAACAAA
  	AAGG.CTAAC
  651	GGTGGCATTC AGTTCGCCCG TTACCGATTT GGCACAACAA
  	...

• This corresponds to the amino acid sequence <SEQ ID 518; ORF117>:

• 1	..SGNNLNAKAA EVSSANGTLA VSANNDINIS AGINTTHVDD
  	ASKHTGRSGG
  51	GNKLVITDKA QSHHETAQSS TFEGKQVVLQ AGNDANILGS
  	NVISDNGTQI
  101	QAGNHVRIGT TQTQSQSETY HQTQKSGLMS AGIGFTIGSK
  	TNTQENQSQS
  151	NEHTGSTVGS LKGDTTIVAG KHYEQIGSTV SSPEGNNTIY
  	AQSIDIQAAH
  201	NKLNSNTTQT YEQKXLTVAF SSPVTDLAQQ ...

• Computer analysis of this amino acid sequence gave the following results:
• Homology with the pspA Putative Secreted Protein of N. meningitidis (Accession Number AF030941)
• ORF117 and pspA protein show 45% aa identity in 224aa overlap:

• Orf117:	4	NLNAKAAEVSSANGTLAVSANNDINISAGINTTHVDDASKHTGRSGGGNKLVITDKAQSH	63
  		++  +AAEV S  G L ++A  DI + AG   T  +DA K+TGRSGGG K  +T   ++
  pspA:	1173	DIRIRAAEVGSEQGRLKLAAGRDIKVEAGKAHTETEDALKYTGRSGGGIKQKMTRHLKNQ	1232
  Orf117:	64	HETAQSSTFEGKQVVLQAGNDANILGSNVISDNGTQIQAGNHVRIGTTQTQSQSETYHQT	123
  		+  A S T +GK+++L +G D  + GSN+I+DN T + A N++ +   +T+S+S   ++
  pspA:	1233	NGQAVSGTLDGKEIILVSGRDITVTGSNIIADNHTILSAKNNIVLKAAETRSRSAEMNKK	1292
  Orf117:	124	QKSGLM-SAGIGFTIGSKTNTQENQSQSNEHTGSTVGSLKGDTTIVAGKHYEQIGSTVSS	182
  		+KSGLM S GIGFT GSK +TQ N+S++  HT S VGSL G+T I AGKHY Q GST+SS
  pspA:	1293	EKSGLMGSGGIGFTAGSKKDTQTNRSETVSHTESVVGSLNGNTLISAGKHYTQTGSTISS	1352
  Orf117:	183	PEGNNTIYAQSIDIQAAHNKLNSNTTQTYEQKXLTVAFSSPVTD	226
  		P+G+  I +  I I AA N+ +  + Q YEQK +TVA S PV +
  pspA:	1353	PQGDVGISSGKISIDAAQNRYSQESKQVYEQKGVTVAISVPVVN	1396

  
  Homology with a Predicted ORF from N. gonorrhoeae
• ORF117 shows 90% identity over a 230aa overlap with a predicted ORF (ORF117ng) from N. gonorrhoeae:
• 
• An ORF117ng nucleotide sequence <SEQ ID 519> was predicted to encode a protein having amino acid sequence <SEQ ID 520>:

• 1	..LLVQTEKDGL HNEQTFGEKK VFSENGKLHN YWRARRKGHD
  	ETGHREQNYT
  51	LPEEITRDIS LGSFAYESHS KALSRHAPSQ GTELPQSNRD
  	NIRTAKSNGI
  101	SLPYTPNSFT PLPGSSLYII NPANKGYLVE TDPRFANYRQ
  	WLGSDYMLGS
  151	LKLDPNNLHK RLGDGYYEQR LINEQIAELT GHRRLDGYQN
  	DEEQFKALMD
  201	NGATAARSMN LSVGIALSAE QAAQLTSDIV WLVQKEVKLP
  	DGGTQTVLMP
  251	QVYVRVKNGG IDGKGALLSG SNTQINVSGS LKNSGTIAGR
  	NALIINTDTL
  301	DNIGGRIHAQ KSAVTATQDI NNIGGILSAE QTLLLNAGNN
  	INNQSTAKSS
  351	QNAQGSSTYL DRMAGIYITG KEKGVLAAQA GKDINIIAGQ
  	ISNQSDQGQT
  401	RLQAGRDINL DTVQTGKYQE IHFDADNHTI RGSTNEVGSS
  	IQTKGDVTLL
  451	SGNNLNAKAA EVGSAKGTLA VYAKNDITIS SGIHAGQVDD
  	ASKHTGRSGG
  501	GNKLVITDKA QSHHETAQSS TFEGKQVVLQ AGNDANILGS
  	NVISDNGTRI
  551	QAGNHVRIGT TQTQSQSETY HQTQKSGLMS AGIGFTIGSK
  	TNTQENQSQS
  601	NEHTGSTVGS LKGDTTIVAS KHYEQTGSNV SSPEGNNLIS
  	TQSMDIGAAQ
  651	NQLNSKTTQT YEQKGLTVAF SSPVTDLAQQ AIAVAHKAAK
  	QFDKAKTTAL
  701	MPWRLPMQVG RLFKQAKAPK K*

• Further work revealed the following gonococcal partial DNA sequence <SEQ ID 521>:

• 1	TTGCTTGTGC AAACAGAAAA AGACGGTTTG CATAACGAGC
  	AAACCTTTGG
  51	CGAGAAGAAA GTCTTCAGCG AAAATGGTAA GTTGCACAAC
  	TACTGGCGTG
  101	CGCGTCGTAA AGGACATGAT GAAACAGGGC ATCGTGAACA
  	AAATTATACT
  151	TTGCCGGAGG AAATCACACG CGACATTTCA CTGGGTTCAT
  	TTGCCTATGA
  201	ATCGCATAGC AAAGCATTAA GCCGTCATGC GCCCAGCCAA
  	GGCACTGAGT
  251	TGCCACAAAG TAACCGGGAT AATATCCGTA CTGCGAAAAG
  	CAACGGTATT
  301	TCGCTACCCT ATACGCCCAA TTCTTTTACC CCATTACCCG
  	GCAGCAGCTT
  351	ATACATTATC AATCCTGCCA ATAAAGGCTA TCTTGTTGAA
  	ACCGATCCAC
  401	GCTTTGCCAA CTACCGTCAA TGGTTGGGTA GTGACTATAT
  	GCTGGGCAGC
  451	CTCAAACTAG ACCCAAACAA TTTACATAAA CGTTTGGGTG
  	ATGGTTATTA
  501	CGAGCAACGT TTAATCAATG AACAAATCGC AGAGCTGACA
  	GGGCATCGTC
  551	GTTTAGACGG TTATCAAAAC GACGAAGAAC AATTTAAAGC
  	CTTAATGGAT
  601	AATGGCGCGA CTGCGGCACG TTCGATGAAT CTCAGCGTTG
  	GCATTGCATT
  651	AAGTGCCGAG CAAGCAGCGC AACTGACCAG CGATATTGTT
  	TGGTTGGTAC
  701	AAAAAGAAGT TAAACTTCCT GATGGCGGCA CACAAACCGT
  	ATTGATGCCA
  751	CAGGTTTATG TACGCGTTAA AAATGGCGGC ATAGACGGTA
  	AAGGTGCATT
  801	GTTGTCAGGC AGCAATACAC AAATCAATGT TTCAGGCAGC
  	CTGAAAAACT
  851	CAGGCACGAT TGCAGGGCGC AATGCGCTTA TTATCAATAC
  	CGATACGCTA
  901	GACAATATCG GTGGGCGTAT TCATGCGCAA AAATCAGCGG
  	TTACGGCCAC
  951	ACAAGACATC AATAATATTG GCGGCATTCT TTCTGCCGAA
  	CAGACATTAT
  1001	TGCTCAATGC GGGTAACAAC ATCAACAACC AAAGCACGGC
  	CAAGAGCAGT
  1051	CAAAATGCAC AAGGTAGCAG CACCTACCTA GACCGAATGG
  	CAGGTATTTA
  1101	TATCACAGGC AAAGAAAAAG GTGTTTTAGC AGCGCAGGCA
  	GGCAAAGACA
  1151	TCAACATCAT TGCCGGTCAA ATCAGCAATC AATCAGATCA
  	AGGGCAAACC
  1201	CGGCTGCAGG CAGGACGCGA CATTAACCTG GATACGGTAC
  	AAACCGGCAA
  1251	ATATCAAGAA ATCCATTTTG ATGCCGATAA CCATACCATC
  	CGAGGTTCAA
  1301	CGAACGAAGT CGGCAGCAGC ATTCAAACAA AAGGCGATGT
  	TACCCtatTG
  1351	TCAGGGAATA ATCTCAATGC CAAAGCTGCC GAAGTCGGCA
  	GCGCAAAAGG
  1401	CACACTTGCC GTGTATGCTA AAAATGACAT TACTATCAGC
  	TCAGGCATCC
  1451	ATGCCGGCCA AGTTGATGAT GCGTCCAAAC ATACAGGCAG
  	AAGCGGCGGC
  1501	GGTAATAAAT TAGTCATTAC CGATAAAGCC CAAAGTCATC
  	ACGAAACTGC
  1551	TCAAAGCAGC ACCTTTGAAG GCAAGCAAGT TGTATTGCAG
  	GCAGGAAACG
  1601	ATGCCAACAT CCTTGGCAGT AATGTTATTT CCGATAATGG
  	CACCCGGATT
  1651	CAAGCAGGCA ATCATGTTCG CATTGGTACA ACCCAAACTC
  	AAAGCCAAAG
  1701	CGAAACCTAT CATCAAACCC AAAAATCAGG ATTGATGAGT
  	GCAGGTATCG
  1751	GCTTCACTAT TGGCAGCAAG ACAAACACAC AAGAAAACCA
  	ATCCCAAAGC
  1801	AACGAACATA CAGGCAGTAC CGTAGGCAGC CTGAAAGGCG
  	ATACCACCAT
  1851	TGTTGCAAGC AAACACTACG AACAAACCGG CAGCAACGTT
  	TCCAGCCCTG
  1901	AGGGCAACAA CCTTATCAGC ACGCAAAGTA TGGATATTGG
  	CGCAGCACAA
  1951	AACCAATTAA ACAGCAAAAC CACCCAAACC TACGAACAAA
  	AAGGCTTAAC
  2001	GGTGGCATTC AGTTCGCCCG TTACCGATTT GGCACAACAA
  	GCGATTGCCG
  2051	TAGCACACAA AGCAGCAAAC AAGTCGGACA AAGCAAAAAC
  	GACCGCGTTA
  2101	ATGCCATGGC GGCTGCCAAT GCAGGTTGGC AGGCCTATCA
  	AACAGGCAAA
  2151	GGCGCACAAA ACTTAG

• This corresponds to the amino acid sequence <SEQ ID 522; ORF117ng-1>:

• 1	LLVQTEKDGL HNEQTFGEKK VFSENGKLHN YWRARRKGHD
  	ETGHREQNYT
  51	LPEEITRDIS LGSFAYESHS KALSRHAPSQ GTELPQSNRD
  	NIRTAKSNGI
  101	SLPYTPNSFT PLPGSSLYII NPANKGYLVE TDPRFANYRQ
  	WLGSDYMLGS
  151	LKLDPNNLHK RLGDGYYEQR LINEQIAELT GHRRLDGYQN
  	DEEQFKALMD
  201	NGATAARSMN LSVGIALSAE QAAQLTSDIV WLVQKEVKLP
  	DGGTQTVLMP
  251	QVYVRVKNGG IDGKGALLSG SNTQINVSGS LKNSGTIAGR
  	NALIINTDTL
  301	DNIGGRIHAQ KSAVTATQDI NNIGGILSAE QTLLLNAGNN
  	INNQSTAKSS
  351	QNAQGSSTYL DRMAGIYITG KEKGVLAAQA GKDINIIAGQ
  	ISNQSDQGQT
  401	RLQAGRDINL DTVQTGKYQE IHFDADNHTI RGSTNEVGSS
  	IQTKGDVTLL
  451	SGNNLNAKAA EVGSAKGTLA VYAKNDITIS SGIHAGQVDD
  	ASKHTGRSGG
  501	GNKLVITDKA QSHHETAQSS TFEGKQVVLQ AGNDANILGS
  	NVISDNGTRI
  551	QAGNHVRIGT TQTQSQSETY HQTQKSGLMS AGIGFTIGSK
  	TNTQENQSQS
  601	NEHTGSTVGS LKGDTTIVAS KHYEQTGSNV SSPEGNNLIS
  	TQSMDIGAAQ
  651	NQLNSKTTQT YEQKGLTVAF SSPVTDLAQQ AIAVAHKAAN
  	KSDKAKTTAL
  701	MPWRLPMQVG RPIKQAKAHK T*

• ORF117ng-1 shows the same 90% identity over a 230aa overlap with ORF117. In addition, it shows homology with a secreted N. meningitidis protein in the database:

• gi|2623258 (AF030941) putative secreted protein [Neisseria meningitidis]
  Length = 2273
  Score = 604 bits (1541), Expect = e−172
  Identities = 325/678 (47%), Positives = 449/678 (65%), Gaps = 22/678 (3%)

  Query:	1	LLVQTEKDGLHNEQTFGEKKVFSENGKLHNYWRARRKGHDETGHREQNYTLPEEITRDIS	60
  		L+V T +  L N++T G K + ++ G LH Y R  +KG D TG+    Y    E++  I
  Sbjct:	739	LIVGTPESALDNDETLGTKTI-TDKGDLHRYHRHHKKGRDSTGYSRSPYEPAPEVS-SIR	796
  Query:	61	LGSFAYESHSKALSRHAPSQGTELPQSNRDNIRTAKSNGISLPYTPNSFTPLPGSSLYII	120
  		+G  AY+ +       AP Q +++P +    +     NGI   +T      LP SSL+ I
  Sbjct:	797	MGISAYKGY-------APQQASDIPGTV---VPVVAENGIHPTFT------LPNSSLFAI	840
  Query:	121	NPANKGYLVETDPRFANYRQWLGSDYMLGSLKLDPNNLHKRLGDGYYEQRLINEQIAELT	180
  		P NKGYL+ETDP F +YR+WLGS YML +L+ DPN++HKRLGDGYYEQ+L+NEQIA+LT
  Sbjct:	841	APNNKGYLIETDPAFTDYRKWLGSGYMLAALQQDPNHIHKRLGDGYYEQKLVNEQIAKLT	900
  Query:	181	GHRRLDGYQNDEEQFKALMDNGATAARSMNLSVGIALSAEQAAQLTSDIVWLVQKEVKLP	240
  		G+RRLDGY NDEEQFKALMDNG T A+ + L+ GIALSAEQ A+LTSDIVWL  + V LP
  Sbjct:	901	GYRRLDGYTNDEEQFKALMDNGITIAKELQLTPGIALSAEQVARLTSDIVWLENETVTLP	960
  Query:	241	DGGTQTVLMPQVYVRVKNGGIDGKGALLSGSNTQINVSGSLKN-SGTIAGRNALIINTDT	299
  		DG TQTVL P+VYVR +   ++G+GALLSGS   I  SG+++N  G IAGR ALI+N
  Sbjct:	961	DGTTQTVLKPKVYVRARPKDMNGQGALLSGSVVDIG-SGAIENRGGLIAGREALILNAQN	1019
  Query:	300	LDNIGGRIHAQKSAVTATQDINNIGGILSAEQTLLLNAGNNINNQSTAKSSQNAQGSSTY	359
  		+ N+ G +  +     A  DI N G I  AE  LLL A NNI ++S  +S+QN QGS
  Sbjct:	1020	IKNLQGDLQGKNIFAAAGSDITNTGSI-GAENALLLKASNNIESRSETRSNQNEQGSVRN	1078
  Query:	360	LDRMAGIYITGKEKGVLAAQAGKDINIIAGQISNQSDQGQTRLQAGRDINLDTVQTGKYQ	419
  		+ R+AGIY+TG++ G +   AG +I + A +++NQS+ GQT L AG DI  DT    + Q
  Sbjct:	1079	IGRVAGIYLTGRQNGSVLLDAGNNIVLTASELTNQSEDGQTVLNAGGDIRSDTTGISRNQ	1138
  Query:	420	EIHFDADNHTIRGSTNEVGSSIQTKGDVTLLSGNNLNAKAAEVGSAKGTLAVYAKNDITI	479
  		FD+DN+ IR   NEVGS+I+T+G+++L +  ++  +AAEVGS +G L + A  DI +
  Sbjct:	1139	NTIFDSDNYVIRKEQNEVGSTIRTRGNLSLNAKGDIRIRAAEVGSEQGRLKLAAGRDIKV	1198
  Query:	480	SSGIHAGQVDDASKHTGRSGGGNKLVITDKAQSHHETAQSSTFEGKQVVLQAGNDANILG	539
  		+G    + +DA K+TGRSGGG K  +T   ++ +  A S T +GK+++L +G D  + G
  Sbjct:	1199	EAGKAHTETEDALKYTGRSGGGIKQKMTRHLKNQNGQAVSGTLDGKEIILVSGRDITVTG	1258
  Query:	540	SNVISDNGTRIQAGNHVRIGTTQTQSQSETYHQTQKSGLM-SAGIGFTIGSKTNTQENQS	598
  		SN+I+DN T + A N++ +   +T+S+S   ++ +KSGLM S GIGFT GSK +TQ N+S
  Sbjct:	1259	SNIIADNHTILSAKNNIVLKAAETRSRSAEMNKKEKSGLMGSGGIGFTAGSKKDTQTNRS	1318
  Query:	599	QSNEHTGSTVGSLKGDTTIVASKHYEQTGSNVSSPEGNNLISTQSMDIGAAQNQLNSKTT	658
  		++  HT S VGSL G+T I A KHY QTGS +SSP+G+  IS+  + I AAQN+ + ++
  Sbjct:	1319	ETVSHTESVVGSLNGNTLISAGKHYTQTGSTISSPQGDVGISSGKISIDAAQNRYSQESK	1378
  Query:	659	QTYEQKGLTVAFSSPVTD	676
  		Q YEQKG+TVA S PV +
  Sbjct:	1379	QVYEQKGVTVAISVPVVN	1396

• Based on this analysis, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 63
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 523>:

• 1	ATGATTTACA TCGTACTGTT TCTAGCTGTC GTCCTCGCCG
  	TTGTCGCCTA
  51	CAACATGTAT CAGGAAAACC AATACCGCAA AAAAGTGCGC
  	GACCAGTTCG
  101	GACACTCCGA CAAAGATGCC CTGCTCAACA GCAwAACCAG
  	CCATGTCCGC
  151	GACGGCAAAC CGTCCGGCGG GTCAGTCATG ATGCCGAAAC
  	CCCAACCGGC
  201	GGTCAAAAAA ACGGCAAAAC CCCAAGACCC CGyCATGCGC
  	AACCTGCAAG
  251	AACAGGATGC CGTCTACATC GCCAAGCAGA AACAGGCAAA
  	AGCCTCCCCG
  301	TTCAAAACCG AAATCGAAAC CGCCTTGGAA GAAAGCGGCA
  	TTATCGGCAA
  351	CTCCGCCCAC ACCGTTTCCG AACCCCAAAC CGGACATTCC
  	GCAACGAAAC
  401	CTGCCGACGC GTCGGCAAAA CCTGCACCCG TTCCGCAAAC
  	ACCTGCAAAA
  451	CCGCTGATTA CGCTCAAAGA ACTGTCAAAA GTCGAATTAT
  	CCTGGTTTGA
  501	CGTGCGCATC GACTTCATCT CCTAT...

• This corresponds to the amino acid sequence <SEQ ID 524; ORF119>:

• 1	MIYIVLFLAV VLAVVAYNMY QENQYRKKVR DQFGHSDKDA
  	LLNSXTSHVR
  51	DGKPSGGSVM MPKPQPAVKK TAKPQDPXMR NLQEQDAVYI
  	AKQKQAKASP
  101	FKTEIETALE ESGIIGNSAH TVSEPQTGHS ATKPADASAK
  	PAPVPQTPAK
  151	PLITLKELSK VELSWFDVRI DFISY...

• Further work revealed the complete nucleotide sequence <SEQ ID 525>:

• 1	ATGATTTACA TCGTACTGTT TCTAGCTGTC GTCCTCGCCG
  	TTGTCGCCTA
  51	CAACATGTAT CAGGAAAACC AATACCGCAA AAAAGTGCGC
  	GACCAGTTCG
  101	GACACTCCGA CAAAGATGCC CTGCTCAACA GCAAAACCAG
  	CCATGTCCGC
  151	GACGGCAAAC CGTCCGGCGG GTCAGTCATG ATGCCGAAAC
  	CCCAACCGGC
  201	GGTCAAAAAA ACGGCAAAAC CCCAAGACCC CGCCATGCGC
  	AACCTGCAAG
  251	AACAGGATGC CGTCTACATC GCCAAGCAGA AACAGGCAAA
  	AGCCTCCCCG
  301	TTCAAAACCG AAATCGAAAC CGCCTTGGAA GAAAGCGGCA
  	TTATCGGCAA
  351	CTCCGCCCAC ACCGTTTCCG AACCCCAAAC CGGACATTCC
  	GCACCGAAAC
  401	CTGCCGACGC GCCGGCAAAA CCTGCACCCG TTCCGCAAAC
  	ACCTGCAAAA
  451	CCGCTGATTA CGCTCAAAGA ACTGTCAAAA GTCGAATTAC
  	CCTGGTTTGA
  501	CGTGCGCTTC GACTTCATCT CCTATATCGC GCTGACCGAA
  	GCCAAAGAAC
  551	TGCACGCACT GCCGCGCCTT TCCAACCGCT GCCGCTACCA
  	GATTGTCGGC
  601	TGCACCATGG ACGACCATTT CCAGATTGCC GAACCCATCC
  	CGGGCATCCG
  651	CTATCAGGCA TTTATCGTGG GTATTCAGGC AGTCAGCCGC
  	AACGGACTTG
  701	CCTCGCAGGA AGAACTCTCC GCATTCAACC GCCAGGTGGA
  	CGCATTCGCA
  751	CAAAGCATGG GCGGTCAGAC GCTGCACACC GACCTTGCCG
  	CCTTTATCGA
  801	AGTGGCTTCC GCACTGGACG CATTCTGCGC GCGCGTCGAC
  	CAGACCATCG
  851	CCATCCATTT GGTTTCCCCG ACCAGCATCA GCGGCGTAGA
  	ACTGCGTTCC
  901	GCCGTAACGG GCGTGGGTTT CGTTTTGGAA GACGACGGCG
  	CGTTCCACTA
  951	TACCGACACG TCGGGCTCGA CCATGTTCTC CATCTGCTCG
  	CTCAACAACG
  1001	AGCCGTTTAC CAACGCCCTT TTGGACAACC AGTCCTACAA
  	AGGCTTCAGT
  1051	ATGCTGCTCG ACATCCCGCA CTCTCCGGCA GGCGAAAAAA
  	CCTTCGACGA
  1101	TTTGTTTATG GATTTGGCGG TACGCCTGTC CGGCCAGTTG
  	AACCTGAATC
  1151	TGGTCAACGA CAAAATGGAA GAAGTTTCGA CCCAATGGCT
  	CAAAGACGTG
  1201	CGCACTTATG TATTGGCGCG TCAGTCCGAG ATGCTCAAAG
  	TCGGTATCGA
  1251	ACCGGGCGGC AAAACCGCAT TGCGCCTGTT CTCCTAA

• This corresponds to the amino acid sequence <SEQ ID 526; ORF119-1>:

• 1	MIYIVLFLAV VLAVVAYNMY QENQYRKKVR DQFGHSDKDA
  	LLNSKTSHVR
  51	DGKPSGGSVM MPKPQPAVKK TAKPQDPAMR NLQEQDAVYI
  	AKQKQAKASP
  101	FKTEIETALE ESGIIGNSAH TVSEPQTGHS APKPADAPAK
  	PAPVPQTPAK
  151	PLITLKELSK VELPWFDVRF DFISYIALTE AKELHALPRL
  	SNRCRYQIVG
  201	CTMDDHFQIA EPIPGIRYQA FIVGIQAVSR NGLASQEELS
  	AFNRQVDAFA
  251	QSMGGQTLHT DLAAFIEVAS ALDAFCARVD QTIAIHLVSP
  	TSISGVELRS
  301	AVTGVGFVLE DDGAFHYTDT SGSTMFSICS LNNEPFTNAL
  	LDNQSYKGFS
  351	MLLDIPHSPA GEKTFDDLFM DLAVRLSGQL NLNLVNDKME
  	EVSTQWLKDV
  401	RTYVLARQSE MLKVGIEPGG KTALRLFS*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF119 shows 93.7% identity over. a 175aa overlap with an ORF (ORF119a) from strain A of N. meningitidis:
• 
• The complete length ORF119a nucleotide sequence <SEQ ID 527> is:

• 1	ATGATTTACA TCGTACTGTT CCTCGCCGCC GTCCTCGCCG
  	TTGTCGCCTA
  51	CAATATGTAT CAGGAAAACC AATACCGCAA AAAAGTGCGC
  	GACCAGTTCG
  101	GGCACTCCGA CAAAGATGCC CTGCTCAACA GCAAAACCAG
  	CCATGTCCGC
  151	GACGGCAAAC CGTCCGGCGG GCCAGTCATG ATGCCGAAAC
  	CCCAACCGGC
  201	GGTCAAAAAA ACGGCAAAAT CCCAAGACCC CGCCATGCGC
  	AACCTGCAAG
  251	AGCAGGATGC CGTCTACATC GCCAAGCAGA AACAGGCAAA
  	AGCCTCCCCG
  301	TTCAAAACCG AAATCGAAAC CGCCTTGGAA GAAAGCGGCA
  	TTATCGGCAA
  351	CTCCGCCCAC ACCGTTCCCG AACCCCAAAC CGGACATTCC
  	GCACCAAAAC
  401	CTGCCGACGC GCCGGCAAAA CCTGTTCCCG TTCCGCAAAC
  	GCCGGCAAAA
  451	CCGCTGATTA CGCTCAAAGA GCTGTCGAAG GTCGAGCTGC
  	CCTGGTTTGA
  501	CGTGCGCTTC GACTTCATCT CTTATATCGC GCTGACCGAA
  	GCCAAAGAAC
  551	TGCACGCACT GCCGCGCCTT TCCAACCGCT GCCGCTACCA
  	GATTGTCGGC
  601	TGCACCATGG ACGACCATTT CCAGATTGCC GAACCCATCC
  	CGGGCATCCG
  651	CTATCAGGCA TTTATCGTGG GTATTCAGGC AGTCAGCCGC
  	AACGGACTTG
  701	CCTCGCAGGA AGAACTCTCC GCATTCAACC GCCAGGTGGA
  	TGCATTCGCA
  751	CACAGCATGG GCGGTCAGAC GCTGCACACC GACCTTGCCG
  	CCTTTATCGA
  801	AGTGGCTTCC GCACTGGACG CATTCTGCGC GCGCGTCGAC
  	CAGACTATCG
  851	CCATCCATTT GGTTTCCCCG ACCAGCATCA GCGGCGTAGA
  	ACTGCGTTCC
  901	GCCGTAACGG GCGTGGGTTT CGTTTTGGAA GACGACGGCG
  	CGTTCCACTA
  951	TACCGACACG TCGGGCTCGA CCATGTTCTC CATCTGCTCG
  	CTCAACAACG
  1001	AGCCGTTTAC CAATGCCCTT TTGGACAACC AGTCCTATAA
  	AGGCTTCAGT
  1051	ATGCTGCTCG ACATCCCGCA CTCTCCGGCA GGCGAAAAAA
  	CCTTCGACGA
  1101	TTTGTTTATG GATTTGGCGG TACGCCTGTC CGGCCAGTTG
  	AACCTGAATC
  1151	TGGTCAACGA CAAAATGGAA GAAGTTTCGA CCCAATGGCT
  	CAAAGACGTG
  1201	CGCACTTATG TATTGGCTCG TCAGTCCGAG ATGCTCAAAG
  	TCGGTATCGA
  1251	ACCGGGCGGC AAAACCGCAT TGCGCCTGTT CTCCTAA

• This encodes a protein having amino acid sequence <SEQ ID 528>:

• 1	MIYIVLFLAA VLAVVAYNMY QENQYRKKVR DQFGHSDKDA
  	LLNSKTSHVR
  51	DGKPSGGPVM MPKPQPAVKK TAKSQDPAMR NLQEQDAVYI
  	AKQKQAKASP
  101	FKTEIETALE ESGIIGNSAH TVPEPQTGHS APKPADAPAK
  	PVPVPQTPAK
  151	PLITLKELSK VELPWFDVRF DFISYIALTE AKELHALPRL
  	SNRCRYQIVG
  201	CTMDDHFQIA EPIPGIRYQA FIVGIQAVSR NGLASQEELS
  	AFNRQVDAFA
  251	HSMGGQTLHT DLAAFIEVAS ALDAFCARVD QTIAIHLVSP
  	TSISGVELRS
  301	AVTGVGFVLE DDGAFHYTDT SGSTMFSICS LNNEPFTNAL
  	LDNQSYKGFS
  351	MLLDIPHSPA GEKTFDDLFM DLAVRLSGQL NLNLVNDKME
  	EVSTQWLKDV
  401	RTYVLARQSE MLKVGIEPGG KTALRLFS*

• ORF119a and ORF119-1 show 98.6% identity in 428 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF119 shows 93.1% identity over a 175aa overlap with a predicted ORF (ORF119ng) from N. gonorrhoeae:
• 
• The complete length ORF119ng nucleotide sequence <SEQ ID 529> is:

• 1	ATGATTTACA TCGTACTGTT CCTCGCCGCC GTCCTCGCCG
  	TTGTCGCCTA
  51	CAATATGTAT CAGGAAAACC AATACCGCAA AAAAGTGCGC
  	GACCAGTTCG
  101	GACACTCCGA CAAAGATGCC CTGCTCAACA GCAAAACCAG
  	CCATGTCCGC
  151	GACGGCAAAC CGTCCGGCGG GCCAGTCATG ATGCCGAAAC
  	CCCAACCGGC
  201	GGTCAAAAAA CCGGCCAAAC CCCAAGACTC CGCCATGCGC
  	AACCTGCAAG
  251	AACAGGATGC CGTCTACATC GCCAAGCAGA AACAGGCAAA
  	AGCCTCCCCG
  301	TTCAAAACCG AAATCGAAAC CGCCTTGGAA GAAATCGGCA
  	TTATCGGCAA
  351	CTCCGCCCAC ACCGTTTCCG AACCCCAAAC CGGACATTCC
  	GCACCGAAAC
  401	CTGCCGACGC GCCGGCAAAA CCCGTTCCCG TTCCGCAAAC
  	GCCGGCAAAA
  451	CCGCTGATTA CGCTCAAAGA GCTGTCGAAG GTCGAGCTGC
  	CCTGGTTTGA
  501	CGTGCGCTtc gACTTCATCT CCTATATCGC GCTGACCGAA
  	GCCAAAGAAC
  551	TGCACGCACT GCCGCGCCTT tccAACCGCT GCCGCTACCA
  	GATTGTCGGC
  601	TGCACCATGG ACGACCATTT CCAGATTGCC GAACCCATCC
  	CGGGCATCCG
  651	CTATCAGGCA TTTATCGTGG GTATCCAGGC AGTCAGCCGC
  	AACGGACTTG
  701	CCTCGCAGGA AGAACTCTCC GCATTCAACC GCCAGGCGGA
  	CGCATTCGCA
  751	CAAAGCATGG GCGGTCAGAC GCTGCACACC GACCTTGCCG
  	CCTTTATCGA
  801	AGTGGCTTCC GCACTGGACG CATTCTGCGC GCGCGTCGAC
  	CAGACCATCG
  851	CCATCCATTT GGTTTCGCCG ACCAGCATCA GCGGCGTAGA
  	ACTGCGTTCC
  901	GCCGTAACGG GCGTGGGTTT CGTTTTGGAA GACGACGGCG
  	CGTTCCACTA
  951	TACCGACACG TCGGGCTCGA CCATGTTCTC CATCTGCTCG
  	CTCAACAACG
  1001	AGCCGTTTAC CAATGCCCTT TTGGACAACC AGTCCTACAA
  	AGGCTTCAGT
  1051	ATGCTGCTCG ACATCCCGCA CTCTCCGGCA GGCGAAAAAA
  	CCTTCGACGA
  1101	TTTGTTTATG GATTTGGCGG TACGCCTGTC CGGTCAGTTG
  	AACCTGAATC
  1151	TGGTCAACGA CAAAATGGAA GAAGTTTCGA CCCAATGGCT
  	CAAAGACGTA
  1201	CGCACTTATG TATTGGCGCG TCAGTCCGAG ATGCTCAAAG
  	TCGGTATCGA
  1251	ACCGGGCGGC AAAACCGCCC TGCGCCTGTT TTCATAA

• This encodes a protein having amino acid sequence <SEQ ID 530>:

• 1	MIYIVLFLAA VLAVVAYNMY QENQYRKKVR DQFGHSDKDA
  	LLNSKTSHVR
  51	DGKPSGGPVM MPKPQPAVKK PAKPQDSAMR NLQEQDAVYI
  	AKQKQAKASP
  101	FKTEIETALE EIGIIGNSAH TVSEPQTGHS APKPADAPAK
  	PVPVPQTPAK
  151	PLITLKELSK VELPWFDVRF DFISYIALTE AKELHALPRL
  	SNRCRYQIVG
  201	CTMDDHFQIA EPIPGIRYQA FIVGIQAVSR NGLASQEELS
  	AFNRQADAFA
  251	QSMGGQTLHT DLAAFIEVAS ALDAFCARVD QTIAIHLVSP
  	TSISGVELRS
  301	AVTGVGFVLE DDGAFHYTDT SGSTMFSICS LNNEPFTNAL
  	LDNQSYKGFS
  351	MLLDIPHSPA GEKTFDDLFM DLAVRLSGQL NLNLVNDKME
  	EVSTQWLKDV
  401	RTYVLARQSE MLKVGIEPGG KTALRLFS*

• ORF119ng and ORF119-1 show 98.4% identity over 428 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 64
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 531>

• 1	..GCGCGGCACG GCACGGAAGA TTTCTTCATG AACAACAGCG
  	ACAC.ATCAG
  51	GCAGATAGTC GAAAGCACCA CCGGTACGAT GAAGCTGCTG
  	ATTTCCTCCA
  101	TCGCCCTGAT TTCATTGGTA GTCGGCGGCA TCGGCGTGAT
  	GAACATCATG
  151	CTGGTGTCCG TTACCGAGCG CACCAAAGAA ATCGGCATAC
  	GGATGGCAAT
  201	CGGCGCGCGG CGCGGCAATA TTTyGCAGCA GTTTTTGATT
  	GAGGCGGTGT
  251	TAATCTGCGT CATCGGCGGT TTGGTCGGCG TGGGTTTGTC
  	CGCCGCCGTC
  301	AGCCTCGTGT TCAATCATTT TGTAACCGAC TTCCCGATGG
  	ACATTTCCGC
  351	CATGTCCGTC ATCGGCGCGG TCGCCTGTTC GACCGGAATC
  	GGCATCGCGT
  401	TCGGCTTTAT GCCTGCCAAT AAAGCAGCCA AACTCAATCC
  	GATAGACGCA
  451	TTGGCACAGG ATTGA

• This corresponds to the amino acid sequence <SEQ ID 532; ORF134>:

• 1	..ARHGTEDFFM NNSDXIRQIV ESTTGTMKLL ISSIALISLV
  	VGGIGVMNIM
  51	LVSVTERTKE IGIRMAIGAR RGNIXQQFLI EAVLICVIGG
  	LVGVGLSAAV
  101	SLVFNHFVTD FPMDISAMSV IGAVACSTGI GIAFGFMPAN
  	KAAKLNPIDA
  151	LAQD*

• Further work revealed the complete nucleotide sequence <SEQ ID 533>:

• 1	ATGTCGGTGC AAGCAGTATT GGCGCACAAA ATGCGTTCGC
  	TTCTGACGAT
  51	GCTCGGCATC ATCATCGGTA TCGCGTCGGT GGTTTCCGTC
  	GTCGCATTGG
  101	GCAATGGTTC GCAGAAAAAA ATCCTTGAAG ACATCAGTTC
  	GATAGGGACG
  151	AACACCATCA GCATCTTCCC GGGGCGCGGC TTCGGCGACA
  	GGCGCAGCGG
  201	CAGGATTAAA ACCCTGACCA TAGACGACGC AAAAATCATC
  	GCCAAACAAA
  251	GCTACGTTGC TTCCGCCACG CCCATGACTT CGAGCGGCGG
  	CACGCTGACT
  301	TACCGCAACA CCGACCTGAC CGCCTCGCTT TACGGCGTGG
  	GCGAACAATA
  351	TTTCGACGTG CGCGGACTGA AGCTGGAAAC GGGGCGGCTG
  	TTTGACGAAA
  401	ACGATGTGAA AGAAGACGCG CAGGTCGTCG TCATCGACCA
  	AAATGTCAAA
  451	GACAAACTCT TTGCGGACTC GGATCCGTTG GGTAAAACCA
  	TTTTGTTCAG
  501	GAAACGCCCC TTGACCGTCA TCGGCGTGAT GAAAAAAGAC
  	GAAAACGCTT
  551	TCGGCAATTC CGACGTGCTG ATGCTTTGGT CGCCCTATAC
  	GACGGTGATG
  601	CACCAAATCA CAGGCGAGAG CCACACCAAC TCCATCACCG
  	TCAAAATCAA
  651	AGACAATGCC AATACCCAGG TTGCCGAAAA AGGGCTGACC
  	GATCTGCTCA
  701	AAGCGCGGCA CGGCACGGAA GATTTCTTCA TGAACAACAG
  	CGACAGCATC
  751	AGGCAGATAG TCGAAAGCAC CACCGGTACG ATGAAGCTGC
  	TGATTTCCTC
  801	CATCGCCCTG ATTTCATTGG TAGTCGGCGG CATCGGCGTG
  	ATGAACATCA
  851	TGCTGGTGTC CGTTACCGAG CGCACCAAAG AAATCGGCAT
  	ACGGATGGCA
  901	ATCGGCGCGC GGCGCGGCAA TATTTTGCAG CAGTTTTTGA
  	TTGAGGCGGT
  951	GTTAATCTGC GTCATCGGCG GTTTGGTCGG CGTGGGTTTG
  	TCCGCCGCCG
  1001	TCAGCCTCGT GTTCAATCAT TTTGTAACCG ACTTCCCGAT
  	GGACATTTCC
  1051	GCCATGTCCG TCATCGGCGC GGTCGCCTGT TCGACCGGAA
  	TCGGCATCGC
  1101	GTTCGGCTTT ATGCCTGCCA ATAAAGCAGC CAAACTCAAT
  	CCGATAGACG
  1151	CATTGGCACA GGATTGA

• This corresponds to the amino acid sequence <SEQ ID 534; ORF134-1>:

• 1	MSVQAVLAHK MRSLLTMLGI IIGIASVVSV VALGNGSQKK
  	ILEDISSIGT
  51	NTISIFPGRG FGDRRSGRIK TLTIDDAKII AKQSYVASAT
  	PMTSSGGTLT
  101	YRNTDLTASL YGVGEQYFDV RGLKLETGRL FDENDVKEDA
  	QVVVIDQNVK
  151	DKLFADSDPL GKTILFRKRP LTVIGVMKKD ENAFGNSDVL
  	MLWSPYTTVM
  201	HQITGESHTN SITVKIKDNA NTQVAEKGLT DLLKARHGTE
  	DFFMNNSDSI
  251	RQIVESTTGT MKLLISSIAL ISLVVGGIGV MNIMLVSVTE
  	RTKEIGIRMA
  301	IGARRGNILQ QFLIEAVLIC VIGGLVGVGL SAAVSLVFNH
  	FVTDFPMDIS
  351	AMSVIGAVAC STGIGIAFGF MPANKAAKLN PIDALAQD*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with the Hypothetical Protein o648 of E. coli (Accession Number AE000189)
• ORF134 and o648 protein show 45% aa identity in 153aa overlap:

• Orf134:   2 RHGTEDFFMNNSDXIRQIVESTTGTMKXXXXXXXXXXXVVGGIGVMNIMLVSVTERTKEI  61
  RHG +DFFN   D + + VE TT T++            VVGGIGVMNIMLVSVTERT+EI
  o648:   496 RHGKKDFFTWNMDGVLKTVEKTTRTLQLFLTLVAVISLVVGGIGVMNIMLVSVTERTREI 555
  Orf134:  62 GIRMAIGARRGNIXQQFLIEAXXXXXXXXXXXXXXXXXXXXXFNHFVTDFPMDISAMSVI 121
  GIRMA+GAR  ++ QQFLIEA                        F+ + +   S ++++
  o648:   556 GIRMAVGARASDVLQQFLIEAVLVCLVGGALGITLSLLIAFTLQLFLPGWEIGFSPLALL 615
  Orf134: 122 GAVACSTGIGIAFGFMPANKAAKLNPIDALAQD                            154
  A  CST  GI FG++PA  AA+L+P+DALA++
  o648:   616 LAFLCSTVTGILFGWLPARNAARLDPVDALARE                            648

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF134 shows 98.7% identity over a 154aa overlap with an ORF (ORF134a) from strain A of N. meningitidis:
• 
• The complete length ORF134a nucleotide sequence <SEQ ID 535> is:

• 1	ATGTCGGTGC AAGCAGTATT GGCGCACAAA ATGCGTTCGC
  	TTCTGACGAT
  51	GCTCGGCATC ATCATCGGTA TCGCTTCGGT TGTCTCCGTC
  	GTCGCATTGG
  101	GCAACGGTTC GCAGAAAAAA ATCCTTGAAG ACATCAGTTC
  	GATAGGGACG
  151	AACACCATCA GCATCTTCCC AGGGCGCGGC TTCGGCGACA
  	GGCGCAGCGG
  201	CAGGATTAAA ACCCTGACCA TAGACGACGC AAAAATCATC
  	GCCAAACAAA
  251	GCTACGTTGC TTCCGCCACG CCCATGACTT CGAGCGGCGG
  	CACGCTGACT
  301	TACCGCAATA CCGACCTGAC CGCTTCTTTG TACGGTGTGG
  	GCGAACAATA
  351	TTTCGACGTG CGCGGGCTGA AGCTGGAAAC GGGGCGGCTG
  	TTTGACGAAA
  401	ACGATGTGAA AGAAGACGCG CAGGTCGTCG TCATCGACCA
  	AAATGTCAAA
  451	GACAAACTCT TTGCGGACTC GGATCCGTTG GGTAAAACCA
  	TTTTGTTCAG
  501	GAAACGCCCC TTGACCGTCA TCGGCGTGAT GAAAAAAGAC
  	GAAAACGCTT
  551	TCGGCAATTC CGACGTGCTG ATGCTTTGGT CGCCCTATAC
  	GACGGTGATG
  601	CACCAAATCA CAGGCGAGAG CCACACCAAC TCCATCACCG
  	TCAAAATCAA
  651	AGACAATGCC AATACCCAGG TTGCCGAAAA AGGGCTGACC
  	GATCTGCTCA
  701	AAGCGCGGCA CGGCACGGAA GATTTCTTCA TGAACAACAG
  	CGACAGCATC
  751	AGGCAGATAG TCGAAAGCAC CACCGGTACG ATGAAGCTGC
  	TGATTTCCTC
  801	CATCGCCCTG ATTTCATTGG TAGTCGGCGG CATCGGCGTG
  	ATGAACATCA
  851	TGCTGGTGTC CGTTACCGAG CGCACCAAAG AAATCGGCAT
  	ACGGATGGCA
  901	ATCGGCGCGC GGCGCGGCAA TATTTTGCAG CAGTTTTTGA
  	TTGAGGCGGT
  951	GTTAATCTGC GTCATCGGCG GTTTGGTCGG CGTGGGTTTG
  	TCCGCCGCCG
  1001	TCAGCCTCGT GTTCAATCAT TTTGTAACCG ACTTCCCGAT
  	GGACATTTCC
  1051	GCCATGTCCG TCATCGGCGC GGTCGCCTGT TCGACCGGAA
  	TCGGCATCGC
  1101	GTTCGGCTTT ATGCCTGCCA ATAAAGCAGC CAAACTCAAT
  	CCGATAGATG
  1151	CATTGGCGCA GGATTGA

• This encodes a protein having amino acid sequence <SEQ ID 536>:

• 1	MSVQAVLAHK MRSLLTMLGI IIGIASVVSV VALGNGSQKK
  	ILEDISSIGT
  51	NTISIFPGRG FGDRRSGRIK TLTIDDAKII AKQSYVASAT
  	PMTSSGGTLT
  101	YRNTDLTASL YGVGEQYFDV RGLKLETGRL FDENDVKEDA
  	QVVVIDQNVK
  151	DKLFADSDPL GKTILFRKRP LTVIGVMKKD ENAFGNSDVL
  	MLWSPYTTVM
  201	HQITGESHTN SITVKIKDNA NTQVAEKGLT DLLKARHGTE
  	DFFMNNSDSI
  251	RQIVESTTGT MKLLISSIAL ISLVVGGIGV MNIMLVSVTE
  	RTKEIGIRMA
  301	IGARRGNILQ QFLIEAVLIC VIGGLVGVGL SAAVSLVFNH
  	FVTDFPMDIS
  351	AMSVIGAVAC STGIGIAFGF MPANKAAKLN PIDALAQD*

• ORF134a and ORF134-1 show 100.0% identity in 388 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF134 shows 96.8% identity over a 154aa overlap with a predicted ORF (ORF 134.ng) from N. gonorrhoeae:
• 
• The complete length ORF134ng nucleotide sequence <SEQ ID 537> is:

• 1	ATGTCGGTGC AAGCAGTATT GGCGCACAAA ATGCGTTCGC
  	TTCTGACCAT
  51	GCTCGGCATC ATCATCGGTA TCGCTTCGGT TGTCTCCGTC
  	GTCGCGCTGG
  101	GCAACGGTTC GCAGAAAAAA ATCCTCGAAG ACATCAGTTC
  	GATGGGGACG
  151	AACACCATCA GCATCTTCCC CGGGCGCGGC TTCGGCGACA
  	GGCGCAGCGG
  201	CAAAATCAAA ACCCTGACCA TAGACGACGC AAAAATCATC
  	GCCAAACAAA
  251	GCTACGTTGC CTCCGCCACG CCCATGACTT CGAGCGGCGG
  	CACGCTGACC
  301	TACCGCAATA CCGACCTGAC CGCTTCTTTG TACGGTGTGG
  	GCGAACAATA
  351	TTTCGACGTG CGCGGGCTGA AGCTGGAAAC GGGGCGGCTG
  	TTTGATGAGA
  401	ACGATGTGAA AGAAGACGCG CAAGTCGTCG TCATCGACCA
  	AAATGTCAAA
  451	GACAAACTCT TTGCGGACTC GGATCCGTTG GGTAAAACCA
  	TTTTGTTCAG
  501	GAAACGCCCC TTGACCGTCA TCGGCGTGAT GAAAAAAGAC
  	GAAAACGCTT
  551	TCGGCAATTC CGACGTGCTG ATGCTTTGGT CGCCCTATAC
  	GACGGTGATG
  601	CACCAAATCA CAGGCGAGAG CCACACCAAC TCCATCACCG
  	TCAAAATCAA
  651	AGACAATGCC AATACCCGGG TTGCCGAAAA AGGGCTGGCC
  	GAGCTGCTCA
  701	AAGCACGGCA CGGCACGGAA GACTTCTTTA TGAACAACAG
  	CGACAGCATC
  751	AGGCAGATGG TCGAAAGCAC CACCGGTACG ATGAAGCTGC
  	TGATTTCCTC
  801	CATCGCCCTG ATTTCATTGG TAGTCGGCGG CATCGGTGTG
  	ATGAACATTA
  851	TGCTGGTGTC CGTTACCGAG CGCACCAAAG AAATCGGCAT
  	ACGGATGGCA
  901	ATCGGCGCGC GGCGCGGCAA TATTTTGCAG CAGTTTTTGA
  	TTGAGGCGGT
  951	GTTAATCTGC ATCATCGGAG GCTTGGTCGG CGTAGGTTTG
  	TCCGCCGCCG
  1001	TCAGCCTCGT GTTCAATCAT TTTGTAACCG ATTTCCCGAT
  	GGACATTTCG
  1051	GCGGCATCCG TTATCGGGGC GGTCGCCTGT TCGACCGGAA
  	TCGGCATCGC
  1101	GTTCGGCTTT ATGCCTGCCA ATAAGGCAGC CAAACTCAAT
  	CCGATAGATG
  1151	CATTGGCGCA GGATTGA

• This encodes a protein having amino acid sequence <SEQ ID 538>:

• 1	MSVQAVLAHK MRSLLTMLGI IIGIASVVSV VALGNGSQKK
  	ILEDISSMGT
  51	NTISIFPGRG FGDRRSGKIK TLTIDDAKII AKQSYVASAT
  	PMTSSGGTLT
  101	YRNTDLTASL YGVGEQYFDV RGLKLETGRL FDENDVKEDA
  	QVVVIDQNVK
  151	DKLFADSDPL GKTILFRKRP LTVIGVMKKD ENAFGNSDVL
  	MLWSPYTTVM
  201	HQITGESHTN SITVKIKDNA NTRVAEKGLA ELLKARHGTE
  	DFFMNNSDSI
  251	RQMVESTTGT MKLLISSIAL ISLVVGGIGV MNIMLVSVTE
  	RTKEIGIRMA
  301	IGARRGNILQ QFLIEAVLIC IIGGLVGVGL SAAVSLVFNH
  	FVTDFPMDIS
  351	AASVIGAVAC STGIGIAFGF MPANKAAKLN PIDALAQD*

• ORF134ng and ORF134-1 show 97.9% identity in 388 aa overlap:
• 
• ORF134ng also shows homology to an E. coli ABC transporter:

• sp|P75831|YBJZ_ECOLI HYPOTHETICAL ABC TRANSPORTER ATP-BINDING
  PROTEIN YBJZ >gi5 (AE000189) o648; similar to YBBA_HAEIN SW: P45247
  [Escherichia coli] Length = 648
  Score = 297 bits (753), Expect = 6e−80
  Identities = 162/389 (41%), Positives = 230/389 (58%), Gaps = 1/389 (0%)

  Query:	1	MSVQAVLAHKMRSLLTMLXXXXXXXXXXXXXXLGNGSQKKILEDISSMGTNTISIFPGRG	60
  		M+ +A+ A+KMR+LLTML              +G+ +++ +L DI S+GTNTI ++PG+
  Sbjct:	260	MAWRALAANKMRTLLTMLGIIIGIASVVSIVVVGDAAKQMVLADIRSIGTNTIDVYPGKD	319
  Query:	61	FGDRRSGKIKTLTIDDAKIIAKQSYVASATPMTSSGGTLTYRNTDLTASLYGVGEQYFDV	120
  		FGD      + L  DD   I KQ +VASATP  S    L Y N D+ AS  GV   YF+V
  Sbjct:	320	FGDDDPQYQQALKYDDLIAIQKQPWVASATPAVSQNLRLRYNNVDVAASANGVSGDYFNV	379
  Query:	121	RGLKLETGRLFDENDVKEDAQVVVIDQNVKDKLFAD-SDPLGKTILFRKRPLTVIGVMKK	179
  		G+    G  F++  +   AQVVV+D N + +LF   +D +G+ IL    P  VIGV ++
  Sbjct:	380	YGMTFSEGNTFNQEQLNGRAQVVVLDSNTRRQLFPHKADVVGEVILVGNMPARVIGVAEE	439
  Query:	180	DENAFGNSDVLMLWSPYTTVMHQITGESHTNSITVKIKDNANTRVAEKGLAELLKARHGT	239
  		++ FG+S VL +W PY+T+  ++ G+S  NSITV++K+  ++  AE+ L  LL  REG
  Sbjct:	440	KQSMFGSSKVLRVWLPYSTMSGRVMGQSWLNSITVRVKEGFDSAEAEQQLTRLLSLRHGK	499
  Query:	240	EDFFMNNSDSIRQMVESTTGTMKXXXXXXXXXXXVVGGIGVMNIMLVSVTERTKEIGIRM	299
  		+DFF  N D + + VE TT T++           VVGGIGVMNIMLVSVTERT+EIGIRM
  Sbjct:	500	KDFFTWNMDGVLKTVEKTTRTLQLFLTLVAVISLVVGGIGVMNIMLVSVTERTREIGIRM	559
  Query:	300	AIGARRGNILQQFLIEXXXXXXXXXXXXXXXXXXXXXXFNHFVTDFPMDISAASVIGAVA	359
  		A+GAR  ++LQQFLIE                         F+  + +  S  +++ A
  Sbjct:	560	AVGARASDVLQQFLIEAVLVCLVGGALGITLSLLIAFTLQLFLPGWEIGFSPLALLLAFL	619
  Query:	360	CSTGIGIAFGFMPANKAAKLNPIDALAQD	388
  		CST  GI FG++PA  AA+L+P+DALA++
  Sbjct:	620	CSTVTGILFGWLPARNAARLDPVDALARE	648

• Based on this analysis, including the presence of the leader peptide and transmembrane regions in the gonococcal protein, it is prediceted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 65
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 539>:

• 1	..GGGACGGGAG CGATGCTGCT GCTGTTTTAC GCGGTAACGA
  	T.CTGCCTTT
  51	GGCCACTGGC GTTACCCTGA GTTACACCTC GTCGATTTTT
  	TTGGCGGTAT
  101	TTTCCTTCCT GATTTTGAAA GAACGGATTT CCGTTTACAC
  	GCAGGCGGTG
  151	CTGCTCCTTG GTTTTGCCGG CGTGGTATTG CTGCTTAATC
  	CCTCGTTCCG
  201	CAGCGGTCAG GAAACGGCGG CACTCGCCGG GCTGGCGGGC
  	GGCGCGATGT
  251	CCGGCTGGGC GTATTTGAAA GTGCGCGAAC TGTCTTTGGC
  	GGGCGAACCC
  301	GGCTGGCGCG TCGTGTTTTA CCTTTCCGTG ACAGGTGTGG
  	CGATGTCGTC
  351	GGTTTGGGCG ACGCTGACCG GCTGGCACAC CCTGTCCTTT
  	CCATCGGCAG
  401	TTTATCTGTC GTGCATCGGC GTGTCCGCGC TGATTGCCCA
  	ACTGTCGATG
  451	ACGCGCGCCT ACAAAGTCGG CGACAAATTC ACGGTTGCCT
  	CGCTTTCCTA
  501	TATGACCGTC GTTTTTTCCG CTCTGTCTGC CGCATTTTTT
  	CTGGGCGAAG
  551	AGCTTTTCTG GCAGGAAATA CTCGGTATGT GCATCATCAT
  	CCTCAGCGGT
  601	ATTTTGA

• This corresponds to the amino acid sequence <SEQ ID 540; ORF135>:

• 1	..GTGAMLLLFY AVTILPLATG VTLSYTSSIF LAVFSFLILK
  	ERISVYTQAV
  51	LLLGFAGVVL LLNPSFRSGQ ETAALAGLAG GAMSGWAYLK
  	VRELSLAGEP
  101	GWRVVFYLSV TGVAMSSVWA TLTGWHTLSF PSAVYLSCIG
  	VSALIAQLSM
  151	TRAYKVGDKF TVASLSYMTV VFSALSAAFF LGEELFWQEI
  	LGMCIIISAV
  201	F*

• Further work revealed the complete nucleotide sequence <SEQ ID 541>:

• 1	ATGGATACCG CAAAAAAAGA CATTTTAGGA TCGGGCTGGA
  	TGCTGGTGGC
  51	GGCGGCCTGC TTTACCATTA TGAACGTATT GATTAAAGAG
  	GCATCGGCAA
  101	AATTTGCCCT CGGCAGCGGC GAATTGGTCT TTTGGCGCAT
  	GCTGTTTTCA
  151	ACCGTTGCGC TCGGGGCTGC CGCCGTATTG CGTCGGGACA
  	mCTTCCGCAC
  201	GCCCCATTGG AAAAACCACT TAAACCGCAG TATGGTCGGG
  	ACGGGGGCGA
  251	TGCTGCTGCT GTTTTACGCG GTAACGCATC TGCCTTTGGC
  	CACTGGCGTT
  301	ACCCTGAGTT ACACCTCGTC GATTTTTTTG GCGGTATTTT
  	CCTTCCTGAT
  351	TTTGAAAGAA CGGATTTCCG TTTACACGCA GGCGGTGCTG
  	CTCCTTGGTT
  401	TTGCCGGCGT GGTATTGCTG CTTAATCCCT CGTTCCGCAG
  	CGGTCAGGAA
  451	ACGGCGGCAC TCGCCGGGCT GGCGGGCGGC GCGATGTCCG
  	GCTGGGCGTA
  501	TTTGAAAGTG CGCGAACTGT CTTTGGCGGG CGAACCCGGC
  	TGGCGCGTCG
  551	TGTTTTACCT TTCCGTGACA GGTGTGGCGA TGTCGTCGGT
  	TTGGGCGACG
  601	CTGACCGGCT GGCACACCCT GTCCTTTCCA TCGGCAGTTT
  	ATCTGTCGTG
  651	CATCGGCGTG TCCGCGCTGA TTGCCCAACT GTCGATGACG
  	CGCGCCTACA
  701	AAGTCGGCGA CAAATTCACG GTTGCCTCGC TTTCCTATAT
  	GACCGTCGTT
  751	TTTTCCGCTC TGTCTGCCGC ATTTTTTCTG GGCGAAGAGC
  	TTTTCTGGCA
  801	GGAAATACTC GGTATGTGCA TCATCATCCT CAGCGGTATT
  	TTGAGCAGCA
  851	TCCGCCCCAC TGCCTTCAAA CAGCGGCTGC AATCCCTGTT
  	CCGCCAAAGA
  901	TAA

• This corresponds to the amino acid sequence <SEQ ID 542; ORF135-1>:

• 1	MDTAKKDILG SGWMLVAAAC FTIMNVLIKE ASAKFALGSG
  	ELVFWRMLFS
  51	TVALGAAAVL RRDXFRTPHW KNHLNRSMVG TGAMLLLFYA
  	VTHLPLATGV
  101	TLSYTSSIFL AVFSFLILKE RISVYTQAVL LLGFAGVVLL
  	LNPSFRSGQE
  151	TAALAGLAGG AMSGWAYLKV RELSLAGEPG WRVVFYLSVT
  	GVAMSSVWAT
  201	LTGWHTLSFP SAVYLSCIGV SALIAQLSMT RAYKVGDKFT
  	VASLSYMTVV
  251	FSALSAAFFL GEELFWQEIL GMCIIILSGI LSSIRPTAFK
  	QRLQSLFRQR
  301	*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF135 shows 99.0% identity over a 197aa overlap with an ORF (ORF135a) from strain A of N. meningitidis:
• 
• The complete length ORF135a nucleotide sequence <SEQ ID 543> is:

• 1	ATGGATACCG CAAAAAAAGA CATTTTAGGA TCGGGCTGGA
  	TGCTGGTGGC
  51	GGCGGCCTGC TTTACCATTA TGAACGTATT GATTAAAGAG
  	GCATCGGCAA
  101	AATTTGCCCT CGGCAGCGGC GAATTGGTCT TTTGGCGCAT
  	GCTGTTTTCA
  151	ACCGTTGCGC TCGGGGCTGC CGCCGTATTG CGTCGGGACA
  	CCTTCCGCAC
  201	GCCCCATTGG AAAAACCACT TAAACCGCAG TATGGTCGGG
  	ACGGGGGCGA
  251	TGCTGCTGCT GTTTTACGCG GTAACGCATC TGCCTTTGGC
  	CACCGGCGTT
  301	ACCCTGAGTT ACACCTCGTC GATTTTTTTG GCGGTATTTT
  	CCTTCCTGAT
  351	TTTGAAAGAA CGGATTTCCG TTTACACGCA GGCGGTGCTG
  	CTCCTTGGTT
  401	TTGCCGGCGT GGTATTGCTG CTTAATCCCT CGTTCCGCAG
  	CGGTCAGGAA
  451	ACGGCGGCAC TCGCCGGGCT GGCGGGCGGC GCGATGTCCG
  	GCTGGGCGTA
  501	TTTGAAAGTG CGCGAACTGT CTTTGGCGGG CGAACCCGGC
  	TGGCGCGTCG
  551	TGTTTTACCT TTCCGTGACA GGTGTGGCGA TGTCATCGGT
  	TTGGGCGACG
  601	CTGACCGGCT GGCACACCCT GTCCTTTCCA TCGGCAGTTT
  	ATCTGTCGTG
  651	CATCGGCGTG TCCGCGCTGA TTGCCCAACT GTCGATGACG
  	CGCGCCTACA
  701	AAGTCGGCGA CAAATTCACG GTTGCCTCGC TTTCCTATAT
  	GACCGTCGTT
  751	TTTTCCGCTC TGTCTGCCGC ATTTTTTCTG GCCGAAGAGC
  	TTTTCTGGCA
  801	GGAAATACTC GGTATGTGCA TCATCATCCT CAGCGGTATT
  	TTGAGCAGCA
  851	TCCGCCCCAC TGCCTTCAAA CAGCGGCTGC AATCCCTGTT
  	CCGCCAAAGA
  901	TAA

• This encodes a protein having amino acid sequence <SEQ ID 544>:

• 1	MDTAKKDILG SGWMLVAAAC FTIMNVLIKE ASAKFALGSG
  	ELVFWRMLFS
  51	TVALGAAAVL RRDTFRTPHW KNHLNRSMVG TGAMLLLFYA
  	VTHLPLATGV
  101	TLSYTSSIFL AVFSFLILKE RISVYTQAVL LLGFAGVVLL
  	LNPSFRSGQE
  151	TAALAGLAGG AMSGWAYLKV RELSLAGEPG WRVVFYLSVT
  	GVAMSSVWAT
  201	LTGWHTLSFP SAVYLSCIGV SALIAQLSMT RAYKVGDKFT
  	VASLSYMTVV
  251	FSALSAAFFL AEELFWQEIL GMCIIILSGI LSSIRPTAFK
  	QRLQSLFRQR
  301	*

• ORF135a and ORF135-1 show 99.3% identity in 300 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF135 shows 97% identity over a 201aa overlap with a predicted ORF (ORF135ng) from N. gonorrhoeae:
• 
• An ORF135ng nucleotide sequence <SEQ ID 545> was predicted to encode a protein having amino acid sequence <SEQ ID 546>:

• 1	MPSEKAFRRH LRTASFQGLH LHHFHQKVGK CGIIGFGIHI
  	FPTLLPAAQG
  51	ILDIQLGLFR IDFAALAVYR RTQVDFIHTV IDGIASDQAF
  	SEVVQILRRL
  101	NLGHFTDTHL IAQARRFIAD FGNIRPMRRG EAKTFCRCFR
  	FDGIDGIHGD
  151	FRQCGHINRL APGKDCRNGK RDKVFFHTRH YNQVCLEKTN
  	CSARKIKFRH
  201	QKQAKTHSTS LAARFTIRPS LSQRPFMDTA KKDILGSGWM
  	LVAAACFTVM
  251	NVLIKEASAK FALGSGELVF WRMLFSTVTL GAAAVLRRDT
  	FRTPHWKNHL
  301	NRSMVGTGAM LLLFYAVTHL PLTTGVTLSY TSSIFLAVFS
  	FLILKERISV
  351	YTQAVLLLGF AGVVLLLNPS FRSGQEPAAL AGLAGGAMSG
  	WAYLKVRELS
  401	LAGEPGWRVV FYLSATGVAM SSVWATLTGW HTLSFPSAVY
  	LSGIGVSALI
  451	AQLSMTRAYK VGDKFTVASL SYMTVVFSAL SAAFFLGEEL
  	FWQEILGMCI
  501	IISAAF*

• Further work revealed the following gonococcal sequence <SEQ ID 547>:

• 1	ATGGATACCG CAAAAAAAGA CATTTTAGGA TCGGGCTGGA
  	TGCTGGTGGC
  51	GGCGGCCTGC TTCACCGTTA TGAACGTATT GATTAAAGAG
  	GCATCGGCAA
  101	AATTTGCCCT CGGCAGCGGC GAATTGGTCT TTTGGCGCAT
  	GCTGTTTTCA
  151	ACCGTTACGC TCGGTGCTGC CGCCGTATTG CGGCGCGACA
  	CCTTCCGCAC
  201	GCCCCATTGG AAAAACCACT TAAACCGCAG TATGGTCGGG
  	ACGGGGGCGA
  251	TGCTGCTGCT GTTTTACGCG GTAACGCATC TGCCTTTGAC
  	AACCGGCGTT
  301	ACCCTGAGTT ACACCTCGTC GATTTTTttg GCGGTATTTT
  	CCTTCCTCAT
  351	TTTGAAAGAA CGGATTTCCG TTTACACGCA GGCGGTGCTG
  	CTCCTTGGTT
  401	TTGCCGGCGT GGTATTGCTG CTTAATCCCT CGTTCCGCAG
  	CGGTCAGGAA
  451	CCGGCGGCAC TCGCCGGGCT GGCGGGCGGC GCGATGTCCG
  	GCTGGGCGTA
  501	TTTGAAAGTG CGCGAACTGT CTTTGGCGGG CGAACCCGGC
  	TGGCGCGTCG
  551	TGTTTTACCT TTCCGCAACC GGCGTGGCGA TGTCGTCggt
  	ttgggcgacg
  601	Ctgaccggct ggCACAcccT GTCCTTTcca tcggcagttt
  	ATCtgtCGGG
  651	CATCGGCGTG tccgcgCtgA TTGCCCAaCT GtcgatgAcg
  	cGCGcctaca
  701	aaGTCGGCGA CAAATTCACG GTTGCCTCGC tttcctaTAt
  	gaccgtcGTC
  751	TTTTCCGCCC TGTCTGCCGC ATTTTTTCTg ggcgaagagc
  	tttTCtggCA
  801	GGAAATACTC GGTATGTGCA TCATTAtccT CAGCGGCATT
  	TTGAGCAGCA
  851	TCCGCCCCAT TGCCTTCAAA CAGCGGCTGC AAGCCCTCTT
  	CCGCCAAAGA
  901	TAA

• This corresponds to the amino acid sequence <SEQ ID 548; ORF135ng-1>:

• 1	MDTAKKDILG SGWMLVAAAC FTVMNVLIKE ASAKFALGSG
  	ELVFWRMLFS
  51	TVTLGAAAVL RRDTFRTPHW KNHLNRSMVG TGAMLLLFYA
  	VTHLPLTTGV
  101	TLSYTSSIFL AVFSFLILKE RISVYTQAVL LLGFAGVVLL
  	LNPSFRSGQE
  151	PAALAGLAGG AMSGWAYLKV RELSLAGEPG WRVVFYLSAT
  	GVAMSSVWAT
  201	LTGWHTLSFP SAVYLSGIGV SALIA QLSMT RAYKVGDKFT
  	VASLSYMTVV
  251	FSALSAAFFL GEELFWQEIL GMCIIILSGI LSSIRPIAFK
  	QRLQALFRQR
  301	*

• ORF135ng-1 and ORF135-1 show 97.0% identity in 300 aa overlap:
• 
• Based on this analysis, including the presence of several putative transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 66
• The following DNA sequence was identified in N. meningitidis <SEQ ID 549>:

• 1	ATGAAGCGGC GTATAGCCGT CTTCGTCCTG TTCCCGCAGA
  	TAATCCGAGT
  51	TTTGGGACAA CTGTTGCCGA AAATCGTCAA TACAGTTCCG
  	GCACATCGGA
  101	TGCTCTTCCA GATTTTCGGG ATGTTCTTTT TCTTCATACA
  	CCAGCAATAT
  151	CTGCCCGGGA TCGCCGAAAT CGATTCCCCA TGCGGCATCG
  	TGTTCGGTGC
  201	GCTCCTCTTC CGTCATCTGC CCGCGCATTG CCTGTATGGT
  	AAAGCCGCCG
  251	TAGGGGATGC CgTTGCACAC GAACATCCAG TCGCTGATGT
  	CGTCAACCGG
  301	AACGCAAACG cTTTCGCCTT GTTCGACATT GGTCAGTTCG
  	CCsGGTTCAT
  351	TGTTCAGCAC ACCGTAAATA TAAAGACCGT CAAAATAAAT
  	ATCGTCGATC
  401	CACATATGTT CGCAAATTTC GCCGTCTTCG CCGTCTTGGA
  	AAAAAGGGAC
  451	TTTGACCATG GCAAAATCCA AGGCGGAAAT AATGCGGCGG
  	CGTTCCCAAA
  501	AAAGcTCGCG CCAAAAATAT TTGAATGTTT TACGGGCGCG
  	TTCGTCGGCA
  551	CGGTTTACCG GTTCGTCTGC CTGTTCTACA TAATAAATGA
  	CGGAATCGCC
  601	CATCATATCT GCTCCTCAAC GTGTACGGTA TCTGTTTGCA
  	CCTTACTGCG
  651	GCTTTCTgcC kTCGGCATCC GATTCGGATT TGAAAAGTTC
  	mmrwyATTCG
  701	GAATAG

• This corresponds to the amino acid sequence <SEQ ID 550; ORF136>:

• 1	MKRRIAVFVL FPQIIRVLGQ LLPKIVNTVP AHRMLFQIFG
  	MFFFFIHQQY
  51	LPGIAEIDSP CGIVFGALLF RHLPAHCLYG KAAVGDAVAH
  	EHPVADVVNR
  101	NANAFALFDI GQFAXFIVQH TVNIKTVKIN IVDPHMFANF
  	AVFAVLEKRD
  151	FDHGKIQGGN NAAAFPKKLA PKIFECFTGA FVGTVYRFVC
  	LFYIINDGIA
  201	HHSAPQRVRY LFAPYCGFLP SASDSDLKSS XXSE*

• Further work revealed the complete nucleotide sequence <SEQ ID 551>:

• 1	ATGATGAAGC GGCGTATAGC CGTCTTCGTC CTGTTCCCGC
  	AGATAATCCG
  51	AGTTTTGGGA CAACTGTTGC CGAAAATCGT CAATACAGTT
  	CCGGCACATC
  101	GGATGCTCTT CCAGATTTTC GGGATGTTCT TTTTCTTCAT
  	ACACCAGCAA
  151	TATCTGCCCG GGATCGCCGA AATCGATTCC CCATGCGGCA
  	TCGTGTTCGG
  201	TGCGCTCCTC TTCCGTCATC TGCCCGCGCA TTGCCTGTAT
  	GGTAAAGCCG
  251	CCGTAGGGGA TGCCGTTGCA CACGAACATC CAGTCGCTGA
  	TGTCGTCAAC
  301	CGGAACGCAA ACGCTTTCGC CTTGTTCGAC ATTGGTCAGT
  	TCGCCGGGTT
  351	CATTGTTCAG CACACCGTAA ATATAAAGAC CGTCAAAATA
  	AATATCGTCG
  401	ATCCACATAT GTTCGCAAAT TTCGCCGTCT TCGCCGTCTT
  	GGAAAAAAGG
  451	GACTTTGACC ATGGCAAAAT CCAAGGCGGA AATAATGCGG
  	CGGCGTTCCC
  501	AAAAAAGCTC GCGCCAAAAA TATTTGAATG TTTTACGGGC
  	GCGTTCGTCG
  551	GCACGGTTTA CCGGTTCGTC TGCCTGTTCT ACATAATAAA
  	TGACGGAATC
  601	GCCCATCATT CTGCTCCTCA ACGTGTACGG TATCTGTTTG
  	CACCTTACTG
  651	CGGCTTTCTG CCTTCGGCAT CCGATTCGGA TTTGAAAAGT
  	TCCAAATATT
  701	CGGAATAG

• This corresponds to the amino acid sequence <SEQ ID 552; ORF136-1>:

• 1	MMKRRIAVFV LFPQIIRVLG QLLPKIVNTV PAHRMLFQIF
  	GMFFFFIHQQ
  51	YLPGIAEIDS PCGIVFGALL FRHLPAHCLY GKAAVGDAVA
  	HEHPVADVVN
  101	RNANAFALFD IGQFAGFIVQ HTVNIKTVKI NIVDPHMFAN
  	FAVFAVLEKR
  151	DFDHGKIQGG NNAAAFPKKL APKIFECFTG AFVGTVYRFV
  	CLFYIINDGI
  201	AHHSAPQRVR YLFAPYCGFL PSASDSDLKS SKYSE*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF136 shows 71.7% identity over. a 237aa overlap with an ORF (ORF136a) from strain A of N. meningitidis:
• 
• The complete length ORF136a nucleotide sequence <SEQ ID 553> is:

• 1	ATGATGAAGC GGCGTATAGC CGTCTTCGTC CTGCTCATGC
  	AGAAAATCCG
  51	GATTTTGGGA CAACTGTTGC CGAAAATCGT CAATACAGTT
  	CCGGCACATC
  101	GGATGCTCTT CCAGATNTTC GGGATGTTCT TTTTCTTCAT
  	ACACCAGCAA
  151	TACCTGCCCG GGATCGCCGA AATCGATTCC CCATGCGGCA
  	TCGTGTTCGG
  201	TACGCTCCTC TTCCGTCATC NGTCCACGCA TTGCCTGTAT
  	GGTAAAGCCG
  251	CCGTAGGGAA TGCCGTTGCA CACGAACATC CAGTCGCTGA
  	TGTCGTCAAC
  301	CGGAACGCAA ACGCTTTCGC CTTGTTCGAC ATTGGTCAGT
  	TCGCCGGGTT
  351	CATTGTTCAG CACGCCATAA ATGTAAAGAC CGTCAAAATA
  	AATATCGTCG
  401	ATCCACATAT GTTCGCAAAT TTCGCCNTCT TCGCCGTCTT
  	GGAAAAAAGG
  451	GCTTTGACCA TGGCAAAATC TAAGGNGNNA NNGATGCGGC
  	GGCGTTCCCA
  501	AAAAAGCTCG CGCCAAAAAT ATTTGAATGT TTTGCGGGCG
  	CGTTCGCCGG
  551	CACGGTTTAC CGGTTTGTCT GCCTGTTCTA CATAATAAAT
  	GACGGAATCG
  601	CCCATCATAT CTGCTCCTCA ACGTGTACGG TATCTGTTTG
  	CACCTTACTG
  651	CGGCTTTCTG CCTTCGGCAT CCGATTCGGA TTTGAAAAGT
  	TCCAAATATT
  701	CGGAATAG

• This encodes a protein having amino acid sequence <SEQ ID 554>:

• 1	MMKRRIAVFV LLMQKIRILG QLLPKIVNTV PAHRMLFQXF
  	GMFFFFIHQQ
  51	YLPGIAEIDS PCGIVFGTLL FRHXSTHCLY GKAAVGNAVA
  	HEHPVADVVN
  101	RNANAFALFD IGQFAGFIVQ HAINVKTVKI NIVDPHMFAN
  	FAXFAVLEKR
  151	ALTMAKSKXX XMRRRSQKSS RQKYLNVLRA RSPARFTGLS
  	ACST**MTES
  201	PIISAPQRVR YLFAPYCGFL PSASDSDLKS SKYSE*

• ORF136a and ORF136-1 show 73.1% identity in 238 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF136 shows 92.3% identity over a 234aa overlap with a predicted ORF (ORF136ng) from N. gonorrhoeae:
• 
• The complete length ORF136ng nucleotide sequence <SEQ ID 555> is:

• 1	ATGATGAAGC GGCGTATAGC CGTCTTCGTC CTGCTCATGC
  	AGAAAATCCG
  51	GATTTTGGGA CAACTGTTGC CGAAAATCGT CAATACAGTT
  	CCGGCACATC
  101	GGATGCTCTT CCAAATTTTC GGGATGTTCT TTTTCTTCAT
  	ACACCGGCAA
  151	TACCTGCCCG GGATCGCCGA AATCGATTCC CCAGGCGGTA
  	TCGTGTTCGG
  201	TACGCTCCTC TTCCGTCATC TGTCCGCGCA TTGCCTGTAC
  	GGTAAAGCCG
  251	CCGTAGGGGA TGCCGTTGCA CACGAACATC CAGTCGCTGA
  	TGTCGCCAAC
  301	CGGAACGCAA ACGCTTTCGC CTTGTTCGAC ATTGGTCAGT
  	CCGCCGGGTT
  351	CATTGTTCAG CACACCGTAA ATATAAAGAC CGTCAAAATA
  	AATATCGTCG
  401	ATCCACATAT GTTCGCAAAT TTCGCCGTCT TCGCCGTCTT
  	GGAAAAAAGG
  451	GACTTTGACC ATGGCAAAAT CCAAGGCGGA AATAATGCGG
  	CGGCGTTCCC
  501	AAAAAAGCTC GCGCCAAAAG TATTTGAATG TTTTACGGGC
  	GCGTTCGCCG
  551	GCACGGTTTA CCGGTTCGTC TGCCTGTTCT ACATAATAAA
  	TGACGGAATC
  601	GCCCATCATA CTGCTCCTCA ACGTGTACGG TATCTGTTTG
  	CACCTTACCG
  651	CGGTTTTCTA CCTCCGGCAT CCGATTCGGA TTTGAAAAGT
  	TCCAAATATT
  701	CGGAATAG

• This encodes a protein having amino acid sequence <SEQ ID 556>:

• 1	MMKRRIAVFV LLMQKIRILG QLLPKIVNTV PAHRMLFQIF
  	GMFFFFIHRQ
  51	YLPGIAEIDS PGGIVFGTLL FRHLSAHCLY GKAAVGDAVA
  	HEHPVADVAN
  101	RNANAFALFD IGQSAGFIVQ HTVNIKTVKI NIVDPHMFAN
  	FAVFAVLEKR
  151	DFDHGKIQGG NNAAAFPKKL APKVFECFTG AFAGTVYRFV
  	CLFYIINDGI
  201	AHHTAPQRVR YLFAPYRGFL PPASDSDLKS SKYSE*

• ORF136ng and ORF136-1 show 93.6% identity in 235 aa overlap:
• 
• Based on the presence of the putative transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 67
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 557>:

• 1	ATGGAAAATA TGGTAACGTT TTCAAAAATC AGACCGCTTT
  	TGGCAATCGC
  51	CGCCGCCGCG TTGCTTGCCG CC.TGCGGAC GGCGGGAAAT
  	AATGCTGTCC
  101	GCAAGCCGGT GCAAACCGCC AAACCCGCCG CAGTGGTCGG
  	TTTGGCACTC
  151	GGTGGCGGCG CATCTAAAGG ATTTGCCCAT GTAGGTATTA
  	TTAAGGTTTT
  201	GAAAGAAAAC GGTATTCCTG TGAAGGTGGT TACCGGCACC
  	TCCGCAGGTT
  251	CGATTGTCGG CAACCTTTTT GCATCGGGTA TGTCGCCCGA
  	CCGCCTCGAA
  301	TTGGAAGCCG AAATTTTAGG CAAAACCGAT TTGGTCGATT
  	TAACCTTGTC
  351	CACCAATGGG TTTATCAAAG GCGCAAAGCT GCAAAATTAC
  	ATCAACCGAA
  401	AACTCCGCGG CATGCAGATT CAGCAGTTTC CCATCAAATT
  	TGCCGCC..

• This corresponds to the amino acid sequence <SEQ ID 558; ORF137>:

• 1	MENMVTFSKI RPLLAIAAAA LLAAXRTAGN NAVRKPVQTA
  	KPAAVVGLAL
  51	GGGASKGFAH VGIIKVLKEN GIPVKVVTGT SAGSIVGNLF
  	ASGMSPDRLE
  101	LEAEILGKTD LVDLTLSTNG FIKGAKLQNY INRKLRGMQI
  	QQFPIKFAA..

• Further work revealed the complete nucleotide sequence <SEQ ID 559>:

• 1	ATGGAAAATA TGGTAACGTT TTCAAAAATC AGACCGCTTT
  	TGGCAATCGC
  51	CGCCGCCGCG TTGCTTGCCG CCTGCGGCAC GGCGGGAAAT
  	AATGCTGTCC
  101	GCAAGCCGGT GCAAACCGCC AAACCCGCCG CAGTGGTCGG
  	TTTGGCACTC
  151	GGTGGCGGCG CATCTAAAGG ATTTGCCCAT GTAGGTATTA
  	TTAAGGTTTT
  201	GAAAGAAAAC GGTATTCCTG TGAAGGTGGT TACCGGCACA
  	TCGGCAGGTT
  251	CGATTGTCGG CAGCCTTTTT GCATCGGGTA TGTCGCCCGA
  	CCGCCTCGAA
  301	TTGGAAGCCG AAATTTTAGG CAAAACCGAT TTGGTCGATT
  	TAACCTTGTC
  351	CACCAGTGGT TTTATCAAAG GCGAAAAGCT GCAAAATTAC
  	ATCAACCGAA
  401	AAGTCGGCGG CAGGCAGATT CAGCAGTTTC CCATCAAATT
  	TGCCGCCGTT
  451	GCTACTGATT TTGAAACCGG CAAGGCCGTC GCTTTCAATC
  	AGGGGAATGC
  501	CGGGCAGGCT GTGCGCGCTT CCGCCGCCAT TCCCAATGTG
  	TTCCAACCCG
  551	TTATCATCGG CAGGCATACA TATGTTGACG GCGGTCTGTC
  	GCAGCCCGTG
  601	CCCGTCAGTG CCGCCCGGCG GCAGGGGGCG AATTTCGTGA
  	TTGCCGTCGA
  651	TATTTCCGCC CGTCCGGGCA AAAACATCAG CCAAGGTTTC
  	TTCTCTTATC
  701	TCGATCAGAC GCTGAACGTA ATGAGCGTTT CTGCGTTGCA
  	AAATGAGTTG
  751	GGGCAGGCGG ATGTGGTTAT CAAACCGCAG GTTTTGGATT
  	TGGGTGCAGT
  801	CGGCGGATTC GATCAGAAAA AACGCGCCAT CCGGTTGGGT
  	GAGGAGGCAG
  851	CACGTGCCGC ATTGCCTGAA ATCAAACGCA AACTGGCGGC
  	ATACCGTTAT
  901	TGA

• This corresponds to the amino acid sequence <SEQ ID 560; ORF137-1>:

• 1	MENMVTFSKI RPLLAIAAAA LLAACGTAGN NAVRKPVQTA
  	KPAAVVGLAL
  51	GGGASKGFAH VGIIKVLKEN GIPVKVVTGT SAGSIVGSLF
  	ASGMSPDRLE
  101	LEAEILGKTD LVDLTLSTSG FIKGEKLQNY INRKVGGRQI
  	QQFPIKFAAV
  151	ATDFETGKAV AFNQGNAGQA VRASAAIPNV FQPVIIGRHT
  	YVDGGLSQPV
  201	PVSAARRQGA NFVIAVDISA RPGKNISQGF FSYLDQTLNV
  	MSVSALQNEL
  251	GQADVVIKPQ VLDLGAVGGF DQKKRAIRLG EEAARAALPE
  	IKRKLAAYRY
  301	*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF137 shows 93.3% identity over a 149aa overlap with an ORF (ORF137a) from strain A of N. meningitidis:
• 
• The complete length ORF137a nucleotide sequence <SEQ ID 561> is:

• 1	ATGGAAAATA TGGTAACGTT TTCAAAAATC AGACCGCTTT
  	TGGCAATCGC
  51	CGCCGCCGCG TTGCTTGCCG CCTGCGGCAC GGCGGGAAAT
  	AATGCTGCCC
  101	GCAAGCCGGT GCAAACCGCC AAACCCGCCG CAGTGGTCGG
  	TTTGGCACTC
  151	GGTGGCGGCG CATCTAAAGG ATTTGCCCAT GTAGGTATTA
  	TTAAGGTTTT
  201	GAAAGAAAAC GGTATTCCTG TGAAGGTGGT TACCGGCACA
  	TCGGCAGGTT
  251	CGATAGTCGG CAGCCTTTTT GCATCGGGTA TGTCGCCCGA
  	CCGCCTCGAA
  301	TTGGAAGCCG AAATTTTAGG TAAAACCGAT TTGGTCGATT
  	TAACCTTGTC
  351	CACCAGTGGT TTTATCAAAG GCGAAAAGCT GCAAAATTAC
  	ATCAACCGAA
  401	AAGTCGGCGG CAGGCGGATT CAGCAGTTTC CCATCAAATT
  	TGCCGCCGTT
  451	GCTACTGATT TTGAAACCGG CAAGGCCGTC GCTTTCAATC
  	AAGGGAATGC
  501	CGGGCAGGCT GTGCGCGCTT CCGCCGCCAT TCCCAATGTG
  	TTCCAACCCG
  551	TTATCATCGG CAGGCATACA TATGTTGACG GCGGTCTGTC
  	GCAGCCCGTG
  601	CCCGTCAGTG CCGCCCGGCG GCANGNNNNG NATNTCGTGA
  	TTGCCGTCGA
  651	TATTTCCGCC CGTCCGAGCA AAAACATCAG CCAAGGCTTC
  	TTCTCTTATC
  701	TCGATCAGAC GCTGAACGTA ATGAGCGTTT CCGCGTTGCA
  	AAATGAGTTG
  751	GGGCAGGCGG ATGTGGTTAT CAAACCGCAG GTTTTGGATT
  	TGGGTGCAGT
  801	CGGCGGATTC GATCAGAAAA AACGCGCCAT CCGGTTGGGT
  	GAGGAGGCAG
  851	CACGTGCCGC ATTGCCTGAA ATCAAACGCA AACTGGCGGC
  	ATACCGTTAT
  901	TGA

• This encodes a protein having amino acid sequence <SEQ ID 562>:

• 1	MENMVTFSKI RPLLAIAAAA LLAACGTAGN NAARKPVQTA
  	KPAAVVGLAL
  51	GGGASKGFAH VGIIKVLKEN GIPVKVVTGT SAGSIVGSLF
  	ASGMSPDRLE
  101	LEAEILGKTD LVDLTLSTSG FIKGEKLQNY INRKVGGRRI
  	QQFPIKFAAV
  151	ATDFETGKAV AFNQGNAGQA VRASAAIPNV FQPVIIGRHT
  	YVDGGLSQPV
  201	PVSAARRXXX XXVIAVDISA RPSKNISQGF FSYLDQTLNV
  	MSVSALQNEL
  251	GQADVVIKPQ VLDLGAVGGF DQKKRAIRLG EEAARAALPE
  	IKRKLAAYRY
  301	*

• ORF137a and ORF137-1 show 97.3% identity in 300 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF137 shows 89.9% identity over a 149aa overlap with a predicted ORF (ORF137ng) from N. gonorrhoeae:
• 
• The complete length ORF137ng nucleotide sequence <SEQ ID 563> is:

• 1	ATGGAAAATA TGGTAACGTT TTCAAAAATC AGATCATTTT
  	TGGCAATCGC
  51	CGCCGCCGCG TTGCTTGCCG CCTGCGGTAC GGCGGGAAAC
  	AATGCCGCCC
  101	GCAAGCCGGT GCAAACCGCC AAACCCGCCG CAGTGGTCGC
  	TTTGGCACTC
  151	GGTGGCGGCG CATCTAAAGG ATTTGCCCAT ATAGGAATTG
  	TTAAGGTTTT
  201	GAAAGAAAAC GGTATTCCTG TGAAGGTGGT TACCGGCACA
  	TCGGCAGGTT
  251	CGATAGTCGG CAGCCTTTTG GCATCGGGTA TGTCGCCCGA
  	CCGCCTCGAA
  301	TTGGAAGCCG AGATTTTAGG TAAAACCGAT TTAGTCGATT
  	TAACCTTGTC
  351	CACCAGTGGT TTTATCAAAG GCGAAAAGCT GCAAAATTAC
  	ATCAACCGAA
  401	AAGTCGGCGG CAGGCAGATT CAGCAGTTTC CCATCAAATT
  	TGCCGCCGTT
  451	GCCACTGATT TTGAAACCGG CAAGGCCGTC GCTTTCAATC
  	AAGGGAATGC
  501	CGGGCAGGCG GTTCGTGCTT CCGCCGCCAT TCCCAATGTG
  	TTCCAGCCAG
  551	TCATCATCGG CAGGCACAAA TATGTTGACG GCGGTCTGTC
  	GCAGCCCGTG
  601	CCCGTCAGTG CCGCTCGGCG GCAGGGGGCG AATTTCGTGA
  	TTGCCGTCGA
  651	TATTTCCGCA CGTCCGAGCA AAAATGTCGG TCAAGGTTTC
  	TTCTCTTATC
  701	TCGATCAGAC GCTGAACGTG ATGAGCGTTT CCGTGTTGCA
  	AAACGAGTTG
  751	gggcAGGCGG ATGTGGTTAT CAAACCGCag gtTTTGGATT
  	TGGGTGCAGT
  801	CGGCGGATTC GATCAGAAAA AGCGCGCCAT CCGGTTGGGC
  	GAGGAGGCAG
  851	CACGTGCCGC ATTGCCTGAA ATCAAACGCA AACTGGCGGC
  	ATACCGTTAT
  901	TGA

• This encodes a protein having amino acid sequence <SEQ ID 564>:

• 1	MENMVTFSKI RSFLAIAAAA LLAACGTAGN NAARKPVQTA
  	KPAAVVALAL
  51	GGGASKGFAH IGIVKVLKEN GIPVKVVTGT SAGSIVGSLL
  	ASGMSPDRLE
  101	LEAEILGKTD LVDLTLSTSG FIKGEKLQNY INRKVGGRQI
  	QQFPIKFAAV
  151	ATDFETGKAV AFNQGNAGQA VRASAAIPNV FQPVIIGRHK
  	YVDGGLSQPV
  201	PVSAARRQGA NFVIAVDISA RPSKNVGQGF FSYLDQTLNV
  	MSVSVLQNEL
  251	GQADVVIKPQ VLDLGAVGGF DQKKRAIRLG EEAARAALPE
  	IKRKLAAYRY
  301	*

• ORF137ng and ORF137-1 show 96.0% identity in 300 aa overlap:
• 
• Based on the presence of a predicted prokaryotic membrane lipoprotein lipid attachment site (underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 68
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 565>:

• 1	ATGTTTCGTT TACAATTCAG GCTGTTTCCC CCTTTGCGAA
  	CCGCCATGCA
  51	CATCCTGTTG ACCGCCCTGC TCAAATGCCT CTCCCTGcTG
  	CCGCTTTCCT
  101	GTCTGCACAC GCTGGGAAAC CGGCTCGGAC ATCTGGCGTT
  	TTACCTTTTA
  151	AAGGAAGACC GCGCGCGCAT CGTCGCCmAT ATGCGGCAGG
  	CGGGTTTGAA
  201	CCCCGACCCC AAAACGGTCA AAGCCGTTTT TGCGGAAACG
  	GCAAAAGGCG
  251	GTTTGGAACT TGCCCCCGCG TTTTTCAGAA AACCGGAAGA
  	CATAGAAACA
  301	ATGTTCAAAG CGGTACACGG CTGGGAACAT GTGCAGCAGG
  	CTTTGGACAA
  351	ACACGAAGGG CTGCTATTC..

• This corresponds to the amino acid sequence <SEQ ID 566; ORF138>:

• 1	MFRLQFRLFP PLRTAMHILL TALLKCLSLL PLSCLHTLGN
  	RLGHLAFYLL
  51	KEDRARIVAX MRQAGLNPDP KTVKAVFAET AKGGLELAPA
  	FFRKPEDIET
  101	MFKAVHGWEH VQQALDKHEG LLF

• Further work revealed the complete nucleotide sequence <SEQ ID 567>:

• 1	ATGTTTCGTT TACAATTCAG GCTGTTTCCC CCTTTGCGAA
  	CCGCCATGCA
  51	CATCCTGTTG ACCGCCCTGC TCAAATGCCT CTCCCTGCTG
  	CCGCTTTCCT
  101	GTCTGCACAC GCTGGGAAAC CGGCTCGGAC ATCTGGCGTT
  	TTACCTTTTA
  151	AAGGAAGACC GCGCGCGCAT CGTCGCCAAT ATGCGGCAGG
  	CGGGTTTGAA
  201	CCCCGACCCC AAAACGGTCA AAGCCGTTTT TGCGGAAACG
  	GCAAAAGGCG
  251	GTTTGGAACT TGCCCCCGCG TTTTTCAGAA AACCGGAAGA
  	CATAGAAACA
  301	ATGTTCAAAG CGGTACACGG CTGGGAACAT GTGCAGCAGG
  	CTTTGGACAA
  351	ACACGAAGGG CTGCTATTCA TCACGCCGCA CATCGGCAGC
  	TACGATTTGG
  401	GCGGACGCTA CATCAGCCAG CAGCTTCCGT TCCCGCTGAC
  	CGCCATGTAC
  451	AAACCGCCGA AAATCAAAGC GATAGACAAA ATCATGCAGG
  	CGGGCAGGGT
  501	TCGCGGCAAA GGAAAAACCG CGCCTACCAG CATACAAGGG
  	GTCAAACAAA
  551	TCATCAAAGC CCTGCGTTCG GGCGAAGCAA CCATCGTCCT
  	GCCCGACCAC
  601	GTCCCCTCCC CTCAAGAAGG CGGGGAAGGC GTATGGGTGG
  	ATTTCTTCGG
  651	CAAACCTGCC TATACCATGA CGCTGGCGGC AAAATTGGCA
  	CACGTCAAAG
  701	GCGTGAAAAC CCTGTTTTTC TGCTGCGAAC GCCTGCCTGG
  	CGGACAAGGT
  751	TTCGATTTGC ACATCCGCCC CGTCCAAGGG GAATTGAACG
  	GCGACAAAGC
  801	CCATGATGCC GCCGTGTTCA ACCGCAATGC CGAATATTGG
  	ATACGCCGTT
  851	TTCCGACGCA GTATCTGTTT ATGTACAACC GCTACAAAAT
  	GCCGTAA

• This corresponds to the amino acid sequence <SEQ ID 568; ORF138-1>:

• 1	MFRLQFRLFP PLRTAMHILL TALLKCLSLL PLSCLHTLGN
  	RLGHLAFYLL
  51	KEDRARIVAN MRQAGLNPDP KTVKAVFAET AKGGLELAPA
  	FFRKPEDIET
  101	MFKAVHGWEH VQQALDKHEG LLFITPHIGS YDLGGRYISQ
  	QLPFPLTAMY
  151	KPPKIKAIDK IMQAGRVRGK GKTAPTSIQG VKQIIKALRS
  	GEATIVLPDH
  201	VPSPQEGGEG VWVDFFGKPA YTMTLAAKLA HVKGVKTLFF
  	CCERLPGGQG
  251	FDLHIRPVQG ELNGDKAHDA AVFNRNAEYW IRRFPTQYLF
  	MYNRYKMP*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF138 shows 99.2% identity over a 123aa overlap with an ORF (ORF138a) from strain A of N. meningitidis:
• 
• The complete length ORF138a nucleotide sequence <SEQ ID 569> is:

• 1	ATGTTTCGTT TACAATTCAG GCTGTTTCCC CCTTTGCGAA
  	CCGCCATGCA
  51	CATCCTGTTG ACCGCCCTGC TCAAATGCCT CTCCCTGCTG
  	CCGCTTTCCT
  101	GTCTGCACAC GCTGGGAAAC CGGCTCGGAC ATCTGGCGTT
  	TTACCTTTTA
  151	AAGGAAGACC GCGCGCGCAT CGTCGCCAAT ATGCGTCAGG
  	CAGGCATGAA
  201	TCCCGACCCC AAAACGGTCA AAGCCGTTTT TGCGGAAACG
  	GCAAAAGGCG
  251	GTTTGGAACT TGCCCCCGCG TTTTTCAGAA AACCGGAAGA
  	CATAGAAACA
  301	ATGTTCAAAG CGGTACACGG CTGGGAACAT GTGCAGCAGG
  	CTTTGGACAA
  351	ACACGAAGGG CTGCTATTCA TCACGCCGCA CATCGGCAGC
  	TACGATTTGG
  401	GCGGACGCTA CATCAGCCAG CAGCTTCCGT TCCCGCTGAC
  	CGCCATGTAC
  451	AAACCGCCGA AAATCAAAGC GATAGACAAA ATCATGCAGG
  	CGGGCAGGGT
  501	TCGCGGCAAA GGAAAAACCG CGCCTACCAG CATACAAGGG
  	GTCAAACAAA
  551	TCATCAAAGC CCTGCGTTCG GGCGAAGCAA CCATCGTCCT
  	GCCCGACCAC
  601	GTCCCCTCCC CTCAAGAAGG CGGGGAAGGC GTATGGGTGG
  	ATTTCTTCGG
  651	CAAACCTGCC TATACCATGA CGCTGGCGGC AAAATTGGCA
  	CACGTCAAAG
  701	GCGTGAAAAC CCTGTTTTTC TGCTGCGAAC GCCTGCCTGG
  	CGGACAAGGT
  751	TTCGATTTGC ACATCCGCCC CGTCCAAGGG GAATTGAACG
  	GCGACAAAGC
  801	CCATGATGCC GCCGTGTTCA ACCGCAATGC CGAATATTGG
  	ATACGCCGTT
  851	TTCCGACGCA GTATCTGTTT ATGTACAACC GCTACAAAAT
  	GCCGTAA

• This encodes a protein having amino acid sequence <SEQ ID 570>:

• 1 MFRLQFRLFP PLRTAMHILL TALLKCLSLL PLSCLHTLGN RLGHLAFYLL
  51 KEDRARIVAN MRQAGLNPDP KTVKAVFAET AKGGLELAPA FFRKPEDIET
  101 MFKAVHGWEH VQQALDKHEG LLFITPHIGS YDLGGRYISQ QLPFPLTAMY
  151 KPPKIKAIDK IMQAGRVRGK GKTAPTSIQG VKQIIKALRS GEATIVLPDH
  201 VPSPQEGGEG VWVDFFGKPA YTMTLAAKLA HVKGVKTLFF CCERLPGGQG
  251 FDLHIRPVQG ELNGDKAHDA AVFNRNAEYW IRRFPTQYLF MYNRYKMP*

• ORF138a and ORF138-1 show 99.7% identity over a 298aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF138 shows 94.3% identity over a 123aa overlap with a predicted ORF (ORF138ng) from N. gonorrhoeae:
• 
• The complete length ORF138ng nucleotide sequence <SEQ ID 571> is:

• 1	ATGTTTCGTT TACAATTCAG GCTGTTTCCC CCTTTGCGAA
  	CCGCCATGCA
  51	CATCCTGTTG ACCGCCCTGC TCAAATGCCT CTCCCTGCTG
  	TCGCTTTCCT
  101	GTCTGCACAC GCTGGGAAAC CGGCTCGGAC ATCTGGCGTT
  	TTACCTTTTA
  151	AAGGAAGACC GCGCGCGCAT CGTCGCCAAT ATGCGGCAGG
  	CGGGTTTGAA
  201	CCCCGACACG CAGACGGTCA AAGCCGTTTT TGCGGAAACG
  	GCAAAATGCG
  251	GTTTGGAACT TGCCCCCGCG TTTTTCAAAA AACCGGAAGA
  	CATCGAAACA
  301	ATGTTCAAAG CGGTACACGG CTGGGAACAC GTGCAGCAGG
  	CTTTGGACAA
  351	GGGCGAAGGG CTGCTGTTCA TCACGCCGCA CATCGGCAGC
  	TACGATTTGG
  401	GCGGACGCTA CATCAGCCAG CAGCTTCCGT TCCACCTGAC
  	CGCCATGTAC
  451	AAGCCGCCGA AAATCAAAGC GATAGACAAA ATCATGCAGG
  	CGGGCAGGGT
  501	GCGCGGCAAA GGCAAAACcg cgcccaccgg catACAAGGG
  	GTCAAACAAA
  551	tcatcaAGGC CCTGCGCGCG GGCGAGGCAA CCAtcATCCT
  	GCCCGACCAC
  601	GTCCCTTCTC CGCAGGAagg cggCGGCGTG TGGGCGGATT
  	TTTTCGGCAA
  651	ACCTGCATAC acCATGACAC TGGCGGCAAA ATTGGCACAC
  	GTCAAAGGCG
  701	TGAAAACCCT GTTTTTCTGC TGCGAACGCC TGCCCGACGG
  	ACAAGGCTTC
  751	GTGTTGCACA TCCGCCCCGT CCAAGGGGAA TTGAACGGCA
  	ACAAAGCCCA
  801	CGATGCCGCC GTGTTCAACC GCAATACCGA ATATTGGATA
  	CGCCGTTTTC
  851	CGACGCAGTA TCTGTTTATG TACAACCGCT ATAAAACGCC
  	GTAA

• This encodes a protein having amino acid sequence <SEQ ID 572>:

• 1	MFRLQFRLFP PLRTAMHILL TALLKCLSLL SLSCLHTLGN
  	RLGHLAFYLL
  51	KEDRARIVAN MRQAGLNPDT QTVKAVFAET AKCGLELAPA
  	FFKKPEDIET
  101	MFKAVHGWEH VQQALDKGEG LLFITPHIGS YDLGGRYISQ
  	QLPFHLTAMY
  151	KPPKIKAIDK IMQAGRVRGK GKTAPTGIQG VKQIIKALRA
  	GEATIILPDH
  201	VPSPQEGGGV WADFFGKPAY TMTLAAKLAH VKGVKTLFFC
  	CERLPDGQGF
  251	VLHIRPVQGE LNGNKAHDAA VFNRNTEYWI RRFPTQYLFM
  	YNRYKTP*

• ORF138ng and ORF138-1 show 94.3% identity over 299aa overlap:
• 
• In addition, ORF138ng is homologous to htrB protein from Pseudomonas fluorescens:

• gnl|PID|e334283 (Y14568) htrB [Pseudomonas fluorescens] Length = 253
  Score = 80.8 bits (196), Expect = 9e−15
  Identities = 49/151 (32%), Positives = 79/151 (51%), Gaps = 6/151 (3%)

  Query:	101	MFKAVHGWEHVQQALDKGEGLLFITPHIGSYD-LGGRYISQQLPFHLTAMYKPPKIKAID	159
  		+ + V G E +++AL  G+G++ IT H+G+++ L   Y SQ  P      Y+PPK+KA+D
  Sbjct:	94	LVREVEGLEVLKEALASGKGVVGITSHLGNWEVLNHFYCSQCKPI---IFYRPPKLKAVD	150
  Query:	160	KIMQAGRVRGKGKTAPTGIQGVKQIIKALRAGEATIILPDHVPSPQEGGGVWADFFGKPA	219
  		++++  RV+   K A +  +G+  +IK +R G    I  D  P P E  G++  FF   A
  Sbjct:	151	ELLRKQRVQLGNKVAASTKEGILSVIKEVRKGGQVGIPAD--PEPAESAGIFVPFFATQA	208
  Query:	220	YTMTLAAKLAHVKGVKTLFFCCERLPDGQGF	250
  		T      +        +F    RLPDG G+
  Sbjct:	209	LTSKFVPNMLAGGKAVGVFLHALRLPDGSGY	239

• Based on this analysis, including the presence of a putative transmembrane domain in the gonococcal protein, it was predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF138-1 (57 kDa) was cloned in the pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 14A shows the results of affinity purification of the GST-fusion protein. Purified GST-fusion protein was used to immunise mice, whose sera were used for ELISA (positive result) and FACS analysis (FIG. 14B). These experiments confirm that ORF138-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 69
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 573>:

• 1	..GCGTGGTCGG CCGGCGAATC GTGGCGTGTG TTAATGGAAA
  	GTGAAACGTG
  51	GCATGCGGTG TGGAATACTT TGCGCTTCTC GGCGGCGGCG
  	GTGTATGCGG
  101	CAGCGGTTTT GGGTGTGGTG TATGCGGCGC CGGCGCGGCG
  	GTCGGCGTGG
  151	ATGCGCGGGC TGATGTTTTA GCCGTTTATG GTGTCGCCGG
  	TTTGTGTTTC
  201	GGCGGGCGTG CTGCTGCTTT ATCCGCAGTG GACGGCTTCG
  	TTGCCGTTGC
  251	TGCTGGCGAT GTATGCGCTG CTGGCGTATC CGTTTGTGGC
  	AAAAGATGTT
  301	TTATCAGCCT GGGATGCACT GCCGCCGGAT TACGGCAGGG
  	CGGCGGCGGG
  351	TTTGGGTGCA AACGGCTTTC AGACGGCATG CCGCATCACG
  	TTCCCCCTCT
  401	TGAAACCGGC GTTGCGGCGC GGTCTGACTT TGGCGGCGGC
  	AACCTGCGTG
  451	GGCGAATTTG CGGCGACATT GTTTCTGTCG CGTCCGGAAT
  	GGCAGACGCT
  501	GACGACTTTG ATTTATGCCT ATTTGGGACG CGCGGGTGAG
  	GATAATTACG
  551	CGCGGGCGAT GGTGCTG..

• This corresponds to the amino acid sequence <SEQ ID 574; ORF139>:

• 1	..AWSAGESWRV LMESETWHAV WNTLRFSAAA VYAAAVLGVV
  	YAAPARRSAW
  51	MRGLMFXPFM VSPVCVSAGV LLLYPQWTAS LPLLLAMYAL
  	LAYPFVAKDV
  101	LSAWDALPPD YGRAAAGLGA NGFQTACRIT FPLLKPALRR
  	GLTLAAATCV
  151	GEFAATLFLS RPEWQTLTTL IYAYLGRAGE DNYARAMVL..

• Further work revealed the complete nucleotide sequence <SEQ ID 575>:

• 1	ATGGATGGAC GGCGTTGGGT GGTATGGGGT GCTTTTGCCC
  	TGCTGCCTTC
  51	GGCTTTTTTG GCGGTAATGG TCGTTGCGCC TTTGTGGGCG
  	GTGGCGGCGT
  101	ATGACGGTTT GGCGTGGCGC GCGGTGCTGT CGGATGCCTA
  	TATGCTCAAA
  151	CGTTTGGCGT GGACGGTATT TCAGGCAGCG GCAACCTGTG
  	TGCTGGTGCT
  201	GCCTTTGGGC GTGCCTGTCG CGTGGGTGCT GGCGCGGCTG
  	GCGTTTCCGG
  251	GGCGGGCTTT GGTGCTGCGC CTGCTGATGC TGCCTTTTGT
  	GATGCCCACG
  301	TTGGTGGCGG GCGTGGGCGT GCTGGCCCTG TTCGGGGCGG
  	ACGGGCTGTT
  351	GTGGCGCGGC AGGCAGGATA CGCCGTATCT GTTGTTGTAC
  	GGCAATGTGT
  401	TTTTCAACCT TCCTGTGTTG GTCAGGGCGG CGTATCAGGG
  	GTTTGTGCAA
  451	GTGCCTGCGG CACGGCTTCA GACGGCACGG ACGTTGGGCG
  	CGGGGGCGTG
  501	GCGGCGGTTT TGGGACATTG AAATGCCCGT TTTGCGCCCG
  	TGGCTTGCCG
  551	GCGGCGTGTG CCTTGTCTTT CTGTATTGTT TTTCCGGGTT
  	CGGGCTGGCG
  601	CTGCTGCTGG GCGGCAGCCG TTATGCCACG GTCGAAGTGG
  	AAATTTACCA
  651	GTTGGTCATG TTCGAACTCG ATATGGCGGT TGCTTCGGTG
  	CTGGTGTGGC
  701	TGGTGTTGGG GGTAACGGCG GCGGCAGGGT TGCTGTATGC
  	GTGGTTCGGC
  751	AGGCGCGCGG TTTCGGATAA GGCGGTTTCC CCTGTGATGC
  	CGTCGCCGCC
  801	GCAGTCGGTC GGGGAATATG TGCTGCTGGC GTTTGCGGCG
  	GCGGTGTTGT
  851	CTGTGTGCTG CCTGTTTCCT TTGTTGGCAA TTGTTGTGAA
  	AGCGTGGTCG
  901	GCCGGCGAAT CGTGGCGTGT GTTAATGGAA AGTGAAACGT
  	GGCAGGCGGT
  951	GTGGAATACT TTGCGCTTCT CGGCGGCGGC GGTGTATGCG
  	GCGGCGGTTT
  1001	TGGGTGTGGT GTATGCGGCG GCGGCGCGGC GGTCGGCGTG
  	GATGCGCGGG
  1051	CTGATGTTTT TGCCGTTTAT GGTGTCGCCG GTTTGTGTTT
  	CGGCGGGCGT
  1101	GCTGCTGCTT TATCCGCAGT GGACGGCTTC GTTGCCGTTG
  	CTGCTGGCGA
  1151	TGTATGCGCT GCTGGCGTAT CCGTTTGTGG CAAAAGATGT
  	TTTATCAGCC
  1201	TGGGATGCAC TGCCGCCGGA TTACGGCAGG GCGGCGGCGG
  	GTTTGGGTGC
  1251	AAACGGCTTT CAGACGGCAT GCCGCATCAC GTTCCCCCTC
  	TTGAAACCGG
  1301	CGTTGCGGCG CGGTCTGACT TTGGCGGCGG CAACCTGCGT
  	GGGCGAATTT
  1351	GCGGCGACAT TGTTTCTGTC GCGTCCGGAA TGGCAGACGC
  	TGACGACTTT
  1401	GATTTATGCC TATTTGGGAC GCGCGGGTGA GGATAATTAC
  	GCGCGGGCGA
  1451	TGGTGCTGAC ATTGCTGTTG GCGGCGTTCG CGCTGGGTAT
  	TTTCCTGCTG
  1501	TTGGACGGCG GCGAAGGCGG AAAACAGACG GAAACGTTAT
  	AA

• This corresponds to the amino acid sequence <SEQ ID 576; ORF139-1>:

• 1	MDGRRWVVWG AFALLPSAFL AVMVVAPLWA VAAYDGLAWR
  	AVLSDAYMLK
  51	RLAWTVFQAA ATCVLVLPLG VPVAWV LARL AFPGRALVLR
  	LLMLPFVMPT
  101	LVAGVGVLAL FGADGLLWRG RQDTPYLLLY GNVFFNLPVL
  	VRAAYQGFVQ
  151	VPAARLQTAR TLGAGAWRRF WDIEMPVLRP WLAGGVCLVF
  	LYCFSGFGLA
  201	LLLGGSRYAT VEVEIYQLVM FELDMAVASV LVWLVLGVTA
  	AAGLLYAWFG
  251	RRAVSDKAVS PVMPSPPQSV GEYVLLAFAA AVLSVCCLFP
  	LLAIVVKAWS
  301	AGESWRVLME SETWQAVWNT LRFSAAAVYA AAVLGVVYAA
  	AARRSAWMRG
  351	LMFLPFMVSP VCVSAGVLLL YPQWTASLPL LLAMYALLAY
  	PFVAKDVLSA
  401	WDALPPDYGR AAAGLGANGF QTACRITFPL LKPALRRGLT
  	LAAATCVGEF
  451	AATLFLSRPE WQTLTTLIYA YLGRAGEDNY ARAM VLTLLL
  	AAFALGIFLL
  501	LDGGEGGKQT ETL*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF139 shows 94.7% identity over a 189aa overlap with an ORF (ORF139a) from strain A of N. meningitidis:
• 
• The complete length ORF139a nucleotide sequence <SEQ ID 577> is:

• 1	ATGGATGGAC GGCGTTGGGC GGTATGGGGT GCTTTTGCCC
  	TGCTGCCTTC
  51	GGCTTTTTTG GCGGCAATGG TCGTTGCGCC TTTGTGGGCG
  	GTGGCGGCGT
  101	ATGACGGTTT GGCGTGGCGC GCGGTGCTGT CGGATGCCTA
  	TATGCTCAAA
  151	CGTTTGGCGT GGACGGTATT TCAGGCAGCG GCAACCTGTG
  	TGCTGGTGCT
  201	GCCTTTGGGC GTGCCTGTCG CGTGGGTGCT GGCGCGGCTG
  	GCGTTTCCGG
  251	GGCGGGCTTT GGTGCTGCGC CTGCTGATGC TGCCTTTTGT
  	GATGCCCACG
  301	TTGGTGGCGG GCGTGGGCGT GCTGGCTCTG TTCGGGGCGG
  	ACGGCCTGTN
  351	GTGGCGCGGC TGGCAGGATA CGCCGTATCT GTTGTTGTAC
  	GGCAATGTGT
  401	TTTTTNACCT TCCTGTGTTG GTCAGGGCGG CATATCAGGG
  	GTTTGTGCAA
  451	GTGCCTGCGG CACGGCTTCA GACGGCACNG ACATTGGGCG
  	CGGGGGCGTG
  501	GCGGCGGTTT TGGGACATTG AAATGCCCGT TTTGCGCCCG
  	TGGCTTGCCG
  551	GCGGCGTGTG CCTTGTCTTC CTGTATTGTT TTTCGGGGTT
  	CGGGCTGGCA
  601	TTGCTGCTGG GCGGCAGCCG TTATGCCACG GTCGAAGTGG
  	AAATTTACCA
  651	GTTGGTCATG TTCGAACTCG ATATGGCGGT TGCTTCGGTG
  	CTNGTGTGGC
  701	TGGTGTNGGG GGTAACNGCG GCGGCAGGGT TGCTGTATGC
  	GTGGTTCGGC
  751	AGGCGCGCGG TTTCGGATAA GGCNGTTTCC CCTGTGATGC
  	CGTCGCCGCC
  801	GCAGTCGGTC GGGGAATATG TGCTNCTGGC GTTTGCGGCG
  	GCGGTGTNGT
  851	CTGTGTGCTG CCTGTTTCNT TTGTTGGCAA TTGTTGTGAA
  	AGCGTGGTCG
  901	GCCGGCGAAT CGTGGCGTGT GTTAATGGAA AGTGAAACGT
  	GGCAGGCGGT
  951	GTGGAATACT NTGCGCTTCT CGGCGGCGGC GGTGTATGCG
  	GCGGCGGTTT
  1001	TGGGTGTGGT GTATGCGGCG GCGGCGCGGC GGTCGGCGTG
  	GATGCGCGGG
  1051	CTGATGTTTT TGCCGTTTAT GGTGTCGCCG GTTTGTGTTT
  	CGGCGGGCGT
  1101	GCTGCTGCTT NATCCGCAGT GGACGGCTTC GTTGCCGCTG
  	CTGCTGGCGA
  1151	TGTATGCGCT GCTGGCGTAT CCGTTTGTGG CAAAAGATGT
  	TTTATCAGCC
  1201	TGNGATGCAC TGCCGCCGGA TTACGGCAGG GCGGCGGCGG
  	GTTTGGGTGC
  1251	AAACGGCTTT CAGACGGCAT GCCGCATCAC GTTCCCCCTC
  	TTGAAACCGG
  1301	CGTTGCGGCG CGGTCTGACT TTGGCGGCGG CAACCTGCGT
  	GGGCGAATTT
  1351	GCGGCAACCT TGTTCNTGTC GCGTCNCGAG TGGCAGACGC
  	TGACGACTTT
  1401	GATTTATGCC TATNTGGGAC GCGCGGGTGA NGATAATTAC
  	GCGCGGGCGA
  1451	TGGTGCTGAC ATTGCTGTTG GCGGCGTTCG CGCTGGGTAT
  	NTTCCTGCTG
  1501	TTGGACGGCG GCGAAGGCGG AAAACGGACG GAAACGTTAT
  	AA

• This encodes a protein having amino acid sequence <SEQ ID 578>:

• 1	MDGRRWAVWG AFALLPSAFL AAMVVAPLWA VAAYDGLAWR
  	AVLSDAYMLK
  51	RLAWTVFQAA ATCVLVLPLG VPVAWV LARL AFPGRALVLR
  	LLMLPFVMPT
  101	LVAGVGVLAL FGADGLXWRG WQDTPYLLLY GNVFFXLPVL
  	VRAAYQGFVQ
  151	VPAARLQTAX TLGAGAWRRF WDIEMPVLRP WLAGGVCLVF
  	LYCFSGFGLA
  201	LLLGGSRYAT VEVEIYQLVM FELDMAVASV LVWLVXGVTA
  	AAGLLYAWFG
  251	RRAVSDKAVS PVMPSPPQSV GEYVLLAFAA AVXSVCCLFX
  	LLAIVVKAWS
  301	AGESWRVLME SETWQAVWNT XRFSAAAVYA AAVLGVVYAA
  	AARRSAWMRG
  351	LMFLPFMVSP VCVSAGVLLL XPQWTASLPL LLAMYALLAY
  	PFVAKDVLSA
  401	XDALPPDYGR AAAGLGANGF QTACRITFPL LKPALRRGLT
  	LAAATCVGEF
  451	AATLFXSRXE WQTLTTLIYA YXGRAGXDNY ARAMVLTLLL
  	AAFALGXFLL
  501	LDGGEGGKRT ETL*

• ORF139a and ORF139-1 show 96.5% homology over a 514aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF139 shows 95.2% identity over a 189aa overlap with a predicted ORF (ORF139ng) from N. gonorrhoeae:
• 
• The complete length ORF139ng nucleotide sequence <SEQ ID 579> is predicted to encode a protein having amino acid sequence <SEQ ID 580>:

• 1	MDGRCWAVRG AFSLLPSAFL AVMVVAPLWA VAAYDGLAWR
  	AVLSDAYMLK
  51	RLAWTVFQAA ATCVLVLPLG VPVAWVLARL AFPGRALVLR
  	LLMLPFVMPT
  101	LVAGVGVLAL FGADGLLWRG RQDTPYLLLY GNVFFNLPVL
  	VRAAYQGFAQ
  151	VPAARLQTAR TLGAGAWRPF WDIEMPVLRP WLAGGVCLVF
  	LYCFSGFGLA
  201	LLLGGSRYAT VEVEIYQLVM FELDMAGASA LVWLVLGVTA
  	AAGLLYAWFG
  251	RRAVSDKAVS PVMPSPPQSV GEYVLLAFSV AVLSVCCLFP
  	LSAIVVKAWS
  301	AGESRRVLME SETWQAVWNT LRFSAAAVFA AAVLGVVYAA
  	AARRLVWMRG
  351	LVFLPFMVSP VCVSAGVLLL YPGWTASLPL LLAMYALLAY
  	PFVAKDVLSA
  401	WDALPPDYGR AAAGLGANGF QTACRITFPL LKPALRRGLT
  	LAAATCVGEF
  451	AATLFLSRPE WQTLTTLIYA YLGRAGEDNY ARAMVLTLLL
  	SAFAVCIFLL
  501	LDNGEGGKRT ETL*

• Further work revealed a variant gonococcal DNA sequence <SEQ ID 581>:

• 1	ATGGATGGAC GGTGTTGGGC GGTACGGGGT GCTTTTTCCC
  	TGCTGCCTTC
  51	GGCTTTTTTG GCGGTAATGG TCGTTGCGCC TTTGTGGGCG
  	GTGGCGGCGT
  101	ATGACGGTTT GGCGTGGCGC GCGGTGCTGT CGGATGCCTA
  	TATGCTCAAA
  151	CGTTTGGCGT GGACGGTGTT TCAGGCGGCG GCAACCTGTG
  	TGCTGGTGCT
  201	GCCTTTGGGC GTGCCTGTCG CGTGGGTGCT GGCGCGGCTG
  	GCGTTCCCGG
  251	GGCGGGCTTT GGTGCTGCGC CTGCTGATGC TGCCGTTTGT
  	GATGCCCACG
  301	CTGGTGGCGG GCGTGGGCGT GCTGGCTCTG TTCGGGGCGG
  	ACGGGCTGTT
  351	GTGGCGCGGC CGGCAGGATA CGCCGTATCT GTTGTTGTAC
  	GGCAATGTGT
  401	TTTTCAACCT GCCCGTGTTG GTCAGGGCGG CGTATCAGGG
  	GTTTGCTCAA
  451	GTGCCTGCGG CACGGCTTCA GACGGCACGG ACGTTGGGCG
  	CGGGGGCGTG
  501	GCGGCGGTTT TGGGACATTG AAATGCCCGT TTTGCGCCCG
  	TGGCTTGCCG
  551	GCGGCGTGTG CCTTGTCTTC CTGTATTGTT TTTCGGGGTT
  	CGGGCTGGCA
  601	TTGCTGTTGG GCGGCAGCCG TTATGCCACG GTCGAAGTGG
  	AAATTTACCA
  651	GTTGGTTATG TTCGAACTCG ATATGGCGGG GGCTTCGGCG
  	CTGGTGTGGC
  701	TGGTGTTGGG GGTAACGGCG GCGGCAGGGT TGCTGTATGC
  	GTGGTTCGGC
  751	AGGCGCGCGG TTTCGGATAA GGCGGTTTCC CCCGTGATGC
  	CGTCGCCGCC
  801	GCAATCGGTG GGGGAATATG TATTGCTGGC ATTTTCGGTG
  	GCGGTGTTGT
  851	CCGTGTGCTG CCTGTTTCCT TTGTCGGCAA TTGTTGTGAA
  	AGCGTGGTCG
  901	GCCGGCGAAT CGCGGCGTGT GTTAATGGAA AGTGAAACGT
  	GGCAGGCAGT
  951	GTGGAATACt ttGCGCTTTT CGGCGGCGGC GGTGTTTGCG
  	GCGGCGGTTT
  1001	TGGGTGTGGT GTATGCGGCG GCGGCGCGGC GGCTGGTGTG
  	GATGCGCGGA
  1051	CTGGTGTTTT TACCGTTTAT GGTGTCGCCG GTTTGTGTTT
  	CGGCGGGCGT
  1101	GCTGCTGCTT TATCCGGGGT GGACGGCTTC GTTACCGCTG
  	CTGCTGGCGA
  1151	TGTATGCGCT GCTGGCGTAT CCGTTTGTGG CAAAAGATGT
  	TTTATCGGCC
  1201	TGGGATGCAC TGCCGCCGGA TTACGGCAGG GCGGCGGCAG
  	GTTTGGGCGC
  1251	AAACGGCTTT CAGACGGCAT GCCGTATCAC GTTCCCCCTC
  	TTGAAACCGG
  1301	CGTTGCGGCG CGGTCTGACT TTGGCGGCGG CGACGTGTGT
  	GGGCGAATTT
  1351	GCGGCAACCT TGTTCCTGTC GCGTCCGGAA TGGCAGACGT
  	TGACGACTTT
  1401	GATTTATGCC TATTTGGGGC GTGCGGGTGA GGACAATTAT
  	GCGCGGGCAA
  1451	TGGTGTTGAC ATTGCTGTTG TCGGCATTTG CGGTGTGCAT
  	TTTCCTGCTG
  1501	TTGGACAACG GCGAAGGCGg aaaACGGACG GAAACGTTAT
  	AA

• This corresponds to the amino acid sequence <SEQ ID 582; ORF139ng-1>:

• 1	MDGRCWAVRG AFSLLPSAFL AVMVVAPLWA VAAYDGLAWR
  	AVLSDAYMLK
  51	RLAWTVFQAA ATCVLVLPLG VPVAWVL ARL AFPGRALVLR
  	LLMLPFVMPT
  101	LVAGVGVLAL FGADGLLWRG RQDTPYLLLY GNVFFNLPVL
  	VRAAYQGFAQ
  151	VPAARLQTAR TLGAGAWRRF WDIEMPVLRP WLAGGVCLVF
  	LYCFSGFGLA
  201	LLLGGSRYAT VEVEIYQLVM FELDMAGASA LVWLVLGVTA
  	AAGLLYAWFG
  251	RRAVSDKAVS PVMPSPPQSV GEYVLLAFSV AVLSVCCLFP
  	LSAIVVKAWS
  301	AGESRRVLME SETWQAVWNT LRFSAAAVFA AAVLGVVYAA
  	AARRLVWMRG
  351	LVFLPFMVSP VCVSAGVLLL YPGWTASLPL LLAMYALLAY
  	PFVAKDVLSA
  401	WDALPPDYGR AAAGLGANGF QTACRITFPL LKPALRRGLT
  	LAAATCVGEF
  451	AATLFLSRPE WQTLTTLIYA YLGRAGEDNY ARAMVLTLLL
  	SAFAVCIFLL
  501	LDNGEGGKRT ETL*

• ORF139ng-1 and ORF139-1 show 95.9% identity over 513aa overlap:
• 
• Based on the presence of a predicted binding-protein-dependent transport systems inner membrane component signature (underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 70
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 583>:

• 1	ATGGACGGCT GGACACAGAC GCTGTCCGCG CAAACCCTGT
  	TGGGCATTTC
  51	GGCGGCGGCA ATCATCCTCA TTCTGATTTT AATCGTCAGA
  	TTCCGCATCC
  101	ACGCGCTGCT GACACTGGTC ATCGTCAGCC TGCTGACGGC
  	TTTGGCAACC
  151	GGTTTGCCCA CAGGCAGCAT TGTCAAAGAC ATACTGGTCA
  	AAAACTTCGG
  201	CGGCACGCTC GGCGGCGTGG CGCTTCTGGT CGGCCTGGGC
  	GCGATGCTCG
  251	AACGTTTGGT C...

• This corresponds to the amino acid sequence <SEQ ID 584; ORF140>:

• 1	MDGWTQTLSA QTLLGISAAA IILILILIVR FRIHALLTLV
  	IVSLLTALAT
  51	GLPTGSIVKD ILVKNFGGTL GGVALLVGLG AMLERLV..

• Further work revealed the complete nucleotide sequence <SEQ ID 585>:

• 1	ATGGACGGCT GGACACAGAC GCTGTCCGCG CAAACCCTGT
  	TGGGCATTTC
  51	GGCGGCGGCA ATCATCCTCA TTCTGATTTT AATCGTCAAA
  	TTCCGCATCC
  101	ACGCGCTGCT GACACTGGTC ATCGTCAGCC TGCTGACGGC
  	TTTGGCAACC
  151	GGTTTGCCCA CAGGCAGCAT TGTCAACGAC ATACTGGTCA
  	AAAACTTCGG
  201	CGGCACGCTC GGCGGCGTGG CGCTTCTGGT CGGCCTGGGC
  	GCGATGCTCG
  251	GACGTTTGGT CGAAACATCC GGCGGCGCAC AGTCGCTGGC
  	GGACGCGCTG
  301	ATCCGGATGT TCGGCGAAAA ACGCGCACCG TTCGCGCTGG
  	GCGTTGCCTC
  351	GCTGATTTTC GGCTTCCCGA TTTTCTTCGA TGCCGGACTA
  	ATCGTCATGC
  401	TGCCCATCGT GTTCGCCACC GCACGGCGCA TGAAACAGGA
  	CGTACTGCCC
  451	TTCGCGCTTG CCTCCATCGG CGCATTTTCC GTCATGCACG
  	TCTTCCTGCC
  501	GCCCCATCCG GGCCCGATTG CCGCTTCCGA ATTTTACGGC
  	GCGAACATCG
  551	GCCAAGTTTT GATTTTGGGT CTGCCGACCG CCTTCATCAC
  	ATGGTATTTC
  601	AGCGGCTATA TGCTCGGCAA AGTGTTGGGG CGCACCATCC
  	ATGTTCCCGT
  651	TCCCGAACTG CTCAGCGGCG GCACGCAAGA CAACGACCTG
  	CCGAAAGAAC
  701	CTGCCAAAGC AGGAACGGTC GTCGCCATCA TGCTGATTCC
  	CATGCTGCTG
  751	ATTTTCCTGA ATACCGGCGT ATCGGCCCTC ATCAGCGAAA
  	AACTCGTAAG
  801	TGCGGACGAA ACCTGGGTTC AGACGGCAAA AATAATCGGT
  	TCGACACCGA
  851	TCGCCCTTCT GATTTCCGTA TTGGTCGCAC TGTTTGTCTT
  	GGGACGCAAA
  901	CGCGGCGAAA GCGGCAGCGC GTTGGAAAAA ACCGTGGACG
  	GCGCACTCGC
  951	CCCCGTCTGT TCCGTGATTC TGATTACCGG CGCGGGCGGT
  	ATGTTCGGCG
  1001	GCGTTTTGCG CGCTTCCGGC ATCGGCAAGG CACTCGCCGA
  	CAGCATGGCG
  1051	GATTTGGGCA TTCCCGTCCT TTTGGGCTGT TTCCTTGTCG
  	CCTTGGCACT
  1101	GCGTATCGCG CAAGGTTCGG CAACCGTCGC CCTGACCACC
  	GCCGCCGCGC
  1151	TGATGGCTCC TGCCGTTGCC GCCGCCGGCT TTACCGACTG
  	GCAGCTCGCC
  1201	TGTATCGTAT TGGCAACGGC GGCAGGTTCG GTCGGTTGCA
  	GCCACTTCAA
  1251	CGACTCCGGC TTCTGGCTGG TCGGCCGTCT CTTGGACATG
  	GACGTACCGA
  1301	CCACGCTGAA AACCTGGACG GTCAACCAAA CCCTCATCGC
  	ACTCATCGGC
  1351	TTTGCCTTGT CCGCACTGCT GTTCGCCATC GTCTGA

• This corresponds to the amino acid sequence <SEQ ID 586; ORF140-1>:

• 1	MDGWTQTLSA QTLLGISAAA IILILILIVK FRIHALLTLV
  	IVSLLTALAT
  51	GLPTGSIVND ILVKNFGGTL  GGVALLVGLG AMLGRLV ETS
  	GGAQSLADAL
  101	IRMFGEKRAP FALGVASLIF GFPIFFDAGL IVMLPIVFAT
  	ARRMKQDVLP
  151	FALASIGAFS VMHVFLPPHP GPIAASEFYG ANIGQVLILG
  	LPTAFITWYF
  201	SGYMLGKVLG RTIHVPVPEL LSGGTQDNDL PKEPAKAGTV
  	VAIMLIPMLL
  251	IFLNTGVSAL ISEKLVSADE TWVQTAKIIG STPIALLISV
  	LVALFVLGRK
  301	RGESGSALEK TVDGALAPVC SVILITGAGG MFGGVLRASG
  	IGKALADSMA
  351	DLGIPVLLGC FLVALALRIA QGSATVALTT AAALMAPAVA
  	AAGFTDWQLA
  401	CIVLATAAGS VGCSHFNDSG FWLVGRLLDM DVPTTLKTWT
  	VNQTLIALIG
  451	FALSALLFAI V*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF140 shows 95.4% identity over a 87aa overlap with an ORF (ORF140a) from strain A of N. meningitidis:
• 
• The complete length ORF140a nucleotide sequence <SEQ ID 587> is:

• 1	ATGGACGGCT GGACACAGAC GCTGTCCGCG CAAACCCTGT
  	TGGGCATTTC
  51	GGCGGCGGCA ATCATCCTCA TTCTGATTTT AATCGTCAAA
  	TTCCGCATCC
  101	ACGCGCTGCT GACACTGGTC ATCGTCAGCC TGCTGACGGC
  	TTTGGCAACC
  151	GGTTTGCCCA CAGGCAGCAT TGTCAACGAC GTACTGGTCA
  	AAAACTTCGG
  201	CGGCACGCTC GGCGGCGTGG CGCTTCTGGT CGGCCTGGGC
  	GCGATGCTCG
  251	GACGTTTGGT CGAAACATCC GGCGGCGCAC AGTCGCTGGC
  	GGACGCGCTG
  301	ATCCGGATGT TCGGCGAAAA ACGCGCACCG TTCGCGCTGG
  	GCGTTGCCTC
  351	GCTGATTTTC GGCTTCCCGA TTTTCTTCGA TGCCGGACTA
  	ATCGTCATGC
  401	TGCCCATCGT GTTCGCCACC GCACGGCGCA TGAAACAGGA
  	CGTACTGCCC
  451	TTCGCGCTTG CCTCCATCGG CGCATTTTCC GTCATGCACG
  	TCTTCCTGCC
  501	GCCCCATCCG GGCCCGATTG CCGCTTCCGA ATTTTACGGC
  	GCGAACATCG
  551	GCCAAGTTTT GATTTTGGGT CTGCCGACCG CCTTCATCAC
  	ATGGTATTTC
  601	AGCGGCTATA TGCTCGGCAA AGTGTTGGGG CGCACCATCC
  	ATGTTCCCGT
  651	TCCCGAACTG CTCAGCGGCG GCACGCAAGA CAACGACCTG
  	CCGAAAGAAC
  701	CTGCCAAAGC AGGAACGGTC GTCGCCATCA TGCTGATTCC
  	CATGCTGCTG
  751	ATTTTCCTGA ATACCGGCGT ATCGGCCCTC ATCAGCGAAA
  	AACTCGTAAG
  801	TGCGGACGAA ACCTGGGTTC AGACGGCAAA AATAATCGGT
  	TCGACACCGA
  851	TCGCCCTTCT GATTTCCGTA TTGGTCGCAC TGTTTGTCTT
  	GGGACGCAAA
  901	CGCGGCGAAA GCGGCAGCGC GTTGGAAAAA ACCGTGGACG
  	GCGCACTCGC
  951	CCCCGTCTGT TCCGTGATTC TGATTACCGG CGCGGGCGGT
  	ATGTTCGGCG
  1001	GCGTTTTGCG CGCTTCCGGC ATCGGCAAGG CACTCGCCGA
  	CAGCATGGCG
  1051	GATTTGGGCA TTCCCGTCCT TTTGGGCTGT TTCCTTGTCG
  	CCTTGGCACT
  1101	GCGTATCGCG CAAGGTTCGG CAACCGTCGC CCTGACCACC
  	GCCGCCGCGC
  1151	TGATGGCTCC TGCCGTTGCC GCCGCCGGCT TTACCGACTG
  	GCAGCTCGCC
  1201	TGTATCGTAT TGGCAACGGC GGCAGGTTCG GTCGGTTGCA
  	GCCACTTCAA
  1251	CGACTCCGGC TTCTGGCTGG TCGGCCGCCT CTTGGACATG
  	GACGTACCGA
  1301	CCACGCTGAA AACCTGGACG GTCAACCAAA CCCTCATCGC
  	ACTCATCGGC
  1351	TTTGCCTTGT CCGCACTGCT GTTCGCCATC GTCTGA

• This encodes a protein having amino acid sequence <SEQ ID 588>:

• 1	MDGWTQTLSA QTLLGISAAA IILILILIVK FRIHALLTLV
  	IVSLLTALAT
  51	GLPTGSIVND VLVKNFGGTL  GGVALLVGLG AMLGRLV ETS
  	GGAQSLADAL
  101	IRMFGEKRAP FALGVASLIF GFPIFFDAGL IVMLPIVFAT
  	ARRMKQDVLP
  151	FALASIGAFS VMHVFLPPHP GPIAASEFYG ANIGQVLILG
  	LPTAFITWYF
  201	SGYMLGKVLG RTIHVPVPEL LSGGTQDNDL PKEPAKAGTV
  	VAIMLIPMLL
  251	IFLPNTGVSAL ISEKLVSADE TWVQTAKIIG STPIALLISV
  	LVALFVLGRK
  301	RGESGSALEK TVDGALAPVC SVILITGAGG MFGGVLRASG
  	IGKALADSMA
  351	DLGIPVLLGC FLVALALRIA QGSATVALTT AAALMAPAVA
  	AAGFTDWQLA
  401	CIVLATAAGS VGCSHFNDSG FWLVGRLLDM DVPTTLKTWT
  	VNQTLIALIG
  451	FALSALLFAI V*

• ORF140a and ORF140-1 show 99.8% identity over a 461aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF140 shows 92% identity over a 87aa overlap with a predicted ORF (ORF140ng) from N. gonorrhoeae:
• 
• The complete length ORF140ng nucleotide sequence <SEQ ID 589> was predicted to encode a protein having amino acid sequence <SEQ ID 590>:

• 	1	MDGRTQTLSA QTLLGISAAA IILILILIVK FRIRALLTLV
  		IASLLTALAT
  	51	GLPTGSIVND VLVKNFGGTL  GGVALLVGLG AMLGRLV ETS
  		GGAQSLADAL
  	101	IRMFGEKRAP FAPGVASLIF GFPIFFDAGL IVMLPIVFAT
  		ARRMKQDVLP
  	151	FALASVGAFS VMHVFLPPHP GPIAASEFYG ANIGQVLILG
  		LPTAFITWYF
  	201	SGYMLGKVLG RAIHVPVPEL LSGGTQDSDP PKEPAKAGTV
  		VAVMLIPMLL
  	251	IFLNTGVSAL ISEKLVSADE TWVQTAKMIG STPVALLISV
  		LAALLVLGRK
  	301	RGESGSTLEK TVDGALAPAC SVILITGAGG MFGGVLRASG
  		IGKALADSMA
  	351	DLGIPVLLGC FLVALALRIA QGSATVALTT AAALMAPAVA
  		AAGFTDWQLA
  	401	CIVLATAAGS VGCSHFNDSG FWLVGRLSDM DVPTTLKTWT
  		VNQTLIAFIG
  	451	FALSALLFAI V*

• Further work revealed a variant gonococcal DNA sequence <SEQ ID 591>:

• 1	ATGGACGGCC GGACACAGAC GCTGTCCGCG CAAACCTTGT
  	TGGGCATTTC
  51	GGCGGCGGCA ATCATCCTCA TTCTGATTTT AATCGTCAAA
  	TTCCGCATCC
  101	GCGCGCTGCT GACACTGGTC ATCGCCAGCC TGCTGACGGC
  	TTTGGCAACC
  151	GGTTTGCCCA CAGGCAGCAT CGTCAACGAC GTACTGGTCA
  	AAAACTTCGG
  201	CGGCACGCTC GGCGGCGTGG CGCTTCTGGT CGGTCTGGGC
  	GCAATGCTCG
  251	GACGTTTGGT AGAAACATCC GGCGGCGCAC AGTCGCTGGC
  	GGACGCGCTG
  301	ATCCGGATGT TCGGCGAAAA ACGCGCACCG TTCGCTCCGG
  	GCGTTGCCTC
  351	GCTGATTTTC GGCTTCCCGA TTTTCTTCGA TGCCGGACTA
  	ATCGTCATGC
  401	TGCCCATCGT ATTCGCCACC GCACGGCGCA TGAAACAGGA
  	CGTACTGCCC
  451	TTCGCGCTTG CCTCCGTCGG CGCATTTTCC GTCATGCACG
  	TCTTCCTGCC
  501	GCCCCATCCG GGCCCGATTG CCGCTTCCGA ATTTTACGGC
  	GCGAACATCG
  551	GCCAGGTTTT GATTTTGGGT CTGCCGACCG CCTTCATCAC
  	ATGGTATTTC
  601	AGCGGCTATA TGCTCGGCAA AGTGTTGGGG CGCGCCATCC
  	ATGTTCCCGT
  651	TCCCGAACTG CTCAGCGGCG GCACGCAAGA CAGCGACCCG
  	CCGAAAGAAC
  701	CTGCCAAAGC AGGAACGGTC GTCGCCGTCA TGCTGATTCC
  	CATGCTGCTG
  751	ATTTTCCTGA ATACCGGCGT ATCAGCCCTC ATCAGCGAAA
  	AACTCGTAAG
  801	TGCGGACGAA ACTTGGGTTC AGACGGCAAA AATGATCGGT
  	TCGACACCTG
  851	TCGCCCTTCT GATTTCCGTA TTGGCCGCAC TGTTGGTCTT
  	GGGACGCAAA
  901	CGCGGCGAAA GCGGCAGCAC GTTGGAAAAA ACCGTGGACG
  	GCGCACTCGC
  951	CCCCGCCTGT TCCGTGATTC TGATTACCGG CGCGGGCGGT
  	ATGTTCGGCG
  1001	GCGTTTTGCG CGCTTCCGGC ATCGGCAAGG CACTCGCCGA
  	CAGCATGGCG
  1051	GATTTGGGCA TTCCCGTCCT TTTGGGCTGC TTCCTTGTCG
  	CCTTGGCACT
  1101	GCGTATCGCG CAAGGTTCGG CAACCGTCGC CCTGACCACA
  	GCCGCCGCGC
  1151	TGATGGCTCC TGCCGTTGCC GCCGCCGGCT TTACCGACTG
  	GCAGCTCGCC
  1201	TGTATCGTAT TGGCAACGGC GGCAGGTTCG GTCGGTTGCA
  	GCCACTTCAA
  1251	CGACTCCGGC TTCTGGCTGG TCGGCCGCCT CTTGGATATG
  	GACGTACCGA
  1301	CCACGCTGAA AACCTGGACG GTCAACCAAA CCCTCATCGC
  	ATTCATCGGC
  1351	TTTGCCTTGT CCGCACTGCT GTTTGCCATC GTCTGA

• This corresponds to the amino acid sequence <SEQ ID 592; ORF140ng-1>:

• 1	MDGRTQTLSA QTLLGISAAA IILILILIVK FRIRALLTLV
  	IASLLTALAT
  51	GLPTGSIVND VLVKNFGGTL  GGVALLVGLG AMLGRLV ETS
  	GGAQSLADAL
  101	IRMFGEKRAP FAPGVASLIF GFPIFFDAGL IVMLPIVFAT
  	ARRMKQDVLP
  151	FALASVGAFS VMHVFLPPHP GPIAASEFYG ANIGQVLILG
  	LPTAFITWYF
  201	SGYMLGKVLG RAIHVPVPEL LSGGTQDSDP PKEPAKAGTV
  	VAVMLIPMLL
  251	IFLTGVSAL ISEKLVSADE TWVQTAKMIG STPVALLISV
  	LAALLVLGRK
  301	RGESGSTLEK TVDGALAPAC SVILITGAGG MFGGVLRASG
  	IGKALADSMA
  351	DLGIPVLLGC FLVALALRIA QGSATVALTT AAALMAPAVA
  	AAGFTDWQLA
  401	CIVLATAAGS VGCSHFNDSG FWLVGRLLDM DVPTTLKTWT
  	VNQTLIAFIG
  451	FALSALLFAI V*

• ORF140ng-1 and ORF140-1 show 96.3% identity over 461aa overlap:
• 
• Furthermore, ORF140ng-1 is homologous to an E. coli protein:

• gi|882633 (U29579) ORF_o454 [Escherichia coli] >gi|1789097 (AE000358) o454;
  This 454 aa ORF is 34% identical (9 gaps) to 444 residues of an approx.
  456 aa protein GNTP_BACLI SW: P46832 [Escherichia coli] Length = 454
  Score = 210 bits (529), Expect = 1e−53
  Identities = 130/384 (33%), Positives = 194/384 (49%), Gaps = 19/384 (4%)

  Query:	88	ETSGGAQSLADALIRMFGEKRAPFAPGVASLIFGFPIFFDAGLIVMLPIVFATARRMKQD	147
  		E SGGA+SLA+   R  G+KR   A  +A+   G P+FFD G I++ PI++  A+  K
  Sbjct:	80	EHSGGAESLANYFSRKLGDKRTIAALTLAAFFLGIPVFFDVGFIILAPIIYGFAKVAKIS	139
  Query:	148	VLPFALASVGAFSVMHVFLPPHPGPIAASEFYGANIGQVLILGLPTAFITWYFSGYMLGK	207
  		L F L   G    +HV +PPHPGP+AA+    A+IG + I+G+  + I    GY   K
  Sbjct:	140	PLKFGLPVAGIMLTVHVAVPPHPGPVAAAGLLHADIGWLTIIGIAIS-IPVGVVGYFAAK	198
  Query:	208	VLGRAIHVPVPELL----------SGGTQDSDPPKEPAKAGTVVAVMLIPMLLIFLNTGV	257
  		++ +  +    E+L           G T+ SD    P  A  V ++++IP+ +I   T
  Sbjct:	199	IINKRQYAMSVEVLEQMQLAPASEEGATKLSDKINPPGVA-LVTSLIVIPIAIIMAGT--	255
  Query:	258	SALISEKLVSADETWVQTAKMIGSTPXXXXXXXXXXXXXXGRKRGESGSTLEKTVDGALA	317
  		+S  L+      + T ++IGS                  +RG S       +  AL
  Sbjct:	256	---VSATLMPPSHPLLGTLQLIGSPMVALMIALVLAFWLLALRRGWSLQHTSDIMGSALP	312
  Query:	318	PACSVILITGAGGMFGGVLRASGIGKALADSMADLGIPVLLGCFLVALALRIAQGSXXXX	377
  		A  VIL+TGAGG+FG VL  SG+GKALA+ +  + +P+L   F+++LALR +QGS
  Sbjct:	313	TAAVVILVTGAGGVFGKVLVESGVGKALANMLQMIDLPLLPAAFIISLALRASQGS--AT	370
  Query:	378	XXXXXXXXXXXXXXXGFTDWQLACIVLATAAGSVGCSHFNDSGFWLVGRLLDMDVPTTLK	437
  		G   Q   + LA   G +G SH NDSGFW+V + L + V   LK
  Sbjct:	371	VAILTTGGLLSEAVMGLNPIQCVLVTLAACFGGLGASHINDSGFWIVTKYLGLSVADGLK	430
  Query:	438	TWTVNQTLIAFIGFALSALLFAIV	461
  		TWTV  T++ F GF ++  ++A++
  Sbjct:	431	TWTVLTTILGFTGFLITWCVWAVI	454

• Based on this analysis, including the identification of the presence of a putative leader sequence (double-underlined) and several putative transmembrane domains (single-underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 71
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 593>:

• 1	..GATTTCGGCA TATCGCCCGT GTATCTTTGG GTTGCCGCCG
  	CGTTCAAACA
  51	TTTGCTGTCG CCGTGGGCTG CCGACTCATA CGATGTCGCA
  	CGCTTTGCAG
  101	GCGTATTTTT TGCCGTTATC GGACTGACTT CCTGCGGCTT
  	TGCCGGTTTC
  151	AACTTTTTGG GCAGACACCA CGGGCGCAC. GTCGTCCTGA
  	TTCTCATCGG
  201	CTGTATCGGG CTGATTCCAG TTGCCCATTT CCTCAACCCC
  	GCTGCCGCCG
  251	CCTTTGCCGC CGCCGGACTG GTGCTGCACG GTTATTCTTT
  	GGCTCGCCGG
  301	CGCGTGATTG CCGCCTCTTT TCTGCTCGGT ACGGGCTGGA
  	CGCTGATGTC
  351	GTTGGCAGCA GCTTATCCGG CAGCATTTGC CCTGATGCTG
  	CCCTTGCCCG
  401	TACTGATGTT TTTCCGTCCG ..

• This corresponds to the amino acid sequence <SEQ ID 594; ORF141>:

• 1	..DFGISPVYLW VAAAFKHLLS PWAADSYDVA RFAGVFFAVI
  	GLTSCGFAGF
  51	NFLGRHHGRX VVLILIGCIG LIPVAHFLNP AAAAFAAAGL
  	VLHGYSLARR
  101	RVIAASFLLG TGWTLMSLAA AYPAAFALML PLPVLMFFRP
  	..

• Further work revealed the complete nucleotide sequence <SEQ ID 595>:

• 1	ATGCTGACCT ATACCCCGCC CGATGCCCGC CCGCCCGCCA
  	AAACCCACGA
  51	AAAGCCGTGG CTGCTGCTGT TGATGGCGTT TGCCTGGTTG
  	TGGCCCGGCG
  101	TGTTTTCCCA CGATTTGTGG AATCCTGACG AACCTGCCGT
  	CTATACCGCC
  151	GTCGAAGCAC TGGCAGGCAG CCCCACCCCC TTGGTTGCCC
  	ATCTGTTCGG
  201	TCAAACCGAT TTCGGCATAC CGCCCGTGTA TCTTTGGGTT
  	GCCGCCGCGT
  251	TCAAACATTT GCTGTCGCCG TGGGCTGCCG ACTCATACGA
  	TGCCGCACGC
  301	TTTGCAGGCG TATTTTTTGC CGTTATCGGA CTGACTTCCT
  	GCGGCTTTGC
  351	CGGTTTCAAC TTTTTGGGCA GACACCACGG GCGCAgCGTC
  	GTCCTGATTC
  401	TCATCGGCTG TATCGGGCTG ATTCCAGTTG CCCATTTCCT
  	CAACCCCGCT
  451	GCCGCCGCCT TTGCCGCCGC CGGACTGGTG CTGCACGGTT
  	ATTCTTTGGC
  501	TCGCCGGCGC GTGATTGCCG CCTCTTTTCT GCTCGGTACG
  	GGCTGGACGC
  551	TGATGTCGTT GGCAGCAGCT TATCCGGCAG CATTTGCCCT
  	GATGCTGCCC
  601	TTGCCCGTAC TGATGTTTTT CCGTCCGTGG CAAAGCAGGC
  	GTTTGATGTT
  651	GACGGCAGTC GCCTCACTTG CCTTTGCCCT GCCGCTTATG
  	ACCGTTTACC
  701	CGCTGCTCTT GGCAAAAACG CAGCCCGCGC TGTTCGCGCA
  	ATGGCTCGAC
  751	TATCACGTTT TCGGTACGTT CGGCGGCGTG CGGCACGTTC
  	AGACGGCATT
  801	CAGTTTGTTT TACTATCTGA AAAACCTGCT TTGGTTTGCA
  	TTGCCCGCGC
  851	TGCCGCTGGC GGTTTGGACG GTTTGCCGCA CGCGCCTGTT
  	TTCGACCGAC
  901	TGGGGGATTT TGGGCGTCGT CTGGATGCTT GCCGTTTTGG
  	TGCTGCTTGC
  951	CGTCAATCCG CAGCGTTTTC AGGATAACCT CGTCTGGCTG
  	CTTCCGCCGC
  1001	TTGCCCTGTT CGGCGCGGCG CAACTGGACA GCCTGAGGCG
  	CGGCGCGGCG
  1051	GCGTTTGTCA ACTGGTTCGG CATTATGGCG TTCGGACTGT
  	TTGCCGTGTT
  1101	CCTGTGGACG GGCTTTTTCG CCATGAATTA CGGCTGGCCC
  	GCCAAGCTTG
  1151	CCGAACGCGC CGCCTATTTC AGCCCGTATT ATGTTCCTGA
  	TATCGATCCC
  1201	ATTCCGATGG CGGTTGCCGT ACTGTTCACA CCCTTGTGGC
  	TGTGGGCGAT
  1251	TACCCGGAAA AACATACGCG GCAGGCAGGC GGTTACCAAC
  	TGGGCGGCAG
  1301	GCGTTACCCT GACCTGGGCT TTGCTGATGA CGCTGTTCCT
  	GCCGTGGCTG
  1351	GACGCGGCGA AAAGCCACGC GCCGGTCGTC CGGAGTATGG
  	AGGCATCGCT
  1401	TTCCCCGGAA TTGAAACGGG AGCTTTCAGA CGGCATCGAG
  	TGTATCGGCA
  1451	TAGGCGGCGG CGACCTGCAC ACGCGGATTG TTTGGACGCA
  	GTACGGCACA
  1501	TTGCCGCACC GCGTCGGCGA TGTACAATGC CGCTACCGCA
  	TCGTCCTCCT
  1551	GCCCCAAAAT GCGGATGCGC CGCAAGGCTG GCAGACGGTT
  	TGGCAGGGTG
  1601	CGCGTCCGCG CAACAAAGAC AGTAAGTTCG CACTGATACG
  	GAAAATCGGG
  1651	GAAAATATAT AA

• This corresponds to the amino acid sequence <SEQ ID 596; ORF141-1>:

• 1	MLTYTPPDAR PPAKTHEKPW LLLLMAFAWL WPGVFSHDLW
  	NPDEPAVYTA
  51	VEALAGSPTP LVAHLFGQTD FGIPPVYLWV AAAFKHLLSP
  	WAADSYDAAR
  101	FAGVFFAVIG LTSCGFAGFN FLGRHHGRSV VLILIGCIGL
  	IPVAHFLNPA
  151	AAAFAAAGLV LHGYSLARRR VIAASFLLGT GWTLMSLAAA
  	YPAAFALMLP
  201	LPVLMFFRPW QSRRLMLTAV ASLAFALPLM TV YPLLLAKT
  	QPALFAQWLD
  251	YHVFGTFGGV RHVQTAFSLF YYLKNLLWFA LPALPLAVWT
  	VCRTRLFSTD
  301	WGILGVVWML AVLVLLAVNP QRFQDNLVWL LPPLALFGAA
  	QLDSLRRGAA
  351	AFVNWFGIMA FGLFAVFLWT GFFAMNYGWP AKLAERAAYF
  	SPYYVPDIDP
  401	IPMAVAVLFT PLWLWAITRK NIRGRQAVTN WAAGVTLTWA
  	LLMTLFLPWL
  451	DAAKSHAPVV RSMEASLSPE LKRELSDGIE CIGIGGGDLH
  	TRIVWTQYGT
  501	LPHRVGDVQC RYRIVLLPQN ADAPQGWQTV WQGARPRNKD
  	SKFALIRKIG
  551	ENI*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF141 shows 95.0% identity over a 140aa overlap with an ORF (ORF141a) from strain A of N. meningitidis:
• 
• The complete length ORF141a nucleotide sequence <SEQ ID 597> is:

• 1	ATGCTGACCT ATACCCCGCC CGATGCCCGC CCGCCCGCCA
  	AAACCCACGA
  51	AAAGCCGTGG CTGTTGCTGT TGATGGCGTT TGCCTGGTTG
  	TGGCCCGGCG
  101	TGTTTTCCCA CGATTTGTGG AATCCTGACG AACCTGCCGT
  	CTATACCGCC
  151	GTCGAAGCAC TGGCAGGCAG CCCCACCCCT TTGGTTGCCC
  	ATCTGTTCGG
  201	TCAAATCGAT TTCGGCATAC CGCCCGTGTA TCTTTGGGTT
  	GCCGCCGCGT
  251	TCAAACATTT GCTGTCGCCG TGGGCTGCCG ACCCGTATGA
  	TGCCGCACGC
  301	TTTGCCGGCG TGTTTTTCGC CGTTGTCGGA CTGACTTCCT
  	GCGGCTTTGC
  351	CGGTTTCAAC TTTTTGGGCA GACACCACGG GCGCAGCGTC
  	GTCCTGATTC
  401	TCATCGGCTG TATCGGGCTG ATTCCGACCG TACACTTTCT
  	CAACCCCGCT
  451	GCCGCCGCCT TTGCCGCCGC CGGACTGGTG CTGCACGGTT
  	ATTCTTTGGC
  501	TCGCCGGCGC GTGATTGCCG CCTCTTTTCT GCTCGGTACG
  	GGTTGGACGC
  551	TGATGTCGTT GGCAGCAGCT TATCCGGCGG CATTTGCCCT
  	GATGCTGCCC
  601	CTGCCCGTGC TGATGTTTTT CCGTCCGTGG CAAAGCAGGC
  	GTTTGATGTT
  651	GACGGCAGTC GCCTCGCTTG CCTTTGCCCT GCCGCTTATG
  	ACCGTTTACC
  701	CGCTGCTCTT GGCAAAAACG CAGCCCGCGC TGTTCGCGCA
  	ATGGCTCGAC
  751	GATCACGTTT TCGGTACGTT CGGCGGCGTG CGGCACATTC
  	AGACGGCATT
  801	CAGTTTGTTT TACTATCTGA AAAACCTGCT TTGGTTTGCA
  	TTGCCTGCGC
  851	TGCCGCTGGC GGTTTGGACG GTTTGCCGCA CGCGCCTGTT
  	TTCGACCGAC
  901	TGGGGGATTT TGGGCGTCGT CTGGATGCTT GCCGTTTTGG
  	TGCTGCTTGC
  951	CGTCAATCCG CAGCGTTTTC AGGATAACCT CGTCTGGCTG
  	CTTCCGCCGC
  1001	TTGCCCTGTT CGGCGCGGCG CAACTGGACA GCCTGAGACG
  	CGGCGCGGCG
  1051	GCGTTTGTCA ACTGGTTCGG CATTATGGCG TTCGGACTGT
  	TTGCCGTGTT
  1101	CCTGTGGACG GGCTTTTTCG CCATGAATTA CGGCTGGCCC
  	GCCAAGCTTG
  1151	CCGAACGCGC CGCCTATTTC AGCCCGTATT ATGTTCCTGA
  	TATCGATCCC
  1201	ATTCCGATGG CGGTTGCCGT ACTGTTCACA CCCTTGTGGC
  	TGTGGGCGAT
  1251	TACCCGCAAA AACATACGCG GCAGGCAGGC GGTTACCAAC
  	TGGGCGGCAG
  1301	GCGTTACCCT GACCTGGGCT TTGCTGATGA CGCTGTTCCT
  	GCCGTGGCTG
  1351	GACGCGGCGA AAAGCCACGC GCCCGTCGTC CGGAGTATGG
  	AGGCATCGCT
  1401	TTCCCCGGAA TTAAAACGGG AGCTTTCAGA CGGCATCGAG
  	TGTATCGACA
  1451	TAGGCGGCGG CGACCTACAC ACGCGGATTG TTTGGACGCA
  	GTACGGCACA
  1501	TTGCCGCACC GCGTCGGCGA TGTACAATGC CGCTACCGCA
  	TCGTCCGCTT
  1551	GCCCCAAAAC GCGGATGCGC CGCAAGGCTG GCAGACGGTC
  	TGGCAGGGTG
  1601	CGCGCCCGCG CAACAAAGAC AGTAAGTTCG CACTGATACG
  	GAAAACCGGG
  1651	GAAAATATAT TAAAAACAAC AGATTGA

• This encodes a protein having amino acid sequence <SEQ ID 598>:

• 1	MLTYTPPDAR PPAKTHEKPW LLLLMAFAWL WPGVFSHDLW
  	NPDEPAVYTA
  51	VEALAGSPTP LVAHLFGQID FGIPPVYLWV AAAFKHLLSP
  	WAADPYDAAR
  101	FAGVFFAVVG LTSCGFAGFN FLGRHHGRSV VLILIGCIGL
  	IPTVHFLNPA
  151	AAAFAAAGLV LHGYSLARRR VIAASFLLGT GWTLMSLAAA
  	YPAAFALMLP
  201	LPVLMFFRPW QSRRLMLTAV ASLAFALPLM TV YPLLLAKT
  	QPALFAQWLD
  251	DHVFGTFGGV RHIQTAFSLF YYLKNLLWFA LPALPLAVWT
  	VCRTRLFSTD
  301	WGILGVVWML AVLVLLAVNP QRFQDNLVWL LPPLALFGAA
  	QLDSLRRGAA
  351	AFVNWFGIMA FGLFAVFLWT GFFAMNYGWP AKLAERAAYF
  	SPYYVPDIDP
  401	IPMAVAVLFT PLWLWAITRK NIRGRQAVTN WAAGVTLTWA
  	LLMTLFLPWL
  451	DAAKSHAPVV RSMEASLSPE LKRELSDGIE CIDIGGGDLH
  	TRIVWTQYGT
  501	LPHRVGDVQC RYRIVRLPQN ADAPQGWQTV WQGARPRNKD
  	SKFALIRKTG
  551	ENILKTTD*

• ORF141a and ORF141-1 show 98.2% identity in 553 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF141 shows 95% identity over a 140aa overlap with a predicted ORF (ORF141ng) from N. gonorrhoeae:
• 
• An ORF141ng nucleotide sequence <SEQ ID 599> was predicted to encode a protein having amino acid sequence <SEQ ID 600>:

• 1	MPSEAVSARP LCEYLLHLAI RPFLLTLMLT YTPPDARPPA
  	KTHEKPWLLL
  51	LMAFAWLWPG VFSHDLWNPA EPAVYTAVEA LAGSPTPLVA
  	HLFGQTDFGI
  101	PPVYLWVAAA FKHLLSPWAA HPYDAARFAG VFFAVIGLTS
  	CGFAGFNFLG
  151	RHHGRSVVLI HIGCIGLIPV AHFFNPAAAA FAAAGLVLHG
  	YSLARRRVIA
  201	ASFLLGTGWT LMSLAAAYPA AFALMLPLPV LMFF RPWQSR
  	RLMLTAVASL
  251	AFALPLMTVY PLLLAKTQPA LFAQWLNYHV FGTFGGVRHI
  	QRAFSLFHYL
  301	KNLLWFAPPG LPLAVWTVCR TRLFSTDWGI LGIVWMLAVL
  	VLLAFNPQRF
  351	QDNLVWLLPP LALFGAAQLD SLRRGAAAFV NWFGIMAFGL
  	FAVFLWTGFF
  401	AMNYGWPAKL AERAAYFSPY YVPDIDPIPM AVAVLFTPLW
  	LWAITRKNIR
  451	GRQAVTNWAA GVTLTWALLM TLFLPWLDAA KSHAPVVRSM
  	EASFSPELKR
  501	ELSDGIECIG IGGGDLHTRI VWTQYGTLPH RVGDVRCRYR
  	IVRLPQNADA
  551	PQGWQTVWQG ARPRNKDSKF ALIRKIGENI LKTTD*

• Further work revealed the following gonococcal DNA sequence <SEQ ID 601>:

• 1	ATGCTGACCT ATACCCCGCC CGATGCCCGC CCGCCCGCCA
  	AAACCCACGA
  51	AAAACCGTGG CTGCTGCTGT TGATGGCGTT TGCCTGGCTG
  	TGGCCCGGCG
  101	TGTTTTCCCA CGATTTGTGG AATCCTGCCG AACCTGCCGT
  	CTATACCGCC
  151	GTCGAAGCAC TGGCAGGCAG CCCCACCCCC TTGGTTGCCC
  	ATCTGTTCGG
  201	TCAAACCGAT TTCGGCATAC CGCCCGTGTA TCTTTGGGTT
  	GCCGCCGCAT
  251	TCAAACATTT GCTGTCGCCG TGGGCAGCCG ACCCGTATGA
  	TGCCGCACGC
  301	TTTGCAGGCG TATTTTTTGC CGTTATCGGA CTGACTTCTT
  	GCGGCTTTGC
  351	CGGTTTCAAC TTTTTGGGCA GACACCACGG GCGCAGCGTT
  	GTTTTAATCC
  401	ATATCGGCTG TATCGGGCTG ATTCCGGTTG CCCATTTCCT
  	CAATCCcgcc
  451	gccgccgcct tTGCCGCCGC CGGACTGGTG CTGCacggct
  	actcgctgGC
  501	ACGCCGGCGC GTGATtgccg cctctTtccT GCTCGGTACG
  	GGTTGGACGT
  551	TGATGTCGCT GGCGGCAGCT TATCCGGCGG CGTTTGCGCT
  	GATGCTGCCC
  601	CTGCCCGTGC TGATGTTTTT CCGTCCGTGG CAAAGCAGGC
  	GTTTGATGTT
  651	GACGGCAGTC GCCTCGCTTG CCTTTGCCCT GCCGCTTATG
  	ACCGTTTACC
  701	CGCTGCTCtt gGCAAAAACG CAGCCCGCGC TGTTTGCGCA
  	ATGGCTCAAC
  751	TATCACGTTT TCGGTACGTt cggcgGCGTG CGGCAcaTTC
  	AGAggGCatT
  801	Cagtttgttt cactatctgA AAaatctgct ttggttcgca
  	ccgcccgggC
  851	TGCCGCTGGC GGTTTGGACG GTTTGCCGCA CACGCCTGTT
  	TTCGACCGAC
  901	TGGGGGATTT TGGGCATTGT CTGGATGCTT GCCGTTTTGG
  	TGCTGCTCGC
  951	CTTTAATCCG CAGCGTTTTC AAGACAACCT CGTCTGGCTG
  	CTGCCGCCGC
  1001	TTGCCCTGTT CGGCGCGGCG CAACTGGACA GCCTGAGGCG
  	CGGCGCGGCG
  1051	GCTTTTGTCA ACTGGTTCGG CATTATGGCG TTCGGGCTGT
  	TTGCCGTGTT
  1101	CCTGTGGACG GGCTTTTTCG CCATGAATTA CGGCTGGCCC
  	GCCAAGCTTG
  1151	CCGAACGCGC CGCCTACTTC AGCCCGTATT ACGTTCCCGA
  	CATCGATCCC
  1201	ATTCCGATGG CGGTTGCCGT ACTGTTCACA CCCTTGTGGC
  	TGTGGGCGAT
  1251	TACCCGGAAA AACATACGCG GCAGGCAGGC GGTTACCAAC
  	TGGGCGGCAG
  1301	GCGTTACCCT GACCTGGGCT TTGCTGATGA CGCTGTTCCT
  	GCCGTGGCTG
  1351	GACGCGGCGA AAAGCCACGC GCCCGTCGTC CGGAGTATGG
  	AGGCATCGTT
  1401	TTCCCCGGAA TTAAAACGGG AGCTTTCAGA CGGCATCGAG
  	TGTATCGGCA
  1451	TAGGCGGCGG CGACCTGCAC ACGCGGATTG TTTGGACGCA
  	GTACGGCACA
  1501	TTGCCGCACC GCGTCGGCGA TGTCCGTTGC CGCTACCGTA
  	TCGTCCGCCT
  1551	GCCCCAAAAC GCGGATGCGC CGCAAGGCTG GCAGACGGTC
  	TGGCAGGGTG
  1601	CGCGCCCGCG CAACAAAGAC AGTAAGTTTG CACTGATACG
  	GAAAATCGGG
  1651	GAAAATATAT TAAAAACAAC AGATTGA

• This corresponds to the amino acid sequence <SEQ ID 602; ORF141ng-1>:

• 1	MLTYTPPDAR PPAKTHEKPW LLLLMAFAWL WPGVFSHDLW
  	NPAEPAVYTA
  51	VEALAGSPTP LVAHLFGQTD FGIPPVYLWV AAAFKHLLSP
  	WAADPYDAAR
  101	FAGVFFAVIG LTSCGFAGFN FLGRHHGRSV VLIHIGCIGL
  	IPVAHFLNPA
  151	AAAFAAAGLV LHGYSLARRR VIAASFLLGT GWTLMSLAAA
  	YPAAFALMLP
  201	LPVLMFFRPW QSRRLMLTAV ASLAFALPLM TV YPLLLAKT
  	QPALFAQWLN
  251	YHVFGTFGGV RHIQRAFSLF HYLKNLLWFA PPGLPLAVWT
  	VCRTRLFSTD
  301	WGILGIVWML AVLVLLAFNP QRFQDNLVWL LPPLALFGAA
  	QLDSLRRGAA
  351	AFVNWFGIMA FGLFAVFLWT GFFAMNYGWP AKLAERAAYF
  	SPYYVPDIDP
  401	IPMAVAVLFT PLWLWAITRK NIRGRQAVTN WAAGVTLTWA
  	LLMTLFLPWL
  451	DAAKSHAPVV RSMEASFSPE LKRELSDGIE CIGIGGGDLH
  	TRIVWTQYGT
  501	LPHRVGDVRC RYRIVRLPQN ADAPQGWQTV WQGARPRNKD
  	SKFALIRKIG
  551	ENILKTTD*

• ORF141ng-1 and ORF141-1 show 97.5% identity in 553 aa overlap:
• 
• Based on the presence of several putative transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 72
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 603>:

• 1	..CAATCCGCCA AATGGTTATC GGGCCAAACT CTAGTCGGCA
  	CAGCAATTGG
  51	GATACGCGGG CAGATAAAGC TTGGCGGCAA CCTGCATTAC
  	GATATATTTA
  101	CCGGCCGCGC ATTGAAAAAG CCCGAATTTT TCCAATCAAG
  	GAAATGGGCA
  151	AGCGGTTTTC AGGTAGGCTA TACGTTTTAA

• This corresponds to the amino acid sequence <SEQ ID 604; ORF142>:

• 1	..QSAKWLSGQT LVGTAIGIRG QIKLGGNLHY DIFTGRALKK
  	PEFFQSRKWA
  51	SGFQVGYTF*

• Further work revealed the complete nucleotide sequence <SEQ ID 605>:

• 1	ATGGATAATT CGGGTAGTGA GGCGACAGGA AAATACCAAG
  	GAAATATCAC
  51	TTTCTCTGCC GACAATCCTT TGGGACTGAG TGATATGTTC
  	TATGTAAATT
  101	ATGGACGTTC GATTGGCGGT ACGCCCGATG AGGAAAGTTT
  	TGACGGCCAT
  151	CGCAAAGAAG GCGGATCAAA CAATTACGCC GTACATTATT
  	CAGCCCCTTT
  201	CGGTAAATGG ACATGGGCAT TCAATCACAA TGGCTACCGT
  	TACCATCAGG
  251	CAGTTTCCGG ATTATCGGAA GTCTATGACT ATAATGGAAA
  	AAGTTACAAT
  301	ACTGATTTCG GCTTCAACCG CCTGTTGTAT CGTGATGCCA
  	AACGCAAAAC
  351	CTATCTCGGT GTAAAACTGT GGATGAGGGA AACAAAAAGT
  	TACATTGATG
  401	ATGCCGAACT GACTGTACAA CGGCGTAAAA CTGCGGGTTG
  	GTTGGCAGAA
  451	CTTTCCCACA AAGAATATAT CGGTCGCAGT ACGGCAGATT
  	TTAAGTTGAA
  501	ATATAAACGC GGCACCGGCA TGAAAGATGC TCTGCGCGCG
  	CCTGAAGAAG
  551	CCTTTGGCGA AGGCACGTCA CGTATGAAAA TTTGGACGGC
  	ATCGGCTGAT
  601	GTAAATACTC CTTTTCAAAT CGGTAAACAG CTATTTGCCT
  	ATGACACATC
  651	CGTTCATGCA CAATGGAACA AAACCCCGCT AACATCGCAA
  	GACAAACTGG
  701	CTATCGGCGG ACACCACACC GTACGTGGCT TCGACGGTGA
  	AATGAGTTTG
  751	TCTGCCGAGC GGGGATGGTA TTGGCGCAAC GATTTGAGCT
  	GGCAATTTAA
  801	ACCAGGCCAT CAGCTTTATC TTGGGGCTGA TGTAGGACAT
  	GTTTCAGGAC
  851	AATCCGCCAA ATGGTTATCG GGCCAAACTC TAGTCGGCAC
  	AGCAATTGGG
  901	ATACGCGGGC AGATAAAGCT TGGCGGCAAC CTGCATTACG
  	ATATATTTAC
  951	CGGCCGCGCA TTGAAAAAGC CCGAATTTTT CCAATCAAGG
  	AAATGGGCAA
  1001	GCGGTTTTCA GGTAGGCTAT ACGTTTTAA

• This corresponds to the amino acid sequence <SEQ ID 606; ORF142-1>:

• 1	MDNSGSEATG KYQGNITFSA DNPLGLSDMF YVNYGRSIGG
  	TPDEESFDGH
  51	RKEGGSNNYA VHYSAPFGKW TWAFNHNGYR YHQAVSGLSE
  	VYDYNGKSYN
  101	TDFGFNRLLY RDAKRKTYLG VKLWMRETKS YIDDAELTVQ
  	RRKTAGWLAE
  151	LSHKEYIGRS TADFKLKYKR GTGMKDALRA PEEAFGEGTS
  	RMKIWTASAD
  201	VNTPFQIGKQ LFAYDTSVHA QWNKTPLTSQ DKLAIGGHHT
  	VRGFDGEMSL
  251	SAERGWYWRN DLSWQFKPGH QLYLGADVGH VSGQSAKWLS
  	GQTLVGTAIG
  301	IRGQIKLGGN LHYDIFTGRA LKKPEFFQSR KWASGFQVGY
  	TF*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF142 shows 88.1% identity over a 59aa overlap with a predicted ORF (ORF142ng) from N. gonorrhoeae:
• 
• The complete length ORF142ng nucleotide sequence <SEQ ID 607> is:

• 1	ATGGATAATT CGGGTAGTGA GGCGACAGGA AAATACCAAG
  	GAAATATCAC
  51	TTTCTCTGCC GACAATCCTT TTGGACTGAG TGATATGTTC
  	TATGTAAATT
  101	ATGGACGTTC AATTGGCGGT ACGCCCGATG AGGAAAATTT
  	TGACGGCCAT
  151	CGCAAAGAAG GCGGATCAAA CAATTACGCC GTACATTATT
  	CAGCCCCTTT
  201	CGGTAAATGG ACATGGGCAT TCAATCACAA TGGCTACCGT
  	TACCATCAGG
  251	CGGTTTCCGG ATTATCGGAA GTCTATGACT ATAATGGAAA
  	AAGTTACAAC
  301	ACTGATTTCG GCTTCAACCG CCTGTTGTAT CGTGATGCCA
  	AACGCAAAAC
  351	CTATCTCAGT GTAAAACTGT GGACGAGGGA AACAAAAAGT
  	TACATTGATG
  401	ATGCCGAACT GACTGTACAA CGGCGTAAAA CCACAGGTTG
  	GTTGGCAGAA
  451	CTTTCCCACA AAGGATATAT CGGTCGCAGT ACGGCAGATT
  	TTAAGTTGAA
  501	ATATAAACAC GGCACCGGCA TGAAAGATGC TCTGCGCGCG
  	CCTGAAGAAG
  551	CCTTTGGCGA AGGCACGTCA CGTATGAAAA TTTGGACGGC
  	ATCGGCTGAT
  601	GTAAATACTC CTTTTCAAAT CGGTAAACAG CTATTTGCCT
  	ATGACACATC
  651	CGTTCATGCA CAATGGAACA AAACCCCGCT AACATCGCAA
  	GACAAACTGG
  701	CTATCGGCGG ACACCACACC GTACGTGGCT TCGACGGTGA
  	AATGAGTTTG
  751	CCTGCCGAGC GGGGATGGTA TTGGCGCAAC GATTTGAGCT
  	GGCAATTTAA
  801	ACCAGGCCAT CAGCTTTATC TTGGGGCTGA TGTAGGACAT
  	GTTTCAGGAC
  851	AATCCGCCAA ATGGTTATCG GGCCAAACTC TAGCCGGCAC
  	AGCAATTGGG
  901	ATACGCGGGC AGATAAAGCT TGGCGGCAAC CTGCATTACG
  	ATATATTTAC
  951	CGGCCGTGCA TTGAAAAAGC CCGAATATTT TCAGACGAAG
  	AAATGGGTAA
  1001	CGGGGTTTCA GGTGGGTTAT TCGTTTTGA

• This encodes a protein having amino acid sequence <SEQ ID 608>:

• 1	MDNSGSEATG KYQGNITFSA DNPFGLSDMF YVNYGRSIGG
  	TPDEENFDGH
  51	RKEGGSNNYA VHYSAPFGKW TWAFNHNGYR YHQAVSGLSE
  	VYDYNGKSYN
  101	TDFGFNRLLY RDAKRKTYLS VKLWTRETKS YIDDAELTVQ
  	RRKTTGWLAE
  151	LSHKGYIGRS TADFKLKYKH GTGMKDALRA PEEAFGEGTS
  	RMKIWTASAD
  201	VNTPFQIGKQ LFAYDTSVHA QWNKTPLTSQ DKLAIGGHHT
  	VRGFDGEMSL
  251	PAERGWYWRN DLSWQFKPGH QLYLGADVGH VSGQSAKWLS
  	GQTLAGTAIG
  301	IRGQIKLGGN LHYDIFTGRA LKKPEYFQTK KWVTGFQVGY
  	SF*

• The underlined sequence (aromatic-Xaa-aromatic amino acid motif) is usually found at the C-terminal end of outer membrane proteins.
• ORF142ng and ORF142-1 show 95.6% identity over 342aa overlap:
• 
• In addition, ORF142ng is homologous to the HecB protein of E. chrysanthemi:

• gi|1772622 (L39897) HecB [Erwinia chrysanthemi] Length = 558
  Score = 119 bits (295), Expect = 3e−26
  Identities = 88/346 (25%), Positives = 151/346 (43%),
  Gaps = 22/346 (6%)

  Query:	2	DNSGSEATGKYQGNITFSADNPFGLSDMFYVNYGRSIGGTPDEENFDGHRKEGGSNNYAV	61
  		DNSG ++TG+ Q N + + DN FGL+D ++++ G S   +    + D    + G
  Sbjct:	230	DNSGQKSTGEEQLNGSLALDNVFGLADQWFISAGHS---SRFATSHDAESLQAG------	280
  Query:	62	HYSAPFGKWTWAFNHNGYRYHQAVSGLSEVYDYNGKSYNTDFGFNRLLYRDAKRKTYLSV	121
  		+S P+G W   +N++  RY          +   G S    F  +R+++RD   KT ++
  Sbjct:	281	-FSMPYGYWNLGYNYSQSRYRNTFINRDFPWHSTGDSDTHRFSLSRVVFRDGTMKTAIAG	339
  Query:	122	KLWTRETKSYIDDAELTVQRRKTTGWLAELSHKGYIGRSTADFKLKYKHGTGMKDALRAP	181
  		R   +Y++ + L    RK +     ++H   +    A F   Y  G     +
  Sbjct:	340	TFSQRTGNNYLNGSLLPSSSRKLSSVSLGVNHSQKLWGGLATFNPTYNRGVRWLGSETDT	399
  Query:	182	EEAFGEGTSRMKIWTASADVNTPFQIGKQLFAYDTSVHAQWNKTPLTSQDKLAIGGHHTV	241
  		+++  E  +    WT SA    P         Y  S++ Q++   L   ++L +GG  ++
  Sbjct:	400	DKSADEPRAEFNKWTLSASYYHPV---TDSITYLGSLYGQYSARALYGSEQLTLGGESSI	456
  Query:	242	RGFDGEMSLPAERGWYWRNDLSWQFKP----GHQLYLGA-DVGHVSGQSAKWLSGQTLAG	296
  		RGF  E      RG YWRN+L+WQ       G+  ++ A D GH+        +  +L G
  Sbjct:	457	RGF-REQYTSGNRGAYWRNELNWQAWQLPVLGNVTFMAAVDGGHLYNHKQDNSTAASLWG	515
  Query:	297	TAIGIRGQIKLGGNLHYDIFTGRALKKPEYFQTKKWVTGFQVGYSF	342
  		A+G+    +    L   +  G  +  P + Q    V G++VG SF
  Sbjct:	516	GAVGMTVASRW---LSQQVTVGWPISYPAWLQPDTMVVGYRVGLSF	558

• On the basis of this analysis, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 73
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 609>:

• 1	ATGCGGACGA AATGGTCAGC AGTGAGAAGC TGCTTACTTG
  	GgCGGACACC
  51	GCCGACATCG ATACCGCTTT GAACCTGTTG TACCGTTTGC
  	AAAAACTCGA
  101	ATTCCTCTAT GGCGATGAAA ACGGTCATTC AGACGGCATC
  	AATTTGwCGG
  151	ACGAGCAATT GCCGTTGCTG ATGGAACAAT TGTCCGGCAG
  	CGGTAAGGCG
  201	TTATTGGTCG ATCGGAACGG TCTGTATCTT GCCAACGCCA
  	ATTTCCATCA
  251	TGAGGCGGCG GAAGAGTTGG GGTTGTTGGC GGCAGAAGTC
  	GCACAGATGG
  301	AAAAGAAATA CCGGCTGCTG ATTAAGAACA AC..

• This corresponds to the amino acid sequence <SEQ ID 610; ORF143>:

• 1	MRTKWSAVRS CTWADTADID TALNLLYRLQ KLEFLYGDEN
  	GHSDGINLXD
  51	EQLPLLMEQL SGSGKALLVD RNGLYLANAN FHHEAAEELG
  	LLAAEVAQME
  101	KKYRLLIKNN ..

• Further work revealed the complete nucleotide sequence <SEQ ID 611>:

• 1	ATGGAATCAA CACTTTCACT ACAAGCAAAT TTATATCCCC
  	GCCTGACTCC
  51	TGCCGGTGCA TTTTATGCCG TATCCAGCGA TGCCCCCAGT
  	GCCGGTAAAA
  101	CTTTGTTGCA CAGCCTGTTG AAAGCAGATG CGGACGAAAT
  	GGTCAGCAGT
  151	GAGAAGCTGC TTACTTGGGC GGACACCGCC GACATCGATA
  	CCGCTTTGAA
  201	CCTGTTGTAC CGTTTGCAAA AACTCGAATT CCTCTATGGC
  	GATGAAAACG
  251	GTCATTCAGA CGGCATCAAT TTGTCGGACG AGCAATTGCC
  	GTTGCTGATG
  301	GAACAATTGT CCGGCAGCGG TAAGGCGTTA TTGGTCGATC
  	GGAACGGTCT
  351	GTATCTTGCC AACGCCAATT TCCATCATGA GGCGGCGGAA
  	GAGTTGGGGT
  401	TGTTGGCGGC AGAAGTCGCA CAGATGGAAA AGAAATACCG
  	GCTGCTGATT
  451	AAGAACAACC TGTATATCAA CAATAACGCT TGGGGCGTTT
  	GCGATCCTTC
  501	CGGTCAGAGC GAATTGACAT TTTTCCCATT GTATATCGGT
  	TCAACCAAAT
  551	TTATTTTGGT TATCGGCGGC ATTCCCGATT TGGGCAAAGA
  	GGCATTTGTT
  601	ACTTTGGTAA GGATTTTATA CCGCCGTTAC AGCAACCGCG
  	TGTAA

• This corresponds to the amino acid sequence <SEQ ID 612; ORF143-1>:

• 1	MESTLSLQAN LYPRLTPAGA FYAVSSDAPS AGKTLLHSLL
  	KADADEMVSS
  51	EKLLTWADTA DIDTALNLLY RLQKLEFLYG DENGHSDGIN
  	LSDEQLPLLM
  101	EQLSGSGKAL LVDRNGLYLA NANFHHEAAE ELGLLAAEVA
  	QMEKKYRLLI
  151	KNNLYINNNA WGVCDPSGQS ELTFFPLYIG STKFILVIGG
  	IPDLGKEAFV
  201	TLVRILYRRY SNRV*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF143 shows 92.4% identity over a 105aa overlap with an ORF (ORF143a) from strain A of N. meningitidis:
• 
• The complete length ORF143a nucleotide sequence <SEQ ID 613> is:

• 1	ATGGAATCAA CANTTTCACT ACAAGCAAAT TTATATCNCC
  	GCCTGACTCC
  51	TGCCGGTGCA TTTTATGCCG TATCCAGCGA TGNCCCCAGT
  	GCCGGTAAAA
  101	CTTTGTTGCA CAGCCTGTTG AAAGCGGATG CGGACGAAAT
  	GGTNAGCAGT
  151	GAGAAGCTGC TTACCTGGGC GGANACCGCC GACATCGATA
  	CCGCTTTGAA
  201	CCTGTTGTAC CGTTTGCAAA AACTCGAATT CCTCTATGGC
  	GATGAAAACG
  251	GTCATTCAGA CGGCATCAAT TTGTCGGACG AGCAATTGCC
  	GTTGCTGATG
  301	GAACAATTGT CCGGCAGCGG TAAGGCGTTA TTGGTCGATC
  	GGAACGGTCT
  351	GTATCTTGCC AACGCCAATT TCCATCATGA GGCGGCGGAA
  	GAGTTGGGGT
  401	TGTTGGCGGC AGAAGTCGCA CAGATGGAAA AGAAATACCG
  	GCTGCNNATT
  451	AAGAACAACC TGTATATCAA CAATAACGCT TGGGGCGTTT
  	GCGATCCTTC
  501	CGGTCAGAGC GAATTGACAT TTTTCCCATT GTATATCGGT
  	TCAACCAAAT
  551	TTATTTTGGT TATCGGCGGC ATTCCCGATT TGGGCAAAGA
  	GGCATTTGTT
  601	ACTTTGGTAA GGATNTTATA CCNCCNGTTA CAGCAACCGC
  	GTGTAAAACT
  651	TGGGAGAGAG GANGGGTTAT GCAGCAATTA TTGA

• This encodes a protein having amino acid sequence <SEQ ID 614>:

• 1	MESTXSLQAN LYXRLTPAGA FYAVSSDXPS AGKTLLHSLL
  	KADADEMVSS
  51	EKLLTWAXTA DIDTALNLLY RLQKLEFLYG DENGHSDGIN
  	LSDEQLPLLM
  101	EQLSGSGKAL LVDRNGLYLA NANFHHEAAE ELGLLAAEVA
  	QMEKKYRLXI
  151	KNNLYINNNA WGVCDPSGQS ELTFFPLYIG STKFILVIGG
  	IPDLGKEAFV
  201	TLVRXLYXXL QQPRVKLGRE XGLCSNY*

• ORF143a and ORF143-1 show 97.1% identity in 207 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF143 shows 95.5% identity over a 110aa overlap with a predicted ORF (ORF143ng) from N. gonorrhoeae:
• 
• An ORF143ng nucleotide sequence <SEQ ID 615> was predicted to encode a protein having amino acid sequence <SEQ ID 616>:

• 1	MRTKWSAVRS CSRADTADID TALNLLYRLQ KLEFLYGDEN
  	GHSDGINLSD
  51	EQLPLLMEQL SGSGKALLVD RNGLYLANAN FHHESAEELG
  	LLAAEVAQME
  101	KKYRLLIRNN LYINNNAWGV CDPSGQSELT FFPLYIGSTK
  	FILVIAGIPD
  151	LSKGGICYFG KDFIPPLQQP RVKLGTGGIM RQLLISILED
  	LNNTSTDIIA
  201	SAVISTDGLP MATMLPSHLN SDRVGAISAT LLALGSRSVQ
  	ELACGELEQV
  251	MIKGKSGYIL LSQAGKDAVL VLVAKETGRL GLILLDAKRA
  	ARHIAEAI*

• Further work revealed the following gonococcal DNA sequence <SEQ ID 617>:

• 1	ATGGAATCAA CACTTTCACT ACAAGCGAAT TTATATCCCT
  	GCCTGACTCC
  51	TGCCGGTGCA TTTTATGCCG TATCCAGCGA TGCCCCCAGT
  	GCCGGTAAAA
  101	CTTTGTTGCG CAGCCTGTTG AAAGCGGATG CGGACGAAGT
  	GGTCAGCAGT
  151	GAGAAGCTGC TCGCGGCGGA CACCGCCGAC ATCGATACCG
  	CTTTGAACCT
  201	GTTGTACCGT TTGCAAAAAC TCGAATTCCT CTATGGCGAT
  	GAAAACGGTC
  251	ATTCAGACGG CATCAATTTG TCGGACGAGC AATTGCCGTT
  	GCTGATGGAA
  301	CAATTGTCCG GCAGCGGTAA GGCATTATTG GTCGATCGGA
  	ACGGTCTGTA
  351	TCTTGCCAAC GCCAATTTCC ATCATGAGTC GGCGGAAGAG
  	TTGGGGTTGT
  401	TGGCGGCAGA AGTCGCACAG ATGGAAAAGA AATACCGGCT
  	GCTGATTAGG
  451	AACAACCTGT ATATCAACAA TAACGCTTGG GGCGTTTGCG
  	ATCCTTCCGG
  501	TCAGAGCGAA TTGACATTTT TCCCATTGTA TATCGGTTCA
  	ACCAAATTTA
  551	TTTTGGTTAT CGCCGGCATT CCCGATTTGA GCAAAGAGGC
  	ATTTGTTACT
  601	TTGGTAAGGA TTTTATACCG CCGTTACAGC AACCGCGTGT
  	AA

• This corresponds to the amino acid sequence <SEQ ID 618; ORF143ng-1>:

• 1	MESTLSLQAN LYPCLTPAGA FYAVSSDAPS AGKTLLRSLL
  	KADADEVVSS
  51	EKLLAADTAD IDTALNLLYR LQKLEFLYGD ENGHSDGINL
  	SDEQLPLLME
  101	QLSGSGKALL VDRNGLYLAN ANFHHESAEE LGLLAAEVAQ
  	MEKKYRLLIR
  151	NNLYINNNAW GVCDPSGQSE LTFFPLYIGS TKFILVIAGI
  	PDLSKEAFVT
  201	LVRILYRRYS NRV*

• ORF143ng-1 and ORF143-1 show 95.8% identity in 214 aa overlap:
• 
• Based on the presence of the putative transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 74
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 619>:

• 1	ATGACCTTTT TACAACGTTT GCAAGGTTTG GCAGACAATA
  	AAATCTGTGC
  51	GTTTGCATGG TTCGTCGTCC GCCGCTTTGA TGAAGAACGC
  	GTACCGCAGr
  101	CGGCGGCAAG CATGACGTTT ACGACGCTGC TGGCACTCGT
  	CCCCGTGCTG
  151	ACCGTGATGG TGGCGGTCGC TTCGATTTTC CCCGTGTTCG
  	ACCGCTGGTC
  201	GGATTCGTTC GTCTCCTTCG TCAACCAAAC CATTGTGCCG
  	CA.GGCGCGG
  251	ACATGGTGTT CGACTATATC AATGCGTTCC GCGAGCAGGC
  	GAACCGGCTG
  301	ACGGCAATCG GCAGCGTGAT GCTGGTCGTT ACCTCGCTGA
  	TGCTGATTCG
  351	GACGATAGAC AATACGTTCA ACCGCATCTG GaCGGGTCAA
  	wTyCCAGCGT
  401	CCGTGGATG..

• This corresponds to the amino acid sequence <SEQ ID 620; ORF144>:

• 1	MTFLQRLQGL ADNKICAFAW FVVRRFDEER VPQXAASMTF
  	TTLLALVPVL
  51	TVMVAVASIF PVFDRWSDSF VSFVNQTIVP XGADMVFDYI
  	NAFREQANRL
  101	TAIGSVMLVV TSLMLIRTID NTFNRIWRVX XQRPWM...

• Further work revealed the complete nucleotide sequence <SEQ ID 621>:

• 1	ATGACCTTTT TACAACGTTT GCAAGGTTTG GCAGACAATA
  	AAATCTGTGC
  51	GTTTGCATGG TTCGTCGTCC GCCGCTTTGA TGAAGAACGC
  	GTACCGCAGG
  101	CGGCGGCAAG CATGACGTTT ACGACGCTGC TGGCACTCGT
  	CCCCGTGCTG
  151	ACCGTGATGG TGGCGGTCGC TTCGATTTTC CCCGTGTTCG
  	ACCGCTGGTC
  201	GGATTCGTTC GTCTCCTTCG TCAACCAAAC CATTGTGCCG
  	CAGGGCGCGG
  251	ACATGGTGTT CGACTATATC AATGCGTTCC GCGAGCAGGC
  	GAACCGGCTG
  301	ACGGCAATCG GCAGCGTGAT GCTGGTCGTT ACCTCGCTGA
  	TGCTGATTCG
  351	GACGATAGAC AATACGTTCA ACCGCATCTG GCGGGTCAAT
  	TCCCAGCGTC
  401	CGTGGATGAT GCAGTTTCTC GTCTATTGGG CTTTACTGAC
  	GTTCGGGCCG
  451	CTGTCTTTGG GCGTGGGCAT TTCCTTTATG GTCGGCTCGG
  	TACAGGATGC
  501	CGCGCTTGCC TCAGGTGCGC CGCAGTGGTC GGGCGCGTTG
  	CGAACGGCGG
  551	CGACGCTGAC CTTCATGACG CTTTTGCTGT GGGGGCTGTA
  	CCGCTTCGTG
  601	CCAAACCGCT TCGTTCCCGC GCGGCAGGCG TTTGTCGGGG
  	CTTTGGCAAC
  651	AGCGTTTTGT CTGGAAACCG CGCGCTCCCT CTTCACTTGG
  	TATATGGGCA
  701	ATTTCGACGG CTACCGCTCG ATTTACGGCG CGTTTGCCGC
  	CGTGCCGTTT
  751	TTTCTGTTGT GGCTGAACCT GTTGTGGACG CTGGTCTTGG
  	GCGGCGCGGT
  801	GCTGACTTCT TCACTCTCCT ACTGGCAGGG AGAAGCGTTC
  	CGCAGGGGCT
  851	TCGACTCGCG CGGACGGTTT GACGACGTGT TGAAAATCCT
  	GCTGCTTCTG
  901	GATGCGGCGC AAAAAGAAGG CAAAGCCTTG CCTGTTCAGG
  	AGTTCAGACG
  951	GCATATCAAT ATGGGCTACG ACGAGTTGGG CGAGCTTTTG
  	GAAAAGCTGG
  1001	CGCGGCACGG CTACATCTAT TCCGGCAGAC AGGGTTGGGT
  	GTTGAAAACG
  1051	GGGGCGGATT CGATTGAGTT GAACGAACTC TTCAAGCTCT
  	TCGTTTACCG
  1101	TCCGTTGCCT GTGGAAAGGG ATCATGTGAA CCAAGCTGTC
  	GATGCGGTAA
  1151	TGACACCGTG TTTGCAGACT TTGAACATGA CGCTGGCAGA
  	GTTTGACGCT
  1201	CAGGCGAAAA AACGGCAGTA G

• This corresponds to, the amino acid sequence <SEQ ID 622; ORF144-1>:

• 1	MTFLQRLQGL ADNKICAFAW FVVRRFDEER VPQAAASMTF
  	TTLLALVPVL
  51	TVMVAVASIF PVFDRWSDSF VSFVNQTIVP QGADMVFDYI
  	NAFREQANRL
  101	TAIGSVMLVV TSLMLIRTID NTFNRIWRVN SQRPWMMQFL
  	VYWALLTFGP
  151	LSLGVGISFM VGSVQDAALA SGAPQWSGAL RTAATLTFMT
  	LLLWGLYRFV
  201	PNRFVPARQA FVGALATAFC LETARSLFTW YMGNFDGYRS
  	IYGAFAAVPF
  251	FLLWLNLLWT LVLGGAVLTS SLSYWQGEAF RRGFDSRGRF
  	DDVLKILLLL
  301	DAAQKEGKAL PVQEFRRHIN MGYDELGELL EKLARHGYIY
  	SGRQGWVLKT
  351	GADSIELNEL FKLFVYRPLP VERDHVNQAV DAVMTPCLQT
  	LNMTLAEFDA
  401	QAKKRQ*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF144 shows 96.3% identity over a 136aa overlap with an ORF (ORF144a) from strain A of N. meningitidis:
• 
• The complete length ORF144a nucleotide sequence <SEQ ID 623> is:

• 1	ATGACCTTTT TACAACGTTT GCAAGGTTTG GCAGACAATA
  	AAATCTGTGC
  51	GTTTGCATGG TTCGTCGTCC GCCGCTTTGA TGAAGAACGC
  	GTACCGCAGG
  101	CGGCGGCAAG CATGACGTTT ACGACACTGC TGGCACTCGT
  	CCCCGTGCTG
  151	ACCGTGATGG TGGCGGTCGC TTCGATTTTC CCCGTGTTCG
  	ACCGNTGGTC
  201	GGATTCGTTC GTCTCCTTCG TCAACCAAAC CATTGTGCCG
  	CAGGGCGCGG
  251	ACATGGTNTT CGACTATATC AATGCGTTCC GCGAGCAGGC
  	GAACCGGCTG
  301	ACGGCAATCG GCAGCGTGAT GCTGGTCGTT ACCTCGCNGA
  	TGCTGATTCG
  351	GACGATAGAC AATACGTTCA ACCGCATCTG GCGGGTCAAT
  	TCCCAGCGTC
  401	CGTGGATGAT GCAGTTTCTC GTCTATTGGG CTTTACTGAC
  	GTTCGGGCCG
  451	CTGTCTTTGG GCGTGGGCAT TTCCTTTATN GTCGGCTCGG
  	TACAGGATGC
  501	CGCGCTTGCC TCAGGTGCGC CGCAGTGGTC GGGCGCGTTG
  	CGAACGGCGG
  551	CGACGCTGAN CTTCATGACG CTTTTGCTGT GGGGGCTGTA
  	CCGCTNCGTG
  601	CCAAACCGCT TCGTTCCCGC GCGGCANGCG TTTGTCGGGG
  	CTTTGGCAAC
  651	AGCGTTCTGT CTGGAAACCG CGCGTTCCCT CTTTACTTGG
  	TATATGGGCA
  701	ATTTCGACGG CTACCGCTCG ATTTACGGNG CGTTTGCCGC
  	CGTGCCGTTT
  751	TTTCTGTTGT GGCTGAACCT GTTGTGGACG CTGGTCTTGG
  	GCGGCGCGGT
  801	GCTGACTTCT TCACTCTCCT ACTGGCAGGG AGAAGCGTTC
  	CGCAGGGNCT
  851	TCGACTCGCG CGGACGGTTT GACGACGTGT TGAAAATCCT
  	GCTGCTTCTG
  901	GATGCGGCGC AAAAAGAAGG CNAAGCCTTG CCTGTTCAGG
  	AGTTCAGACG
  951	GCATATCAAT ATGGGCTACG ACGAGTTGGG CGAGCTTTTG
  	GAAAAGCTGG
  1001	CGCGGCACGG CTACATCTAT TCCGGCAGAC AGGGTTGGGT
  	GTTGAAAACG
  1051	GGGGCGGATT CGATTGAGTT GAACGAACTC TTCAAGCTCT
  	TCGTTTACCG
  1101	TCCGTTGCCT GTGGAAAGGG ATCATGTGAA CCAAGCTGTC
  	GATGCGGTAA
  1151	TGATGCCGTG TTTGCAGACT TTGAACATGA CGCTGGCAGA
  	GTTTGACGCT
  1201	CAGGCGAAAA AACAGCAGCA ATCTTGA

• This encodes a protein having amino acid sequence <SEQ ID 624>:

• 1	MTFLQRLQGL ADNKICAFAW FVVRRFDEER VPQAAASMTF
  	TTLLALVPVL
  51	TVMVAVASIF PVFDRWSDSF VSFVNQTIVP QGADMVFDYI
  	NAFREQANRL
  101	TAIGSVMLVV TSXMLIRTID NTFNRIWRVN SQRPWMMQFL
  	VYWALLTFGP
  151	LSLGVGISFX VGSVQDAALA SGAPQWSGAL RTAATLXFMT
  	LLLWGLYRXV
  201	PNRFVPARXA FVGALATAFC LETARSLFTW YMGNFDGYRS
  	IYGAFAAVPF
  251	FLLWLNLLWT LVLGGAVLTS SLSYWQGEAF RRXFDSRGRF
  	DDVLKILLLL
  301	DAAQKEGXAL PVQEFRRHIN MGYDELGELL EKLARHGYIY
  	SGRQGWVLKT
  351	GADSIELNEL FKLFVYRPLP VERDHVNQAV DAVMMPCLQT
  	LNMTLAEFDA
  401	QAKKQQQS*

• ORF144a and ORF144-1 show 97.8% identity in 406 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF144 shows 91.2% identity over a 136aa overlap with a predicted ORF (ORF144ng) from N. gonorrhoeae:
• 
• The complete length ORF144ng nucleotide sequence <SEQ ID 625> is predicted to encode a protein having amino acid sequence <SEQ ID 626>:

• 1	MTFLQCWQGS ADNKICAFAW FVIRRFSEER VPQAAASMTF
  	TTLLALVPVL
  51	TVMVAVASIF PVFDRWSDSF VSFVNQTIVP QGADMVFDYI
  	DAFRDQANRL
  101	TAIGSVMLVV TSLMLIRTID NAFNRIWRVN TQRPWMMQFL
  	VYWALLTFGP
  151	LSLGVGISFM VGSVQDSVLS SGAQQWADAL KTAARLAFMT
  	LLLWGLYRFV
  201	PNRFVPARQA FVGALITAFC LETARFLFTW YMGNFDGYRS
  	IYGAFAAVPF
  251	FLLWLNLLWT LVLGGAVLTS SLSYWQGEAF RRGFDSRGRF
  	DDVLKILLLL
  301	DAAQKEGRTL SVQEFRRHIN MGYDELGELL EKLARYGYIY
  	SGRQGWVLKT
  351	GADSIELSEL FKLFVYRPLP VERDHVNQAV DAVMTPCLQT
  	LNMTLAEFDA
  401	QAKKQQQS*

• Further work revealed the following gonococcal DNA sequence <SEQ ID 627>:

• 1	ATGACCTTTT TACAACGTTG GCAAGGTTTG GCGGACAATA
  	AAATCTGTGC
  51	ATTTGCATGG TTCGTCATCC GCCGTTTCAG TGAAGAGCGC
  	GTACCGCAGG
  101	CAGCGGCGAG CATGACGTTT ACGACACTGC TGGCACTCGT
  	CCCCGTACTG
  151	ACCGTAATGG TCGCGGTCGC TTCGATTTTC CCCGTGTTCG
  	ACCGCTGGTC
  201	GGATTCGTTC GTCTCCTTCG TCAACCAAAC CATTGTGCCG
  	CAGGGCGCGG
  251	ATATGGTGTT CGACTATATC GACGCATTCC GCGATCAGGC
  	AAACCGGCTG
  301	ACCGCCATCG GCAGCGTGAT GCTGGTCGTA ACCTCGCTGA
  	TGCTGATTCG
  351	GACGATAGAC AATGCGTTCA ACCGCATCTG GCGGGTTAAC
  	ACGCAACGCC
  401	CCTGGATGAT GCAGTTCCTC GTTTATTGGG CGTTGCTGAC
  	TTTCGGGCCT
  451	TTGTCTTTGG GTGTGGGCAT TTCCTTTATG GTCGGGTCGG
  	TTCAAGACTC
  501	CGTACTCTCC TCCGGAGCGC AACAATGGGC GGACGCGTTG
  	AAGACGGCGG
  551	CAAGGCTGGC TTTCATGACG CTTTTGCTGT GGGGGCTGTA
  	CCGCTTCGTG
  601	CCCAACCGCT TCGTGCCCGC CCGGCAGGCG TTTGTCGGAG
  	CTTTGATTAC
  651	GGCATTCTGC CTGGAGACGG CACGTTTCCT GTTCACCTGG
  	TATATGGGCA
  701	ATTTCGACGG CTACCGCTCG ATTTACGGCG CATTTGCCGC
  	CGTGCCGTTT
  751	TTCCTGCTGT GGTTAAACCT GCTGTGGACG CTGGTCTTGG
  	GCGGGGCGGT
  801	GCTGACTTCG TCGCTGTCTT ATTGGCAGGG CGAGGCCTTC
  	CGCAGGGGAT
  851	TCGACTCGCG CGGACGGTTT GACGACGTGT TGAAAATCCT
  	GCTGCTTCTG
  901	GATGCGGCGC AAAAAGAAGG CCGAACCCTG TCCGTTCAGG
  	AGTTCAGACG
  951	GCATATCAAT ATGGGTTACG ATGAATTGGG CGAGCTTTTG
  	GAAAAGCTGG
  1001	CGCGGTACGG CTATATCTAT TCCGGCAGAC AGGGCTGGGT
  	TTTGAAAACG
  1051	GGGGCGGATT CGATTGAGTT GAGCGAACTC TTCAAGCTCT
  	TCGTGTACCG
  1101	CCCGTTGCct gtggaAAGGG ATCATGTGAA CCAAGCTGtc
  	gaTGCGGTAA
  1151	TGAcgccgtG TTTGCAGACT TTGAACATGA CGCTGGCGGA
  	GTTTGACGCT
  1201	CAGgcgAAAA AACAGCAGCA GTCTTGA

• This encodes a variant of ORF144ng, having the amino acid sequence <SEQ ID 628; ORF144ng-1>:

• 1	MTFLQCWQGL ADNKICAFAW FVIRRFSEER VPQAAASMTF
  	TTLLALVPVL
  51	TVMVAVASIF PVFDRWSDSF VSFVNQTIVP QGADMVFDYI
  	DAFRDQANRL
  101	TAIGSVMLVV TSLMLIRTID NAFNRIWRVN TQRPWMMQFL
  	VYWALLTFGP
  151	LSLGVGISFM VGSVQDSVLS SGAQQWADAL KTAARLAFMT
  	LLLWGLYRFV
  201	PNRFVPARQA FVGALITAFC LETARFLFTW YMGNFDGYRS
  	IYGAFAAVPF
  251	FLLWLNLLWT LVLGGAVLTS SLSYWQGEAF RRGFDSRGRF
  	DDVLKILLLL
  301	DAAQKEGRTL SVQEFRRHIN MGYDELGELL EKLARYGYIY
  	SGRQGWVLKT
  351	GADSIELSEL FKLFVYRPLP VERDHVNQAV DAVMTPCLQT
  	LNMTLAEFDA
  401	QAKKQQQS*

• ORF144ng-1 and ORF144-1 show 94.1% identity in 406 aa overlap:
• 
• On this basis of this analysis, including the identification of several putative transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 75
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 629>:

• 1	..AGACACGCCC GCCGCATCCG CATCGACACC GCCATCAACC
  	CCGAACTGGA
  51	AGCCCTCGCC GAACACCTCC ACTACCAATG GCAGGGCTTC
  	CTCTGGCTCA
  101	GCACCGATAT GCGTCAGGAA ATTTCCGCCC TCGTCATCCT
  	GCTGCAACGC
  151	ACCCGCCGCA AATGGCTGGA TGCCCACGAA CGCCAACACC
  	TGCGCCAAAG
  201	CCTGCTTGAA ACACGGGAAC ACGGCTGA

• This corresponds to the amino acid sequence <SEQ ID 630; ORF146>:

• 1	..RHARRIRIDT AINPELEALA EHLHYQWQGF LWLSTDMRQE
  	ISALVILLQR
  51	TRRKWLDAHE RQHLRQSLLE TREHG*

• Further work revealed the complete nucleotide sequence <SEQ ID 631>:

• 1	ATGAACACCT CGCAACGCAA CCGCCTCGTC AGCCGCTGGC
  	TCAACTCCTA
  51	CGAACGCTAC CGCTACCGCC GCCTCATCCA CGCCGTCCGG
  	CTCGGCGGGG
  101	CCGTCCTGTT CGCCACCGCC TCCGCCCGGC TGCTCCACCT
  	CCAACACGGC
  151	GAGTGGATAG GGATGACCGT CTTCGTCGTC CTCGGCATGC
  	TCCAGTTTCA
  201	AGGGGCGATT TACTCCAAGG CGGTGGAACG TATGCTCGGC
  	ACGGTCATCG
  251	GGCTGGGCGC GGGTTTGGGC GTTTTATGGC TGAACCAGCA
  	TTATTTCCAC
  301	GGCAACCTCC TCTTCTACCT CACCGTCGGC ACGGCAAGCG
  	CACTGGCCGG
  351	CTGGGCGGCG GTCGGCAAAA ACGGCTACGT CCCTATGCTG
  	GCAGGGCTGA
  401	CGATGTGTAT GCTCATCGGC GACAACGGCA GCGAATGGCT
  	CGACAGCGGA
  451	CTCATGCGCG CCATGAACGT CCTCATCGGC GCGGCCATCG
  	CCATCGCCGC
  501	CGCCAAACTG CTGCCGCTGA AATCCACACT GATGTGGCGT
  	TTCATGCTTG
  551	CCGACAACCT GGCCGACTGC AGCAAAATGA TTGCCGAAAT
  	CAGCAACGGC
  601	AGGCGCATGA CCCGCGAACG CCTCGAGGAG AACATGGCGA
  	AAATGCGCCA
  651	AATCAACGCA CGCATGGTCA AAAGCCGCAG CCATCTCGCC
  	GCCACATCGG
  701	GCGAAAGCCG CATCAGCCCC GCCATGATGG AAGCCATGCA
  	GCACGCCCAC
  751	CGTAAAATCG TCAACACCAC CGAGCTGCTC CTGACCACCG
  	CCGCCAAGCT
  801	GCAATCTCCC AAACTCAACG GCAGCGAAAT CCGGCTGCTT
  	GACCGCCACT
  851	TCACACTGCT CCAAACCGAC CTGCAACAAA CCGTCGCCCT
  	TATCAACGGC
  901	AGACACGCCC GCCGCATCCG CATCGACACC GCCATCAACC
  	CCGAACTGGA
  951	AGCCCTCGCC GAACACCTCC ACTACCAATG GCAGGGCTTC
  	CTCTGGCTCA
  1001	GCACCAATAT GCGTCAGGAA ATTTCCGCCC TCGTCATCCT
  	GCTGCAACGC
  1051	ACCCGCCGCA AATGGCTGGA TGCCCACGAA CGCCAACACC
  	TGCGCCAAAG
  1101	CCTGCTTGAA ACACGGGAAC ACGGCTGA

• This corresponds to the amino acid sequence <SEQ ID 632; ORF146-1>:

• 1	MNTSQRNRLV SRWLNSYERY RYRRLIHAVR LGGAVLFATA
  	SARLLHLQHG
  51	EWIGMTVFVV LGMLQFQGAI YSKAVERMLG TVIGLGAGLG
  	VLWLNQHYFH
  101	GNLLFYLTVG TASALAGWAA VGKNGYVPML AGLTMCMLIG
  	DNGSEWLDSG
  151	LMRAMNVLIG AAIAIAAAKL LPLKSTLMWR FMLADNLADC
  	SKMIAEISNG
  201	RRMTRERLEE NMAKMRQINA RMVKSRSHLA ATSGESRISP
  	AMMEAMQHAH
  251	RKIVNTTELL LTTAAKLQSP KLNGSEIRLL DRHFTLLQTD
  	LQQTVALING
  301	RHARRIRIDT AINPELEALA EHLHYQWQGF LWLSTNMRQE
  	ISALVILLQR
  351	TRRKWLDAHE RQHLRQSLLE TREHG*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF146 shows 98.6% identity over a 74aa overlap with an ORF (ORF146a) from strain A of N. meningitidis:
• 
• The complete length ORF146a nucleotide sequence <SEQ ID 633> is:

• 1	ATGAACACCT CGCAACGCAA CCGCCTCGTC AGCCGCTGGC
  	TCAACTCCTA
  51	CGAACGCTAC CGCTACCGCC GCCTCATCCA CGCCGTCCGG
  	CTCGGCGGGG
  101	CCGTCCTGTT CGCCACCGCC TCCGCCCGGC TGCTCCACCT
  	CCAACACGGC
  151	GAGTGGATAG GGATGACCGT CTTCGTCGTC CTCGGCATGC
  	TCCAGTTTCA
  201	AGGGGCGATT TACTCCAAGG CGGTGGAACG TATGCTCGGC
  	ACGGTCATCG
  251	GGCTGGGCGC GGGTTTGGGC GTTTTATGGC TGAACCAGCA
  	TTATTTCCAC
  301	GGCAACCTCC TCTTCTACCT CACCGTCGGC ACGGCAAGCG
  	CACTGGCCGG
  351	CTGGGCGGCG GTCGGCAAAA ACGGCTACGT CCCTATGCTG
  	GCGGGGCTGA
  401	CGATGTGCAT GCTCATCGGC GACAACGGCA GCGAATGGTT
  	CGACAGCGGC
  451	CTGATGCGCG CGATGAACGT CCTCATCGGC GCGGCCATCG
  	CCATCGCCGC
  501	CGCCAAACTG CTGCCGCTGA AATCCACACT GATGTGGCGT
  	TTCATGCTTG
  551	CCGACAACCT GACCGACTGC AGCAAAATGA TTGCCGAAAT
  	CAGCAACGGC
  601	AGGCGCATGA CCCGCGAACG CCTCGAAGAG AACATGGCGA
  	AAATGCGCCA
  651	AATCAACGCA CGCATGGTCA AAAGCCGCAG CCACCTCGCC
  	GCCACATCGG
  701	GCGAAAGCCG CATCAGCCCC GCCATGATGG AAGCCATGCA
  	GCACGCCCAC
  751	CGTAAAATTG TCAACACCAC CGAGCTGCTC CTGACCACCG
  	CCGCCAAGCT
  801	GCAATCTCCC AAACTCAACG GCAGCGAAAT CCGGCTGCTT
  	GACCGCCACT
  851	TCACACTGCT CCAAACCGAC CTGCAACAAA CCGTCGCCCT
  	TATCAACGGC
  901	AGACACGCCC GCCGCATCCG CATCGACACC GCCATCAACC
  	CCGAACTGGA
  951	AGCCCTCGCC GAACACCTCC ACTACCAATG GCAGGGCTTC
  	CTCTGGCTCA
  1001	GCACCAATAT GCGTCAGGAA ATTTCCGCCC TCGTCATCCT
  	GCTGCAACGC
  1051	ACCCGCCGCA AATGGCTGGA TGCCCACGAA CGCCAACACC
  	TGCGCCAAAG
  1101	CCTGCTTGAA ACACGGGAAC ACAGTTGA

• This encodes a protein having amino acid sequence <SEQ ID 634>:

• 1	MNTSQRNRLV SRWLNSYERY RYRRLIHAVR LGGAVLFATA
  	SARLLHLQHG
  51	EWIGMTVFVV LGMLQFQGAI YSKAVERMLG TVIGLGAGLG
  	VLWLNQHYFH
  101	GNLLFYLTVG TASALAGWAA VGKNGYVPML AGLTMCMLIG
  	DNGSEWFDSG
  151	LMRAMNVLIG AAIAIAAAKL LPLKSTLMWR FMLADNLTDC
  	SKMIAEISNG
  201	RRMTRERLEE NMAKMRQINA RMVKSRSHLA ATSGESRISP
  	AMMEAMQHAH
  251	RKIVNTTELL LTTAAKLQSP KLNGSEIRLL DRHFTLLQTD
  	LQQTVALING
  301	RHARRIRIDT AINPELEALA EHLHYQWQGF LWLSTNMRQE
  	ISALVILLQR
  351	TRRKWLDAHE RQHLRQSLLE TREHS*

• ORF146a and ORF146-1 show 99.5% identity in 374 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF146 shows 97.3% identity over a 75aa overlap with a predicted ORF (ORF146ng) from N. gonorrhoeae:
• 
• An ORF146ng nucleotide sequence <SEQ ID 635> was predicted to encode a protein having amino, acid sequence <SEQ ID 636>:

• 1	MSGVRFPSPA PIPSTDPPSG SLCFFTFPLQ TASDMNSSQR
  	KRLSGRWLNS
  51	YERYRHRRLI HAVRLGGTVL FATALARLLH LQHGEWIGMT
  	VFVVLGMLQF
  101	QGAIYSNAVE RMLGTVIGLG AGLGVLWLNQ HYFHGNLLFY
  	LTIGTASALA
  151	GWAAVGKNGY VPMLAGLTMC MLIGDNGSEW LDSGLMRAMN
  	VLIGAAIAIA
  201	AAKLLPLKST LMWRFMLADN LADCSKMIAE ISNGRRMTRE
  	RLEQNMVKMR
  251	QINARMVKSR SHLAATSGES RISPSMMEAM QHAHRKIVNT
  	TELLLTTAAK
  301	LQSPKLNGSE IRLLDRHFTL LQTDLQQTAA LINGRHARRI
  	RIDTAINPEL
  351	EALAEHLHYQ WQGFLWLSTN MRQEISALVI PLQRTRRKWL
  	DAHERQHLRQ
  401	SLLETREHG*

• Further work revealed the following gonococcal DNA sequence <SEQ ID 637>:

• 1	ATGAACTCCT CGCAACGCAA ACGCCTTTCC GgccGCTGGC
  	TCAACTCCTA
  51	CGAACGCTac cGCCaccGCC GCCTCATACA TGCCGTGCGG
  	CTCGGCggaa
  101	ccgtCCTGTT CGCCACCGCA CTCGCCCGgc tACTCCACCT
  	CCAacacggc
  151	gAATGGATAG GGAtgaCCGT CTTCGTCGTC CTCGGCATGC
  	TCCAGTTCCA
  201	AGGCgcgatt tActccaacg cggtgGAacg taTGctcggt
  	acggtcatcg
  251	ggctgGGCGC GGGTTTGGgc gTTTTATGGC TGAACCAGCA
  	TTAtttccac
  301	ggcaacCTcc tcttctacct gaccatcggc acggcaagcg
  	cactggccgg
  351	ctGGGCGGCG GTCGGCAAAA acggctacgt ccctatgctg
  	GCGGGGctgA
  401	CGATGTGCAT gctcatcggc gACAACGGCA GCGAATGGCT
  	CGACAGCGGC
  451	CTGATGCGCG CGATGAACGT CCTCATCGGC GCCGCCATCG
  	CCATTGCCGC
  501	CGCCAAACTG CTGCCGCTGA AATCCACACT GATGTGGCGT
  	TTCATGCTTG
  551	CCGACAACCT GGCCGACTGC AGCAAAATGA TTGCCGAAAT
  	CAGCAACGGC
  601	AGGCGTATGA CGCGCGAACG TTTGGAGCAG AATATGGTCA
  	AAATGCGCCA
  651	AATCAACGCA CGCATGGTCA AAAGCCGCAG CCACCTCGCC
  	GCCACATCGG
  701	GCGAAAGCCG CATCAGCCCC TCCATGATGG AAGCCATGCA
  	GCACGCCCAC
  751	CGCAAAATCG TCAACACCAC CGAGCTGCTC CTGACCACCG
  	CCGCCAAGCT
  801	GCAATCTCCC AAACTCAACG GCAGCGAAAT CCGGCTGCTC
  	GACCGCCACT
  851	TCACACTGCT CCAAACCGAC CTGCAACAAA CCGCCGCCCT
  	CATCAACGGC
  901	AGACACGCCC GCCGCATCCG CATCGACACC GCCATCAACC
  	CCGAACTGGA
  951	AGCCCTCGCC GAACACCTCC ACTACCAATG GCAGGGCTTC
  	CTCTGGCTCA
  1001	GCACCAATAT GCGTCAGGAA ATTTCCGCCC TCGTCATCCT
  	GCTGCAACGC
  1051	ACCCGCCGCA AATGGCTGGA TGCCCACGAA CGCCAACACC
  	TGCGCCAAAG
  1101	CCTGCTTGAA ACACGGGAAC ACGGCTGA

• This corresponds to the amino acid sequence <SEQ ID 638; ORF146ng-1>:

• 1	MNSSQRKRLS GRWLNSYERY RHRRLIHAVR LGGTVLFATA
  	LARLLHLQHG
  51	EWIGMTVFVV LGMLQFQGAI YSNAVERMLG TVIGLGAGLG
  	VLWLNQHYFH
  101	GNLLFYLTIG TASALAGWAA VGKNGYVPML AGLTMCMLIG
  	DNGSEWLDSG
  151	LMRAMNVLIG AAIAIAAAKL LPLKSTLMWR FMLADNLADC
  	SKMIAEISNG
  201	RRMTRERLEQ NMVKMRQINA RMVKSRSHLA ATSGESRISP
  	SMMEAMQHAH
  251	RKIVNTTELL LTTAAKLQSP KLNGSEIRLL DRHFTLLQTD
  	LQQTAALING
  301	RHARRIRIDT AINPELEALA EHLHYQWQGF LWLSTNMRQE
  	ISALVILLQR
  351	TRRKWLDAHE RQHLRQSLLE TREHG*

• ORF146ng-1 and ORF146-1 show 96.5% identity in 375 aa overlap
• 
• Furthermore, ORF146ng-1 shows homology with a hypothetical E. coli protein:

• sp|P33011|YEEA_ECOLI HYPOTHETICAL 40.0 KD PROTEIN IN
  COBU-SBMC INTERGENIC REGION
  >gi|1736674|gnl|PID|d1016553 (D90838) ORF_ID: o348#20;
  similar to [SwissProt Accession Number P33011] [Escherichia coli]
  >gi|1736682|gnl|PID|d1016560 (D90839) ORF_ID: o348#20;
  similar to [SwissProt Accession Number P33011] [Escherichia coli]
  >gi|1788318 (AE000292) f352; 100% identical to fragment YEEA_ECOLI
  SW: P33011 but has 203 additional C-terminal residues [Escherichia coli]
  Length = 352 Score = 109 bits (271), Expect = 2e−23
  Identities = 89/347 (25%), Positives = 150/347 (42%), Gaps = 21/347 (6%)

  Query:	20	YRHRRLIHAVRLGGTVLFATALARLLHLQHGEWIGMTVFVVLGMLQFQGAIYSNAVERML	79
  		YRH R++H  R+    L    + RL  +    W  +T+ V++G + F G +   A ER+
  Sbjct:	15	YRHYRIVHGTRVALAFLLTFLIIRLFTIPESTWPLVTMVVIMGPISFWGNVVPRAFERIG	74
  Query:	80	GTVIGLGAGLGVLWLNQHYFHGNLLFYLTIGTASALAGWAAVGKNGYVPMLAGLTMCMLI	139
  		GTV+G   GL  L L         L  +    A  L GW A+GK  Y  +L G+T+ +++
  Sbjct:	75	GTVLGSILGLIALQLE---LISLPLMLVWCAAAMFLCGWLALGKKPYQGLLIGVTLAIVV	131
  Query:	140	GDNGSEWLDSGLMRAMNVLIGXXXXXXXXKLLPLKSTLMWRFMLADNLADCSKMIAEISN	199
  		G    E +D+ L R+ +V++G         + P ++ + WR  LA +L + +++     +
  Sbjct:	132	GSPTGE-IDTALWRSGDVILGSLLAMLFTGIWPQRAFIHWRIQLAKSLTEYNRVYQSAFS	190
  Query:	200	GRRMTRERLEQNMVKMRQINARMVKSRSHLAATSGESRISPSMMEAMQHAHRKIVNXXXX	259
  		+ R RLE ++ K+       VK R  +A  S E+RI  S+ E +Q  +R +V
  Sbjct:	191	PNLLERPRLESHLQKLL---TDAVKMRGLIAPASKETRIPKSIYEGIQTINRNLVCMLEL	247
  Query:	260	XXXXXXXXQSPK---LNGSEIRLLDRHFXXXXXXXXXXAALINGRHARRIRIDTAINPEL	316
  		+      LN  ++R  D              AL  G           +N  +
  Sbjct:	248	QINAYWATRPSHFVLLNAQKLR--DTQHMMQQILLSLVHALYEGNPQPVFANTEKLNDAV	305
  Query:	317	EALAEHL--HYQWQ-------GFLWLSTNMRQEISALVILLQRTRRK	354
  		E L + L  H+  +       G++WL+     ++  L  L+ R  RK
  Sbjct:	306	EELRQLLNNHHDLKVVETPIYGYVWLNMETAHQLELLSNLICRALRK	352

• On the basis of this analysis, including the identification of several transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 76
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 639>

• 1	..GCCGAAGACA CGCGCGTTAC CGCACAGCTT TTGAGCGCGT
  	ACGGCATTCA
  51	GGGCAAACTC GTCAGTGTGC GCGAACACAA CGAACGGCAG
  	ATGGCGGACA
  101	AGATTGTCGG CTATCTTTCA GACGGCATGG TTGTGGCACA
  	GGTTTCCGAT
  151	GCGGGTACGC CGGCCGTGTG CGACCCGGGC GCGAAACTCG
  	CCCGCCGCGT
  201	GCGTGAGGCC GGGTTTAAAG TCGTTCCCGT CGTGGGCGCA
  	AC.GCGGTGA
  251	TGGCGGCTTT GAGCGTGGCC GGTGTGGAAG GATCCGATTT
  	TTATTTCAAC
  301	GGTTTTGTAC CGCCGAAATC GGGAGAACGC AGGAAACTGT
  	TTGCCAAATG
  351	GGTGCGGGCG GCGTTTCCTA TCGTCATGTT TGAAACGCCG
  	CACCGCATCG
  401	GTGCAGCGCT TGCCGATATG GCGGAACTGT TCCCCGAACG
  	CCGATTAATG
  451	CTGGCGCGCG AAATTACGAA AACGTTTGAA ACGTTCTTAA
  	GCGGCACGGT
  501	TGGGGAAATT CAGACGGCAT TGTCTGCCGA CGGCGACCAA
  	TCGCGCGGCG
  551	AGATGGTGTT GGTGCTTTAT CCGGCGCAGG ATGAAAAACA
  	CGAAGGCTTG
  601	TCCGAGTCCG CGCAAAACAT CATGAAAATC CTCACAGCCG
  	AGCTGCCGAC
  651	CAAACAGGCG GCGGAGCTTG CTGCCAAAAT CACGGGCGAG
  	GGAAAGAAAG
  701	CTTTGTACGA T..

• This corresponds to the amino acid sequence <SEQ ID 640; ORF147>:

• 1	..AEDTRVTAQL LSAYGIQGKL VSVREHNERQ MADKIVGYLS
  	DGMVVAQVSD
  51	AGTPAVCDPG AKLARRVREA GFKVVPVVGA XAVMAALSVA
  	GVEGSDFYFN
  101	GFVPPKSGER RKLFAKWVRA AFPIVMFETP HRIGAALADM
  	AELFPERRLM
  151	LAREITKTFE TFLSGTVGEI QTALSADGDQ SRGEMVLVLY
  	PAQDEKHEGL
  201	SESAQNIMKI LTAELPTKQA AELAAKITGE GKKALYD..

• Further work revealed the complete nucleotide sequence <SEQ ID 641>:

• 1	ATGTTTCAGA AACATTTGCA GAAAGCCTCC GACAGCGTCG
  	TCGGAGGGAC
  51	ATTATACGTG GTTGCCACGC CCATCGGCAA TTTGGCGGAC
  	ATTACCCTGC
  101	GCGCTTTGGC GGTATTGCAA AAGGCGGACA TCATCTGTGC
  	CGAAGACACG
  151	CGCGTTACCG CACAGCTTTT GAGCGCGTAC GGCATTCAGG
  	GCAAACTCGT
  201	CAGTGTGCGC GAACACAACG AACGGCAGAT GGCGGACAAG
  	ATTGTCGGCT
  251	ATCTTTCAGA CGGCATGGTT GTGGCACAGG TTTCCGATGC
  	GGGTACGCCG
  301	GCCGTGTGCG ACCCGGGCGC GAAACTCGCC CGCCGCGTGC
  	GTGAGGCCGG
  351	GTTTAAAGTC GTTCCCGTCG TGGGCGCAAG CGCGGTGATG
  	GCGGCTTTGA
  401	GCGTGGCCGG TGTGGAAGGA TCCGATTTTT ATTTCAACGG
  	TTTTGTACCG
  451	CCGAAATCGG GAGAACGCAG GAAACTGTTT GCCAAATGGG
  	TGCGGGCGGC
  501	GTTTCCTATC GTCATGTTTG AAACGCCGCA CCGCATCGGT
  	GCGACGCTTG
  551	CCGATATGGC GGAACTGTTC CCCGAACGCC GATTAATGCT
  	GGCGCGCGAA
  601	ATTACGAAAA CGTTTGAAAC GTTCTTAAGC GGCACGGTTG
  	GGGAAATTCA
  651	GACGGCATTG TCTGCCGACG GCAACCAATC GCGCGGCGAG
  	ATGGTGTTGG
  701	TGCTTTATCC GGCGCAGGAT GAAAAACACG AAGGCTTGTC
  	CGAGTCCGCG
  751	CAAAACATCA TGAAAATCCT CACAGCCGAG CTGCCGACCA
  	AACAGGCGGC
  801	GGAGCTTGCT GCCAAAATCA CGGGCGAGGG AAAGAAAGCT
  	TTGTACGATC
  851	TGGCTCTGTC TTGGAAAAAC AAATAG

• This corresponds to the amino acid sequence <SEQ ID 642; ORF147-1>:

• 1	MFQKHLQKAS DSVVGGTLYV VATPIGNLAD ITLRALAVLQ
  	KADIICAEDT
  51	RVTAQLLSAY GIQGKLVSVR EHNERQMADK IVGYLSDGMV
  	VAQVSDAGTP
  101	AVCDPGAKLA RRVREAGFKV VPVVGASAVM AALSVAGVEG
  	SDFYFNGFVP
  151	PKSGERRKLF AKWVRAAFPI VMFETPHRIG ATLADMAELF
  	PERRLMLARE
  201	ITKTFETFLS GTVGEIQTAL SADGNQSRGE MVLVLYPAQD
  	EKHEGLSESA
  251	QNIMKILTAE LPTKQAAELA AKITGEGKKA LYDLALSWKN
  	K*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with Hypothetical Protein ORF286 of E. coli (Accession Number U 18997)
• ORF147 and E. coli ORF286 protein show 36% aa identity in 237aa overlap:

• Orf147:	1	AEDTRVTAQLLSAYGIQGKLVSVREHNERQMADKIVGYLSDGMVVAQVSDAGTPAVCDPG	60
  		AEDTR T  LL  +GI  +L ++ +HNE+Q A+ ++  L +G  +A VSDAGTP + DPG
  Orf286:	43	AEDTRHTGLLLQHFGINARLFALHDHNEQQKAETLLAKLQEGQNIALVSDAGTPLINDPG	102
  Orf147:	61	AKLARRVREXXXXXXXXXXXXXXXXXXXXXXXEGSDFYFNGFVPPKSGERRKLFAKWVRA	120
  		L R  RE                           F + GF+P KS  RR
  Orf286:	103	YHLVRTCREAGIRVVPLPGPCAAITALSAAGLPSDRFCYEGFLPAKSKGRRDALKAIEAE	162
  Orf147:	121	AFPIVMFETPHRIGAALADMAELFPERR-LMLAREITKTFETFLSGTVGEIQTALSADGD	179
  		++ +E+ HR+  +L D+  +  E R ++LARE+TKT+ET     VGE+   +  D +
  Orf286:	163	PRTLIFYESTHRLLDSLEDIVAVLGESRYVVLARELTKTWETIHGAPVGELLAWVKEDEN	222
  Orf147:	180	QSRGEMVLVLYPAQDEKHEGLSESAQNIMKILTAELPTKQAAELAAKITGEGKKALY	236
  		+ +GEMVL++      + E L   A   + +L AELP K+AA LAA+I G  K ALY
  Orf286:	223	RRKGEMVLIV-EGHKAQEEDLPADALRTLALLQAELPLKKAAALAAEIHGVKKNALY	278

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF147 shows 96.6% identity over a 237aa overlap with ORF75a from strain A of N. meningitidis:
• 
• ORF147a is identical to ORF75a, which includes aa 56-292 of ORF75.
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF147 shows 94.1% identity over a 237aa overlap with a predicted ORF (ORF147ng) from N. gonorrhoeae:
• 
• An ORF147ng nucleotide sequence <SEQ ID 643> was predicted to encode a protein having amino acid sequence <SEQ ID 644>:

• 1	MSVFQTAFFM FQKHLQKASD SVVGGTLYVV ATPIGNLADI
  	TLRALAVLQK
  51	ADIICAEDTR VTAQLLSAYG IQGRLVSVRE HNERQMADKV
  	IGFLSDGLVV
  101	AQVSDAGTPA VCDPGAKLAR RVREAGFKVV PVVGASAVMA
  	ALSVAGVAES
  151	DFYFNGFVPP KSGERRKLFA KWVRAAFPVV MFETPHRIGA
  	TLADMAELFP
  201	ERRLMLAREI TKTFETFLSG TVGEIQTALA ADGNQSRGEM
  	VLVLYPAQDE
  251	KHEGLSESAQ NAMKILAAEL PTKQAAELAA KITGEGKKAL
  	YDLALSWKNK
  301	*

• Further work revealed the following gonococcal DNA sequence <SEQ ID 645>:

• 1	ATGTTTCAGA AACACTTGCA GAAAGCCTCC GACAGCGTCG
  	TCGGAGGGAC
  51	ATTATACGTG GTTGCCACGC CCATCGGCAA TTTGGCAGAC
  	ATTACCCTGC
  101	GCGCTTTGGC GGTATTGCAA AAGGCGGACA TCATTTGTGC
  	CGAAGACACG
  151	CGCGTTACTG CGCAGCTTTT GAGCGCGTAC GGCATTCAGG
  	GCAGGTTGGT
  201	CAGTGTGCGC GAACACAACG AGCGGCAGAT GGCGGACAAG
  	GTAATCGGTT
  251	TCCTTTCAGA CGGCCTGGTT GTGGCGCAGG TTTCCGATGC
  	GGGTACGCCG
  301	GCCGTGTGCG ACCCGGGCGC GAAACTCGCC CGCCGCGTGC
  	GCGAAGCAGG
  351	GTTCAAAGTC GTTCCCGTCG TGGGCGCAAG CGCGGTAATG
  	GCGGCGTTGA
  401	GTGTGGCCGG TGTGGCGGAA TCCGATTTTT ATTTCAACGG
  	TTTTGTACCG
  451	CCGAAATCGG GCGAACGTAG GAAATTGTTT GCCAAATGGG
  	TGCGGGCGGC
  501	ATTTCCTGTC GTCATGTTTG AAACGCCGCA CCGAATCGGG
  	GCAACGCTTG
  551	CCGATATGGC GGAATTGTTC CCCGAACGCC GTCTGATGCT
  	GGCGCGCGAA
  601	ATCACGAAAA CGTTTGAAAC GTTCTTAAGC GGCACGGTTG
  	GGGAAATTCA
  651	GACGGCATTG GCGGCGGACG GCAACCAATC GCGCGGCGAG
  	ATGGTGTTGG
  701	TGCTTTATCC GGCGCAGGAT GAAAAACACG AAGGCTTGTC
  	CGAGTCTGCG
  751	CAAAATGCGA TGAAAATCCT TGCGGCCGAG CTGCCGACCA
  	AGCAGGCGGC
  801	GGAGCTTGCC GCCAAGATTA CAGGTGAGGG CAAAAAGGCT
  	TTGTACGATT
  851	TGGCACTGTC GTGGAAAAAC AAATGA

• This corresponds to the amino acid sequence <SEQ ID 646; ORF147ng-1>:

• 1	MFQKHLQKAS DSVVGGTLYV VATPIGNLAD ITLRALAVLQ
  	KADIICAEDT
  51	RVTAQLLSAY GIQGRLVSVR EHNERQMADK VIGFLSDGLV
  	VAQVSDAGTP
  101	AVCDPGAKLA RRVREAGFKV VPVVGASAVM AALSVAGVAE
  	SDFYFNGFVP
  151	PKSGERRKLF AKWVRAAFPV VMFETPHRIG ATLADMAELF
  	PERRLMLARE
  201	ITKTFETFLS GTVGEIQTAL AADGNQSRGE MVLVLYPAQD
  	EKHEGLSESA
  251	QNAMKILAAE LPTKQAAELA AKITGEGKKA LYDLALSWKN
  	K*

• ORF147ng shows homology to a hypothetical E. coli protein:

• sp|P45528|YRAL_ECOLI HYPOTHETICAL 31.3 KD PROTEIN IN AGAI-MTR
  INTERGENIC REGION (F286)
  >gi|606086 (U18997) ORF_f286 [Escherichia coli]
  >gi|1789535 (AE000395) hypothetical 31.3 kD protein in agai-mtr intergenic region
  [Escherichia coli] Length = 286
  Score = 218 bits (550), Expect = 3e−56
  Identities = 128/284 (45%), Positives = 171/284 (60%), Gaps = 4/284 (1%)

  Query:	4	KHLQKASDSVVGGTLYVVATPIGNLADITLRALAVLQKADIICAEDTRVTAQLLSAYGIQ	63
  		K  Q A +S   G LY+V TPIGNLADIT RAL VLQ  D+I AEDTR T  LL  +GI
  Sbjct:	2	KQHQSADNSQ--GQLYIVPTPIGNLADITQRALEVLQAVDLIAAEDTRHTGLLLQHFGIN	59
  Query:	64	GRLVSVREHNERQMADKVIGFLSDGLVVAQVSDAGTPAVCDPGAKLARRVREAGFKVVPV	123
  		RL ++ +HNE+Q A+ ++  L +G  +A VSDAGTP + DPG  L R  REAG +VVP+
  Sbjct:	60	ARLFALHDHNEQQKAETLLAKLQEGQNIALVSDAGTPLINDPGYHLVRTCREAGIRVVPL	119
  Query:	124	VGASAVMAALSVAGVAESDFYFNGFVPPKSGERRKLFAKWVRAAFPVVMFETPHRIGATL	183
  		G  A + ALS AG+    F + GF+P KS  RR            ++ +E+ HR+  +L
  Sbjct:	120	PGPCAAITALSAAGLPSDRFCYEGFLPAKSKGRRDALKAIEAEPRTLIFYESTHRLLDSL	179
  Query:	184	ADMAELFPERR-LMLAREITKTFETFLSGTVGEIQTALAADGNQSRGEMVLVLYPAQDEK	242
  		D+  +  E R ++LARE+TKT+ET     VGE+   +  D N+ +GEMVL++      +
  Sbjct:	180	EDIVAVLGESRYVVLARELTKTWETIHGAPVGELLAWVKEDENRRKGEMVLIV-EGHKAQ	238
  Query:	243	HEGLSESAQNAMKILAAELPTKQAAELAAKITGEGKKALYDLAL	286
  		E L   A   + +L AELP K+AA LAA+I G  K ALY  AL
  Sbjct:	239	EEDLPADALRTLALLQAELPLKKAAALAAEIHGVKKNALYKYAL	282

• Based on the computer analysis and the presence of a putative transmembrane domain in the gonococcal protein, it is predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 77
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 647>
• 
• This corresponds to the amino acid sequence <SEQ ID 648; ORF1>:
• 
• Further sequencing analysis revealed the complete nucleotide sequence <SEQ ID 649>:

• 1	ATGAAAACAA CCGACAAACG GACAACCGAA ACACACCGCA
  	AAGCCCCGAA
  51	AACCGGCCGC ATCCGCTTCT CGCCTGCTTA CTTAGCCATA
  	TGCCTGTCGT
  101	TCGGCATTCT TCCCCAAGCC TGGGCGGGAC ACACTTATTT
  	CGGCATCAAC
  151	TACCAATACT ATCGCGACTT TGCCGAAAAT AAAGGCAAGT
  	TTGCAGTCGG
  201	GGCGAAAGAT ATTGAGGTTT ACAACAAAAA AGGGGAGTTG
  	GTCGGCAAAT
  251	CAATGACAAA AGCCCCGATG ATTGATTTTT CTGTGGTGTC
  	GCGTAACGGC
  301	GTGGCGGCAT TGGTGGGCGA TCAATATATT GTGAGCGTGG
  	CACATAACGG
  351	CGGCTATAAC AACGTTGATT TTGGTGCGGA AGGAAGAAAT
  	CCCGATCAAC
  401	ATCGTTTTAC TTATAAAATT GTGAAACGGA ATAATTATAA
  	AGCAGGGACT
  451	AAAGGCCATC CTTATGGCGG CGATTATCAT ATGCCGCGTT
  	TGCATAAATT
  501	TGTCACAGAT GCAGAACCTG TTGAAATGAC CAGTTATATG
  	GATGGGCGGA
  551	AATATATCGA TCAAAATAAT TACCCTGACC GTGTTCGTAT
  	TGGGGCAGGC
  601	AGGCAATATT GGCGATCTGA TGAAGATGAG CCCAATAACC
  	GCGAAAGTTC
  651	ATATCATATT GCAAGTGCGT ATTCTTGGCT CGTTGGTGGC
  	AATACCTTTG
  701	CACAAAATGG ATCAGGTGGT GGCACAGTCA ACTTAGGTAG
  	TGAAAAAATT
  751	AAACATAGCC CATATGGTTT TTTACCAACA GGAGGCTCAT
  	TTGGCGACAG
  801	TGGCTCACCA ATGTTTATCT ATGATGCCCA AAAGCAAAAG
  	TGGTTAATTA
  851	ATGGGGTATT GCAAACGGGC AACCCCTATA TAGGAAAAAG
  	CAATGGCTTC
  901	CAGCTGGTTC GTAAAGATTG GTTCTATGAT GAAATCTTTG
  	CTGGAGATAC
  951	CCATTCAGTA TTCTACGAAC CACGTCAAAA TGGGAAATAC
  	TCTTTTAACG
  1001	ACGATAATAA TGGCACAGGA AAAATCAATG CCAAACATGA
  	ACACAATTCT
  1051	CTGCCTAATA GATTAAAAAC ACGAACCGTT CAATTGTTTA
  	ATGTTTCTTT
  1101	ATCCGAGACA GCAAGAGAAC CTGTTTATCA TGCTGCAGGT
  	GGTGTCAACA
  1151	GTTATCGACC CAGACTGAAT AATGGAGAAA ATATTTCCTT
  	TATTGACGAA
  1201	GGAAAAGGCG AATTGATACT TACCAGCAAC ATCAATCAAG
  	GTGCTGGAGG
  1251	ATTATATTTC CAAGGAGATT TTACGGTCTC GCCTGAAAAT
  	AACGAAACTT
  1301	GGCAAGGCGC GGGCGTTCAT ATCAGTGAAG ACAGTACCGT
  	TACTTGGAAA
  1351	GTAAACGGCG TGGCAAACGA CCGCCTGTCC AAAATCGGCA
  	AAGGCACGCT
  1401	GCACGTTCAA GCCAAAGGGG AAAACCAAGG CTCGATCAGC
  	GTGGGCGACG
  1451	GTACAGTCAT TTTGGATCAG CAGGCAGACG ATAAAGGCAA
  	AAAACAAGCC
  1501	TTTAGTGAAA TCGGCTTGGT CAGCGGCAGG GGTACGGTGC
  	AACTGAATGC
  1551	CGATAATCAG TTCAACCCCG ACAAACTCTA TTTCGGCTTT
  	CGCGGCGGAC
  1601	GTTTGGATTT AAACGGGCAT TCGCTTTCGT TCCACCGTAT
  	TCAAAATACC
  1651	GATGAAGGGG CGATGATTGT CAACCACAAT CAAGACAAAG
  	AATCCACCGT
  1701	TACCATTACA GGCAATAAAG ATATTGCTAC AACCGGCAAT
  	AACAACAGCT
  1751	TGGATAGCAA AAAAGAAATT GCCTACAACG GTTGGTTTGG
  	CGAGAAAGAT
  1801	ACGACCAAAA CGAACGGGCG GCTCAACCTT GTTTACCAGC
  	CCGCCGCAGA
  1851	AGACCGCACC CTGCTGCTTT CCGGCGGAAC AAATTTAAAC
  	GGCAACATCA
  1901	CGCAAACAAA CGGCAAACTG TTTTTCAGCG GCAGACCAAC
  	ACCGCACGCC
  1951	TACAATCATT TAAACGACCA TTGGTCGCAA AAAGAGGGCA
  	TTCCTCGCGG
  2001	GGAAATCGTG TGGGACAACG ACTGGATCAA CCGCACATTT
  	AAAGCGGAAA
  2051	ACTTCCAAAT TAAAGGCGGA CAGGCGGTGG TTTCCCGCAA
  	TGTTGCCAAA
  2101	GTGAAAGGCG ATTGGCATTT GAGCAATCAC GCCCAAGCAG
  	TTTTTGGTGT
  2151	CGCACCGCAT CAAAGCCACA CAATCTGTAC ACGTTCGGAC
  	TGGACGGGTC
  2201	TGACAAATTG TGTCGAAAAA ACCATTACCG ACGATAAAGT
  	GATTGCTTCA
  2251	TTGACTAAGA CCGACATCAG CGGCAATGTC GATCTTGCCG
  	ATCACGCTCA
  2301	TTTAAATCTC ACAGGGCTTG CCACACTCAA CGGCAATCTT
  	AGTGCAAATG
  2351	GCGATACACG TTATACAGTC AGCCACAACG CCACCCAAAA
  	CGGCAACCTT
  2401	AGCCTCGTGG GCAATGCCCA AGCAACATTT AATCAAGCCA
  	CATTAAACGG
  2451	CAACACATCG GCTTCGGGCA ATGCTTCATT TAATCTAAGC
  	GACCACGCCG
  2501	TACAAAACGG CAGTCTGACG CTTTCCGGCA ACGCTAAGGC
  	AAACGTAAGC
  2551	CATTCCGCAC TCAACGGTAA TGTCTCCCTA GCCGATAAGG
  	CAGTATTCCA
  2601	TTTTGAAAGC AGCCGCTTTA CCGGACAAAT CAGCGGCGGC
  	AAGGATACGG
  2651	CATTACACTT AAAAGACAGC GAATGGACGC TGCCGTCAGG
  	CACGGAATTA
  2701	GGCAATTTAA ACCTTGACAA CGCCACCATT ACACTCAATT
  	CCGCCTATCG
  2751	CCACGATGCG GCAGGGGCGC AAACCGGCAG TGCGACAGAT
  	GCGCCGCGCC
  2801	GCCGTTCGCG CCGTTCGCGC CGTTCCCTAT TATCCGTTAC
  	ACCGCCAACT
  2851	TCGGTAGAAT CCCGTTTCAA CACGCTGACG GTAAACGGCA
  	AATTGAACGG
  2901	TCAGGGAACA TTCCGCTTTA TGTCGGAACT CTTCGGCTAC
  	CGCAGCGACA
  2951	AATTGAAGCT GGCGGAAAGT TCCGAAGGCA CTTACACCTT
  	GGCGGTCAAC
  3001	AATACCGGCA ACGAACCTGC AAGCCTCGAA CAATTGACGG
  	TAGTGGAAGG
  3051	AAAAGACAAC AAACCGCTGT CCGAAAACCT TAATTTCACC
  	CTGCAAAACG
  3101	AACACGTCGA TGCCGGCGCG TGGCGTTACC AACTCATCCG
  	CAAAGACGGC
  3151	GAGTTCCGCC TGCATAATCC GGTCAAAGAA CAAGAGCTTT
  	CCGACAAACT
  3201	CGGCAAGGCA GAAGCCAAAA AACAGGCGGA AAAAGACAAC
  	GCGCAAAGCC
  3251	TTGACGCGCT GATTGCGGCC GGGCGCGATG CCGTCGAAAA
  	GACAGAAAGC
  3301	GTTGCCGAAC CGGCCCGGCA GGCAGGCGGG GAAAATGTCG
  	GCATTATGCA
  3351	GGCGGAGGAA GAGAAAAAAC GGGTGCAGGC GGATAAAGAC
  	ACCGCCTTGG
  3401	CGAAACAGCG CGAAGCGGAA ACCCGGCCGG CTACCACCGC
  	CTTCCCCCGC
  3451	GCCCGCCGCG CCCGCCGGGA TTTGCCGCAA CTGCAACCCC
  	AACCGCAGCC
  3501	CCAACCGCAG CGCGACCTGA TCAGCCGTTA TGCCAATAGC
  	GGTTTGAGTG
  3551	AATTTTCCGC CACGCTCAAC AGCGTTTTCG CCGTACAGGA
  	CGAATTAGAC
  3601	CGCGTATTTG CCGAAGACCG CCGCAACGCC GTTTGGACAA
  	GCGGCATCCG
  3651	GGACACCAAA CACTACCGTT CGCAAGATTT CCGCGCCTAC
  	CGCCAACAAA
  3701	CCGACCTGCG CCAAATCGGT ATGCAGAAAA ACCTCGGCAG
  	CGGGCGCGTC
  3751	GGCATCCTGT TTTCGCACAA CCGGACCGAA AACACCTTCG
  	ACGACGGCAT
  3801	CGGCAACTCG GCACGGCTTG CCCACGGCGC CGTTTTCGGG
  	CAATACGGCA
  3851	TCGACAGGTT CTACATCGGC ATCAGCGCGG GCGCGGGTTT
  	TAGCAGCGGC
  3901	AGCCTTTCAG ACGGCATCGG AGGCAAAATC CGCCGCCGCG
  	TGCTGCATTA
  3951	CGGCATTCAG GCACGATACC GCGCCGGTTT CGGCGGATTC
  	GGCATCGAAC
  4001	CGCACATCGG CGCAACGCGC TATTTCGTCC AAAAAGCGGA
  	TTACCGCTAC
  4051	GAAAACGTCA ATATCGCCAC CCCCGGCCTT GCATTCAACC
  	GCTACCGCGC
  4101	GGGCATTAAG GCAGATTATT CATTCAAACC GGCGCAACAC
  	ATTTCCATCA
  4151	CGCCTTATTT GAGCCTGTCC TATACCGATG CCGCTTCGGG
  	CAAAGTCCGA
  4201	ACACGCGTCA ATACCGCCGT ATTGGCTCAG GATTTCGGCA
  	AAACCCGCAG
  4251	TGCGGAATGG GGCGTAAACG CCGAAATCAA AGGTTTCACG
  	CTGTCCCTCC
  4301	ACGCTGCCGC CGCCAAAGGC CCGCAACTGG AAGCGCAACA
  	CAGCGCGGGC
  4351	ATCAAATTAG GCTACCGCTG GTAA

• This corresponds to the amino acid sequence <SEQ ID 650; ORF1-1>:

• 1	MKTTDKRTTE THRKAPKTGR IRFSPAYLAI CLSFGILPQA
  	WAGHTYFGIN
  51	YQYYRDFAEN KGKFAVGAKD IEVYNKKGEL VGKSMTKAPM
  	IDFSVVSRNG
  101	VAALVGDQYI VSVAHNGGYN NVDFGAEGRN PDQHRFTYKI
  	VKRNNYKAGT
  151	KGHPYGGDYH MPRLHKFVTD AEPVEMTSYM DGRKYIDQNN
  	YPDRVRIGAG
  201	RQYWRSDEDE PNNRESSYHI ASAYSWLVGG NTFAQNGSGG
  	GTVNLGSEKI
  251	KHSPYGFLPT GGSFGDSGSP MFIYDAQKQK WLINGVLQTG
  	NPYIGKSNGF
  301	QLVRKDWFYD EIFAGDTHSV FYEPRQNGKY SFNDDNNGTG
  	KINAKHEHNS
  351	LPNRLKTRTV QLFNVSLSET AREPVYHAAG GVNSYRPRLN
  	NGENISFIDE
  401	GKGELILTSN INQGAGGLYF QGDFTVSPEN NETWQGAGVH
  	ISEDSTVTWK
  451	VNGVANDRLS KIGKGTLHVQ AKGENQGSIS VGDGTVILDQ
  	QADDKGKKQA
  501	FSEIGLVSGR GTVQLNADNQ FNPDKLYFGF RGGRLDLNGH
  	SLSFHRIQNT
  551	DEGAMIVNHN QDKESTVTIT GNKDIATTGN NNSLDSKKEI
  	AYNGWFGEKD
  601	TTKTNGRLNL VYQPAAEDRT LLLSGGTNLN GNITQTNGKL
  	FFSGRPTPHA
  651	YNHLNDHWSQ KEGIPRGEIV WDNDWINRTF KAENFQIKGG
  	QAVVSRNVAK
  701	VKGDWHLSNH AQAVFGVAPH QSHTICTRSD WTGLTNCVEK
  	TITDDKVIAS
  751	LTKTDISGNV DLADHAHLNL TGLATLNGNL SANGDTRYTV
  	SHNATQNGNL
  801	SLVGNAQATF NQATLNGNTS ASGNASFNLS DHAVQNGSLT
  	LSGNAKANVS
  851	HSALNGNVSL ADKAVFHFES SRFTGQISGG KDTALHLKDS
  	EWTLPSGTEL
  901	GNLNLDNATI TLNSAYRHDA AGAQTGSATD APRRRSRRSR
  	RSLLSVTPPT
  951	SVESRFNTLT VNGKLNGQGT FRFMSELFGY RSDKLKLAES
  	SEGTYTLAVN
  1001	NTGNEPASLE QLTVVEGKDN KPLSENLNFT LQNEHVDAGA
  	WRYQLIRKDG
  1051	EFRLHNPVKE QELSDKLGKA EAKKQAEKDN AQSLDALIAA
  	GRDAVEKTES
  1101	VAEPARQAGG ENVGIMQAEE EKKRVQADKD TALAKQREAE
  	TRPATTAFPR
  1151	ARRARRDLPQ LQPQPQPQPQ RDLISRYANS GLSEFSATLN
  	SVFAVQDELD
  1201	RVFAEDRRNA VWTSGIRDTK HYRSQDFRAY RQQTDLRQIG
  	MQKNLGSGRV
  1251	GILFSHNRTE NTFDDGIGNS ARLAHGAVFG QYGIDRFYIG
  	ISAGAGFSSG
  1301	SLSDGIGGKI RRRVLHYGIQ ARYRAGFGGF GIEPHIGATR
  	YFVQKADYRY
  1351	ENVNIATPGL AFNRYRAGIK ADYSFKPAQH ISITPYLSLS
  	YTDAASGKVR
  1401	TRVNTAVLAQ DFGKTRSAEW GVNAEIKGFT LSLHAAAAKG
  	PQLEAQHSAG
  1451	IKLGYRW*

• Computer analysis of these sequences gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF1 shows 57.8% identity over a 1456aa overlap with an ORF (ORF1a) from strain A of N. meningitidis:
• 

  
• The complete length ORF1a nucleotide sequence <SEQ ID 651> is:

• 1	ATGAAAACAA CCGACAAACG GACAACCGAA ACACACCGCA
  	AAGCCCCGAA
  51	AACCGGCCGC ATCCGCTTCT CGCCTGCTTA CTTAGCCATA
  	TGCCTGTCGT
  101	TCGGCATTCT TCCCCAAGCT TGGGCGGGAC ACACTTATTT
  	CGGCATCAAC
  151	TACCAATACT ATCGCGACTT TGCCGAAAAT AAAGGCAAGT
  	TTGCAGTCGG
  201	GGCGAAAGAT ATTGAGGTNT ACAACAAAAA AGGGGAGTTG
  	GTCGGCAAAT
  251	CAATGACAAA AGCCCCGATG ATTGATTTTT CTGTGGTGTC
  	GCGTAACGGC
  301	GTGGCGGCAT TGGTGGGCGA TCAATATATT GTGAGCGTGG
  	CACATAACGG
  351	CGGCTATAAC AACGTTGATT TTGGTGCGGA AGGAAGNAAT
  	CCCGATCAGC
  401	ACCGTTTTTC TTACCAAATT GTGAAAAGAA ATAATTATAA
  	GCCTGACAAT
  451	TCACACCCTT ACAACGGCGA TTANCATATG CCGCGTTTGC
  	ATAAATTTGT
  501	CACAGATGCA GAACCTGTCG AAATGACGAG TGACATGAGG
  	GGGAATACCT
  551	ATTCCGATAA AGAAAAATAT CCCGAGCGTG TCCGCATCGG
  	CTCAGGACAC
  601	CACTATTGGC GTTATGATGA TGACAAACAC GGCGATTTAT
  	CCTACTCCGG
  651	CGCATGGTTA ATTGGCGGCA ATACACATAT GCAGGGTTGG
  	GGAAATAATG
  701	GCGTANTTAG TTTGAGCGGC GATGTGCGCC ATGCCAACGA
  	CTATGGCCCT
  751	ATGCCGATTG CAGGTGCGGC AGGCGACAGC GGTTCGCCAA
  	TGTTTATTTA
  801	TGACAAAACA AACAATAAAT GGCTGCTCAA CGGAGTTTTA
  	CAAACCGGCT
  851	ACCCTTATTC CGGCAGGGAA AACGGTTTCC AGCTGATACG
  	CAAAGATTGG
  901	TTCTACGATG ACATTTACAG AGGCGATACA CATACCGTCT
  	NTTTTGAACC
  951	GCGCAGTAAC GGACATTTTT CCTTTACATC CAACAACAAC
  	GGTACGGGTA
  1001	CGGTAACAGA AACCAACGAA AAGGTNTCCA ATCCAAAGCT
  	TAAAGTACAG
  1051	ACAGTCCGAC TGTTTGACGA ATCTTTGAAT GAAACTGATA
  	AAGAACCAGT
  1101	TTACGCGGCA GGGGGTGTTA ATCAGTACCG TCCAAGGTTA
  	AACAACGGTG
  1151	AAAACCTTTC TTTTATCGAT TACGGCAACG GCAAACTCAT
  	CTTATCAAAC
  1201	AACATCAACC AAGGCGCGGG CGGTTTGTAT TTTGAAGGTG
  	ATTTTACGGT
  1251	CTCGCCTGAA AACAACGAAA CGTGGCAAGG CGCGGGCGTT
  	CATATCAGTG
  1301	AAGACAGTAC CGTTACTTGG AAAGTAAACG GCGTGGCAAA
  	CGACCGCCTG
  1351	TCCAAAATCG GCAAAGGCAC GCTGCACGTT CAAGCCAAAG
  	GGGAAAACCA
  1401	AGGCTCGATC AGCGTGGGCG ACGGTACAGT CATTTTGGAT
  	CAGCAGGCAG
  1451	ACGATAAAGG CAAAAAACAA GCCTTTAGTG AAATCGGCTT
  	GNTCAGCGGC
  1501	AGGGGTACGG TGCAACTGAA TGCCGATAAT CAGTTCAACC
  	CCGACAAACT
  1551	CTATTTCGGC TTTCGCGGCG GACGTTTGGA TTTAAACGGG
  	CATTCGCTTT
  1601	CGTTCCACCG TATTCAAAAT ACCGATGAAG GGGCGATGAT
  	TGNCNATCAT
  1651	AATGCCACAA CAACATCCAC CGTTACCATT ACAGGGAATG
  	AAAGTATTAC
  1701	ACAACCGAGT GGTAAGAATA TCAATAGACT TAATTACAGC
  	AAAGAAATTG
  1751	CCTACAACGG TTGGTTTGGC GAGAAAGATA CGACCAAAAC
  	GAACGGGCGG
  1801	CTCAACCTTG TTTACCAGCC CGCCGCAGAA GACCGCACCC
  	NGCTGCTTTC
  1851	CGGCGGAACA AATTTAAACG GCAACATCAC GCAAACAAAC
  	GGCAAACTGT
  1901	TTTTCAGCGG CAGACCGACA CCGCACGCCT ACAATCATTT
  	AGGAAGCGGG
  1951	TGGTCAAAAA TGGAAGGTAT CCCACAAGGA GAAATCGTGT
  	GGGACAACGA
  2001	CTGGATCNAC CGCACGTTTA AAGCGGAAAA TTTCCATATT
  	CAGGGCGGGC
  2051	AGGCGGTGAT TTCCCGCAAT GTTGCCAAAG TGGAAGGCGA
  	TTGNCATTTG
  2101	AGCAATCACG CCCAAGCAGT TTTTGGTGTC GCACCGCATC
  	AAAGCCATAC
  2151	AATCTGTACA CGTTCGGACT GGACNGGTCT GACAAATTGT
  	GTCGAANAAA
  2201	NCATTACCGA CGATAAAGTG ATTGCTTCAT TGACTAAGAC
  	NGACNTNAGC
  2251	GGCANTGTNA GNCTNNCCNA TNACGNTNNT TNAAANCTCN
  	CNGGGCNTGC
  2301	NNCACTNAAN GGCAATCTTA GTGCAAATGG CGATACACGT
  	TATACAGTCA
  2351	GCCACAACGC CACCCAAAAC GGCAACCTTA GCCTCGTGGG
  	CAATGCCCAA
  2401	GCAACATTTA ATCAAGCCAC ATTAAACGGC AACNCATCGG
  	NTTCGGGCAA
  2451	TGCTTCATTT AATCTAAGCA ACAACGCCGC ACAAAACGGC
  	AGTCTGACGC
  2501	TTTCCGACAA CGCTAAGGCA AACGTAAGCC ATTCCGCACT
  	CAACGGCAAT
  2551	GTCTCCCTAG CCGATAAGGC AGTATTCCAT TTTGAAAACA
  	GCCGCTTTAC
  2601	CGGACAACTC AGCGGCAGCA AGGANACAGC ATTACACTTA
  	AAAGACAGCG
  2651	AATGGACGCT GCCGTCAGGC ACGGAATTAG GCAATTTAAA
  	CCTTGACAAC
  2701	GCCACCATTA CACTCAATTC CGCCTATCGC CACGATGCTG
  	CAGGCGCGCA
  2751	AACCGGCAGN GTGTCAGACA CGCCGCGCCG CCGTTCGCGC
  	CGTTCCCTAT
  2801	TATCCGTTAC ACCGCCAACT TCGGTAGAAT CCCGTTTCAA
  	CACGCTGACG
  2851	GTAAACGGCA AATTGAACNG TCAAGGAACA TTCCGCTTTA
  	TGTCGGAACT
  2901	CTTCGGCTAC CGAAGCGACA AATTGAAGCT GGCGGAAAGT
  	TCCGAAGGNA
  2951	CTTACACCTT GGCGGTCAAC AATACCGGCA ACGAACCCGT
  	AAGCCTCGAT
  3001	CAATTGACGG TAGTGGAAGG GAAAGACAAC AAACCGCTGT
  	CCGAAAACCT
  3051	TAATTTCACC CTGCAAAACG AACACGTCGA TGCCGGCGCG
  	TGGCGTTACC
  3101	AACTCATCCG CAAAGACGGC GAGTTCCGCC TGCATAATCC
  	GGTCAAAGAA
  3151	CAAGAGCTTT CCGACAAACT CGGCAAGGCA GAAGCCAAAA
  	AACAGGCGGA
  3201	AAAAGACAAC GCGCAAAGCC TTGACGCGCT GATTGCGGCC
  	GGGCGCGATG
  3251	CCGCCGAAAA GACAGAAAGC GTTGCCGAAC CGGCCCGGCN
  	GGCAGGCGGG
  3301	GAAAATGTCG GCATTATGCA GGCGGAGGAA GAGAAAAAAC
  	GGGTGCAGGC
  3351	GGATAAAGAC AGCGCNTTGG CGAAACAGCG CGAAGCGGAA
  	ACCCGGCCGG
  3401	NTACCACCGC CTTCCCCCGC GCCCGCNGCG CCCGCCGGGA
  	TTTGCCGCAA
  3451	CCGCAGCCCC AACCGCAACC TCAACCCCAA CCGCAGCGCG
  	ACCTGATNAG
  3501	CCGTTATGCC AATAGCGGTT TGAGTGAATT TTCCGCCACG
  	CTCAACAGCG
  3551	TTTTCGCCGT ACAGGACGAA TTGGACCGCG TGTTTGCCGA
  	AGACCGCCGC
  3601	AACGCNGTTT GGACAAGCNG CATCCGGNAC ACCAAACACT
  	ACCGTTCGCA
  3651	AGATTTCCGC GCCTACCGCC AACAAACCGA CCTGCGCCAA
  	ATCGGTATGC
  3701	AGAAAAACCT CGGCAGCGGG CGCGTCGGCA TCCTGTTTTC
  	GCACAACCGG
  3751	ACCGAAAACA NCTTCGACGA CGGCATCGGC AACTCGGCAC
  	GGCTTGCCCA
  3801	CGGCGCCGTT TTCGGGCAAT ACGGCATCGG CAGGTTCGAC
  	ATCGGCATCA
  3851	GCACGGGCGC GGGTTTTAGC AGCGGCANTC TNTCAGACGG
  	CATCGGAGGC
  3901	AAAATCCGCC GCCGCGTGCT GCATTACGGC ATTCAGGCAC
  	GATACCGCGC
  3951	CGGTTTCGGC GGATTCGGCA TCGAACCGTA CATCGGCGCA
  	ACGCGCTATT
  4001	TCGTCCAAAA AGCGGATTAC CGCTACGAAA ACGTCAATAT
  	CGCCACCCCC
  4051	GGTCTTGCGT TCAACCGNTA CCGNGCGGGC ATTAAGGCAG
  	ATTATTCATT
  4101	CAAACCGGCG CAACACATNT CCATCACNCC TTATTTNAGC
  	CTGTCCTATA
  4151	CCGATGCCGC TTCGGGCAAA GTCCGAACAC GCGTCAATAC
  	CGCNGTATTG
  4201	GCTCAGGATT TCGGCAAAAC CCGCAGTGCG GAATGGGGCG
  	TAAACGCCGA
  4251	AATCAAAGGT TTCACGCTGT CCNTCCACGC TGCCGCCGCC
  	AAAGGNCCGC
  4301	AACTGGAAGC GCAACACAGC GCGGGCATCA AATTAGGCTA
  	CCGCTGGTAA

• This encodes a protein having amino acid sequence <SEQ ID 652>:

• 1	MKTTDKRTTE THRKAPKTGR IRFSPAYLAI CLSFGILPQA
  	WAGHTYFGIN
  51	YQYYRDFAEN KGKFAVGAKD IEVYNKKGEL VGKSMTKAPM
  	IDFSVVSRNG
  101	VAALVGDQYI VSVAHNGGYN NVDFGAEGXN PDQHRFSYQI
  	VKRNNYKPDN
  151	SHPYNGDXHM PRLHKFVTDA EPVEMTSDMR GNTYSDKEKY
  	PERVRIGSGH
  201	HYWRYDDDKH GDLSYSGAWL IGGNTHMQGW GNNGVXSLSG
  	DVRHANDYGP
  251	MPIAGAAGDS GSPMFIYDKT NNKWLLNGVL QTGYPYSGRE
  	NGFQLIRKDW
  301	FYDDIYRGDT HTVXFEPRSN GHFSFTSNNN GTGTVTETNE
  	KVSNPKLKVQ
  351	TVRLFDESLN ETDKEPVYAA GGVNQYRPRL NNGENLSFID
  	YGNGKLILSN
  401	NINQGAGGLY FEGDFTVSPE NNETWQGAGV HISEDSTVTW
  	KVNGVANDRL
  451	SKIGKGTLHV QAKGENQGSI SVGDGTVILD QQADDKGKKQ
  	AFSEIGLXSG
  501	RGTVQLNADN QFNPDKLYFG FRGGRLDLNG HSLSFHRIQN
  	TDEGAMIXXH
  551	NATTTSTVTI TGNESITQPS GKNINRLNYS KEIAYNGWFG
  	EKDTTKTNGR
  601	LNLVYQPAAE DRTXLLSGGT NLNGNITQTN GKLFFSGRPT
  	PHAYNHLGSG
  651	WSKMEGIPQG EIVWDNDWIX RTFKAENFHI QGGQAVISRN
  	VAKVEGDXHL
  701	SNHAQAVFGV APHQSHTICT RSDWTGLTNC VEXXITDDKV
  	IASLTKTDXS
  751	GXVXLXXXXX XXLXGXAXLX GNLSANGDTR YTVSHNATQN
  	GNLSLVGNAQ
  801	ATFNQATLNG NXSXSGNASF NLSNNAAQNG SLTLSDNAKA
  	NVSHSALNGN
  851	VSLADKAVFH FENSRFTGQL SGSKXTALHL KDSEWTLPSG
  	TELGNLNLDN
  901	ATITLNSAYR HDAAGAQTGX VSDTPRRRSR RSLLSVTPPT
  	SVESRFNTLT
  951	VNGKLNXQGT FRFMSELFGY RSDKLKLAES SEGTYTLAVN
  	NTGNEPVSLD
  1001	QLTVVEGKDN KPLSENLNFT LQNEHVDAGA WRYQLIRKDG
  	EFRLHNPVKE
  1051	QELSDKLGKA EAKKQAEKDN AQSLDALIAA GRDAAEKTES
  	VAEPARXAGG
  1101	ENVGIMQAEE EKKRVQADKD SALAKQREAE TRPXTTAFPR
  	ARXARRDLPQ
  1151	PQPQPQPQPQ PQRDLXSRYA NSGLSEFSAT LNSVFAVQDE
  	LDRVFAEDRR
  1201	NAVWTSXIRX TKHYRSQDFR AYRQQTDLRQ IGMQKNLGSG
  	RVGILFSHNR
  1251	TENXFDDGIG NSARLAHGAV FGQYGIGRFD IGISTGAGFS
  	SGXLSDGIGG
  1301	KIRRRVLHYG IQARYRAGFG GFGIEPYIGA TRYFVQKADY
  	RYENVNIATP
  1351	GLAFNRYRAG IKADYSFKPA QHXSITPYXS LSYTDAASGK
  	VRTRVNTAVL
  1401	AQDFGKTRSA EWGVNAEIKG FTLSXHAAAA KGPQLEAQHS
  	AGIKLGYRW*

• A transmembrane region is underlined.
• ORF1-1 shows 86.3% identity over a 1462aa overlap with ORF1a:
• 

  
• Homology with Adhesion and Penetration Protein Hap Precursor of H. influenzae (Accession Number P45387)
• Amino acids 23-423 of ORF1 show 59% aa identity with hap protein in 450aa overlap:

• orf1	23	FXAAYLAICLSFGILPQAWAGHTYFGINYQYYRDFAENKGKFAVGAKDIEVYNKKGELVG	82
  		F   +L  C+S GI  QAWAGHTYFGI+YQYYRDFAENKGKF VGAK+IEVYNK+G+LVG
  hap	6	FRLNFLTACVSLGIASQAWAGHTYFGIDYQYYRDFAENKGKFTVGAKNIEVYNKEGQLVG	65
  orf1	83	KSMTKAPMIDFSVVSRNGVAALVGVQYIVSVAHNGGYNNVDFGAEGXNIXDQXRXTYKIV	142
  		SMTKAPMIDFSVVSRNGVAALVG QYIVSVAHNGGYN+VDFGAEG N  DQ R TY+IV
  hap	66	TSMTKAPMIDFSVVSRNGVAALVGDQYIVSVAHNGGYNDVDFGAEGRN-PDQHRFTYQIV	124
  orf1	143	KRNNYKAGTKGHPYGGDYHMPRLHKXVTDAEPVEMTSYMDGRKYIDQNNYPDRVRIGAGR	202
  		KRNNY+A  + HPY GDYHMPRLHK VT+AEPV MT+ MDG+ Y D+ NYP+RVRIG+GR
  hap	125	KRNNYQAWERKHPYDGDYHMPRLHKFVTEAEPVGMTTNMDGKVYADRENYPERVRIGSGR	184
  orf1	203	QYWRSDEDEPNNRESSYHIA----------------------------------------	222
  		QYWR+D+DE  N  SSY+++
  hap	185	QYWRTDKDEETNVHSSYYVSGAYRYLTAGNTHTQSGNGNGTVNLSGNVVSPNHYGPLPTG	244
  orf1	223	-----SGSPMFIYDAQKQKWLINGVLQTGNPYIGKSNGFQLVRKDWFYDEIFAGDTHSVF	277
  		SGSPMFIYDA+K++WLIN VLQTG+P+ G+ NGFQL+R++WFY+E+ A DT SVF
  hap	245	GSKGDSGSPMFIYDAKKKQWLINAVLQTGHPFFGRGNGFQLIREEWFYNEVLAVDTPSVF	304
  orf1	278	--YEPRQNGKYSFNDDNNGTGKIN-AKHEHNSLPNRLKTRTVQLFNVSLSETAREPVYHA	334
  		Y P  NG YSF  +N+GTGK+   +   +    + +  TV+LFN SL++TA+E V  A
  hap	305	QRYIPPINGHYSFVSNNDGTGKLTLTRPSKDGSKAKSEVGTVKLFNPSLNQTAKEHV-KA	363
  orf1	335	AGGVNSYRPRLNNGENISFIDEGKGELILTSNINQGAGGLYFQGDFTV-SPENNETWQGA	393
  		A G N Y+PR+  G+NI   D+GKG L + +NINQGAGGLYF+G+F V   +NN TWQGA
  hap	364	AAGYNIYQPRMEYGKNIYLGDQGKGTLTIENNINQGAGGLYFEGNFVVKGKQNNITWQGA	423
  orf1	394	GVHISEDSTVTWKVNGVANDRLSKIGKGTL	423
  		GV I +D+TV WKV+   NDRLSKIG GTL
  hap	424	GVSIGQDATVEWKVHNPENDRLSKIGIGTL	453

• Amino acids 715-1011 of ORF1 show 50% aa identity with hap protein in 258aa overlap:

• Orf1	41	DTRYTVSHNATQ-NGNXSLVXNAQATFNQ-ATLNGNTSASGNASFNLSDHAVQNGSLTLS	98
  		DT+   S   TQ NG+ +L  NA    +  A LNGN +   ++ F LS++A Q G++ LS
  hap	733	DTKVINSIPITQINGSINLTNNATVNIHGLAKLNGNVTLIDHSQFTLSNNATQTGNIKLS	792
  orf1	99	GNAKANVSHSALNGNVSLADKAVFHFESSRFTGQISGGKDTALHLKDSEWTLPSGXELGN	158
  		+A A V+++ LNGNV L D A F  ++S F  QI G KDT + L+++ WT+PS   L N
  hap	793	NHANATVNNATLNGNVHLTDSAQFSLKNSHFWHQIQGDKDTTVTLENATWTMPSDTTLQN	852
  orf1	159	LNLDNATITLNSAYRHDAAGAQTGSATDAPXXXXXXXXXXLLXVTPPTSVESRFNTLTVN	218
  		L L+N+T+TLNSAY        + S+ +AP          L   T PTS E RFNTLTVN
  hap	853	LTLNNSTVTLNSAY--------SASSNNAPRHRRS-----LETETTPTSAEHRFNTLTVN	899
  orf1	219	GKLNGQGTFRFMSELFGYRSDKLKLAESSEGTYTLAVNNTGNEPASLEQLTVVEGKDNKP	278
  		GKL+GQGTF+F S LFGY+SDKLKL+  +EG YTL+V NTG EP +LEQLT++E  DNKP
  hap	900	GKLSGQGTFQFTSSLFGYKSDKLKLSNDAEGDYTLSVRNTGKEPVTLEQLTLIESLDNKP	959
  orf1	279	LSENLNFTLQNEHVDAGA	296
  		LS+ L FTL+N+HVDAGA
  hap	960	LSDKLKFTLENDHVDAGA	977

• Amino acids 1192-1450 of ORF1 show 41% aa identity with hap protein in 259aa overlap:

• Orf1	1	LDRVFAEDRRNAVWTSGIRDTKHYRSQDFRAYRQQTDLRQIGMQKNLGSGRVGILFSHNR	60
  		LDR+F +  ++AVWT+  +D + Y S  FRAY+Q+T+LRQIG+QK L +GR+G +FSH+R
  hap	1135	LDRLFVDQAQSAVWTNIAQDKRRYDSDAFRAYQQKTNLRQIGVQKALANGRIGAVFSHSR	1194
  orf1	61	TENTFDDGIGNSARLAHGAVFGQYGIDRFYXXXXXXXXXXXXXXXXXIGXKXRRRVLHYG	120
  		++NTFD+ + N A L   + F QY                          K  R+ ++YG
  hap	1195	SDNTFDEQVKNHATLTMMSGFAQYQWGDLQFGVNVGTGISASKMAEEQSRKIHRKAINYG	1254
  orf1	121	IQARYRAGFGGFGIEPHIGATRYFVQKADYRYENVNIATPGLAFNRYRAGIKADYSFKPA	180
  		+ A Y+   G  GI+P+ G  RYF+++ +Y+ E V + TP LAFNRY AGI+ DY+F P
  hap	1255	VNASYQFRLGQLGIQPYFGVNRYFIERENYQSEEVRVKTPSLAFNRYNAGIRVDYTFTPT	1314
  orf1	181	QHISITPYLSLSYTDAASGKVRTRVNTAVLAQDFGKTRSAEWGVNAEIKGFTLSLHAAAA	240
  		+IS+ PY  ++Y D ++  V+T VN  VL Q FG+    E G+ AEI  F +S   + +
  hap	1315	DNISVKPYFFVNYVDVSNANVQTTVNLTVLQQPFGRYWQKEVGLKAEILHFQISAFISKS	1374
  orf1	241	KGPQLEAQHSAGIKLGYRW	259
  		+G QL  Q + G+KLGYRW
  hap	1375	QGSQLGKQQNVGVKLGYRW	1393

  
  Homology with a Predicted ORF from N. gonorrhoeae
• The blocks of ORF1 show 83.5%, 88.3%, and 97.7% identities in 467, 298, and 259 aa overlap, respectively with a predicted ORF (ORF1ng) from N. gonorrhoeae:
• 

  
• The complete length ORF1ng nucleotide sequence was identified <SEQ ID 653>:

• 1	ATGAAAACAA CCGACAAACG GACAACCGAA ACACACCGCA
  	AAGCCCCTAA
  51	AACCGGCCGC ATCCGCTTCT CGCCCGCTTA CTTAGCCATA
  	TGCCTGTCGT
  101	TCGGCATTCT GCCCCAAGCC CGGGCGGGAC ACACTTATTT
  	CGGCATCAAC
  151	TACCAATACT ATCGCGACTT TGCCGAAAAT AAAGGCAAGT
  	TTGCAGTCGG
  201	GGCGAAAGAT ATTGAGGTTT ACAACAAAAA AGGGGAGTTG
  	GTCGGCAAAT
  251	CGATGACGAA AGCCCCGATG ATTGATTTTT CTGTGGTATC
  	GCGTAACGGC
  301	GTGGCGGCAT TGGCGGGCGA TCAATATATT GTGAGCGTGG
  	CACATAACGG
  351	CGGCTATAAC AATGTTGATT TTGGTGCGGA GGGAAGCAAT
  	CCCGATCAGC
  401	ACCGCTTTTC TTACCAAATT GTGAAAAGAA ATAATTATAA
  	AGCAGGGACT
  451	AACGGCCATC CTTATGGCGG CGATTATCAT ATGCCGCGTT
  	TGCACAAATT
  501	TGTCACAGAT GCAGAACCTG TTGAGATGAC CAGTTATATG
  	GATGGGTGGA
  551	AATACGCTGA TTTAAATAAA TACCCTGATC GTGTTCGAAT
  	CGGAGCAGGC
  601	AGACAATATT GGCGGTCTGA TGAAGACGAA CCCAATAACC
  	GCGAAAGTTC
  651	ATATCATATT GCAAGCGCAT ATTCTTGGCT CGTCGGTGGC
  	AATACCTTTG
  701	CACAAAATGG ATCAGGTGGT GGCACAGTCA ACTTAGGTAG
  	CGAAAAAATT
  751	AAACATAGCC CATATGGTTT TTTACCAACA GGAGGCTCAT
  	TTGGCGACAG
  801	TGGCTCACCA ATGTTTATCT ATGATGCCCA AAAGCAAAAG
  	TGGTTAATTA
  851	ATGGGGTATT GCAAACAGGC AACCCCTATA TAGGAAAAAG
  	CAATGGCTTC
  901	CAGCTAGTTC GTAAAGATTG GTTCTATGAT GAAATCTTTG
  	CTGGAGATAC
  951	CCATTCAGTA TTCTACGAAC CACATCAAAA TGGGAAATAC
  	TTTTTTAACG
  1001	ACAATAATAA TGGCGCAGGA AAAATCGATG CCAAACATAA
  	ACACTATTCT
  1051	CTACCTTATA GATTAAAAAC ACGAACCGTT CAATTGTTTA
  	ATGTTTCTTT
  1101	ATCCGAGACA GCAAGAGAAC CTGTTTATCA TGCTGCAGGT
  	GGGGTCAACA
  1151	GTTATCGACC CAGACTGAAT AATGGAGAAA ATATTTCCTT
  	TATTGACAAA
  1201	GGAAAAGGTG AATTGATACT TACCAGCAAC ATCAACCAAG
  	GCGCGGGCGG
  1251	TTTGTATTTT GAGGGTAATT TTACGGTCTC GCCTAAAAAC
  	AACGAAACGT
  1301	GGCAAGGCGC GGGCGTTCAT ATCAGTGATG GCAGTACCGT
  	TACTTGGAAA
  1351	GTAAACGGCG TGGCAAACGA CCGCCTGTCC AAAATCGGCA
  	AAGGCACGCT
  1401	GCTGGTTCAA GCCAAAGGGG AAAACCAAGG CTCGGTCAGC
  	GTGGGCGACG
  1451	GTAAAGTCAT CTTAGATCAG CAGGCGGACG ATCAAGGCAA
  	AAAACAAGCC
  1501	TTTAGTGAAA TCGGCTTGGT CAGCGGCAGG GGGACGGTGC
  	AACTGAATGC
  1551	CGATAATCAG TTCAACCCCG ACAAACTCTA TTTCGGCTTT
  	CGCGGCGGAC
  1601	GTTTGGATTT GAACGGGCAT TCGCTTTCGT TCCACCGCAT
  	TCAAAATACC
  1651	GATGAAGGGG CGATGATTGT CAACCACAAT CAAGACAAAG
  	AATCCACCGT
  1701	TACCATTACA GGCAATAAAG ATATTACTAC AACCGGCAAT
  	AACAACAACT
  1751	TGGATAGCAA AAAAGAAATT GCCTACAACG GTTGGTTTGG
  	CGAGAAAGAT
  1801	GCAACCAAAA CGAACGGGCG GCTCAATCTG AATTACCAAC
  	CGGAAGAAGC
  1851	GGATCGCACT TTACTGCTTT CCGGCGGAAC AAATTTAAAC
  	GGCAATATCA
  1901	CGCAAACAAA CGGCAAACTG TTTTTCAGCG GCAGACCGAC
  	ACCGCACGCC
  1951	TACAATCATT TAGGAAGCGG GTGGTCAAAA ATGGAAGGTA
  	TCCCACAAGG
  2001	AGAAATCGTG TGGGACAACG ATTGGATCGA CCGCACATTT
  	AAAGCGGAAA
  2051	ACTTCCATAT TCAGGGCGGA CAAGCGGTGG TTTCCCGCAA
  	TGTTGCCAAA
  2101	GTGGAAGGCG ATTGGCATTT AAGCAATCAC GCCCAAGCAG
  	TTTTCGGTGT
  2151	CGCACCGCAT CAAAGCCACA CAATCTGTAC ACGTTCGGAC
  	TGGACGGGTC
  2201	TGACAAGTTG TACCGAAAAA ACCATTACCG ACGATAAAGT
  	GATTGCTTCA
  2251	TTGAGCAAGA CCGACATCAG AGGCAATGTC AGCCTTGCCG
  	ATCACGCTCA
  2301	TTTAAATCTC ACAGGACTTG CCACACTCAA CGGCAATCTT
  	AGTGCAGGCG
  2351	GAGACACGCA CTATACGGTT ACGCGCAACG CCACCCAAAA
  	CGGCAACCTC
  2401	AGCCTCGTGG GCAATGCCCA AGCAACATTT AATCAAGCCA
  	CATTAAACGG
  2451	CAACACATCG GCTTCGGACA ATGCTTCATT TAATCTAAGC
  	AACAACGCCG
  2501	TACAAAACGG CAGTCTGACG CTTTCCGACA ACGCTAAGGC
  	AAACGTAAGC
  2551	CATTCCGCAC TCAACGGCAA TGTCTCCCTA GCCGATAAGG
  	CAGTATTCCA
  2601	TTTTGAAAAC AGCCGCTTTA CCGGAAAAAT CAGCGGCGGC
  	AAGGATACGG
  2651	CATTACACTT AAAAGACAGC GAATGGACGC TGCCGTCGGG
  	CACGGAATTA
  2701	GGCAATTTAA ACCTTGACAA CGCCACCATT ACACTCAATT
  	CCGCCTATCG
  2751	ACACGATGCG GCAGGCGCGC AAACCGGCAG TGCGGCAGAT
  	GCGCCGCGCC
  2801	GCCGTTCGCG CCGTTCCCTA TTATCCGTTA CGCCGCCAAC
  	TTCGGCAGAA
  2851	TCCCGTTTCA ACACGCTGAC GGTAAACGGC AAATTGAACG
  	GTCAGGGAAC
  2901	ATTCCGCTTT ATGTCGGAAC TCTTCGGCTA CCGCAGCGGC
  	AAATTGAAGC
  2951	TGGCGGAAAG TTCCGAAGGC ACTTACACCT TGGCTGTCAA
  	CAATACCGGC
  3001	AACGAACCCG TAAGTCTCGA GCAATTGACG GTAGTGGAAG
  	GAAAAGACAA
  3051	CACACCGCTG TCCGAAAATC TTAATTTCAC CCTGCaaaAc
  	gaacacgtcg
  3101	atgccggcgc atggCGTTAT CAGCTTATCC gcaaagacgG
  	CGAGTTCCgc
  3151	CTGCATAATC CGGTCAAAGA ACAAGAGCTT TCCGACAAAC
  	TCGGCAAGgc
  3201	gggagaaACA GAggccgccT TGACGGCAAA ACAGGCacaA
  	CTTGCCGCCA
  3251	AAcaacaggc ggaaaAAGAC AACgcgcaaa gccttgAcgc
  	gctgattgcg
  3301	gCcgggcgca atgccaccga AAAGGCAgaa agtgttgccg
  	aaccgGCCCG
  3351	GCAGGCAGGC GGGGAAAAtg ccgGCATTAT GCAGGCGGAG
  	GAAGAGAAAA
  3401	AACGGGTGCA GGCGGATAAA GACACCGCCT TGGCGAAACA
  	GCGCGAAGCG
  3451	GAAACCCGGC CGGCTACCAC CGCCTTCCCC CGCGCCCGCC
  	GCGCCCGCCG
  3501	GGATTTGCCG CAACCGCAGC CCCAACCGCA ACCCCAACCG
  	CAGCGCGACC
  3551	TGATCAGCCG TTATGCCAAT AGCGGTTTGA GTGAATTTTC
  	CGCCACGCTC
  3601	AACAGCGTTT TCGCCGTACA GGACGAATTG GACCGCGTGT
  	TTGCCGAAGA
  3651	CCGCCGCAAC GCCGTTTGGA CAAGCGGCAT CCGGGACACC
  	AAACACTACC
  3701	GTTCGCAAGA TTTCCGCGCC TACCGCCAAC AAACCGACCT
  	GCGCCAAATC
  3751	GGTATGCAGA AAAACCTCGG CAGCGGGCGC GTCGGCATCC
  	TGTTTTCGCA
  3801	CAACCGGACC GGAAACACCT TCGACGACGG CATCGGCAAC
  	TCGGCACGGC
  3851	TTGCCCACGG TGCCGTTTTC GGGCAATACG GCATCGGCAG
  	GTTCGACATC
  3901	GGCATCAGCG CGGGCGCGGG TTTTAGTAGC GGCAGCCTTT
  	CAGACGGCAT
  3951	CAGAGGCAAA ATCCGCCGCC GCGTGCTGCA TTACGGCATT
  	CAGGCAAGAT
  4001	ACCGCGCAGG TTTCGGCGGA TTCGGCATCG AACCGCACAT
  	CGGCGCAACG
  4051	CGCTATTTCG TCCAAAAAGC GGATTACCGA TACGAAAACG
  	TCAATATCGC
  4101	CACCCCGGGC CTTGCATTCA ACCGCTACCG CGCGGGCATT
  	AAGGCAGATT
  4151	ATTCATTCAA ACCGGCGCAA CACATTTCCA TCACGCCTTA
  	TTTGAGCCTG
  4201	TCCTATACCG ATGCCGCTTC CGGCAAAGTC CGAACGCGCG
  	TCAATACCGC
  4251	CGTATTGGCG CAGGATTTCG GCAAAACCCG CAGTGCGGAA
  	TGGGGCGTAA
  4301	ACGCCGAAAT CAAAGGTTTC ACGCTGTCCC TCCACGCTGC
  	CGCCGCCAAG
  4351	GGGCCGCAAT TGGAAGCGCA GCACAGCGCG GGCATCAAAT
  	TAGGCTACCG
  4401	CTGGTAA

• This is predicted to encode a protein having amino acid sequence <SEQ ID 654>:

• 1	MKTTDKRTTE THRKAPKTGR IRFSPAYLAI CLSFGILPQA
  	RAGHTYFGIN
  51	YQYYRDFAEN KGKFAVGAKD IEVYNKKGEL VGKSMTKAPM
  	IDFSVVSRNG
  101	VAALAGDQYI VSVAHNGGYN NVDFGAEGSN PDQHRFSYQI
  	VKRNNYKAGT
  151	NGHPYGGDYH MPRLHKFVTD AEPVEMTSYM DGWKYADLNK
  	YPDRVRIGAG
  201	RQYWRSDEDE PNNRESSYHI ASAYSWLVGG NTFAQNGSGG
  	GTVNLGSEKI
  251	KHSPYGFLPT GGSFGDSGSP MFIYDAQKQK WLINGVLQTG
  	NPYIGKSNGF
  301	QLVRKDWFYD EIFAGDTHSV FYEPHQNGKY FFNDNNNGAG
  	KIDAKHKHYS
  351	LPYRLKTRTV QLFNVSLSET AREPVYHAAG GVNSYRPRLN
  	NGENISFIDK
  401	GKGELILTSN INQGAGGLYF EGNFTVSPKN NETWQGAGVH
  	ISDGSTVTWK
  451	VNGVANDRLS KIGKGTLLVQ AKGENQGSVS VGDGKVILDQ
  	QADDQGKKQA
  501	FSEIGLVSGR GTVQLNADNQ FNPDKLYFGF RGGRLDLNGH
  	SLSFHRIQNT
  551	DEGAMIVNHN QDKESTVTIT GNKDITTTGN NNNLDSKKEI
  	AYNGWFGEKD
  601	ATKTNGGLNL NYPPEEADRT LLLSGGTNLN GNITQTNGKL
  	FFSGRPTPHA
  651	YNHLGSGWSK MEGIPQGEIV WDNDWIDRTF KAENFHIQGG
  	QAVVSRNVAK
  701	VEGDWHLSNH AQAVFGVAPH QSHTICTRSD WTGLTSCTEK
  	TITDDKVIAS
  751	LSKTDVRGNV SLADHAHLNL TGLATFNGNL VQAETRTIRL
  	RANATQNGNL
  801	SLVGNAQATF NQATLNGNTS ASDNASFNLS NNAVQNGSLT
  	LSDNAKANVS
  851	HSALNGNVSL ADKAVFHFEN SRFTGKISGG KDTALHLKDS
  	EWTLPSGTEL
  901	GNLNLDNATI TLNSAYRHDA AGAQTGSAAD APRRRSRRSL
  	LSVTPPTSAE
  951	SRFNTLTVNG KLNGQGTFRF MSELFGYRSG KLKLAESSEG
  	TYTLAVNNTG
  1001	NEPVSLEQLT VVEGKDNTPL SENLNFTLQN EHVDAGAWRY
  	QLIRKDGEFR
  1051	LHNPVKEQEL SDKLGKAGET EAALTAKQAQ LAAKQQAEKD
  	NAQSLDALIA
  1101	AGRNATEKAE SVAEPARQAG GENAGIMQAE EEKKRVQADK
  	DTALAKQREA
  1151	ETRPATTAFP RARRARRDLP QPQPQPQPQP QRDLISRYAN
  	SGLSEFSATL
  1201	NSVFAVQDEL DRVFAEDRRN AVWTSGIRDT KHYRSQDFRA
  	YRQQTDLRQI
  1251	GMQKNLGSGR VGILFSHNRT GNTFDDGIGN SARLAHGAVF
  	GQYGIGRFDI
  1301	GISAGAGFSS GSLSDGIRGK IRRRVLHYGI QARYRAGFGG
  	FGIEPHIGAT
  1351	RYFVQKADYR YENVNIATPG LAFNRYRAGI KADYSFKPAQ
  	HISITPYLSL
  1401	SYTDAASGKV RTRVNTAVLA QDFGKTRSAE WGVNAEIKGF
  	TLSLHAAAAK
  1451	GPQLEAQHSA GIKLGYRW*

• Underlined and double-underlined sequences represent the active site of a serine protease (trypsin family) and an ATP/GTP-binding site motif A (P-loop).
• ORF1-1 and ORF1ng show 93.7% identity in 1471 aa overlap:
• 

  
• In addition, ORF1ng shows 55.7% identity with hap protein (P45387) over a 1455aa overlap:
• 

  

  
• Based on this analysis, it is predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 78
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 655>:

• 1	..AAGGTGTGGC AATTTGTCGA AGA.CCGCTG CGTGCCGTCG
  	TGCCTGCCGA
  51	CAGTTTTGAA CCGACCGCGC AAAAATTGAA CCTGTTTAAG
  	GCGGGTGCGG
  101	CAACCATTTT GTTTTATGAA GATCAAAATG TCGTCAAAGG
  	TTTGCAGGAG
  151	CAGTTCCCTG CTTATGCCGC TAACTTCCCC GTTTGGGCGg
  	ATCAGGCAAA
  201	CGCGATGGTG CAGTATGCCG TTTGGACGAC ACTTGCCGCG
  	GTCGGCGTAG
  251	GTGCAAACCT GCAACATTAC AATCCCTTGC CCGATGCGGC
  	GATTGCCAAA
  301	GCGTGGAATA TCCCCGAAAA CTGGTTGTTG CGCGCACAAA
  	TGGTTATCGG
  351	CGGTATTGAA GGGGCGGCAG GTGAAAAGAC CTTTGAACCC
  	GTTGCAGAAC
  401	GTTTGAAAGT GTTCGGCGCA TAA

• This corresponds to the amino acid sequence <SEQ ID 656; ORF6>:

• 1	. . . KVWQFVEXPL RAWPADSFE PTAQKLNLFK AGAATILFYE  DQNVVKGLQE
  51	QFPAYAANFP VWADQANAMV QYAVWTTLAA VGVGANLQHY NPLPDAAIAK
  101	AWNIPENWLL RAQMVIGGIE GAAGEKTFEP VAERLKVFGA *

• Further sequence analysis revealed a further partial DNA sequence <SEQ ID 657>:

• 1	. . . CTGCGTGCCG TCGTGCCTGC CGACAGTTTT GAACCGACCG CGCAAAAATT
  51	GAACCTGTTT AAGGCGGGTG CGGCAACCAT TTTGTTTTAT GAAGATCAAA
  101	ATGTCGTCAA AGGTTTGCAG GAGCAGTTCC CTGCTTATGC CGCTAACTTC
  151	CCCGTTTGGG CGGATCAGGC AAACGCGATG GTGCAGTATG CCGTTTGGAC
  201	GACACTTGCC GCGGTCGGCG TAGGTGCAAA CCTGCAACAT TACAATCCCT
  251	TGCCCGATGC GGCGATTGCC AAAGCGTGGA ATATCCCCGA AAACTGGTTG
  301	TTGCGCGCAC AAATGGTTAT CGGCGGTATT GAAGGGGCGG CAGGTGAAAA
  351	GACCTTTGAA CCCGTTGCAG AACGTTTGAA AGTGTTCGGC GCATAA

• This corresponds to the amino acid sequence <SEQ ID 658; ORF6-1>:

• 1	. . . LRAVVPADSF EPTAQKLNLF KAGAATILFY EDQNVVKGLQ EQFPAYAANF
  51	PVWADQANAM VQYAVWTTLA AVGVGANLQH YNPLPDAAIA KAWNIPENWL
  101	LRAQMVIGGI EGAAGEKTFE PVAERLKVFG A*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF6 shows 98.6% identity over a 140aa overlap with an ORF (ORF6a) from strain A of N. meningitidis:
• 
• The complete length ORF6a nucleotide sequence <SEQ ID 659> is:

• 1	ATGACCCGTC AATCTCTGCA ACAGGCTGCC GAAAGCCGCC GTTCCATTTA
  51	TTCGTTAAAT AAAAATCTGC CCGTCGGCAA AGATGAAATC GTCCAAATCG
  101	TCGAACACGC CGTTTTGCAC ACACCTTCTT CGTTCAATTC CCAATCTGCC
  151	CGTGTGGTCG TGCTGTTTGG CGAAGAGCAT GATAAGGTGT GGCAATTTGT
  201	CGAAGACGCG CTGCGTGCCG TCGTGCCTGC CGACAGTTTT GAACCGACCG
  251	CGCAAAAATT GAACCTGTTT AAGGCGGGTG CGGCAACTAT TTTGTTTTAT
  301	GAAGATCAAA ATGTCGTCAA AGGTTTGCAG GAGCAGTTCC CTGCTTATGC
  351	CGCCAACTTT CCCGTTTGGG CGGACCAGGC GAACGCGATG GTGCAGTATG
  401	CCGTTTGGAC GACACTTGCC GCGGTCGGCG TAGGTGCAAA CCTGCAACAT
  451	TACAATCCCT TGCCCGATGC GGCGATTGCC AAAGCGTGGA ATATCCCCGA
  501	AAACTGGTTG TTGCGCGCAC AAATGGTTAT CGGCGGTATT GAAGGGGCGG
  551	CAGGTGAAAA GACCTTTGAA CCAGTTGCAG AACGTTTGAA AGTGTTCGGC
  601	GCATAA

• This is predicted to encode a protein having amino acid sequence <SEQ ID 660>:

• 1	MTRQSLQQAA ESRRSIYSLN KNLPVGKDEI VQIVEHAVLH TPSSFNSQSA
  51	RVVVLFGEEH DKVWQFVEDA LRAVVPADSF EPTAQKLNLF KAGAATILFY
  101	EDQNVVKGLQ EQFPAYAANF PVWADQANAM VQYAVWTTLA AVGVGANLQH
  151	YNPLPDAAIA KAWNIPENWL LRAQMVIGGI EGAAGEKTFE PVAERLKVFG
  201	A*

• ORF6a and ORF6-1 show 100.0% identity in 131 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF6 shows 95.7% identity over a 140aa overlap with a predicted ORF (ORF6ng) from N. gonorrhoeae:
• 
• The complete length ORF6ng nucleotide sequence <SEQ ID 661> was identified as:

• 1	ATGGCCGTTG CGTCAAATGT CAGCTTGGAT ATGTCCAATC CTACGGTGTT
  51	ACGCATGGGA TTACCCTTAT ATATTGCGTC CCTAAGAAGG GGCGCAATAT
  101	ATAAGGTGTG GCAATTTGTC GAAGACGCGC TGCGTGCCGT CGTGCCTGCC
  151	GACAGTTTTG AACCGACCGC GCAAAAATTG AAGCTGTTTA AGGCGGGCGC
  201	GGCAACCATT TTGTTTTATG AAGATCAAAA TGTCGTCAAA GGTTTGCAGG
  251	AGCAGTTCCC TGCTTATGCC GCCAACTTTC CCGTTTGGGC GGACCAGGCG
  301	AACGCTATGG TACAGTATGC CGTCTGGACG ACACTTGCCG CGGTCGGTGC
  351	AGGTGCAAAT CTGCAACATT ACAACCCCTT GCCCGATGTG GCGATTGCTA
  401	AAGCGTGGAA TATTCCCGAA AACTGGCTGT TGCGCGCGCA AATGGTTATC
  451	GGTGGTATTG AAGGGGcggc aggtgaaaaa gtctttgaac CCGTTGCgga
  501	acgtttgAAA GTGTTCGGCG CATAA

• This encodes a protein having amino acid sequence <SEQ ID 662>:

• 1	MAVASNVSLD MSNPTVLRMG LPLYIASLRR GAIYKVWQFV EDALRAVVPA
  51	DSFEPTAQKL KLFKAGAATI LFYEDQNVVK GLQEQFPAYA ANFPVWADQA
  101	NAMVQYAVWT TLAAVGAGAN LQHYNPLPDV AIAKAWNIPE NWLLRAQMVI
  151	GGIEGAAGEK VFEPVAERLK VFGA*

• ORF6ng and ORF6-1 show 96.9% identity in 131 aa overlap:
• 
• It is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 79
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 663>

• 1	. . . GGCTACAACT ACCTGTTCGC GCGCGGCAGC CGCATCGCCA ACTACCAAAT
  51	CAACGGCATC CCCGTTGCCG ACGCGCTGGC CGATACGGGt CAATGCCAAC
  101	ACCGCCGCCT ATGAGCGCGT AGAAGTCGTG CGCGGCGTGG CGGGGCTGCT
  151	GGACGGCACG GGCGAGCCTT CCGCCACCGT CAATCTGGTG CGCAAACGCC
  201	TGACCCGCAA GCCATTGTTT GAAGTCCGCG CCGAAGCgGG CAACCGcAAA
  251	CATTTCGGGC TGGACGCGGA CGTATCGGGC AGCCTGAACA CCGAAG.crC
  301	rCTGCGCgGC CGCCTGGTTT CCAcCTTCGG ACGCGGCGAC TCGTGGCGGC
  351	GGCGCGAACG CAGCCGskAT GCCGAACTCT ACGGCATTTT GGAATACGAC
  401	ATCGCACCGC AAACCCGCGT CCACGCArGC ATGGACTACC AGCAGGCGAA
  451	AGAAACCGCC GACGCGCCGC TCAGcTACGC CGTGTACGAC AGCCAAGGTT
  501	ATGCCACCGC CTTCGGCCCG AAAGACAACC CCGCCACAAA TTGGGCGAAC
  551	AGCCACCACC GTGCGCTCAA CCTGTTCGCC GGCATCGAAC ACCGCTTCAA
  601	CCAAGACTGG AAACTCAAAG CCGAATACGA CTAC . . .

• This corresponds to the amino acid sequence <SEQ ID 664; ORF23>:

• 1	. . . GYNYLFARGS RIANYQINGI PVADALADTG NANTAAYERV EVVRGVAGLL
  51	DGTGEPSATV NLVRKRLTRK PLFEVRAEAG NRKHFGLDAD VSGSLNTEXX
  101	LRGRLVSTFG RGDSWRRRER SRXAELYGIL EYDIAPQTRV HAXMDYQQAK
  151	ETADAPLSYA VYDSQGYATA FGPKDNPATN WANSHHRALN LFAGIEHRFN
  201	QDWKLKAEYD Y . . .

• Further work revealed the complete nucleotide sequence <SEQ ID 665>:

• 1	ATGACACGCT TCAAATATTC CCTGCTGTTT GCCGCCCTGT TGCCCGTGTA
  51	CGCGCAGGCC GATGTTTCTG TTTCAGACGA CCCCAAACCG CAGGAAAGCA
  101	CTGAATTGCC GACCATCACC GTTACCGCCG ACCGCACCGC GAGTTCCAAC
  151	GACGGCTACA CTGTTTCCGG CACGCACACC CCGCTCGGGC TGCCCATGAC
  201	CCTGCGCGAA ATCCCGCAGA GCGTCAGCGT CATCACATCG CAACAAATGC
  251	GCGACCAAAA CATCAAAACG CTCGACCGCG CCCTGTTGCA GGCGACCGGC
  301	ACCAGCCGCC AGATTTACGG CTCCGACCGC GCGGGCTACA ACTACCTGTT
  351	CGCGCGCGGC AGCCGCATCG CCAACTACCA AATCAACGGC ATCCCCGTTG
  401	CCGACGCGCT GGCCGATACG GGCAATGCCA ACACCGCCGC CTATGAGCGC
  451	GTAGAAGTCG TGCGCGGCGT GGCGGGGCTG CTGGACGGCA CGGGCGAGCC
  501	TTCCGCCACC GTCAATCTGG TGCGCAAACG CCTGACCCGC AAGCCATTGT
  551	TTGAAGTCCG CGCCGAAGCG GGCAACCGCA AACATTTCGG GCTGGACGCG
  601	GACGTATCGG GCAGCCTGAA CACCGAAGGC ACGCTGCGCG GCCGCCTGGT
  651	TTCCACCTTC GGACGCGGCG ACTCGTGGCG GCGGCGCGAA CGCAGCCGCG
  701	ATGCCGAACT CTACGGCATT TTGGAATACG ACATCGCACC GCAAACCCGC
  751	GTCCACGCAG GCATGGACTA CCAGCAGGCG AAAGAAACCG CCGACGCGCC
  801	GCTCAGCTAC GCCGTGTACG ACAGCCAAGG TTATGCCACC GCCTTCGGCC
  851	CGAAAGACAA CCCCGCCACA AATTGGGCGA ACAGCCGCCA CCGTGCGCTC
  901	AACCTGTTCG CCGGCATCGA ACACCGCTTC AACCAAGACT GGAAACTCAA
  951	AGCCGAATAC GACTACACCC GCAGCCGCTT CCGCCAGCCC TACGGCGTAG
  1001	CAGGCGTGCT TTCCATCGAC CACAACACCG CCGCCACCGA CCTGATTCCC
  1051	GGTTATTGGC ACGCCGACCC GCGCACCCAC AGCGCCAGCG TGTCATTGAT
  1101	CGGCAAATAC CGCCTGTTCG GCCGCGAACA CGATTTAATC GCGGGTATCA
  1151	ACGGTTACAA ATACGCCAGC AACAAATACG GCGAACGCAG CATCATCCCC
  1201	AACGCCATTC CCAACGCCTA CGAATTTTCC CGCACGGGTG CCTACCCGCA
  1251	GCCTGCATCG TTTGCCCAAA CCATCCCGCA ATACGGCACC AGGCGGCAAA
  1301	TCGGCGGCTA TCTCGCCACC CGTTTCCGCG CCGCCGACAA CCTTTCGCTG
  1351	ATTTTGGGCG GACGATACAC CCGTTACCGC ACCGGCAGCT ACGACAGCCG
  1401	CACACAAGGC ATGACCTATG TGTCCGCCAA CCGTTTCACC CCCTACACAG
  1451	GCATCGTGTT CGACCTGACC GGCAACCTGT CTCTTTACGG CTCGTACAGC
  1501	AGCCTGTTCG TCCCGCAATC GCAAAAAGAC GAACACGGCA GCTACCTGAA
  1551	ACCCGTAACC GGCAACAATC TGGAAGCCGG CATCAAAGGC GAATGGCTTG
  1601	AAGGCCGTCT GAACGCATCC GCCGCCGTGT ACCGCGCCCG TAAAAACAAC
  1651	CTCGCCACCG CAGCAGGACG CGACCCGAGC GGCAACACCT ACTACCGCGC
  1701	CGCCAACCAA GCCAAAACCC ACGGCTGGGA AATCGAAGTC GGCGGCCGCA
  1751	TCACGCCCGA ATGGCAGATA CAGGCAGGTT ACAGCCAAAG CAAAACCCGC
  1801	GACCAAGACG GCAGCCGCCT GAACCCCGAC AGCGTACCCG AACGCAGCTT
  1851	CAAACTCTTC ACTGCCTACC ACTTTGCCCC CGAAGCCCCC AGCGGCTGGA
  1901	CCATCGGCGC AGGCGTGCGC TGGCAGAGCG AAACCCACAC CGACCCTGCC
  1951	ACGCTCCGCA TCCCCAACCC CGCCGCCAAA GCCCGCGCCG CCGACAACAG
  2001	CCGCCAAAAA GCCTACGCCG TCGCCGACAT CATGGCGCGT TACCGCTTCA
  2051	ATCCGCGCGC CGAACTGTCG CTGAACGTGG ACAATCTGTT CAACAAACAC
  2101	TACCGCACCC AGCCCGACCG CCACAGCTAC GGCGCACTGC GGACAGTGAA
  2151	CGCGGCGTTT ACCTATCGGT TTAAATAA

• This corresponds to the amino acid sequence <SEQ ID 666; ORF23-1>:

• 1	MTRFKYSLLF AALLPVYAQA DVSVSDDPKP QESTELPTIT VTADRTASSN
  51	DGYTVSGTHT PLGLPMTLRE IPQSVSVITS QQMRDQNIKT LDRALLQATG
  101	TSRQIYGSDR AGYNYLFARG SRIANYQING IPVADALADT GNANTAAYER
  151	VEVVRGVAGL LDGTGEPSAT VNLVRKRLTR KPLFEVRAEA GNRKHFGLDA
  201	DVSGSLNTEG TLRGRLVSTF GRGDSWRRRE RSRDAELYGI LEYDIAPQTR
  251	VHAGMDYQQA KETADAPLSY AVYDSQGYAT AFGPKDNPAT NWANSRHRAL
  301	NLFAGIEHRF NQDWKLKAEY DYTRSRFRQP YGVAGVLSID HNTAATDLIP
  351	GYWHADPRTH SASVSLIGKY RLFGREHDLI AGINGYKYAS NKYGERSIIP
  401	NAIPNAYEFS RTGAYPQPAS FAQTIPQYGT RRQIGGYLAT RFRAADNLSL
  451	ILGGRYTRYR TGSYDSRTQG MTYVSANRFT PYTGIVFDLT GNLSLYGSYS
  501	SLFVPQSQKD EHGSYLKPVT GNNLEAGIKG EWLEGRLNAS AAVYRARKNN
  551	LATAAGRDPS GNTYYRAANQ AKTHGWEIEV GGRITPEWQI QAGYSQSKTR
  601	DQDGSRLNPD SVPERSFKLF TAYHFAPEAP SGWTIGAGVR WQSETHTDPA
  651	TLRIPNPAAK ARAADNSRQK AYAVADIMAR YRFNPRAELS LNVDNLFNKH
  701	YRTQPDRHSY GALRTVNAAF TYRFK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with the Ferric-Pseudobactin Receptor PupB of Pseudomonas putida (Accession Number P38047)
• ORF23 and PupB protein show 32% aa identity in 205aa overlap:

• Orf23	6	FARGSRIANYQINGIPVADALADTGNANTAAYERVEVVRGVAGLLDGTGEPSATVNLVRK	65
  		++RG  I NY+++G+P +   L D  + + A ++RVE+VRG  GL+ G G PSAT+NL+RK
  PupB	215	WSRGFAIQNYEVDGVPTSTRL-DNYSQSMAMFDRVEIVRGATGLISGMGNPSATINLIRK	273
  Orf23	66	RLTRKPLFEVRAEAGNRKHFGLDADVSGSLNTEXXLRGRLVSTFXXXXXXXXXXXXXXAE	125
  		R T +    +  EAGN   +G   DVSG L     +RGR V+ +
  PupB	274	RPTAEAQASITGEAGNWDRYGTGFDVSGPLTETGNIRGRFVADYKTEKAWIDRYNQQSQL	333
  Orf23	126	LYGILEYDIAPQTRVHAXMDYQQAKETADAPLSYAVYD--SQGYATAFGPKDNPATNWAN	183
  		+YGI E+D++  T+      Y   +    D+PL   +    S G  T      N A +W+
  PupB	334	MYGITEFDLSEDTLLTVGFSY--LRSDIDSPLRSGLPTRFSTGERTNLKRSLNAAPDWSY	391
  Orf23	184	SHHRALNLFAGIEHRFNQDWKLKAE	208
  		+ H   + F  IE +    W  K E
  PupB	392	NDHEQTSFFTSIEQQLGNGWSGKIE	416

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF23 shows 95.7% identity over a 211aa overlap with an ORF (ORF23a) from strain A of N. meningitidis:
• 
• The complete length ORF23a nucleotide sequence <SEQ ID 667> is:

• 1	ATGACACGCT TCAAATATTC CCTGCTGTTT GCCGCCCTGT TGCCCGTGTA
  51	CGCGCAGGCC GATGTTTCTG TTTCAGACGA CCCAAAACCG CAGGAAAGCA
  101	CTGAATTGCC GACCATCACC GTTACCGCCG ACCGCACCGC GAGTTCCAAC
  151	GACGGCTACA CTGTTTCCGG CACGCACACC CCGCTCGGGC TGCCCATGAC
  201	CCTGCGCGAA ATCCCGCAGA GCGTCAGCGT CATCACATCG CAACAAATGC
  251	GCGACCAAAA CATCAAAGCG CTCGACCGCG CCCTGTTGCA GGCGACCGGC
  301	ACCAGCCGCC AGATTTACGG CTCCGACCGC GCGGGCTACA ACTACCTGTT
  351	CGCGCGCGGC AGCCGCATCG CCAACTACCA AATCAACGGC ATCCCCGTTG
  401	CCGACGCGCT GGCCGATACG GGCAATGCCA ACACCGCCGC CTATGAGCGC
  451	GTAGAAGTCG TGCGCGGCGT GGCGGGGCTG CTGGACGGCA CGGGCGAGCC
  501	TTCCGCCACC GTCAATCTGG TGCGCAAACG CCCGACCCGC AAGCCATTGT
  551	TTGAAGTCCG CGCCGAAGCG GGCAACCGCA AACATTTCGG GCTGGGCGCG
  601	GACGTATCGG GCAGCCTGAA TGCCGAAGGC ACGCTGCGCG GCCGCCTGGT
  651	TTCCACCTTC GGACGCGGCG ACTCGTGGCG GCAGCGCGAA CGCAGCCGCG
  701	ATGCCGAACT CTACGGCATT TTGGAATACG ACATCGCACC GCAAACCCGC
  751	GTCCACGCAG GCATGGACTA CCAGCAGGCG AAAGAAACCG CCGACGCGCC
  801	GCTCAGCTAC GCCGTGTACG ACAGCCAAGG TTATGCCACC GCCTTCGGCC
  851	CGAAAGACAA CCCCGCCACA AATTGGGCGA ACAGCCGCCA CCGTGCGCTC
  901	AACCTGTTCG CCGGCATCGA ACACCGCTTC AACCAAGACT GGAAACTCAA
  951	AGCCGAATAC GACTACACCC GCAGCCGCTT CCGCCAGCCC TACGGCGTAG
  1001	CAGGCGTGCT TTCCATCGAC CACAACACCG CCGCCACCGA CCTGATTCCC
  1051	GGTTATTGGC ACGCCGACCC GCGCACCCAC AGCGCCAGCG TGTCATTAAT
  1101	CGGCAAATAC CGCCTGTTCG GCCGCGAACA CGATTTAATC GCGGGTATCA
  1151	ACGGTTACAA ATACGCCAGC AACAAATACG GCGAACGCAG CATCATCCCC
  1201	AACGCCATTC CCAACGCCTA CGAATTTTCC CGCACGGGTG CCTACCCGCA
  1251	GCCTGCATCG TTTGCCCAAA CCATCCCGCA ATACGGCACC AGGCGGCAAA
  1301	TCGGCGGCTA TCTCGCCACC CGTTTCCGCG CCGCCGACAA CCTTTCGCTG
  1351	ATACTCGGCG GCAGATACAG CCGTTACCGC ACCGGCAGCT ACGACAGCCG
  1401	CACACAAGGC ATGACCTATG TGTCCGCCAA CCGTTTCACC CCCTACACAG
  1451	GCATCGTGTT CGACCTGACC GGCAACCTGT CGCTTTACGG CTCGTACAGC
  1501	AGCCTGTTCG TCCCGCAATC GCAAAAAGAC GAACACGGCA GCTACCTGAA
  1551	ACCCGTAACC GGCAACAATC TGGAAGCCGG CATCAAAGGC GAATGGCTTG
  1601	AAGGCCGTCT GAACGCATCC GCCGCCGTGT ACCGCGCCCG TAAAAACAAC
  1651	CTCGCCACCG CAGCAGGACG CGACCCGAGC GGCAACACCT ACTACCGCGC
  1701	CGCCAACCAA GCCAAAACCC ACGGCTGGGA AATCGAAGTC GGCGGCCGCA
  1751	TCACGCCCGA ATGGCAGATA CAGGCAGGTT ACAGCCAAGG CAAAACCCGC
  1801	GACCAAGACG GCAGCCGCCT GAACCCCGAC AGCGTACCCG AACGCAGCTT
  1851	CAAACTCTTC ACTGCCTACC ACTTTGCCCC CGAAGCCCCC AGCGGCTGGA
  1901	CCATCGGCGC AGGCGTGCGC TGGCAGAGCG AAACCCACAC CGACCCTGCC
  1951	ACGCTCCGCA TCCCCAACCC CGCCGCCAAA GCCCGCGCCG CCGACAACAG
  2001	CCGCCAAAAA GCCTACGCCG TCGCCGACAT CATGGCGCGT TACCGCTTCA
  2051	ATCCGCGCGC CGAACTGTCG CTGAACGTGG ACAATCTGTT CAACAAACAC
  2101	TACCGCACCC AGCCCGACCG CCACAGCTAC GGCGCACTGC GGACAGTGAA
  2151	CGCGGCGTTT ACCTATCGGT TTAAATAA

• This encodes a protein having amino acid sequence <SEQ ID 668>:

• 1	MTRFKYSLLF AALLPVYAQA DVSVSDDPKP QESTELPTIT VTADRTASSN
  51	DGYTVSGTHT PLGLPMTLRE IPQSVSVITS QQMRDQNIKA LDRALLQATG
  101	TSRQIYGSDR AGYNYLFARG SRIANYQING IPVADALADT GNANTAAYER
  151	VEVVRGVAGL LDGTGEPSAT VNLVRKRPTR KPLFEVRAEA GNRKHFGLGA
  201	DVSGSLNAEG TLRGRLVSTF GRGDSWRQRE RSRDAELYGI LEYDIAPQTR
  251	VHAGMDYQQA KETADAPLSY AVYDSQGYAT AFGPKDNPAT NWANSRHRAL
  301	NLFAGIEHRF NQDWKLKAEY DYTRSRFRQP YGVAGVLSID HNTAATDLIP
  351	GYWHADPRTH SASVSLIGKY RLFGREHDLI AGINGYKYAS NKYGERSIIP
  401	NAIPNAYEFS RTGAYPQPAS FAQTIPQYGT RRQIGGYLAT RFRAADNLSL
  451	ILGGRYSRYR TGSYDSRTQG MTYVSANRFT PYTGIVFDLT GNLSLYGSYS
  501	SLFVPQSQKD EHGSYLKPVT GNNLEAGIKG EWLEGRLNAS AAVYRARKNN
  551	LATAAGRDPS GNTYYRAANQ AKTHGWEIEV GGRITPEWQI QAGYSQSKTR
  601	DQDGSRLNPD SVPERSFKLF TAYHFAPEAP SGWTIGAGVR WQSETHTDPA
  651	TLRIPNPAAK ARAADNSRQK AYAVADIMAR YRFNPRAELS LNVDNLFNKH
  701	YRTQPDRHSY GALRTVNAAF TYRFK*

• ORF23a and ORF23-1 show 99.2% identity in 725 aa overlap:
• 

  
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF23 shows 93.4% identity over a 211 as overlap with a predicted ORF (ORF23.ng) from N. gonorrhoeae:
• 
• The ORF23ng nucleotide sequence <SEQ ID 669> is predicted to encode a protein comprising amino acid sequence <SEQ ID 670>:

• 1	SAVDACRIPG YNYLFARGSR IANYQINGIP VADALADTGN ANTAAYERVE
  51	VVRGVAGLPD GTGEPSATVN LVRKHPTRKP LFEVRAEAGN RKHFGLGADV
  101	SGSLNAEGTL RGRLVSTFGR GDSWRQLERS RDAELYGILE YDIAPQTRVH
  151	AGMDYQQAKE TADAPLSYAV YDSQGYATAF GPKDNPATNW SNSRNRALNL
  201	FAGIEHRFNQ DWKLKAEYDY TRSRFRQPYG VAGVLSIDHS TAATDLIPGY
  251	WHADPRTHSA SMSLTGKYRL FGREHDLIAG INGYKYASNK YGERSIIPNA
  301	IPNAYEFSRT GAYPQPSSFA QTIPQYDTRR QIGGYLATRF RAADNLSLIL
  351	GGRYSRYRAG SYNSRTQGMT YVSANRFTPY TGIVFDLTGN LSLYGSYSSL
  401	FVPQLQKDEH GSYLKPVTGN NLEADIKGEW LEGRLNASAA VYRARKNNLA
  451	TAAGRDQSGN TYYRAANQAK THGWEIEVGG RITPEWQIQA GYSQSKPRDQ
  501	DGSRLNPDSV PERSFKLFTA YHLAPEAPSG RTIGAGVRRQ GETHTDPAAL
  551	RIPNPAAKAR AVANSRQKAY AVADIMARYR FNPRTELSLN VDNLFNKHYR
  601	TQPDRHSYGA LRTVNAAFTY RFK*

• Further work revealed the complete nucleotide sequence <SEQ ID 671>:

• 1	ATGACACGCT TCAAATACTC CCTGCTTTTT GCCGCCCTGC
  	TACCCGTGTA
  51	CGCGCAGGCC GATGTTTCTG TTTCAGACGA CCCCAAACCG
  	CAGGAAAGCA
  101	CCGAATTGCC GACCATCACC GTTACCGCCG ACCGCACCGC
  	GAGTTCCAAC
  151	GACGGCTACA CCGTTTCCGG CACGCACACC CCGTTCGGGC
  	TGCCCATGAC
  201	CCTGCGCGAA ATCCCGCAGA GCGTCAGCGT CATCACATCG
  	CAACAAATGC
  251	GCGACCAAAA CATCAAAACG CTCGACCGCG CCCTGTTGCA
  	GGCGACCGGC
  301	ACCAGCCGCC AGATTTACGG CTCCGACCGC GCGGGCTACA
  	ACTACCTGTT
  351	CGCGCGCGGC AGCCGCATCG CCAACTACCA AATCAACGGC
  	ATCCCCGTTG
  401	CCGACGCGCT GGCCGATACG GGCAATGCCA ACACCGCCGC
  	CTATGAGCGC
  451	GTAGAAGTCG TGCGCGGCGT GGCGGGGCTG CCGGACGGCA
  	CGGGCGAGCC
  501	TTCTGCCACC GTCAATCTGG TACGCAAACA CCCGACCCGC
  	AAGCCATTGT
  551	TTGAAGTCCG CGCCGAAGCC GGCAACCGCA AACATTTCGG
  	GCTGGGCGCG
  601	GACGTATCGG GCAGCCTGAA CGCCGAAGGC ACGCTGCGCG
  	GCCGCCTGGT
  651	TTCCACCTTC GGACGCGGCG ACTCGTGGCG GCAGCTCGAA
  	CGCAGCCGCG
  701	ATGCCGAACT CTACGGCATT TTGGAATACG ACATCGCACC
  	GCAAACCCGC
  751	GTCCACGCAG GCATGGACTA CCAGCAGGCG AAAGAAACCG
  	CAGACGCGCC
  801	GCTCAGCTAC GCCGTGTACG ACAGCCAAGG TTATGCCACC
  	GCCTTCGGCC
  851	CAAAAGACAA CCCCGCCACA AATTGGTCGA ACAGCCGCAA
  	CCGTGCGCTC
  901	AACCTGTTCG CCGGCATAGA ACACCGCTTC AACCAAGACT
  	GGAAACTCAA
  951	AGCCGAATAC GACTACACCC GTAGCCGCTT CCGCCAGCCC
  	TACGGTGTGG
  1001	CAGGCGTACT TTCCATCGAC CACAGCACTG CCGCCACCGA
  	CCTGATTCCC
  1051	GGTTATTGGC ACGCcgatcc GCGCACCCAC AGCGCCAGCA
  	TGTCATTGAC
  1101	CGGCAAATAC CgcctGTTCG GCCGCGAGCA CGATTTAATC
  	GCGGGTATCA
  1151	ACGGCTACAA ATACGCCAGC AACAAATACG GCGAACGCAG
  	CATCATTCCC
  1201	AACGCCATTC CCAACGCCTA CGAATTTTCC CGCACGGGCG
  	CCTATCCGCA
  1251	GCCATCATCG TTTGCCCAAA CCATCCCGCA ATACGACACC
  	AGGCGGCAAA
  1301	TCGGCGGCTA TCTCGCCACC CGTTTCCGCG CCGCCGACAA
  	CCTTTCGCTG
  1351	ATACTCGGCG GCAGATACAG CCGCTACCGC GCAGGCAGCT
  	ACAACAGCCG
  1401	CACACAAGGC ATGACCTATG TGTCCGCCAA CCGTTTCACC
  	CCCTACACAG
  1451	GCATCGTGTT CGATCTGACC GGCAACCTGT CGCTTTACGG
  	CTCGTACAGC
  1501	AGCCTGTTCG TCCCGCAATT GCAAAAAGAC GAACACGGCA
  	GCTACCTGAA
  1551	ACCCGTAACC GGCAACAATC TGGAAGCCGA CATCAAAGGC
  	GAATGGCTTG
  1601	AAGGGCGTCT GAACGCATCC GCCGCCGTGT ACCGCGCCCG
  	TAAAAACAAC
  1651	CTCGCCACCG CAGCAGGACG CGACCAGAGC GGCAACACCT
  	ACTATCGCGC
  1701	CGCCAACCAA GCCAAAACCC ACGGCTGGGA AATCGAAGTC
  	GGCGGCCGCA
  1751	TCACGCCCGA ATGGCAGATA CAGGCAGGCT ACAGCCAAAG
  	CAAACCCCGC
  1801	GACCAAGACG GCAGCCGCCT GAACCCCGAC AGCGTAcCCG
  	AACGCAGCTT
  1851	CAAACTCTTC ACCGCCTACC ACTTAGCCCC CGAAGCCCCC
  	AGCGGCCGGA
  1901	CCATcggTGC GGGTGTGCGC CGGCAGGGCG AAACCCACAC
  	CGACCCAGCC
  1951	GCGCTCCGCA TCCCCAACCC CGCCGCCAAA GCCCGCGCCG
  	TCGCCAACAG
  2001	CCGCCAGAAA GCCTACGCCG TCGCCGACAT CATGGCGCGT
  	TACCGCTTCA
  2051	ATCCGCGCAC CGAACTGTCG CTGAACGTGG ACAACCTGTT
  	CAACAAACAC
  2101	TACCGCACCC AGCCCGACCG CCACAGCTAC GGCGCACTGC
  	GGACAGTGAA
  2151	CGCGGCGTTT ACCTATCGGT TTAAATAA

• This corresponds to the amino acid sequence <SEQ ID 672; ORF23ng-1>:

• 1	MTRFKYSLLF AALLPVYAQA DVSVSDDPKP QESTELPTIT
  	VTADRTASSN
  51	DGYTVSGTHT PFGLPMTLRE IPQSVSVITS QQMRDQNIKT
  	LDRALLQATG
  101	TSRQIYGSDR AGYNYLFARG SRIANYQING IPVADALADT
  	GNANTAAYER
  151	VEVVRGVAGL PDGTGEPSAT VNLVRKHPTR KPLFEVRAEA
  	GNRKHFGLGA
  201	DVSGSLNAEG TLRGRLVSTF GRGDSWRQLE RSRDAELYGI
  	LEYDIAPQTR
  251	VHAGMDYQQA KETADAPLSY AVYDSQGYAT AFGPKDNPAT
  	NWSNSRNRAL
  301	NLFAGIEHRF NQDWKLKAEY DYTRSRFRQP YGVAGVLSID
  	HSTAATDLIP
  351	GYWHADPRTH SASMSLTGKY RLFGREHDLI AGINGYKYAS
  	NKYGERSIIP
  401	NAIPNAYEFS RTGAYPQPSS FAQTIPQYDT RRQIGGYLAT
  	RFRAADNLSL
  451	ILGGRYSRYR AGSYNSRTQG MTYVSANRFT PYTGIVFDLT
  	GNLSLYGSYS
  501	SLFVPQLQKD EHGSYLKPVT GNNLEADIKG EWLEGRLNAS
  	AAVYRARKNN
  551	LATAAGRDQS GNTYYRAANQ AKTHGWEIEV GGRITPEWQI
  	QAGYSQSKPR
  601	DQDGSRLNPD SVPERSFKLF TAYHLAPEAP SGRTIGAGVR
  	RQGETHTDPA
  651	ALRIPNPAAK ARAVANSRQK AYAVADIMAR YRFNPRTELS
  	LNVDNLFNKH
  701	YRTQPDRHSY GALRTVNAAF TYRFK*

• ORF23ng-1 and ORF23-1 show 95.9% identity in 725 aa overlap:
• 

  
• In addition, ORF23ng-1 shows significant homology with an OMP from E. coli:

• sp|P16869|FHUE_ECOLI OUTER-MEMBRANE RECEPTOR FOR FE(III)-COPROGEN,
  FE(III)-FERRIOXAMINE B AND FE(III)-RHODOTRULIC ACID PRECURSOR
  >gi|1651542|gnl|PID|d1015403
  (D90745) Outer membrane protein FhuE precursor [Escherichia coli]
  >gi|1651545|gnl|PID|d1015405 (D90746) Outer membrane protein
  FhuE precursor [Escherichia coli] >gi|1787344 (AE000210)
  outer-membrane receptor for Fe(III)-
  coprogen, Fe(III)-ferrioxamine B and Fe(III)-rhodotrulic acid precursor
  [Escherichia coli] Length = 729
  Score = 332 bits (843), Expect = 3e−90
  Identities = 228/717 (31%), Positives = 350/717 (48%),
  Gaps = 60/717 (8%)

  Query:	38	TITVTADRTASSN--DGYTVSGTHTPFGLPMTLREIPQSVSVITSQQMRDQNIKTLDRAL	95
  		T+ V    TA  +  + Y+V+ T     + MT R+IPQSV++++ Q+M DQ ++TL   +
  Sbjct:	43	TVIVEGSATAPDDGENDYSVTSTSAGTKMQMTQRDIPQSVTIVSQQRMEDQQLQTLGEVM	102
  Query:	96	LQATGTSRQIYGSDRAGYNYLFARGSRIANYQINGIP--------VADALADTGNANTAA	147
  		G S+    SDRA Y   ++RG +I NY ++GIP        + DAL+D      A
  Sbjct:	103	ENTLGISKSQADSDRALY---YSRGFQIDNYMVDGIPTYFESRWNLGDALSDM-----AL	154
  Query:	148	YERVEVVRGVAGLPDGTGEPSATVNLVRKHPTRKPLF-EVRAEAGNRKHFGLGADVSGSL	206
  		+ERVEVVRG  GL  GTG PSA +N+VRKH T +    +V AE G+       AD+   L
  Sbjct:	155	FERVEVVRGATGLMTGTGNPSAAINMVRKHATSREFKGDVSAEYGSWNKERYVADLQSPL	214
  Query:	207	NAEGTLRGRLVSTFGRGDSWRQLERSRDAELYGILEYDIAPQTRVHAGMDYQQAKETADA	266
  		+G +R R+V  +   DSW     S      GI++ D+   T + AG +YQ+    +
  Sbjct:	215	TEDGKIRARIVGGYQNNDSWLDRYNSEKTFFSGIVDADLGDLTTLSAGYEYQRIDVNSPT	274
  Query:	267	PLSYAVYDSQGYATAFGPKDNPATNWSNSRNRALNLFAGIEHRFNQDWKLKAEYDYTRSR	326
  		+++ G + ++    + A +W+ +      +F  ++ +F   W+      ++
  Sbjct:	275	WGGLPRWNTDGSSNSYDRARSTAPDWAYNDKEINKVFMTLKQQFADTWQATLNATHSEVE	334
  Query:	327	F--RQPYGVAGVLSIDHSTAA--TDLIPGY-------WHADPRTHSA-SMSLTGKYRLFG	374
  		F  +  Y  A V   D       ++  PG+       W++  R   A  +   G Y LFG
  Sbjct:	335	FDSKMMYVDAYVNKADGMLVGPYSNYGPGFDYVGGTGWNSGKRKVDALDLFADGSYELFG	394
  Query:	375	REHDLIAGINGYKYASNKYGER--SIIPNAIPNAYEFSRTGAYPQPSSFAQTIPQYDTRR	432
  		R+H+L+ G   Y   +N+Y     +I P+ I + Y F+  G +PQ     Q++ Q DT
  Sbjct:	395	RQHNLMFG-GSYSKQNNRYFSSWANIFPDEIGSFYNFN--GNFPQTDWSPQSLAQDDTTH	451
  Query:	433	QIGGYLATRFRAADNLSLILGGRYSRYRAGSYNSRTQGMTY-VSANRFTPYTGIVFDXXX	491
  		Y ATR   AD L LILG RY+ +R  +       +TY +  N  TPY G+VFD
  Sbjct:	452	MKSLYAATRVTLADPLHLILGARYTNWRVDT-------LTYSMEKNHTTPYAGLVFDIND	504
  Query:	492	XXXXXXXXXXXFVPQLQKDEHGSYLKPVTGNNLEADIKGEWLEGRLNASAAVYRARKNNL	551
  		F PQ  +D  G YL P+TGNN E  +K +W+  RL  + A++R  ++N+
  Sbjct:	505	NWSTYASYTSIFQPQNDRDSSGKYLAPITGNNYELGLKSDWMNSRLTTTLAIFRIEQDNV	564
  Query:	552	ATAAGR---DQSGNTYYRAANQAKTHGWEIEVGGRITPEWQIQAGYSQSKPRDQDGSRLN	608
  		A + G      +G T Y+A +   + G E E+ G IT  WQ+  G ++    D +G+ +N
  Sbjct:	565	AQSTGTPIPGSNGETAYKAVDGTVSKGVEFELNGAITDNWQLTFGATRYIAEDNEGNAVN	624
  Query:	609	PDSVPERSFKLFTAYHLAPEAPSGRTIGAGVRRQGETHTDPAALRIPNPAAKARAVANSR	668
  		P ++P  + K+FT+Y L P  P   T+G GV  Q   +TD        P    RA
  Sbjct:	625	P-NLPRTTVKMFTSYRL-PVMPE-LTVGGGVNWQNRVYTDTV-----TPYGTFRA----E	672
  Query:	669	QKAYAVADIMARYRFNPRTELSLNVDNLFNKHYRTQPDRH-SYGALRTVNAAFTYRF	724
  		Q +YA+ D+  RY+      L  NV+NLF+K Y T  +    YG  R  +   TY+F
  Sbjct:	673	QGSYALVDLFTRYQVTKNFSLQGNVNNLFDKTYDTNVEGSIVYGTPRNFSITGTYQF	729

• Based on this analysis, it was predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF23-1 (77.5 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 15A shows the results of affinity purification of the His-fusion protein, and FIG. 15B shows the results of expression of the GST-fusion in E. coli. Purified His-fusion protein was used to immunise mice, whose sera were used for Western blot (FIG. 15C) and for ELISA (positive result). These experiments confirm that ORF23-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 80
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 673>:

• 1	ATGCGCACGG CAGTGGTTTT GCTGTTGATC ATGCCGATGG
  	CGGCTTCGTC
  51	GGCAATGATG CCGGAAATGG TGTGCGCGGG CGTGTCGCCG
  	GGAACGGCAA
  101	TCATATCCAA GCCGACCGAA CAAACGGCGG TCATGGCTTC
  	GAGTTTGTCC
  151	AGCGTCAgcA CGCCTGCTTC GGCGgcGgCa ATCATACCTT
  	CGTCTTCGGA
  201	AACGGGGATA AACGcGCCAC TCAAACCCCC GACCGCGCTG
  	GAAGCCATCA
  251	TGCCGCCTTT TTTCACGGCA TCGTTCAGCA ATGCCAAAGC
  	TGCTGTTGTG
  301	CCGTGCGTAC CGCAGACGCT CAAGCCCATT TnTTCAAGAA
  	TGCGTGCCAC
  351	TnAGTCGCCG ACGGGG..

• This corresponds to the amino acid sequence <SEQ ID 674; ORF24>:

• 1	MRTAVVLLLI MPMAASSAMM PEMVCAGVSP GTAIISKPTE
  	QTAVMASSLS
  51	SVSTPASAAA IIPSSSETGI NAPLKPPTAL EAIMPPFFTA
  	SFSNAKAAVV
  101	PCVPQTLKPI XSRMRATXSP TG..

• Further work revealed the complete nucleotide sequence <SEQ ID 675>:

• 1	ATGCGCACGG CAGTGGTTTT GCTGTTGATC ATGCCGATGG
  	CGGCTTCGTC
  51	GGCAATGATG CCGGAAATGG TGTGCGCGGG CGTGTCGCCG
  	GGAACGGCAA
  101	TCATATCCAA GCCGACCGAA CAAACGGCGG TCATGGCTTC
  	GAGTTTGTCC
  151	AGCGTCAGCA CGCCTGCTTC GGCGGCGGCA ATCATACCTT
  	CGTCTTCGGA
  201	AACGGGGATA AACGCGCCAC TCAAACCCCC GACCGCGCTG
  	GAAGCCATCA
  251	TGCCGCCTTT TTTCACGGCA TCGTTCAGCA ATGCCAAAGC
  	TGCTGTTGTG
  301	CCGTGCGTAC CGCAGACGCT CAAGCCCATT TCTTCAAGAA
  	TGCGTGCCAC
  351	TGAGTCGCCG ACGGCGGGGG TCGGCGCCAG CGACAAGTCG
  	AGAATACCAA
  401	ACGGGATATT CAGCATTTTT GAGGCTTCGC GGCCGATGAG
  	TTCGCCCACG
  451	CGGGTAATTT TGAAAGCAGT TTTCTTCACT ACTTCCGCAA
  	CTTCGGTCAA
  501	TGTCGTTGCA TCTGAATTTT CCAACGCGGC TTTTACGACA
  	CCTGGGCCGG
  551	ATACGCCGAC ATTGATAACG GCATCCGCTT CGCCCGAACC
  	ATGAAACGCG
  601	CCCGCCATAA ACGGGTTGTC TTCCACCGCG TTGCAGAACA
  	CGACAATTTT
  651	AGCGCAGCCG AAACCTTCGG GCGTGATTTC CGCCGTGCGT
  	TTGACGGTTT
  701	CGCCCGCCAG CTTGACCGCA TCCATATTGA TACCGGCACG
  	CGTACTGCCG
  751	ATATTGATGG AGCTGCACAC AATATCGGTA GTCTTCATCG
  	CTTCGGGAAT
  801	GGAGCGGATT AACACCTCAT CCGAAGGCGA CATCCCTTTT
  	TGCACCAACG
  851	CGGAAAAACC GCCGATAAAA GACACACCGA TGGCTTTGGC
  	AGCTTTATCC
  901	AAAGTTTGCG CCACGCTGAC GTAA

• This corresponds to the amino acid sequence <SEQ ID 676; ORF24-1>:

• 1	MRTAVVLLLI MPMAASSAMM PEMVCAGVSP GTAIISKPTE
  	QTAVMASSLS
  51	SVSTPASAAA IIPSSSETGI NAPLKPPTAL EAIMPPFFTA
  	SFSNAKAAVV
  101	PCVPQTLKPI SSRMRATESP TAGVGASDKS RIPNGIFSIF
  	EASRPMSSPT
  151	RVILKAVFFT TSATSVNVVA SEFSNAAFTT PGPDTPTLIT
  	ASASPEP*NA
  201	PAINGLSSTA LQNTTILAQP KPSGVISAVR LTVSPASLTA
  	SILIPARVLP
  251	ILMELHTISV VFIASGMERI NTSSEGDIPF CTNAEKPPIK
  	DTPMALAALS
  301	KVCATLT*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF24 shows 96.4% identity over a 307 aa overlap with an ORF (ORF24a) from strain A of N. meningitidis:
• 
• The complete length ORF24a nucleotide sequence <SEQ ID 677> is:

• 1	ATGCGCACGG CAGTGGTTTT GCTGTTGATC ATGCCGATGG
  	CGGCTTCGTC
  51	GGCAATGATG CCGGAAATGG TGTGCGCGGG TGTGTCGCCG
  	GGAACGGCAA
  101	TCATATCCAA NCCGACCGAA CAAACGGCGG TCATCGCTTC
  	GAGTTTATCC
  151	AACGTCAGCA CGCCTGCTTC GGCGGCGGCA ATCATACCTT
  	CGTCTTCGGA
  201	NACGGGGATA AACGCGCCAC TCAAACCGCC AACCGCGCTC
  	GAAGCCATCA
  251	TGCCGCCCTT TTTCACGGCA TCGTTCAGCA ATGCCAAAGC
  	TGCTGTTGTG
  301	CCGTGCGTAC CGCAGACGCT CAAACCCATT TCTTCAAGAA
  	TGCGCGCCAC
  351	CGAGTCGCCG ACGGCAGGGG TCGGTGCCAG CGACAAGTCG
  	AGAATACCAA
  401	ACGGGATATT CAGCATTTTT GAGGCTTCGC GGCCGATGAG
  	TTCGCCCACG
  451	CGGGTAATTT TGAAGGCGGT TTTCTTCACA ACTTCGGCAA
  	CTTCGGTCAA
  501	TGTCGTTGCA TCCGAATTTT CCAACGCGGC TTTTACGACA
  	CCCGGGCCGG
  551	ATACGCCGAC ATTAATCACA GCATCCGCTT CGCCTGAGCC
  	GTGAAACGCG
  601	CCCGCCATAN ACGGGTTGTC TTCCNCCGCG TTGCAGAACA
  	CGACGATTTT
  651	GGCGCAGCCG AAACCTTCTA GTGTGATTTC ANCCGTGCGT
  	TTGATGGTTT
  701	CGCCCGCCAG TCTGACCGCG TCCATATTGA TACCGGCGCG
  	CGTACTGCCG
  751	ATATTGATGG AGCTGCACAC GATATCAGTA GTCTTCATCG
  	CTTCGGGAAT
  801	GGAACGGATN AACACCTCGT CAGAAGGCGA CATACCTTTT
  	TGCACCAGCG
  851	CGGAAAAGCC GCCAATAAAA GACACGCCGA TGGCTTTGGC
  	AGCCTTATCC
  901	AAAGTTTGCG CCACGCTGAC GTAA

• This encodes a protein having amino acid sequence <SEQ ID 678>:

• 1	MRTAVVLLLI MPMAASSAMM PEMVCAGVSP GTAIISXPTE
  	QTAVIASSLS
  51	NVSTPASAAA IIPSSSXTGI NAPLKPPTAL EAIMPPFFTA
  	SFSNAKAAVV
  101	PCVPQTLKPI SSRMRATESP TAGVGASDKS RIPNGIFSIF
  	EASRPMSSPT
  151	RVILKAVFFT TSATSVNVVA SEFSNAAFTT PGPDTPTLIT
  	ASASPEP*NA
  201	PAIXGLSSXA LQNTTILAQP KPSSVISXVR LMVSPASLTA
  	SILIPARVLP
  251	ILMELHTISV VFIASGMERX NTSSEGDIPF CTSAEKPPIK
  	DTPMALAALS
  301	KVCATLT*

• It should be noted that this protein includes a stop codon at position 198.
• ORF24a and ORF24-1 show 96.4% identity in 307 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF24 shows 96.7% identity over a 121 aa overlap with a predicted ORF (ORF24ng) from N. gonorrhoeae:
• 
• The complete length ORF24ng nucleotide sequence <SEQ ID 679> is:

• 1	ATGCGCACGG CGGTGGTTTT GCTGTTGATC ATGCCGATGG
  	CGGCTTCGTC
  51	GGCGATGATG CCGGAAATGG TGTGCGCGGG CGTGTCGCCG
  	GGAACGGCAA
  101	TCATGTCCAA ACCAACGGAG CAGACGGCGG TCATGGCTTC
  	GAGTTTGTCC
  151	AGCGTCAACA CGCCTGCCTC GGCGGCGGCA ATCATACCTT
  	CGTCTTCGGA
  201	AACGGGGATA AACGCGCCGC TCAAACCGCC GACCGCGCTG
  	GAAGCCATCA
  251	TGCCGCCCTT TTTCACGGCA TCGTTCAGCA ATGCCAAAGC
  	TGCTGTTGTG
  301	CCGTGCGTAC CGCAGACGCT CAAGCCCATT TCTTCAAGAA
  	TGCGCGCCAC
  351	CGAGTCGCCG ACGGCGGGGG TCGGTGCCAG CGACAAATCG
  	AGAATGCCGA
  401	ACGGGATATT CAGCATTTTT GAGGCTTCGC GACCGATGAG
  	TTCGCCCACG
  451	CGGGTGATTT TGAAAGCGGT TTTCTTCACG ACTTCGGCGA
  	CCTCGGTCAG
  501	GCTGACCGCG TCCGAATTTT CCAGCGCGGC TTTGACCACG
  	CCTGGACCGG
  551	ATACGCCGAC ATTAATCACA GCATCCGCTT CGCCCGAGCC
  	GTGGAACGCA
  601	CCCGCCATAA ACGGATTGTC TTCCACCGCG TTGCAGAACA
  	CGACGATTTT
  651	GGCGCAGCCG AAACCTTCGG GTGTGATTTC AGCCGTGCGT
  	TTGATGGTTT
  701	CGCCTGCCAG CTTGACCGCA TCCATATTGA TACCGGCACG
  	CGTGCTGCCG
  751	ATATTGATGG AGCTGCACAC GATATCGGTA GTTTTCATCG
  	CTTCGGGAAC
  801	GGAACGGATC AACACCTCAT CCGAAGGCGA CATACCTTTT
  	TGCACCAGCG
  851	CGGAAAAGCC GCCGATAAAG GACACGCCGA TGGCTTTGGC
  	TGCCTTGTCC
  901	AAAGTCTGCG CCACGCTGAC ATAA

• This encodes a protein having amino acid sequence <SEQ ID 680>:

• 1	MRTAVVLLLI MPMAASSAMM PEMVCAGVSP GTAIMSKPTE
  	QTAVMASSLS
  51	SVNTPASAAA IIPSSSETGI NAPLKPPTAL EAIMPPFFTA
  	SFSNAKAAVV
  101	PCVPQTLKPI SSRMRATESP TAGVGASDKS RMPNGIFSIF
  	EASRPMSSPT
  151	RVILKAVFFT TSATSVRLTA SEFSSAALTT PGPDTPTLIT
  	ASASPEPWNA
  201	PAINGLSSTA LQNTTILAQP KPSGVISAVR LMVSPASLTA
  	SILIPARVLP
  251	ILMELHTISV VFIASGTERI NTSSEGDIPF CTSAEKPPIK
  	DTPMALAALS
  301	KVCATLT*

• ORF24ng and ORF24-1 show 96.1% identity in 307 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence (first 18 aa—double-underlined) and putative transmembrane domains (single-underlined) in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 81
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 681>:

• 1	..ACCGACGTGC AAAAAGAGTT GGTCGGCGAA CAACGCAAGT
  	GGGCGCAGGA
  51	AAAAATCAGC AACTGCCGAC AAGCCGCCGC GCAGGCAGAC
  	CGGCAGGAAT
  101	ACGCCGAATA CCTCAAGCTG CAATGCGACA CGCGGATGAC
  	GCGCGAACGG
  151	ATACAGTATC TTCGCGGCTA TTCCATCGAT TAG

• This corresponds to the amino acid sequence <SEQ ID 682; ORF25>:
• • 1 . . . TDVQKELVGE QRKWAQEKIS NCRQAAAQAD RQEYAEYLKL QCDTRMTRER
  • 51 IQYLRGYSID *
• Further work revealed the complete nucleotide sequence <SEQ ID 683>:

• 1	ATGTATCGGA AACTCATTGC GCTGCCGTTT GCCCTGCTGC
  	TTGCCGCTTG
  51	CGGCAGGGAA GAACCGCCCA AGGCATTGGA ATGCGCCAAC
  	CCCGCCGTGT
  101	TGCAAGGCAT ACGCGGCAAT ATTCAGGAAA CGCTCACGCA
  	GGAAGCGCGT
  151	TCTTTCGCGC GCGAAGACGG CAGGCAGTTT GTCGATGCCG
  	ACAAAATTAT
  201	CGCCGCCGCC TACGGTTTGG CGTTTTCTTT GGAACACGCT
  	TCGGAAACGC
  251	AGGAAGGCGG GCGCACGTTC TGTATCGCCG ATTTGAACAT
  	TACCGTGCCG
  301	TCTGAAACGC TTGCCGATGC CAAGGCAAAC AGCCCCCTGT
  	TGTACGGGGA
  351	AACTGCTTTG TCGGATATTG TGCGGCAGAA GACGGGCGGC
  	AATGTCGAGT
  401	TTAAAGACGG CGTATTGACG GCAGCCGTCC GCTTCCTGCC
  	CGTCAAAGAC
  451	GGTCAGACGG CATTTGTCGA CAACACGGTC GGTATGGCGG
  	CGCAAACGCT
  501	GTCTGCCGCG CTGCTGCCTT ACGGCGTGAA GAGCATCGTG
  	ATGATAGACG
  551	GCAAGGCGGT GAAAAAAGAA GACGCGGTCA GGATTTTGAG
  	CGGAAAAGCC
  601	CGTGAAGAAG AACCGTCCAA ACCCACGCCC GAAGACATTT
  	TGGAACACAA
  651	TGCCGCCGGC GGCGATGCGG GCGTACCCCA AGCCGCAGAA
  	GGCGCGCCCG
  701	AACCGGAAAT CCTGCATCCT GACGACGGCG AGCGTGCCGA
  	TACCGTTACC
  751	GTATCACGGG GCGAAGTGGA AGAGGCGCGC GTACAAAACC
  	AGCGTGCGGA
  801	ATCCGAAATT ACCAAACTTT GGGGAGGACT CGATACCGAC
  	GTGCAAAAAG
  851	AGTTGGTCGG CGAACAACGC AAGTGGGCGC AGGAAAAAAT
  	CAGCAACTGC
  901	CGACAAGCCG CCGCGCAGGC AGACCGGCAG GAATACGCCG
  	AATACCTCAA
  951	GCTGCAATGC GACACGCGGA TGACGCGCGA ACGGATACAG
  	TATCTTCGCG
  1001	GCTATTCCAT CGATTAG

• This corresponds to the amino acid sequence <SEQ ID 684; ORF25-1>:

• 1	MYRKLIALPF ALLLAACGRE EPPKALECAN PAVLQGIRGN
  	IQETLTQEAR
  51	SFAREDGRQF VDADKIIAAA YGLAFSLEHA SETQEGGRTF
  	CIADLNITVP
  101	SETLADAKAN SPLLYGETAL SDIVRQKTGG NVEFKDGVLT
  	AAVRFLPVKD
  151	GQTAFVDNTV GMAAQTLSAA LLPYGVKSIV MIDGKAVKKE
  	DAVRILSGKA
  201	REEEPSKPTP EDILEHNAAG GDAGVPQAAE GAPEPEILHP
  	DDGERADTVT
  251	VSRGEVEEAR VQNQRAESEI TKLWGGLDTD VQKELVGEQR
  	KWAQEKISNC
  301	RQAAAQADRQ EYAEYLKLQC DTRMTRERIQ YLRGYSID*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF25 shows 98.3% identity over a 60aa overlap with an ORF (ORF25a) from strain A of N. meningitidis:
• 
• The complete length ORF25a nucleotide sequence <SEQ ID 685> is:

• 1	ATGTATCGGA AACTCATTGC GCTGCCGTTT GCCCTGCTGC
  	TTGCCGCTTG
  51	CGGCAGGGAA GAACCGCCCA AGGCATTGGA ATGCGCCAAC
  	CCCGCCGTGT
  101	TGCAANGCAT ACGCNGCAAT ATTCAGGAAA CGCTCACGCA
  	GGAAGCGCGT
  151	TCTTTCGCGC GCGAAGACNG CANGCAGTTT GTCGATGCCG
  	ACNAAATTAT
  201	CGCCGCCGCC TANGNTNNGN NGNTNTCTTT GGAACACGCT
  	TCGGAAACGC
  251	AGGAAGGCGG GCGCACGTTC TGTNTCGCCG ATTTGAACAT
  	TACCGTGCCG
  301	TCTGAAACGC TTGCCGATGC CAAGGCAAAC AGCCCCCTGC
  	TGTACGGGGA
  351	AACCGCTTTG TCGGATATTG TGCGGCAGAA GACGGGCGGC
  	AATGTCGAGT
  401	TTAAAGACGG CGTATTGACG GCAGCCGTCC GCTTCCTACC
  	CGTCAAAGAC
  451	GGTCAGANGG CATTTGTCGA CAACACGGTC GGTATGGCGG
  	CGCAAACGCT
  501	GTCTGCCGCG TTGCTGCCTT ACGGCGTGAA GAGCATCGTG
  	ATGATAGACG
  551	GCAAGGCGGT AAAAAAAGAA GACGCGGTCA GGATTNTGAG
  	CNGANAAGCC
  601	CGTGAANAAG AACCGTCCAA ANCCNNGCCC GAAGACATTT
  	TGGAACATAA
  651	TGCCGCCGGA GGGGATGCAG ACGTACCCCA AGCCGGAGAA
  	GACGCGCCCG
  701	AACCGGAAAT CCTGCATCCT GACGACGGCG AGCGTGCCGA
  	TACCGTTACC
  751	GTATCACGGG GCGAAGTGGA AGAGGCGCGN GTACAAAACC
  	AGCGTGCGGA
  801	ATCCGAAATT ACCAAACTTT GGGGAGGACT CGATACCGAC
  	GTGCAAAAAG
  851	AGTTGGTCGG CGAANAACGC AAGTGGGCGC AGGAAAAAAT
  	CAGCAACTGC
  901	CGACAAGCCG CCGCGCAGGC AGACCGGCAG GAATACGCCG
  	AATACCTCAA
  951	GCTGCAATGC GACACGCGGA TGACGCGCGA ACGGATACAG
  	TATCTTCGCG
  1001	GCTATTCCAT CGATTAG

• This encodes a protein having amino acid sequence <SEQ ID 686>:

• 1	MYRKLIALPF ALLLAACGRE EPPKALECAN PAVLQXIRXN
  	IQETLTQEAR
  51	SFAREDXXQF VDADXIIAAA XXXXXSLEHA SETQEGGRTF
  	CXADLNITVP
  101	SETLADAKAN SPLLYGETAL SDIVRQKTGG NVEFKDGVLT
  	AAVRFLPVKD
  151	GQXAFVDNTV GMAAQTLSAA LLPYGVKSIV MIDGKAVKKE
  	DAVRIXSXXA
  201	REXEPSKXXP EDILEHNAAG GDADVPQAGE DAPEPEILHP
  	DDGERADTVT
  251	VSRGEVEEAR VQNQRAESEI TKLWGGLDTD VQKELVGEXR
  	KWAQEKISNC
  301	RQAAAQADRQ EYAEYLKLQC DTRMTRERIQ YLRGYSID*

• ORF25a and ORF25-1 show 93.5% identity in 338 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF25 shows 100% identity over a 60aa overlap with a predicted ORF (ORF25ng) from N. gonorrhoeae:
• 
• The complete length ORF25ng nucleotide sequence <SEQ ID 687> is:

• 1	ATGTATCGGA AACTCATTGC GCTGCCGTTT GCCCTGCTGC
  	TTGCAGCGTG
  51	CGGCAGGGAA GAACCGCCCA AGGCGTTGGA ATGCGCCAAC
  	CCCGCCGTGT
  101	TGCAGGACAT ACGCGGCAGT ATTCAGGAAA CGCTCACGCA
  	GGAAGCGCGT
  151	TCTTTCGCGC GCGAAGACGG CAGGCAGTTT GTCGATGCCG
  	ACAAAATTAT
  201	CGCCGCCGCC TACGGTTTGG CGTTTTCTTT GGAACACGCT
  	TCGGAAACGC
  251	AGGAAGGCGG GCGCACGTTC TGTATCGCCG ATTTGAACAT
  	TACCGTGCCG
  301	TCTGAAACGC TTGCCGATGC CGAGGCAAAC AGCCCCCTGC
  	TGTATGGGGA
  351	AACGTCTTTG GCAGACATCG TGCAGCAGAA GACGGGCGGC
  	AATGTCGAGT
  401	TTAAAGACGG CGTATTGACG GCAGCCGTCC GCTTCCTGCC
  	CGCCAAAGAC
  451	GCTCGGACGG CATTTATCGA CAACACGGTC GGTATGGCGA
  	CGCAAACGCT
  501	GTCTGCCGCG TTGCTGCCTT ACGGCGTGAA GAGCATCGTG
  	ATGATAGACG
  551	GCAAGGCGGT GACAAAAGAA GACGCGGTCA GGGTTTTGAG
  	CGGCAAAGCC
  601	CGTGAAGAAG AACCGTCCAA ACCCACCCCC GAAGACATTT
  	TGGAACACAA
  651	TGCCGCCGGC GGCGATGCGG GCGTACCCCA AGCCGCAGAA
  	GGCGCACCCG
  701	AACCCGAAAT CCTGCATCCC GACGACGTCG AGCGTGCCGA
  	TACCGTTACC
  751	GTATCACGGG GCGAAGTGGA AGAGGCGCGC GTACAAAACC
  	AACGTGCGGA
  801	ATCCGAAATT ACCAAACTTT GGGGAGGACT CGATACCGAC
  	GTGCAAAAAG
  851	AGTTGGTCGG CGAACAGCGC AAGTGGGCGC AGGAAAAAAT
  	CAGcaactgc
  901	cgACAAGCCG CCGCGCAGGC AGACCGGCAG GAATACGCCG
  	AATACCTCAA
  951	GCTCCAATGC GACACGCGGA TGACGCGCGA ACggaTACAG
  	TATCTTCGCG
  1001	GCTATTCCAT CGATTAG

• This encodes a protein having amino acid sequence <SEQ ID 688>:

• 1	MYRKLIALPF ALLLAACGRE EPPKALECAN PAVLQDIRGS
  	IQETLTQEAR
  51	SFAREDGRQF VDADKIIAAA YGLAFSLEHA SETQEGGRTF
  	CIADLNITVP
  101	SETLADAEAN SPLLYGETSL ADIVQQKTGG NVEFKDGVLT
  	AAVRFLPAKD
  151	ARTAFIDNTV GMATQTLSAA LLPYGVKSIV MIDGKAVTKE
  	DAVRVLSGKA
  201	REEEPSKPTP EDILEHNAAG GDAGVPQAAE GAPEPEILHP
  	DDVERADTVT
  251	VSRGEVEEAR VQNQRAESEI TKLWGGLDTD VQKELVGEQR
  	KWAQEKISNC
  301	RQAAAQADRQ EYAEYLKLQC DTRMTRERIQ YLRGYSID*

• ORF25ng and ORF25-1 show 95.9% identity in 338 aa overlap:
• 
• Based on this analysis, including the presence of a predicted prokaryotic membrane lipoprotein lipid attchment site (underlined) in the gonococcal protein, it was predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF25-1 (37 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 16A shows the results of affinity purification of the GST-fusion protein, and FIG. 16B shows the results of expression of the His-fusion in E. coli. Purified His-fusion protein was used to immunise mice, whose sera were used for Western blot (FIG. 16C), ELISA (positive result), and FACS analysis (FIG. 16D). These experiments confirm that ORF25-1 is a surface-exposed protein, and that it is a useful immunogen.
• FIG. 16E shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF25-1.
Example 82
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 689>

• 1	ATGCAGCTGA TCGACTATTC ACATTCATTT TTCTCGGTTG
  	TGCCACCCTT
  51	TTTGGCACTG GCACTTGCCG TCATTACCCG CCGCGTACTG
  	CTGTCTTTAG
  101	GCATCGGTAT TCTGGwysGC GTTGCCTTTT TGGTCGGCGG
  	CAACCCCGTC
  151	GACGGTCTGA CACACCTGAA AGACATGGTC GTCGGCTTGG
  	CTTGGTCAGA
  201	CGsyGATTGG TCGCTGGGCA AACCAAAAAT CTTGGTTTTC
  	CkGATACTTT
  251	TGGGTATTTT TACTTCCCTG CTGACCTACT CCGGCAGCAA
  	T.........
  	//
  851	.......... .......... .......... ........AC
  	TTCGCTGGTA
  901	TTCGGCGGCA CTTGCGGCGT CTTTGCCGTC GTTCTCTGCA
  	CGCTCGGCAC
  951	GATTAAAACC GCCGACTATC CCAAAGCCGT TTGGCAGGGT
  	GCGAAATCTA
  1001	TGTTCGGCGC AATCGCCATT TTAATCCTCG CTTGGCTCAT
  	CAGTACGGTT
  1051	GTCGGCGAAA TGCACACCGG CGATTACCTC TCCACACTGG
  	TTGCGGGCAA
  1101	CATCCATCCC GGCTTCCTGC CCGTCATCCT CTTCCTGCTC
  	GCCAGCGTGA
  1151	TGGCGTTTGC CACAGGCACA AGCTGGGGGA CGTTCGGCAT
  	TATGCTGCCG
  1201	ATTGCCGCCG CCATGGCGGT CAAAGTCGAA CCCGCGCTGA
  	TTATCCCGTG
  1251	TATGTCCGCA GTAATGGCGG GGGCGGTATG CGGCGACCAC
  	TGCTCGCCCA
  1301	TTTCCGACAC GACCATCCTG TCGTCCACCG GCGCGCGCTG
  	CAACCACATC
  1351	GACCACGTTA CCTCGCAACT GCCTTACGCC TTAACCGTTG
  	CCGCCGCCGC
  1401	CGCATCGGGC TACCTCGCAT TGGGTCTGAC AAAATCCGCG
  	CTGTTGGGCT
  1451	TTGGCACGAC AGGCATTGTA TTGGCGGTGC TGATTTTTCT
  	GTTGAAAGAT
  1501	AAAAAA..

• This corresponds to the amino acid sequence <SEQ ID 690; ORF26>:

• 1	MQLIDYSHSF FSVVPPFLAL ALAVITRRVL LSLGIGILXX
  	VAFLVGGNPV
  51	DGLTHLKDMV VGLAWSDXDW SLGKPKILVF XILLGIFTSL
  	LTYSGSN...
  	//
  251	.......... .......... .......... ..........
  	......TSLV
  301	FGGTCGVFAV VLCTLGTIKT ADYPKAVWQG AKSMFGAIAI
  	LILAWLISTV
  351	VGEMHTGDYL STLVAGNIHP GFLPVILFLL ASVMAFATGT
  	SWGTFGIMLP
  401	IAAAMAVKVE PALIIPCMSA VMAGAVCGDH CSPISDTTIL
  	SSTGARCNHI
  451	DHVTSQLPYA LTVAAAAASG YLALGLTKSA LLGFGTTGIV
  	LAVLIFLLKD
  501	KK..

• Further work revealed the complete nucleotide sequence <SEQ ID 691>:

• 1	ATGCAGCTGA TCGACTATTC ACATTCATTT TTCTCGGTTG
  	TGCCACCCTT
  51	TTTGGCACTG GCACTTGCCG TCATTACCCG CCGCGTACTG
  	CTGTCTTTAG
  101	GCATCGGTAT TCTGGTCGGC GTTGCCTTTT TGGTCGGCGG
  	CAACCCCGTC
  151	GACGGTCTGA CACACCTGAA AGACATGGTC GTCGGCTTGG
  	CTTGGTCAGA
  201	CGGCGATTGG TCGCTGGGCA AACCAAAAAT CTTGGTTTTC
  	CTGATACTTT
  251	TGGGTATTTT TACTTCCCTG CTGACCTACT CCGGCAGCAA
  	TCAGGCGTTT
  301	GCCGACTGGG CAAAACGGCA CATTAAAAAC CGGCGCGGCG
  	CGAAAATGCT
  351	GACCGCCTGC CTCGTGTTCG TAACCTTTAT CGACGACTAT
  	TTCCACAGTC
  401	TCGCCGTCGG TGCGATTGCC CGCCCCGTTA CCGACAAGTT
  	TAAAGTTTCC
  451	CGCACCAAAC TCGCCTACAT CCTCGACTCC ACTGCCGCTC
  	CTATGTGCGT
  501	GCTGATGCCC GTTTCAAGCT GGGGCGCGTC GATTATCGCC
  	ACGCTTGCCG
  551	GACTGCTCGT TACCTACAAA ATCACCGAAT ACACGCCGAT
  	GGGGACGTTT
  601	GTCGCCATGA GCCTGATGAA CTATTACGCA CTGTTTGCCC
  	TGATTATGGT
  651	GTTCGTCGTC GCATGGTTTT CCTTCGACAT CGGCTCGATG
  	GCACGTTTCG
  701	AACAAGCCGC GTTGAACGAA GCCCACGATG AAACTGCCGT
  	TTCAGACGCT
  751	ACCAAAGGTC GTGTTTACGC ACTGATTATT CCCGTTTTGG
  	CCTTAATCGC
  801	CTCAACGGTT TCCGCCATGA TCTACACCGG CGCGCAGGCA
  	AGCGAAACCT
  851	TCAGCATTTT GGGGGCATTT GAAAACACGG ACGTAAACAC
  	TTCGCTGGTA
  901	TTCGGCGGCA CTTGCGGCGT CCTTGCCGTC GTTCTCTGCA
  	CGCTCGGCAC
  951	GATTAAAACC GCCGACTATC CCAAAGCCGT TTGGCAGGGT
  	GCGAAATCTA
  1001	TGTTCGGCGC AATCGCCATT TTAATCCTCG CTTGGCTCAT
  	CAGTACGGTT
  1051	GTCGGCGAAA TGCACACCGG CGATTACCTC TCCACACTGG
  	TTGCGGGCAA
  1101	CATCCATCCC GGCTTCCTGC CCGTCATCCT CTTCCTGCTC
  	GCCAGCGTGA
  1151	TGGCGTTTGC CACAGGCACA AGCTGGGGGA CGTTCGGCAT
  	TATGCTGCCG
  1201	ATTGCCGCCG CCATGGCGGT CAAAGTCGAA CCCGCGCTGA
  	TTATCCCGTG
  1251	TATGTCCGCA GTAATGGCGG GGGCGGTATG CGGCGACCAC
  	TGCTCGCCCA
  1301	TTTCCGACAC GACCATCCTG TCGTCCACCG GCGCGCGCTG
  	CAACCACATC
  1351	GACCACGTTA CCTCGCAACT GCCTTACGCC TTAACCGTTG
  	CCGCCGCCGC
  1401	CGCATCGGGC TACCTCGCAT TGGGTCTGAC AAAATCCGCG
  	CTGTTGGGCT
  1451	TTGGCACGAC AGGCATTGTA TTGGCGGTGC TGATTTTTCT
  	GTTGAAAGAT
  1501	AAAAAACGCG CCAACGCCTG A

• This corresponds to the amino acid sequence <SEQ ID 692; ORF26-1>:

• 1	MQLIDYSHSF FSVVPPFLAL ALAVITRRVL LSLGIGILVG
  	VAFLVGGNPV
  51	DGLTHLKDMV VGLAWSDGDW SLGKPKILVF LILLGIFTSL
  	LTYSGSNQAF
  101	ADWAKRHIKN RRGAKMLTAC LVFVTFIDDY FHSLAVGAIA
  	RPVTDKFKVS
  151	RTKLAYILDS TAAPMCVLMP VSSWGASIIA TLAGLLVTYK
  	ITEYTPMGTF
  201	VAMSLMNYYA LFALIMVFVV AWFSFDI GSM ARFEQAALNE
  	AHDETAVSDA
  251	TKGRVYALII PVLALIASTV SAMIYTGAQA SETFSILGAF
  	ENTDVNTSLV
  301	FGGTCGVLAV VLCTLGTIKT ADYPKAVWQG AKSMFGAIAI
  	LILAWLISTV
  351	VGEMHTGDYL STLVAGNIHP GFLPVILFLL ASVMAFATGT
  	SWGTFGIMLP
  401	IAAAMAVKVE PALIIPCMSA VMAGAVCGDH CSPISDTTIL
  	SSTGARCNHI
  451	DHVTSQLPYA LTVAAAAASG YLALGLTKSA LLGFGTTGIV
  	LAVLIFLLKD
  501	KKRANA*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with the Hypothetical Transmembrane Protein HI1586 of H. influenzae (Accession Number P44263)
• ORF26 and HI1586 show 53% and 49% amino acid identity in 97 and 221 aa overlap at the N-terminus and C-terminus, respectively:

• Orf26	1	MQLIDYSHSFFSVVPPFLALALAVITRRVXXXXXXXXXXXVAFLVGGNPVDGLTHLKDMV	60
  		M+LID+S S +S+VP  LA+ LA+ TRRV              L          +L   V
  HI1586	14	MELIDFSSSVWSIVPALLAIILAIATRRVLVSLSAGIIIGSLMLSDWQIGSAFNYLVKNV	73
  Orf26	61	VGLAWSDXDWSLGKPKILVFXILLGIFTSLLTYSGSN	97
  		V L ++D + +     I++F +LLG+ T+LLT SGSN
  HI1586	74	VSLVYADGEIN-SNMNIVLFLLLLGVLTALLTVSGSN	109
  		//
  Orf26	86	IFTSLLTYSGS--NTSLVFGGTCGVFAVVLCTL--GTIKTADYPKAVWQGAKSMFGXXXX	141
  		+F+ L T+  +   TSLV GG C +    L  +    +   +Y ++   G KSM G
  HI1586	299	VFSVLGTFENTVVGTSLVVGGFCSIIISTLLIILDRQVSVPEYVRSWIVGIKSMSGAIAI	358
  Orf26	142	XXXXXXXSTVVGEMHTGDYLSTLVAGNIHPGFLPVILFLLASVMAFATGTSWGTFGIMLP	201
  		+ +VG+M TG YLS+LV+GNI   FLPVILF+L + MAF+TGTSWGTFGIMLP
  HI1586	359	LFFAWTINKIVGDMQTGKYLSSLVSGNIPMQFLPVILFVLGAAMAFSTGTSWGTFGIMLP	418
  Orf26	202	IAAAMAVKVEPALIIPCMSAVMAGAVCGDHCSPISDTTILSSTGARCNHIDHVTSQXXXX	261
  		IAAAMA    P L++PC+SAVMAGAVCGDHCSP+SDTTILSSTGA+CNHIDHVT+Q
  HI1586	419	IAAAMAANAAPELLLPCLSAVMAGAVCGDHCSPVSDTTILSSTGAKCNHIDHVTTQLPYA	478
  Orf26	262	XXXXXXXXXXXXXXXXXKSALLGFGTTGIVLAVLIFLLKDK	302
  		S L GF  T + L V+IF +K +
  HI1586	479	ATVATATSIGYIVVGFTYSGLAGFAATAVSLIVIIFAVKKR	519

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF26 shows 58.2% identity over a 502aa overlap with an ORF (ORF26a) from strain A of N. meningitidis:
• 
• The complete length ORF26a nucleotide sequence <SEQ ID 693> is:

• 1	ATGCAGCTGA TCGACTATTC ACATTCATTT TTCTCGGTTG
  	TGCCACCCTT
  51	TTTGGCACTG GCACTTGCCG TCATTACCCG CCGCGTACTG
  	CTGTCTTTAG
  101	GCATCGGTAT TCTGGTCGGC GTTGCCTTTT TGGTCGGCGG
  	CAACCCCGTC
  151	GACGGTCTGA CACACCTGAA AGACATGGTC GTCGGCTTGG
  	CTTGGTCAGA
  201	CGGCGATTGG TCGCTGGGCA AACCAAAANT CTTGGTTTTC
  	CTGATACTTT
  251	TGGGTATTTT TACTTCCCTG CTGACCTACT CCGGCAGCAA
  	TCAGGCGTTT
  301	GCCGACTGGG CAAAACGGCA CATTAAAAAC CGGCGCGGCG
  	CGAAAATGCT
  351	GACCGCCTGC CTCGTGTTCG TAACCTTTAT CGACGACTAT
  	TTCCACAGTC
  401	TCGCCGTCGG TGCGNTTGCC CGCCCCGTTA CCGACAAGTT
  	TAAAGTTTCC
  451	CGCGCCAAAC TCGCCTACAT CCTCGACTCC ACTGCCGCGC
  	CTATGTGCGT
  501	GCTGATGCCC GTTTCAAGCT GGGGCGCGTC GATTATCGCC
  	ACGCTTGCCG
  551	GACTGCTCGT TACCTACAAA ATCACCGAAT ACACGCCGAT
  	GGGGACGTTT
  601	GTCGCCATGA GCCTGATGAA CTATTACGCA CTGTTTGCCC
  	TGATTATGGT
  651	GTTCGTCGTC GCATGGTTCT CCTTCGACAT CGGCTCGATG
  	GCACGTTTCG
  701	AACAAGCCGC GTTGAACGAA GCCCACGATG AAACTGCCGT
  	TTCAGACGGC
  751	AGCTGGGGCA GGGTTTACGC ATTGATTATT CCCGTTTTGG
  	CCTTAATCGC
  801	CTCAACGGTT TCCGCCATGA TCTACACCGG TGCACAGGCA
  	AGCGAAACCT
  851	TCAGCATTTT GGGTGCATTT GAAAATACGG ACGTGAACAC
  	TTCGCTGGTA
  901	TTCGGCGGCA CTTGCGGCGT GCTTGCCGTC GTCCTCTGCA
  	CGCTCGGCAC
  951	GATTAAAATC GCCGATTATC CCAAAGCCGT TTGGCAGGGT
  	GCGAAATCCA
  1001	TGTTCGGCGC AATCGCCATT TTAATCCTTG CCTGGCTCAT
  	CAGTACGGTT
  1051	GTCGGCGAAA TGCACACAGG CGACTACCTC TCCACGCTGG
  	TTGCGGGCAA
  1101	CATCCATCCC GGCTTCCTGN CCGTCATCCT TTTCCTGCTC
  	GCCAGCGTGA
  1151	TGGCGTTTGC CACAGGCACA AGCTGGGGGA CGTTCGGCAT
  	CATGCTGCCG
  1201	ATTGCCGCCG CCATGGCGGT CAAAGTCGAT CCCTCACTGA
  	TTATCCCGTG
  1251	TATGTCCGCC GTGATGGCGG GGGCGGTATG CGGCGACCAC
  	TGCTCGCCCA
  1301	TTTCCGACAC GACCATCCTG TCGTCCACCG GCGCGCGCTG
  	CAACCACATC
  1351	GACCACGTTA CNTCGCAACT GCCTTACGCC TTAACCGTTG
  	CCGCCGCCGC
  1401	CGCATCGGGN TACCTCGCAT TGGGTCTGAC AAAATCCGCG
  	CTGTTGGGTT
  1451	TTGGCANGAC AGGCATTGTA TTGGCGGTGC TGATTTTTCT
  	GTTGAAAGAT
  1501	AAAAAACGCG CCAACGCCTG A

• This encodes a protein having amino acid sequence <SEQ ID 694>:

• 1	MQLIDYSHSF FSVVPPFLAL ALAVITRRVL LSLGIGILVG
  	VAFLVGGNPV
  51	DGLTHLKDMV VGLAWSDGDW SLGKPKXLVF LILLGIFTSL
  	LTYSGSNQAF
  101	ADWAKRHIKN RRGAKMLTAC LVFVTFIDDY FHSLAVGAXA
  	RPVTDKFKVS
  151	RAKLAYILDS TAAPMCVLMP VSSWGASIIA TLAGLLVTYK
  	ITEYTPMGTF
  201	VAMSLMNYYA LFALIMVFVV AWFSFDI GSM ARFEQAALNE
  	AHDETAVSDG
  251	SWGRVYALII PVLALIASTV SAMIYTGAQA SETFSILGAF
  	ENTDVNTSLV
  301	FGGTCGVLAV VLCTLGTIKI ADYPKAVWQG AKSMFGAIAI
  	LILAWLISTV
  351	VGEMHTGDYL STLVAGNIHP GFLXVILFLL ASVMAFATGT
  	SWGTFGIMLP
  401	IAAAMAVKVD PSLIIPCMSA VMAGAVCGDH CSPISDTTIL
  	SSTGARCNHI
  451	DHVTSQLPYA LTVAAAAASG YLALGLTKSA LLGFGXTGIV
  	LAVLIFLLKD
  501	KKRANA*

• ORF26a and ORF26-1 show 97.8% identity in 506 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF26 shows 94.8% and 99% identity in 97 and 206 aa overlap at the N-terminus and C-terminus, respectively, with a predicted ORF (ORF26ng) from N. gonorrhoeae:
• 
• The complete length ORF26ng nucleotide sequence <SEQ ID 695> is:

• 1	ATGCAGCTGA TTGACTATTC ACATTCATTT TTCTCGGTTG
  	TGCCACCCTT
  51	TTTGGCACTG GCACTTGCCG TCATTACCCG CCGCGTACTG
  	CTGTCTTTAG
  101	GCATCGGTAT TTTGGTCGGC GTTGCCTTTT TGGTCGGCGG
  	CAACCCCGTC
  151	GACGGTCTGA CACACCTGAA AGACATGGTC GTCGGCTTGG
  	CTTGGGCAGA
  201	CGGCGATTGG TCGCTGGGCA AACCAAAAAT CTTGGTTTTC
  	CTGATACTTT
  251	TGGGCATTTT CACTTCACTG CTGACCTACT CCGGCAGCAA
  	TCAGGCGTTT
  301	GCCGACTGGG CAAAACGGCA CATTAAAAAC CGGTGCGGCG
  	CGAAAATGCT
  351	GACCGCCTGC CTCGTGTTCG TAACCTTTAT CGACGACTAT
  	TTCCACAGCC
  401	TCGCCGTCGG TGCGATTGCC CGCCCCGTTA CCGACAAGTT
  	TAAAGTTTCC
  451	CGCGCCAAAC TCGCCTACAT CCTCGACTCC ACTGCCTCGC
  	CCATGTGCGT
  501	GCTGATGCCC GTTTCAAGCT GGGGCGCGTC GATTATCGCC
  	ACGCTTGCCG
  551	GATTGCTCGT TACCTACAAA ATTACCGAAT ACACGCCGAT
  	GGGGACGTTT
  601	GTCGCCATGA GCCTGATGAA CTATTACGCG CTGTTTGCCC
  	TGATTATGGT
  651	ATTCGTCGTC GCATGGTTCT CCTTCGACAT CGGCTCGAtg
  	gCGCGTTTCG
  701	AACAGGCTGC GTTGAACGAA gcccaggacg aaaccgccgc
  	tTCAGACgCT
  751	ACCAAAGGTC GTGTTTACGC ATTGATTATT CCCGTTTTGG
  	CCTTAATCGC
  801	CTCAACGGTT TCCGCCATGA TCTACACCGG CGCGCAGGCA
  	AGCGAAACCT
  851	TCAGCATTTT GGGGGCATTT GAAAATACCG ACGTAAACAC
  	TTCGCTGGTA
  901	TTCGGCGGCA CTTGCGGCGT GCTTGCCGTC GTCCTCTGCA
  	CGTTCGGCAC
  951	GATTAAAACC GCCGATTATC CCAAAGCCGT GTGGCAGGGT
  	GCGAAATCCA
  1001	TGTTCGGCGC AATCGCCATT TTAATCCTCG CCTGGCTCAT
  	CAGTACGGTT
  1051	GTCGGCGAAA TGCACACGGG CGACTACCTC TCCACGCTGG
  	TTGCGGGCAA
  1101	CATCCATCCC GGCTTCCTGC CCGTCATCCT CTTCCTGCTC
  	GCCAGCGTGA
  1151	TGGCGTTTGC CACAGGCACA AGCTGGGGGA CGTTCGGCAT
  	TATGCTGCCG
  1201	ATTGCCGCCG CCATGGCGGT CAAAGTCGAA CCCGCGCTGA
  	TTAtcccGTG
  1251	TATGTCCGCA GTAATGGCGG GGGCGGTATG CGGCGACCAC
  	TGTTCGCCCA
  1301	TCTCCGACAC GACCATCCTG TCGTCCACCG GCGCGCGCTG
  	CAACCACATC
  1351	GACCACGTTA CCTCGCAACT GCCTTATGCC CTGACGGTTG
  	CCGCCGCCGC
  1401	CGCATCGGGC TACCTCGCAT TGGGTCTGAC AAAATCCGCG
  	CTGTTGGGCT
  1451	TTGGCACGAC CGGTATTGTA TTGGCGGTGC TGATTTTTCT
  	GTTGAAAGAT
  1501	AAAAAACGCG CCGACGTTTG A

• This encodes a protein having amino acid sequence <SEQ ID 696>:

• 1	MQLIDYSHSF FSVVPPFLAL ALAVITRRVL LSLGIGILVG
  	VAFLVGGNPV
  51	DGLTHLKDMV VGLAWADGDW SLGKPKILVF LILLGIFTSL
  	LTYSGSNQAF
  101	ADWAKRHIKN RCGAKMLTAC LVFVTFIDDY FHSLAVGAIA
  	RPVTDKFKVS
  151	RAKLAYILDS TASPMCVLMP VSSWGASIIA TLAGLLVTYK
  	ITEYTPMGTF
  201	VAMSLMNYYA LFALIMVFVV AWFSFDI GSM ARFEQAALNE
  	AQDETAASDA
  251	TKGRVYALII PVLALIASTV SAMIYTGAQA SETFSILGAF
  	ENTDVNTSLV
  301	FGGTCGVLAV VLCTFGTIKT ADYPKAVWQG AKSMFGAIAI
  	LILAWLISTV
  351	VGEMHTGDYL STLVAGNIHP GFLPVILFLL ASVMAFATGT
  	SWGTFGIMLP
  401	IAAAMAVKVE PALIIPCMSA VMAGAVCGDH CSPISDTTIL
  	SSTGARCNHI
  451	DHVTSQLPYA LTVAAAAASG YLALGLTKSA LLGFGTTGIV
  	LAVLIFLLKD
  501	KKRADV*

• ORF26ng and ORF26-1 show 98.4% identity in 505 aa overlap:
• 
• In addition, ORF26 ng shows significant homology to a hypothetical H. influenzae protein:

• sp|P44263|YF86_HAEIN HYPOTHETICAL PROTEIN HI1586 >gi|1074850|pir||C64037
  hypothetical
  protein HI1586 - Haemophilus influenzae (strain Rd KW20) >gi|1574427
  (U32832) H. influenzae predicted coding region HI1586 [Haemophilus
  influenzae] Length = 519
  Score = 538 bits (1370), Expect = e−152
  Identities = 280/507 (55%), Positives = 346/507 (68%), Gaps = 7/507 (1%)

  Query:	1	MQLIDYSHSFFSVVPPFLALALAVITRRXXXXXXXXXXXXXAFLVGGNPVDGLTHLKDMV	60
  		M+LID+S S +S+VP  LA+ LA+ TRR               L          +L   V
  Sbjct:	14	MELIDFSSSVWSIVPALLAIILAIATRRVLVSLSAGIIIGSLMLSDWQIGSAFNYLVKNV	73
  Query:	61	VGLAWADGDWSLGKPKILVFLILLGIFTSLLTYSGSNQAFADWAKRHIKNRCGAKMLTAC	120
  		V L +ADG+ +     I++FL+LLG+ T+LLT SGSN+AFA+WA+  IK R GAK+L A
  Sbjct:	74	VSLVYADGEIN-SNMNIVLFLLLLGVLTALLTVSGSNRAFAEWAQSRIKGRRGAKLLAAS	132
  Query:	121	LVFVTFIDDYFHSLAVGAIARPVTDKFKVSRAKLAYILDSTASPMCVLMPVSSWGASIIA	180
  		LVFVTFIDDYFHSLAVGAIARPVTD+FKVSRAKLAYILDSTA+PMCV+MPVSSWGA II
  Sbjct:	133	LVFVTFIDDYFHSLAVGAIARPVTDRFKVSRAKLAYILDSTAAPMCVMMPVSSWGAYIIT	192
  Query:	181	TLAGLLVTYKITEYTPMGTFVAMSLMNYYALFALIMVFVVAWFSFDIGSMARFEQAALNE	240
  		+ GLL TY ITEYTP+G FVAMS MN+YA+F++IMVF VA+FSFDI SM R E+ AL
  Sbjct:	193	LIGGLLATYSITEYTPIGAFVAMSSMNFYAIFSIIMVFFVAYFSFDIASMVRHEKLALKN	252
  Query:	241	AQDETAASDATKGRVYALIIPVLALIASTVSAMIYTGAQA----SETFSILGAFENTDVN	296
  		+D+      TKG+V  LI+P+L LI +TVS MIYTGA+A     + FS+LG FENT V
  Sbjct:	253	TEDQLEEETGTKGQVRNLILPILVLIIATVSMMIYTGAEALAADGKVFSVLGTFENTVVG	312
  Query:	297	TSLVFGGTCGVL--AVVLCTFGTIKTADYPKAVWQGAKSMFGXXXXXXXXXXXSTVVGEM	354
  		TSLV GG C ++   +++     +   +Y ++   G KSM G           + +VG+M
  Sbjct:	313	TSLVVGGFCSIIISTLLIILDRQVSVPEYVRSWIVGIKSMSGAIAILFFAWTINKIVGDM	372
  Query:	355	HTGDYLSTLVAGNIHPGFLPVILFLLASVMAFATGTSWGTFGIMLPIAAAMAVKVEPALI	414
  		TG YLS+LV+GNI   FLPVILF+L + MAF+TGTSWGTFGIMLPIAAAMA    P L+
  Sbjct:	373	QTGKYLSSLVSGNIPMQFLPVILFVLGAAMAFSTGTSWGTFGIMLPIAAAMAANAAPELL	432
  Query:	415	IPCMSAVMAGAVCGDHCSPISDTTILSSTGARCNHIDHVTSQXXXXXXXXXXXXXXXXXX	474
  		+PC+SAVMAGAVCGDHCSP+SDTTILSSTGA+CNHIDHVT+Q
  Sbjct:	433	LPCLSAVMAGAVCGDHCSPVSDTTILSSTGAKCNHIDHVTTQLPYAATVATATSIGYIVV	492
  Query:	475	XXXKSALLGFGTTGIVLAVLIFLLKDK	501
  		S L GF  T + L V+IF +K +
  Sbjct:	493	GFTYSGLAGFAATAVSLIVIIFAVKKR	519

• Based on this analysis, it is predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 83
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 697>:

• 1	..AAGCAATGGT ATGCCGACGN .AGTATCAAG ACGGAAATGG
  	TTATGGTCAA
  51	CGATGAGCCT GCCAAAATTC TGACTTGGGA TGAAAGCGGC
  	CGATTACTCT
  101	CGGAACTGTC TATCCGCCAC CATCAACGCA ACGGGGTGGT
  	TTTGGAGTGG
  151	TATGAAGATG GTTCTAAAAA GAGCGAAGT. GTTTATCAGG
  	ATGACAAGTT
  201	GGTCAGGAAA ACCCAGTGGG ATAAGGATGG TTATTTAATC
  	GAACCCTGA

• This corresponds to the amino acid sequence <SEQ ID 698; ORF27>:

• 1	..KQWYADXSIK TEMVMVNDEP AKILTWDESG RLLSELSIRH
  	HQRNGVVLEW
  51	YEDGSKKSEX VYQDDKLVRK TQWDKDGYLI EP*

• Further work revealed the complete nucleotide sequence <SEQ ID 699>:

• 1	ATGAAAAAAT TATCTCGGAT TGTATTTTCA ACTGTCCTGT
  	TGGGTTTTTC
  51	GGCCGCTTTG CCGGCGCAGA CCTATTCTGT TTATTTTAAT
  	CAGAACGGAA
  101	AGCTGACGGC GACGATGTCT TCTGCCGCTT ATATCAGGCA
  	ATATAGTGTG
  151	GTGGCGGGTA TTGCGCACGC GCAGGATTTT TATTATCCGT
  	CGATGAAGAA
  201	ATATTCTGAA CCTTATATCG TTGCTTCAAC GCAAATCAAA
  	TCTTTTGTGC
  251	CTACCCTGCA AAACGGTATG TTGATTTTGT GGCATTTTAA
  	TGGTCAGAAA
  301	AAAATGGCGG GGGGCTTCAG CAAGGGTAAG CCGGACGGGG
  	AGTGGGTCAA
  351	CTGGTATCCG AACGGTAAAA AATCTGCCGT TATGCCTTAT
  	AAAAATGGCT
  401	TGAGTGAGGG TACGGGATAC CGCTATTACC GTAACGGCGG
  	CAAGGAAAGC
  451	GAAATCCAGT TTAAGCAAAA TAAGGCAAAC GGCGTATGGA
  	AGCAATGGTA
  501	TGCCGACGGC AGTATCAAGA CGGAAATGGT TATGGTCAAC
  	GATGAGCCTG
  551	CCAAAATTCT GACTTGGGAT GAAAGCGGCC GATTACTCTC
  	GGAACTGTCT
  601	ATCCGCCACC ATCAACGCAA CGGGGTGGTT TTGGAGTGGT
  	ATGAAGATGG
  651	TTCTAAAAAG AGCGAAGCTG TTTATCAGGA TGACAAGTTG
  	GTCAGGAAAA
  701	CCCAGTGGGA TAAGGATGGT TATTTAATCG AACCCTGA

• This corresponds to the amino acid sequence <SEQ ID 700; ORF27-1>:

• 1	MKKLSRIVFS TVLLGFSAAL PAQTYSVYFN QNGKLTATMS
  	SAAYIRQYSV
  51	VAGIAHAQDF YYPSMKKYSE PYIVASTQIK SFVPTLQNGM
  	LILWHFNGQK
  101	KMAGGFSKGK PDGEWVNWYP NGKKSAVMPY KNGLSEGTGY
  	RYYRNGGKES
  151	EIQFKQNKAN GVWKQWYADG SIKTEMVMVN DEPAKILTWD
  	ESGRLLSELS
  201	IRHHQRNGVV LEWYEDGSKK SEAVYQDDKL VRKTQWDKDG
  	YLIEP*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF27 shows 91.5% identity over a 82aa overlap with an ORF (ORF27a) from strain A of N. meningitidis:
• 
• The complete length ORF27a nucleotide sequence <SEQ ID 701> is:

• 1	ATGAAAAAAT TATCTCGGAT TGTATTTTCA ACTGTCCTGT
  	TGGGTTTTTC
  51	GGCCGCTTTG CCGGCGCAGA NCTATTCTGT TTATTTTAAT
  	CAGAACGGGA
  101	AACTGACGGC GACGNTGTCT TCTGCCGCNT ATATCAGGCA
  	ATATAGTGTG
  151	GCGGAGGGTA TTGCGCACGC GCAGGANTTT TANTATCCGT
  	CGATGAAGAA
  201	ATATTCCGAA CCTTATATCG TTGCTTCAAC GCAAATCAAA
  	TCTTTTGTGC
  251	CTACCCTGCA AAACGGTATG TTGATTTTGT GGCATTTTAA
  	NGGTCAGAAA
  301	AAAATGGCNG GGGGCTTCAG CAAGGGTAAG CCGGACGGGG
  	AGTGGGTCAA
  351	CTGGTATCCG AACGGTAAAA AATCTGCCGT TATGCCTTAT
  	AAAAATGGTT
  401	TGAGTGAAGG TACGGGGTNN CGCTATTACC GTAACGGCGG
  	CAAGGAAAGC
  451	GAAATCCAGT TTAAACAGAA TAAGGCAAAC GGCGTATGGA
  	AGCAATGGTA
  501	TGCCGACGGC AATATCAAAA CGGAAATGGT TATGGTCAAT
  	GATGAGCCTG
  551	CCAAAATTCT GACATGGGAT GAAAGCGGTC GATTACTCTC
  	GGAACTGTCT
  601	ATCCATCATC ATNAACGTAA TGGAGTAGTC TTAGAGTGGT
  	ATGAAGATGG
  651	TTCTAAAAAG ANTGAAGCTG TTTATCAGGA TGATAAGTTG
  	GTCAGGAAAA
  701	CCCAGTGGGA TAANGATGGT TATTTAATCG AACCCTGA

• This encodes a protein having amino acid sequence <SEQ ID 702>:

• 1	MKKLSRIVFS TVLLGFSAAL PAQXYSVYFN QNGKLTATXS
  	SAAYIRQYSV
  51	AEGIAHAQXF XYPSMKKYSE PYIVASTQIK SFVPTLQNGM
  	LILWHFXGQK
  101	KMAGGFSKGK PDGEWVNWYP NGKKSAVMPY KNGLSEGTGX
  	RYYRNGGKES
  151	EIQFKQNKAN GVWKQWYADG NIKTEMVMVN DEPAKILTWD
  	ESGRLLSELS
  201	IHHHXRNGVV LEWYEDGSKK XEAVYQDDKL VRKTQWDXDG
  	YLIEP*

• ORF27a and ORF27-1 show 94.7% identity in 245 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF27 shows 96.3% identity over 82 aa overlap with a predicted ORF (ORF27ng) from N. gonorrhoeae:
• 
• The complete length ORF27ng nucleotide sequence <SEQ ID 703> is:

• 1	ATGAAGAAAT TATCTCGGAT TGTATTTTCA ATCGTACTGT
  	TGGGTTTTTC
  51	GGCCGCTTTG CCGGCGCAGA CCTATTCTGT TTATTTTAAT
  	CAGAACGGGA
  101	AACTGACGGC GACGATGTCT TCTGCCGCTT ATATCAGGCA
  	ATATAGTGTG
  151	GCGGCGGGTA TCGCACACGC GCAGGATTTT TATTATCCGT
  	CGATGAAGAA
  201	ATATTCCGAA CCTTATATCG TTGCTTCAAC GCAAATCAAA
  	TCTTTTGTGC
  251	CTACCCTGCA AAACGGTATG TTGATTTTGT GGCATTTTAA
  	TGGTCAGAAA
  301	AAAATGGCGG GGGGCTTCAG CAAGGGTAAG CCGGACGGGG
  	AATGGGTCAA
  351	CTGGTATCCG AACGGTAAAA AATCTGCGGT TATGCCTTAT
  	AAAAATGGCT
  401	TGAGTGAGGG TACGGGATAC CGTTATTACC GTAACGGCGG
  	CAAGGAAAGC
  451	GAAATCCAGT TTAAGCAAAA TAAGGCGAAC GGCGTATGGA
  	AGCAATGGTA
  501	TGCCGATGGA AGTATCAAGA CGGAAATGGT TATGGTCAAC
  	GATGAGCCTG
  551	CCAAAATTCT GACTTGGGAT GAAAGCGGCC GATTACTTTC
  	GGAACTGTCT
  601	ATCCGCCACC ATAAACGCAA CGGGGTGGTT TTGGAGTGGT
  	ATGAAGATGG
  651	TTCTAAAAAG AGCGAGGCTG TTTATCAGGA TGACAAGTTG
  	GTCAGGAAAA
  701	CCCAATGGGA TAAGGATGGT TATTTAATCG AACCCTGA

• This encodes a protein having amino acid sequence <SEQ ID 704>:

• 1	MKKLSRIVFS IVLLGFSAAL PAQTYSVYFN QNGKLTATMS
  	SAAYIRQYSV
  51	AAGIAHAQDF YYPSMKKYSE PYIVASTQIK SFVPTLQNGM
  	LILWHFNGQK
  101	KMAGGFSKGK PDGEWVNWYP NGKKSAVMPY KNGLSEGTGY
  	RYYRNGGKES
  151	EIQFKQNKAN GVWKQWYADG SIKTEMVMVN DEPAKILTWD
  	ESGRLLSELS
  201	IRHHKRNGVV LEWYEDGSKK SEAVYQDDKL VRKTQWDKDG
  	YLIEP*

• ORF27ng and ORF27-1 show 98.8% identity in 245 aa overlap:
• 
• Based on this analysis, including the putative leader sequence in the gonococcal protein, it was predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF27-1 (24.5 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 17A shows the results of affinity purification of the GST-fusion protein, and FIG. 17B shows the results of expression of the His-fusion in E. coli. Purified GST-fusion protein was used to immunise mice, whose sera were used for ELISA, which gave a positive result, confirming that ORF27-1 is a surface-exposed protein and a useful immunogen.
Example 84
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 705>:

• 1	ATGAAATTTA CCAAGCACCC CGTCTGGGCA ATGGCGTTCC
  	GCCCATTTTA
  51	TTCGCTGGCG GCTCTGTACG GCGCATTGTC CGTATTGCTG
  	TGGGGTTTCG
  101	GCTACACGGG AACGCACkAG CTGTCCGGTT TCTATTGGCA
  	CGCGCATGAg
  151	ATGATTTGGG GTTATGCCGG ACTGGTCGTC ATCGCCTTCC
  	TGCTGACCGC
  201	CGTCGCCACT TGGACGGGGC AGCCGCCCAC GCGGGGCGGC
  	GTaTCTGGTC
  251	GGCTTGACTA TCTTTTGGCT GGCTGCGCGG ATTGCCGCCT
  	TTATCCCGGG
  301	TTGGGGTGCG TCGGCAAGCG GCATACTCGG TACGCTGTTT
  	TTCTGGTACG
  351	GCGCGGTGTG CATGGCTTTG CCCGTTATCC GTTCGCAGAA
  	TCAACGCAAC
  401	TATGTTgCCG TGTTCGCGCT GTTCGTCTTG GGCGGCACGC
  	ATGCGGCGTT
  451	CCACGTCCAG CTGCACAACG GCAACCTAGG CGGACTCTTG
  	AGCGGATTGC
  501	AGTCGGGCTT GGTGATG

• This corresponds to the amino acid sequence <SEQ ID 706; ORF47>:

• 1	MKFTKHPVWA MAFRPFYSLA ALYGALSVLL WGFGYTGTHX
  	LSGFYWHAHE
  51	MIWGYAGLVV IAFLLTAVAT WTGQPPTRGG VLVGLTIFWL
  	AARIAAFIPG
  101	WGASASGILG TLFFWYGAVC MALPVIRSQN QRNYVAVFAL
  	FVLGGTHAAF
  151	HVQLHNGNLG GLLSGLQSGL VM

• Further work revealed the complete nucleotide sequence <SEQ ID 707>:

• 1	ATGAAATTTA CCAAGCACCC CGTCTGGGCA ATGGCGTTCC
  	GCCCATTTTA
  51	TTCGCTGGCG GCTCTGTACG GCGCATTGTC CGTATTGCTG
  	TGGGGTTTCG
  101	GCTACACGGG AACGCACGAG CTGTCCGGTT TCTATTGGCA
  	CGCGCATGAG
  151	ATGATTTGGG GTTATGCCGG ACTGGTCGTC ATCGCCTTCC
  	TGCTGACCGC
  201	CGTCGCCACT TGGACGGGGC AGCCGCCCAC GCGGGGCGGC
  	GTTCTGGTCG
  251	GCTTGACTAT CTTTTGGCTG GCTGCGCGGA TTGCCGCCTT
  	TATCCCGGGT
  301	TGGGGTGCGT CGGCAAGCGG CATACTCGGT ACGCTGTTTT
  	TCTGGTACGG
  351	CGCGGTGTGC ATGGCTTTGC CCGTTATCCG TTCGCAGAAT
  	CAACGCAACT
  401	ATGTTGCCGT GTTCGCGCTG TTCGTCTTGG GCGGCACGCA
  	TGCGGCGTTC
  451	CACGTCCAGC TGCACAACGG CAACCTAGGC GGACTCTTGA
  	GCGGATTGCA
  501	GTCGGGCTTG GTGATGGTGT CGGGTTTTAT CGGTCTGATT
  	GGTACGCGGA
  551	TTATTTCGTT TTTTACGTCC AAACGCTTGA ATGTGCCGCA
  	GATTCCCAGT
  601	CCGAAATGGG TGGCGCAGGC TTCGCTGTGG CTGCCCATGC
  	TGACTGCCAT
  651	GCTGATGGCG CACGGTGTGT TGGCTTGGCT GTCTGCCGTT
  	TTTGCCTTTG
  701	CGGCAGGTGT GATTTTTACC GTGCAGGTGT ACCGCTGGTG
  	GTATAAACCC
  751	GTGTTGAAAG AGCCGATGCT GTGGATTCTG TTTGCCGGCT
  	ATCTGTTTAC
  801	CGGATTGGGG CTGATTGCGG TCGGCGCGTC TTATTTCAAA
  	CCCGCTTTCC
  851	TCAATCTGGG TGTGCATCTG ATCGGGGTCG GCGGTATCGG
  	CGTGCTGACT
  901	TTGGGCATGA TGGCGCGTAC CGCGCTTGGT CATACGGGCA
  	ATCCGATTTA
  951	TCCGCCGCCC AAAGCCGTTC CCGTTGCGTT TTGGCTGATG
  	ATGGCGGCAA
  1001	CCGCCGTCCG TATGGTTGCC GTATTTTCTT CCGGCACTGC
  	CTACACGCAC
  1051	AGCATCCGCA CCTCTTCGGT TTTGTTTGCA CTCGCGCTTT
  	TGGTGTATGC
  1101	GTGGAAGTAT ATTCCTTGGC TGATTCGTCC GCGTTCGGAC
  	GGCAGGCCCG
  1151	GTTGA

• This corresponds to the amino acid sequence <SEQ ID 708; ORF47-1>:

• 1	MKFTKHPVWA MAFRPFYSLA ALYGALSVLL WGFGYTGTHE
  	LSGFYWHAHE
  51	MIWGYAGLVV IAFLLTAVAT WTGQPPTRGG VLVGLTIFWL
  	AARIAAFIPG
  101	WGASASGILG TLFFWYGAVC MALPVIRSQN QRNYVAVFAL
  	FVLGGTHAAF
  151	HVQLHNGNLG GLLSGLQSGL VMVSGFIGLI GTRIISFFTS
  	KRLNVPQIPS
  201	PKWVAQASLW LPMLTAMLMA HGVLAWLSAV FAFAAGVIFT
  	VQVYRWWYKP
  251	VLKEPMLWIL FAGYLFTGLG LIAVGASYFK PAFLNLGVHL
  	IGVGGIGVLT
  301	LGMMARTALG HTGNPIYPPP KAVPVAFWLM MAATAVRMVA
  	VFSSGTAYTH
  351	SIRTSSVLFA LALLVYAWKY IPWLIRPRSD GRPG*

• Computer analysis of this amino acid sequence predicts a leader peptide and also gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF47 shows 99.4% identity over a 172aa overlap with an ORF (ORF47a) from strain A of N. meningitidis:
• 
• The complete length ORF47a nucleotide sequence <SEQ ID 709> is:

• 1	ATGAAATTTA CCAAGCACCC CGTTTGGGCA ATGGCGTTCC
  	GCCCGTTTTA
  51	TTCACTGGCG GCTCTGTACG GCGCATTGTC CGTATTGCTG
  	TGGGGTTTCG
  101	GCTACACGGG AACGCACGAG CTGTCCGGTT TCTATTGGCA
  	CGCGCATGAG
  151	ATGATTTGGG GTTATGCCGG ACTGGTCGTC ATCGCCTTCC
  	TGCTGACCGC
  201	CGTCGCCACT TGGACGGGGC AGCCGCCCAC GCGGGGCGGC
  	GTTCTGGTCG
  251	GCTTGACTAT CTTTTGGCTG GCTGCGCGGA TTGCCGCCTT
  	TATCCCGGGT
  301	TGGGGTGCGT CGGCAAGCGG CATACTCGGT ACGCTGTTTT
  	TCTGGTACGG
  351	CGCGGTGTGC ATGGCTTTGC CCGTTATCCG TTCGCAGAAT
  	CAACGCAATT
  401	ATGTTGCCGT GTTCGCGCTG TTCGTCTTGG GCGGTACGCA
  	CGCGGCGTTC
  451	CACGTCCAGC TGCACAACGG CAACCTAGGC GGACTCTTGA
  	GCGGATTGCA
  501	GTCGGGCTTG GTGATGGTGT CGGGTTTTAT CGGTCTGATT
  	GGTACGCGGA
  551	TTATTTCGTT TTTTACGTCC AAACGGTTGA ATGTGCCGCA
  	GATTCCCAGT
  601	CCGAAATGGG TGGCGCAGGC TTCGCTGTGG CTGCCCATGC
  	TGACCGCCAT
  651	GCTGATGGCG CACGGCGTGA TGCCTTGGCT GTCGGCGGCT
  	TTCGCGTTTG
  701	CGGCAGGTGT GATTTTTACC GTGCAGGTGT ACCGCTGGTG
  	GTATAAGCCT
  751	GTGTTGAAAG AGCCGATGCT GTGGATTCTG TTTGCCGGCT
  	ATCTGTTTAC
  801	CGGATTGGGG CTGATTGCGG TCGGCGCGTC TTATTTCAAA
  	CCCGCTTTCC
  851	TCAATCTGGG TGTGCATCTG ATCGGGGTCG GCGGTATCGG
  	CGTGCTGACT
  901	TTGGGCATGA TGGCGCGTAC CGCGCTCGGT CATACGGGCA
  	ATCCGATTTA
  951	TCCGCCGCCC AAAGCCGTTC CCGTTGCGTT TTGGCTGATG
  	ATGGCGGCAA
  1001	CCGCCGTCCG TATGGTTGCC GTATTTTCTT CCGGCACTGC
  	CTACACGCAC
  1051	AGCATACGCA CCTCTTCGGT TTTGTTTGCA CTCGCGCTTT
  	TGGTGTATGC
  1101	GTGGAAGTAT ATTCCTTGGC TGATTCGTCC GCGTTCGGAC
  	GGCAGGCCCG
  1151	GTTGA

• This encodes a protein having amino acid sequence <SEQ ID 710>:

• 1	MKFTKHPVWA MAFRPFYSLA ALYGALSVLL WGFGYTGTHE
  	LSGFYWHAHE
  51	MIWGYAGLVV IAFLLTAVAT WTGQPPTRGG VLVGLTIFWL
  	AARIAAFIPG
  101	WGASASGILG TLFFWYGAVC MALPVIRSQN QRNYVAVFAL
  	FVLGGTHAAF
  151	HVQLHNGNLG GLLSGLQSGL VMVSGFIGLI GTRIISFFTS
  	KRLNVPQIPS
  201	PKWVAQASLW LPMLTAMLMA HGVMPWLSAA FAFAAGVIFT
  	VQVYRWWYKP
  251	VLKEPMLWIL FAGYLFTGLG LIAVGASYFK PAFLNLGVHL
  	IGVGGIGVLT
  301	LGMMARTALG HTGNPIYPPP KAVPVAFWLM MAATAVRMVA
  	VFSSGTAYTH
  351	SIRTSSVLFA LALLVYAWKY IPWLIRPRSD GRPG*

• ORF47a and ORF47-1 show 99.2% identity in 384 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF47 shows 97.1% identity over 172 aa overlap with a predicted ORF (ORF47ng) from N. gonorrhoeae:
• 
• The ORF47ng nucleotide sequence <SEQ ID 711> is predicted to encode a protein comprising amino acid sequence <SEQ ID 712>:

• 1	MKFTKHPVWA MAFRPFYSLA ALYGALSVLL WGFGYTGTHE
  	LSGFYWHAHE
  51	MIWGYAGLVV IAFLLTAVAT WTGQPPTRGG VLVGLTAFWL
  	AARIAAFIPG
  101	WGAAASGILG TLFFWYGAVC MALPVIRSQN RRNYVAVFAI
  	FVLGGTHAAF
  151	HVQLHNGNLG GLLSGLQSGL VMVWGFIGLI GMKIISFFTS
  	KRLKLPQIPS
  201	PKWVAHASLW LPMLNAILMA HRVMPWLSAA FPFAAGVIFT
  	VQVYAGGITP
  251	IEETSCGSVA GICYRLGNSS G

• The predicted leader peptide and transmembrane domains are identical (except for an Ile/Ala substitution at residue 87 and an Leu/Ile substitution at position 140) to sequences in the meningococcal protein (see also Pseudomonas stutzeri orf396, accession number e246540):

• TM segments in ORF47ng

  INTEGRAL	Likelihood = −5.63	Transmembrane	52 - 68
  INTEGRAL	Likelihood = −3.88	Transmembrane	169 - 185
  INTEGRAL	Likelihood = −3.08	Transmembrane	82 - 98
  INTEGRAL	Likelihood = −1.91	Transmembrane	134 - 150
  INTEGRAL	Likelihood = −1.44	Transmembrane	107 - 123
  INTEGRAL	Likelihood = −1.38	Transmembrane	227 - 243

• Further work revealed the complete gonococcal DNA sequence <SEQ ID 713>:

• 1	ATGAAATTTA CCAAACATCC CGTCTGGGCA ATGGCGTTCC
  	GCCCGTTTTA
  51	TTCACTGGCG GCACTGTACG GCGCATTGTC CGTATTGCTG
  	TGGGGTTTCG
  101	GCTACACGGG AACGCACGAG CTGTCCGGTT TCTATTGGCA
  	CGCGCATGAG
  151	ATGATTTGGG GTTATGCCGG TCTCGTCGTC ATCGCCTTCC
  	TGCTGACCGC
  201	CGTCGCCACT TGGACGGGAC AGCCGCCCAC GAGGGGCGGC
  	GTTCTGGTCG
  251	GCTTGACCGC CTTTTGGCTG GCTGCGCGGA TTGCCGCCTT
  	TATCCCGGGT
  301	TGGGGTGCGG CGGCAAGCGG CATACTCGGT ACGCTGTTTT
  	TCTGGTACGG
  351	CGCGGTGTGC ATGGCTTTGC CCGTTATCCG TtcgCAAAAC
  	CGGCGCAACT
  401	ATGtcgCCGT ATTCGCAATA TTTGTGCTGG GCGGTACGCA
  	TGCGgcgTTC
  451	CACGtccAgc tGCACAACGG CAACCTAGGC GGACTCTTGA
  	GCGGATTGCA
  501	GTCGGGCCTG GTTATGGTGT CGGGCTTTAT CGGCCTGATT
  	GGGATGAGGA
  551	TTATTTCGTT TTTTACGTCC AAACGGTTGA ACGTGCCGCA
  	GATTCCCAGT
  601	CCGAAATGGG TGGCGCAGGC TTCGCTGTGG CTACCCATGC
  	TGACCGCCAT
  651	ACTGATGGCG CACGGCGTGA TGCCTTGGCT GTCGGCGGCT
  	TTCGCGTTTG
  701	CGGCGGGCGT GATTTTTACC GTACAGGTGT ACCGCTGGTG
  	GTATAAACCC
  751	GTATTGAAAG AACCGATGCT GTGGATTCTG TTTGCCGGCT
  	ATCTGTTTAC
  801	CGGATTGGGG CTGATTGCGG TCGGCGCGTC TTATTTCAAA
  	CCTGCCTTCC
  851	TCAATCTGGG CGTACATCTG ATCGGGGTCG GCGGTATCGG
  	CGTGCTGACT
  901	TTGGGCATGA TGGCGCGTAC CGCGCTCGGT CATACGGGCA
  	ATTCGATTTA
  951	TCCGCCGCCC AAAGCCGTTC CCGTTGCGTT TTGGCTGATG
  	ATGGCGGCAA
  1001	CCGCCGTCCG TATGGTTGCC GTATTTTCTT CCGGCACTGC
  	CTACACGCAC
  1051	AGCATCCGCA CGTCTTCGGT TTTGTTTGCA CTCGCGCTGC
  	TGGTGTATGC
  1101	GTGGAAATAC ATTCCGTGGC TGATCCGTCC GCGTTCGGAC
  	GGCAGGCCCG
  1151	GTTGA

• This encodes a protein having amino acid sequence <SEQ ID 714; ORF47ng-1>:

• 1	MKFTKHPVWA MAFRPFYSLA ALYGALSVLL WGFGYTGTHE
  	LSGFYWHAHE
  51	MIWGYAGLVV IAFLLTAVAT WTGQPPTRGG VLVGLTAFWL
  	AARIAAFIPG
  101	WGAAASGILG TLFFWYGAVC MALPVIRSQN RRNYVAVFAI
  	FVLGGTHAAF
  151	HVQLHNGNLG GLLSGLQSGL VMVSGFIGLI GMRIISFFTS
  	KRLNVPQIPS
  201	PKWVAQASLW LPMLTAILMA HGVMPWLSAA FAFAAGVIFT
  	VQVYRWWYKP
  251	VLKEPMLWIL FAGYLFTGLG LIAVGASYFK PAFLNLGVHL
  	IGVGGIGVLT
  301	LGMMARTALG HTGNSIYPPP KAVPVAFWLM MAATAVRMVA
  	VFSSGTAYTH
  351	SIRTSSVLFA LALLVYAWKY IPWLIRPRSD GRPG*

• ORF47ng-1 and ORF47-1 show 97.4% identity in 384 aa overlap:
• 
• Furthermore, ORF47ng-1 shows significant homology to an ORF from Pseudomonas stutzeri:

• gnl|PID|e246540 (Z73914) ORF396 protein [Pseudomonas stutzeri]
  Length = 396 Score = 155 bits (389), Expect = 5e−37
  Identities = 121/391 (30%), Positives = 169/391 (42%), Gaps = 21/391 (5%)

  Query:	7	PVWAMAFRPFYSLAALYGALSVLLWGFGYTGTHELSGFY-------WHAHEMIWGYAGLV	59
  		P+W +AFRPF+   +LY  L++ LW   +TG     GF        WH HEM++G+A  +
  Sbjct:	14	PIWRLAFRPFFLAGSLYALLAIPLWVAAWTGLWP--GFQPTGGWLAWHRHEMLFGFAMAI	71
  Query:	60	VIAFLLTAVATWTGQPPTRGGVLVGLTAFWLAARIAAFIPGWGAAASGILGTLFFWYGAV	119
  		V  FLLTAV TWTGQ    G  LVGL A WLAAR+  ++ G  AA    L  LF
  Sbjct:	72	VAGFLLTAVQTWTGQTAPSGNRLVGLAAVWLAARL-GWLFGLPAAWLAPLDLLFLVALVW	130
  Query:	120	CMALPVIRSQNRRNYVAVFAIFVLGGTHAAFXXXXXXXXXXXXXXXXXXXXXMVSGFIGL	179
  		MA  +   + +RNY  V  + ++ G                          +V+  + L
  Sbjct:	131	MMAQMLWAVRQKRNYPIVVVLSLMLGADVLILTGLLQGNDALQRQGVLAGLWLVAALMAL	190
  Query:	180	IGMRIISFFTSKRLNVPQIPSP-KWVAQASLWLPMLTAILMAHGV----MPWLSAAFAFA	234
  		IG R+I FFT + L       P  W+  A L    + A+L A GV     P L   F  A
  Sbjct:	191	IGGRVIPFFTQRGLGKVDAVKPWVWLDVALLVGTGVIALLHAFGVAMRPQPLLGLLFV-A	249
  Query:	235	AGVIFTVQVYRWWYKPVLKEPMLWILFAGYLFTGLGLIAVGASYF-KPAFXXXXXXXXXX	293
  		GV   +++ RW+ K + K  +LW L    L+  +    +   +F   A
  Sbjct:	250	IGVGHLLRLMRWYDKGIWKVGLLWSLHVAMLWLVVAAFGLALWHFGLLAQSSPSLHALSV	309
  Query:	294	XXXXXXXXXMMARTALGHTGNSIYPPPKAVPVAFWLXXXXXXXXXXXXFSSGTAYTHSIR	353
  		M+AR  LGHTG  +  P   +  AF L            F S       +
  Sbjct:	310	GSMSGLILAMIARVTLGHTGRPLQLPAGIIG-AFVL---FNLGTAARVFLSVAWPVGGLW	365
  Query:	354	TSSVLFALALLVYAWKYIPWLIRPRSDGRPG	384
  		++V + LA  +Y W+Y P L+  R DG PG
  Sbjct:	366	LAAVCWTLAFALYVWRYAPMLVAARVDGHPG	396

• Based on this analysis, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 85
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 715>:

• 1	..ATGCCGTCTG AAGGTTCAGA CGGCmTCGGT GyCGGGGAAy
  	CAGAAGyGGT
  51	AGCGCATGCC CAATGAGACT TCGTGGGTTT TGAAGCGGGT
  	GTTTTCCAAG
  101	CGTCCCCAGT TGTGGTAACG GTATCCGGTG TCyAArGTCA
  	GCTTGGGyGT
  151	GATGTCGAAa CCGACACCGG CGATGACACC AAGACCyAmG
  	CTGCTGATrC
  201	TGTkGCTTTC GTGATAGGsA GGTTTGyTGG kmksAsyTTG
  	TAyrATwkkG
  251	CCTssCwsTG kAGmGCCkTk CkyTGGTkkA swGrwArTAG
  	TCGTGGTTTy
  301	TkTTyyCACC GAATGAACyT GATGTTTAAC GTGTCCGTAG
  	GCGACGCGCG
  351	CGCCGATATA GGGTTTGAAT TTATCGTTGA GTTTGAAATC
  	GTAAATGGCG
  401	GACAAGCCGA GAGAAGAAAC GGCGTGGAAG CTGCCGTTTC
  	CCTGATGTTT
  451	TGTTTGGGTT TCTTTGTAGT TGTTGTTTAT CTCTTCAGTA
  	ACTTTTTTAG
  501	TAGAAGAATT ACTTTCTTTC CATTTTCTGT AACTGGCATA
  	ATCTGCCGCT
  551	ATTCTCCAGC CGCCGAAATC ..

• This corresponds to the amino acid sequence <SEQ ID 716; ORF67>:

• 1	..MPSEGSDGXG XGEXEXVAHA QXDFVGFEAG VFQASPVVVT
  	VSGVXXQLGX
  51	DVETDTGDDT KTXAADXVAF VIGRFXGXXL YXXAXXXXAX
  	XWXXXXSRGF
  101	XXHRMNLMFN VSVGDARADI GFEFIVEFEI VNGGQAERRN
  	GVEAAVSLMF
  151	CLGFFVVVVY LFSNFFSRRI TFFPFSVTGI ICRYSPAAEI
  	..

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF67 shows 51.8% identity over 199 aa overlap with a predicted ORF (ORF67ng) from N. gonorrhoeae:
• 
• The ORF67ng nucleotide sequence <SEQ ID 717> is predicted to encode a protein comprising amino acid sequence <SEQ ID 718>:

• 1	MPSETVGSIV NVGVDESVGF SPPFPSIQHF YRFHRIHRIR
  	LFRPPGPMQL
  51	NRHSHGSGNL GRGVWATVLS DKFPCGQVRI PACAGMTNFE
  	IAVLSGMTVR
  101	VFYCARPAPV NGGRLKMPSE GSDGIGIGES EAVAHAQRGF
  	VGFEAGVFQA
  151	SPVVVAVAGV QGQAGRDVYA HARHRAEAQA AAAVAFLIGV
  	FLRMSVRINR
  201	NCCVSITRVG GKSTCYFFSR IDAVSDVSVG DARTDIGFEF
  	VVEFEIVNGG
  251	QAERRNGVEC AVFLMFRLLV FYVKLVAAKS FIILSFQLFY
  	VHGIFIVVPF
  301	PVTGIIRGDA PAAEVVADRH PGVDGMRTDV SEIIAYRAYF
  	VFAWSGWFRI
  351	IVGNAFGGVG *

• Based on the presence of a several putative transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 86
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 719>

• 1	ATGTTTGCTT TTTTAGAAGC CTTTTTTGTC GAATACGGTT
  	ATGCGGCTGT
  51	TTTTTTTGTA TTGGTCATCT GCGGTTTCGG CGTGCCGATT
  	CCCGAGGATT
  101	TGACCTTGGT AACAGGCGGC GTGATTTCGG GTATGGGTTA
  	TACCAATCCG
  151	CATATTATGT TTGCAGTCGG TATGCTCGGC GTATTGGTCG
  	GGGACGGCAT
  201	CATGTTCGCC GCCGGACGAA TTTGGGGGCA GArArTCCTA
  	rGGTTCArAC
  251	CTATTGCGsG CATCATGACG CCGrAACGTT ATGAGCAGGT
  	TCAGGAAAAA
  301	TTCGACAAAT ACGGTAACTG GGTCTTATTT GTCGCCCGTT
  	TCCTGCCCGG
  351	TTTGAGAACG GCCGTATTTG TTACAGCCGG TATCAGCCGC
  	AAGGTTTCAT
  401	ACTTGCGTTT TATCATTATG GATGGACTGG CCGCA...

• This corresponds to the amino acid sequence <SEQ ID 720; ORF78>:

• 1	MFAFLEAFFV EYGYAAVFFV LVICGFGVPI PEDLTLVTGG
  	VISGMGYTNP
  51	HIMFAVGMLG VLVGDGIMFA AGRIWGQXXL XFXPIAXIMT
  	PXRYEQVQEK
  101	FDKYGNWVLF VARFLPGLRT AVFVTAGISR KVSYLRFIIM
  	DGLAA...

• Further work revealed the complete nucleotide sequence <SEQ ID 721>:

• 1	ATGTTTGCTT TTTTAGAAGC CTTTTTTGTC GAATACGGTT
  	ATGCGGCTGT
  51	TTTTTTTGTA TTGGTCATCT GCGGTTTCGG CGTGCCGATT
  	CCCGAGGATT
  101	TGACCTTGGT AACAGGCGGC GTGATTTCGG GTATGGGTTA
  	TACCAATCCG
  151	CATATTATGT TTGCAGTCGG TATGCTCGGC GTATTGGTCG
  	GGGACGGCAT
  201	CATGTTCGCC GCCGGACGAA TTTGGGGGCA GAAAATCCTA
  	AGGTTCAAAC
  251	CTATTGCGCG CATCATGACG CCGAAACGTT ATGAGCAGGT
  	TCAGGAAAAA
  301	TTCGACAAAT ACGGTAACTG GGTCTTATTT GTCGCCCGTT
  	TCCTGCCCGG
  351	TTTGAGAACG GCCGTATTTG TTACAGCCGG TATCAGCCGC
  	AAGGTTTCAT
  401	ACTTGCGTTT TATCATTATG GATGGACTGG CCGCACTGAT
  	TTCCGTCCCT
  451	ATTTGGATTT ATCTGGGCGA ATACGGTGCG CACAACATCG
  	ATTGGCTGAT
  501	GGCGAAAATG CACAGCCTGC AATCGGGTAT TTTTGTTATC
  	TTGGGTATAG
  551	GTGCGACCGT TGTCGCTTGG ATTTGGTGGA AAAAACGCCA
  	ACGTATCCAG
  601	TTTTACCGCA GCAAATTGAA AGAAAAGCGG GCGCAACGCA
  	AAGCCGCCAA
  651	GGCAGCCAAA AAAGCCGCGC AAAGCAAACA ATAA

• This corresponds to the amino acid sequence <SEQ ID 722; ORF78-1>:

• 1	MFAFLEAFFV EYGYAAVFFV LVICGFGVPI PEDLTLVTGG
  	VISGMGYTNP
  51	HIMFAVGMLG VLVGDGIMFA AGRIWGQKIL RFKPIARIMT
  	PKRYEQVQEK
  101	FDKYGNWVLF VARFLPGLRT AVFVTAGISR KVSYLRFIIM
  	DGLAALISVP
  151	IWIYLGEYGA HNIDWLMAKM HSLQSGIFVI LGIGATVVAW
  	IWWKKRQRIQ
  201	FYRSKLKEKR AQRKAAKAAK KAAQSKQ*

• Computer analysis of this amino acid sequence predicts several transmembrane domains, and also gave the following results:
• Homology with the dedA Homologue of H. influenzae (Accession Number P45280)
• ORF78 and the dedA homologue show 58% aa identity in 144aa overlap:

• Orf78:	4	FLEAFFVEYGYAAVFFVLVICGFGVPIPEDLTLVTGGVISGM--GYTNPHIMFAVGMLGV	61
  		FL  FF EYGY AV FVL+ICGFGVPIPED+TLV+GGVI+G+     N H+M  V M+GV
  DedA:	20	FLIGFFTEYGYWAVLFVLIICGFGVPIPEDITLVSGGVIAGLYPENVNSHLMLLVSMIGV	79
  Orf78:	62	LVGDGIMFAAGRIWGQXXLXFXPIAXIMTPKRYEQVQEKFDKYGNWVLFVARFLPGLRTA	121
  		L GD  M+  GRI+G   L F PI  I+T  R   V+EKF +YGN VLFVARFLPGLR
  DedA:	80	LAGDSCMYWLGRIYGTKILRFRPIRRIVTLQRLRMVREKFSQYGNRVLFVARFLPGLRAP	139
  Orf78:	122	VFVTAGISRKVSYLRFIIMDGLAA	145
  		+++ +GI+R+VSY+RF+++D  AA
  DedA:	140	IYMVSGITRRVSYVRFVLIDFCAA	163

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF78 shows 93.8% identity over a 145aa overlap with an ORF (ORF78a) from strain A of N. meningitidis:
• 
• The complete length ORF78a nucleotide sequence <SEQ ID 723> is:

• 1	ATGTTTGCCC TTTTGGAAGC CTTTTTTGTC GAATACGGCT
  	ATGCGGCCGT
  51	GTTTTTCGTT TTGGTCATCT GCGGTTTCGG CGTGCCGATT
  	CCCGAGGATT
  101	TGACCTTGGT AACAGGCGGC GTGATTTCGG GTATGGGTTA
  	TACCAATCCG
  151	CATATTATGT TTGCAGTCGG TATGCTCGGC GTATTGGTCG
  	GGGACGGCAT
  201	CATGTTCGCC GCCGGACGCA TCTGGGGGCA GAAAATCCTC
  	AAGTTCAAAC
  251	CGATTGCGCG CATCATGACG CCGAAACGTT ACGCACAGGT
  	TCAGGAAAAA
  301	TTCGACAAAT ACGGCAACTG GGTGTTATTT GTCGCTCGTT
  	TCCTGCCCGG
  351	TTTGCGGACT GCCGTTTTCG TTACCGCCGG CATCAGCCGC
  	AAAGTATCGT
  401	ATCTGCGCTT TCTGATTATG GACGGGCTTG CCGCGCTGAT
  	TTCCGTGCCC
  451	GTTTGGATTT ACTTGGGCGA GTACGGCGCG CACAACATCG
  	ATTGGCTGAT
  501	GGCGAAAATG CACAGCCTGC AATCCGGCAT CTTCATCGCA
  	TTGGGCGTGC
  551	TGGCGGCGGC GCTGGCGTGG TTCTGGTGGC GCAAACGCCG
  	ACATTATCAG
  601	CTTTACCGCG CACAATTGAG CGAAAAACGC GCCAAACGCA
  	AGGCGGAAAA
  651	GGCAGCGAAA AAAGCGGCAC AGAAGCAGCA GTAA

• This encodes a protein having amino acid sequence <SEQ ID 724>:

• 1	MFALLEAFFV EYGYAAVFFV LVICGFGVPI PEDLTLVTGG
  	VISGMGYTNP
  51	HIMFAVGMLG VLVGDGIMFA AGRIWGQKIL KFKPIARIMT
  	PKRYAQVQEK
  101	FDKYGNWVLF VARFLPGLRT AVFVTAGISR KVSYLRFLIM
  	DGLAALISVP
  151	VWIYLGEYGA HNIDWLMAKM HSLQSGIFIA LGVLAAALAW
  	FWWRKRRHYQ
  201	LYRAQLSEKR AKRKAEKAAK KAAQKQQ*

• ORF78a and ORF78-1 show 89.0% identity in 227 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF78 shows 97.4% identity over 38 aa overlap with a predicted ORF (ORF78ng) from N. gonorrhoeae:
• 
• The ORF78ng nucleotide sequence <SEQ ID 725> is predicted to encode a protein comprising amino acid sequence <SEQ ID 726>:

• 1	..YPVLFVARFL PGLRTAVFVT AGISRKVSYL RFLIMDGLAA
  	LISVPVWIYL
  51	GEYGAHNIDW LMAKMHSLQS GIFIALGVLA AALAWF WWRK
  	RRHYQLYRAQ
  101	LSEKRAKRKA EKAAKKAAQK QQ*

• Further work revealed the complete gonococcal nucleotide sequence <SEQ ID 727>:

• 1	atgtttgccc tttTggaagc CTTTTTTGTC GAAtacggCt
  	atgcGGCCGT
  51	GTTTTTCGTT TTGGTCATCT GCGGTTTCGG CGTGCCGATT
  	CCCGAAGATT
  101	TGACCTTGGT AACGGGCGGC GTGATTTCGG GTATGGGTTA
  	TACCAATCCG
  151	CATATTATGT TTGCGGTCGG TATGCTCGGC GTGTTGGCGG
  	GCGACGGCGT
  201	GATGTTTGCC GCCGGACGCA TCTGGGGGCA GAAAATCCTC
  	AAGTTCAAAC
  251	CGATTGCGCG CATCATGACG CCGAAACGTT ACGCGCAGGT
  	TCAGGAAAAA
  301	TTCGACAAAT ACGGCAACTG GGTTCTGTTT GTCGCCCGTT
  	TCCTGCCGGG
  351	TTTGCGGACT GCCGTTTTCG TTACCGCCGG CATCAGCCGC
  	AAAGTATCGT
  401	ATCTGCGCTT TCTGATTATG GACGGGCTGG CCGCGCTGAT
  	TTCCGTGCCC
  451	GTTTGGATTT ACTTGGGCGA GTACGGCGCG CACAACATCG
  	ATTGGCTGAT
  501	GGCGAAAATG CACAGCCTGC AATCGGGCAT CTTCATCGCA
  	TTGGGCGTGC
  551	TGGCGGCGGC GCTGGCGTGG TTCTGGTGGC GCAAACGCCG
  	ACATTATCAG
  601	CTTTACCGCG CACAATTGAG CGAAAAACGC GCCAAACGCA
  	AGGCGGAAAA
  651	GGCAGCGAAA AAAGCGGCAC AGAAGCAGCA GTAa

• This corresponds to the amino acid sequence <SEQ ID 728; ORF78ng-1>:

• 1	MFALLEAFFV EYGYAAVFFV LVICGFGVPI PEDLTLVTGG
  	VISGMGYTNP
  51	HIMFAVGMLG VLAGDGVMFA AGRIWGQKIL KFKPIARIMT
  	PKRYAQVQEK
  101	FDKYGNWVLF VARFLPGLRT AVFVTAGISR KVSYLRFLIM
  	DGLAALISVP
  151	VWIYLGEYGA HNIDWLMAKM HSLQSGIFIA LGVLAAALAW
  	FWWRKRRHYQ
  201	LYRAQLSEKR AKRKAEKAAK KAAQKQQ*

• ORF78ng-1 and ORF78-1 show 88.1% identity in 227 aa overlap:
• 
• Furthermore, orf78ng-1 shows homology to the dedA protein from H. influenzae:

• sp|P45280|YG29_HAEIN HYPOTHETICAL PROTEIN HI1629 >gi|1073983|pir||D64133
  dedA protein (dedA) homolog - Haemophilus influenzae (strain Rd KW20)
  >gi|1574476 (U32836) dedA protein (dedA) [Haemophilus influenzae]
  Length = 212 Score = 223 bits (563), Expect = 7e−58
  Identities = 108/182 (59%), Positives = 140/182 (76%), Gaps = 2/182 (1%)

  Query:	5	LEAFFVEYGYAAVFFVLVICGFGVPIPEDLTLVTGGVISGM--GYTNPHIMFAVGMLGVL	62
  		L  FF EYGY AV FVL+ICGFGVPIPED+TLV+GGVI+G+     N H+M  V M+GVL
  Sbjct:	21	LIGFFTEYGYWAVLFVLIICGFGVPIPEDITLVSGGVIAGLYPENVNSHLMLLVSMIGVL	80
  Query:	63	AGDGVMFAAGRIWGQKILKFKPIARIMTPKRYAQVQEKFDKYGNWVLFVARFLPGLRTAV	122
  		AGD  M+  GRI+G KIL+F+PI RI+T +R   V+EKF +YGN VLFVARFLPGLR  +
  Sbjct:	81	AGDSCMYWLGRIYGTKILRFRPIRRIVTLQRLRMVREKFSQYGNRVLFVARFLPGLRAPI	140
  Query:	123	FVTAGISRKVSYLRFLIMDGLAALISVPVWIYLGEYGAHNIDWLMAKMHSLQSGIFIALG	182
  		++ +GI+R+VSY+RF+++D  AA+ISVP+WIYLGE GA N+DWL  ++   Q  I+I +G
  Sbjct:	141	YMVSGITRRVSYVRFVLIDFCAAIISVPIWIYLGELGAKNLDWLHTQIQKGQIVIYIFIG	200
  Query:	183	VL	184
  		L
  Sbjct:	201	YL	202

• Based on this analysis, including the presence of putative transmembrane domains, it is predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 87
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 729>:

• 1	ATGAAAAAAT TATTGGCGGC CGTGATGATG GCAGGTTTGG
  	CAGGCGCGGT
  51	TTCCGCCGCC GGAGTCCACG TTGAGGACGG CTGGGCGCGC
  	ACCACCGTCG
  101	AAGGTATGAA AATAGGCGGC GCGTTCATGA AAATCCACAA
  	CGACGAAGCC
  151	AAACAAGACT TTTTGCTCGG CGGAAGCAGC CCCGTTGCCG
  	ACCGCGTCGA
  201	AGTGCATACC CACATCAACG ACAACGGCGT GATGCGGATG
  	CGCGAAGTCG
  251	AAGGCGGCGT GCCTTTGGAA GCGAAATCCG TTACCGAACT
  	CAAACCCGGC
  301	AGCTATCATG TGATGTTTAT GGGTTTGAAA AAACAATTAA
  	AAGAGGGCGA
  351	TAAAATTCCC GTTACCCTGA AATTTAAAAA CGCCAAAGCG
  	CAAACCGTCC
  401	AACTGGAAGT CAAAATCGCG CCGATGCCGG CAATGAACCA
  	C...

• This corresponds to the amino acid sequence <SEQ ID 730; ORF79>:

• 1	MKKLLAAVMM AGLAGAVSAA GVHVEDGWAR TTVEGMKIGG
  	AFMKIHNDEA
  51	KQDFLLGGSS PVADRVEVHT HINDNGVMRM REVEGGVPLE
  	AKSVTELKPG
  101	SYHVMFMGLK KQLKEGDKIP VTLKFKNAKA QTVQLEVKIA
  	PMPAMNH..

• Further work revealed the complete nucleotide sequence <SEQ ID 731>:

• 1	ATGAAAAAAT TATTGGCGGC CGTGATGATG GCAGGTTTGG
  	CAGGCGCGGT
  51	TTCCGCCGCC GGAGTCCACG TTGAGGACGG CTGGGCGCGC
  	ACCACCGTCG
  101	AAGGTATGAA AATAGGCGGC GCGTTCATGA AAATCCACAA
  	CGACGAAGCC
  151	AAACAAGACT TTTTGCTCGG CGGAAGCAGC CCCGTTGCCG
  	ACCGCGTCGA
  201	AGTGCATACC CACATCAACG ACAACGGCGT GATGCGGATG
  	CGCGAAGTCG
  251	AAGGCGGCGT GCCTTTGGAA GCGAAATCCG TTACCGAACT
  	CAAACCCGGC
  301	AGCTATCATG TGATGTTTAT GGGTTTGAAA AAACAATTAA
  	AAGAGGGCGA
  351	TAAAATTCCC GTTACCCTGA AATTTAAAAA CGCCAAAGCG
  	CAAACCGTCC
  401	AACTGGAAGT CAAAATCGCG CCGATGCCGG CAATGAACCA
  	CGGTCATCAC
  451	CACGGCGAAG CGCATCAGCA CTAA

• This corresponds to the amino acid sequence <SEQ ID 732; ORF79-1>:

• 1	MKKLLAAVMM AGLAGAVSAA GVHVEDGWAR TTVEGMKIGG
  	AFMKIHNDEA
  51	KQDFLLGGSS PVADRVEVHT HINDNGVMRM REVEGGVPLE
  	AKSVTELKPG
  101	SYHVMFMGLK KQLKEGDKIP VTLKFKNAKA QTVQLEVKIA
  	PMPAMNHGHH
  151	HGEAHQH*

• Computer analysis of this amino acid sequence revealed a putative leader peptide and also gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF79 shows 94.6% identity over a 147aa overlap with an ORF (ORF79a) from strain A of N. meningitidis:
• 
• The complete length ORF79a nucleotide sequence <SEQ ID 733> is:

• 1	ATGAAANAAC TATTGGCAGC CGTGATGATG GCAGGTTTGG
  	CAGGCGCGGT
  51	TTCCGCCGCC GGAATCCACG TTGAGGACGG CTGGGCGCGC
  	ACCACCGTCG
  101	AAGGTATGAA AATGGGCGGC GCGTTCATGA AAATCCACAA
  	CGACGAAGCC
  151	AAACAAGACT TTTTGCTCGG CGGAAGCAGC CCTGTTGCCG
  	ACCGCGTCGA
  201	AGTGCATACC CATATCAATG ATAACGGTGT GATGCGGATG
  	CGCGAAGTCG
  251	AAGGCGGCGT GCCTTTGGAG GCGAAATCCG TTACCGAACT
  	CAAACCCGGC
  301	AGCTATCATG TCATGTTTAT GGGTNTGAAA AAACAATTAA
  	AAGANGGCGA
  351	CAAGATTCCC GTTACCCTGA AATTTAAAAA CGCCAAAGCA
  	CAAACCGTCC
  401	AACTGGAAGT CAAAACCGCG CCGATGTCGG CAATGGACCA
  	CGGTCATCAC
  451	CACGGCGAAG CGCATCAGCA CTAA

• This encodes a protein having amino acid sequence <SEQ ID 734>:

• 1	MKXLLAAVMM AGLAGAVSAA GIHVEDGWAR TTVEGMKMGG
  	AFMKIHNDEA
  51	KQDFLLGGSS PVADRVEVHT HINDNGVMRM REVEGGVPLE
  	AKSVTELKPG
  101	SYHVMFMGXK KQLKXGDKIP VTLKFKNAKA QTVQLEVKTA
  	PMSAMDHGHH
  151	HGEAHQH*

• ORF79a and ORF79-1 show 94.9% identity in 157 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF79 shows 96.1% identity over 76 aa overlap with a predicted ORF (ORF79ng) from N. gonorrhoeae:
• 
• An ORF79ng nucleotide sequence <SEQ ID 735> was predicted to encode a protein comprising amino acid sequence <SEQ ID 736>:

• 1	..INDNGVMRMR EVKGGVPLEA KSVTELKPGS YHVMFMGLKK
  	QLKEGDKIPV
  51	TLKFKNAKAQ TVQLEVKTAP MSAMNHGHHH GEAHQH*

• Further work revealed the complete gonococcal DNA sequence <SEQ ID 737>:

• 1	ATGAAAAAAT TATTGGCAGC CGTGATGATG GCAGGTTTGG
  	CAGGCGCGGT
  51	TTccgccgCc GGagTccAtG TCGAggACGG CTGGGCGCGc
  	accaCTGtcg
  101	aaggtATgaa aatggGCGGC GCgttCATga aaATCCACAA
  	CGACGaaGcc
  151	atacaaGACt ttgtgcTCgg CGGaagcatg cccgttgccg
  	accgcGTCGA
  201	AGTGCAtaca cacATCAACG ACAACGGCGT GATGCGTATG
  	CGCGAAGTCA
  251	AAGGCGGCGT GCCTTTGGAG GCGAAATCCG TTACCGAACT
  	CAAACCCGGC
  301	AGCTATCACG TGATGTTTAT GGGTTTGAAA AAACAACTGA
  	AAGAGGGCGA
  351	CAAGATTCCC GTTACCCTGA AATTTAAAAA CGCCAAAGCG
  	CAAACCGTCC
  401	AACTGGAAGT CAAAACCGCG CCGATGTCGG CAATGAACCA
  	CGGTCATCAC
  451	CACGGCGAAG CGCATCAGCA CTAA

• This corresponds to the amino acid sequence <SEQ ID 738; ORF79ng-1>:

• 1	MKKLLAAVMM AGLAGAVSAA GVHVEDGWAR TTVEGMKMGG
  	AFMKIHNDEA
  51	IQDFVLGGSM PVADRVEVHT HINDNGVMRM REVKGGVPLE
  	AKSVTELKPG
  101	SYHVMFMGLK KQLKEGDKIP VTLKFKNAKA QTVQLEVKTA
  	PMSAMNHGHH
  151	HGEAHQH*

• ORF79ng-1 and ORF79-1 show 95.5% identity in 157 aa overlap:
• 
• Furthermore, ORF79ng-1 shows significant homology to a protein from Aquifex aeolicus:

• gi|2983695 (AE000731) putative protein [Aquifex aeolicus] Length = 151
  Score = 63.6 bits (152), Expect = 6e−10
  Identities = 38/114 (33%), Positives = 58/114 (50%), Gaps = 1/114 (0%)

  Query:	24	VEDGWARTTVEGMKMGGAFMKIHNDEAIQDFVLGGSMPVADRVEVHTHINDNGVMRMREV	83
  		V+  W      G       M I N+    D+++G    +A RVE+H  + +N V +M
  Sbjct:	27	VKHPWVMEPPPGPNTTMMGMIIVNEGDEPDYLIGAKTDIAQRVELHKTVIENDVAKMVPQ	86
  Query:	84	KGGVPLEAKSVTELKPGSYHVMFMGLKKQLKEGDKIPVTLKFKNAKAQTVQLEV	137
  		+  + +  K   E K   YHVM +GLKK++KEGDK+ V L F+ +   TV+  V
  Sbjct:	87	ER-IEIPPKGKVEFKHHGYHVMIIGLKKRIKEGDKVKVELIFEKSGKITVEAPV	139

• Based on this analysis, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF79-1 (15.6 kDa) was cloned in the pET vector and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 18A shows the results of affinity purification of the His-fusion protein. Purified His-fusion protein was used to immunise mice, whose sera were used for ELISA (positive result) and FACS analysis (FIG. 18B) These experiments confirm that ORF79-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 88
• The following DNA sequence, believed to be complete, was identified in N. meningitidis <SEQ ID 739>:

• 1	ATGACGGTAA CTGCGGCCGA AGGCGGCAAA GCTGCCAAGG
  	CGTTAAAAAA
  51	ATATCTGATT ACGGGCATTT TGGTCTGGCT GCCGATTGCG
  	GTAACGGTTT
  101	GGGTGGTTTC CTATATCGTT TCCGCGTCCG ATCAGCTCGT
  	CAACCTGCTG
  151	CCGAAGCAAT GGCGGCCGCA ATATGTTTTG GGGTTTAATA
  	TCCCGGGGCT
  201	GGGCGTTATC GTTGCCATTG CCGTATTGTT TGTAACCGGA
  	TTGTTTGCCG
  251	CCAACGTATT GGGTCGGCAG ATCCTCGCCG CGTGGGACAG
  	CCTGTTGGGG
  301	CGGATTCCGG TTGTGAAAtC CATCTATTCG AGTGTGAAAA
  	AAGTATCCGA
  351	ATacgTGCTG TCCGACAGCA GCCGTTCGTT TAAAACGCCG
  	GTACTCGTGC
  401	CGTTTCCCCA GCCCGGTATT TGGACGATyG CTTTCGTGTC
  	AGGGCAGGTG
  451	TCGAATGCGG TTAAGGCCGC ATTGCCGAAs GACGGCGATT
  	ATCTTTCCGT
  501	GTATGTTCCG ACCACGCCGA ATCCGACCGG CGGTTACTAT
  	ATTATGGTAA
  551	AGAAAAGCGA TGTGCGCGAA CTCGATATGA GCGTGGACGA
  	AsCATTGAAA
  601	TATGTGATTT CGCTGGGTAT GGTCATCCCT GACGACCTGC
  	CCGTCAAAAC
  651	ATTGGCAsGA CCTATGCCGT CTGAAAAGGC GGATTTGCCC
  	GAACAACAAT
  701	AA

• This corresponds to the amino acid sequence <SEQ ID 740; ORF98>:

• 1	MTVTAAEGGK AAKALKKYLI TGILVWLPIA VTVWVVSYIV
  	SASDQLVNLL
  51	PKQWRPQYVL GFNIPGLGVI VAIAVLFVTG LFAANVLGRQ
  	ILAAWDSLLG
  101	RIPVVKSIYS SVKKVSEYVL SDSSRSFKTP VLVPFPQPGI
  	WTIAFVSGQV
  151	SNAVKAALPX DGDYLSVYVP TTPNPTGGYY IMVKKSDVRE
  	LDMSVDEXLK
  201	YVISLGMVIP DDLPVKTLAX PMPSEKADLP EQQ*

• Further work revealed the complete nucleotide sequence <SEQ ID 741>:

• 1	ATGACGGAAC nTGCGGCCGA AGGCGGCAAA GCTGCCAArG
  	CGTTAAAAAA
  51	ATATCTGATT ACGGGCATTT TGGTCTGGCT GCCGATTGCG
  	GTAACGGTTT
  101	GGGTGGTTTC CTATATCGTT TCCGCGTCCG ATCAGCTCGT
  	CAACCTGCTG
  151	CCGAAGCAAT GGCGGCCGCA ATATGTTTTG GGGTTTAATA
  	TCCCGGGGCT
  201	GGGCGTTATC GTTGCCATTG CCGTATTGTT TGTAACCGGA
  	TTGTTTGCCG
  251	CCAACGTATT GGGTCGGCAG ATCCTCGCCG CGTGGGACAG
  	CCTGTTGGGG
  301	CGGATTCCGG TTGTGAAATC CATCTATTCG AGTGTGAAAA
  	AAGTATCCGA
  351	ATCGCTGCTG TCCGACAGCA GCCGTTCGTT TAAAACGCCG
  	GTACTCGTGC
  401	CGTTTCCCCA GCCCGGTATT TGGACGATTG CTTTCGTGTC
  	AGGGCAGGTG
  451	TCGAATGCGG TTAAGGCCGC ATTGCCGAAG GACGGCGATT
  	ATCTTTCCGT
  501	GTATGTTCCG ACCACGCCGA ATCCGACCGG CGGTTACTAT
  	ATTATGGTAA
  551	AGAAAAGCGA TGTGCGCGAA CTCGATATGA GCGTGGACGA
  	AGCATTGAAA
  601	TATGTGATTT CGCTGGGTAT GGTCATCCCT GACGACCTGC
  	CCGTCAAAAC
  651	ATTGGCAGGA CCTATGCCGT CTGAAAAGGC GGATTTGCCC
  	GAACAACAAT
  701	AA

• This corresponds to the amino acid sequence <SEQ ID 742; ORF98-1>:

• 1	MTEXAAEGGK AAKALKKYLI TGILVWLPIA VTVWVVSYIV
  	SASDQLVNLL
  51	PKQWRPQYVL GFNIPGLGVI VAIAVLFVTG LFA ANVLGRQ
  	ILAAWDSLLG
  101	RIPVVKSIYS SVKKVSESLL SDSSRSFKTP VLVPFPQPGI
  	WTIAFVSGQV
  151	SNAVKAALPK DGDYLSVYVP TTPNPTGGYY IMVKKSDVRE
  	LDMSVDEALK
  201	YVISLGMVIP DDLPVKTLAG PMPSEKADLP EQQ*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF98 shows 96.1% identity over a 233aa overlap with an ORF (ORF98a) from strain A of N. meningitidis:
• 
• The complete length ORF98a nucleotide sequence <SEQ ID 743> is:

• 1	ATGACGGAAC CTGCGGCCGA AGGCGGCAAA GCTGCCAAGG
  	CGTTAAAAAA
  51	ATATCTGATT ACGGGCATTT TGGTCTGGCT GCCGATTGCG
  	GTAACGGTTT
  101	GGGTGGTTTC CTATATCGTT TCCGCGTCCG ATCAGCTCGT
  	CAACCTGCTG
  151	CCGAAGCAAT GGCGGCCGCA ATATGTTTTG GGGTTTAATA
  	TCCCGGGGCT
  201	GGGCGTTATC GTTGCCATTG CCGTATTGTT TGTAACCGGA
  	TTATTTGCCG
  251	CAAACGTATT GGGCCGGCAG ATTCTTGCCG CGTGGGACAG
  	CTTGTTGGGG
  301	CGGATTCCGG TTGTGAAGTC CATCTATTCG AGTGTGAAAA
  	AAGTATCCGA
  351	NTCGTTGCTG TCCGACAGCA GCCGTTCGTT TAAAACACCA
  	GTACTCGTGC
  401	CGTTTCCCCA ATCGGGTATT TGGACAATCG CATTCGTGTC
  	CGGTCAGGTG
  451	TCGAATGCGG TTAAGGCCGC ATTGCCGAAG GACGGCGATT
  	ATCTTTCCGT
  501	GTATGTTCCG ACCACGCCGA ATCCGACCGG CGGTTACTAT
  	ATTATGGTAA
  551	AGAAAAGCGA TGTGCGCGAA CTCGATATGA GCGTGGACGA
  	AGCGTTGAAA
  601	TATGTGATTT CGCTGGGTAT GGTCATCCCT GACGACCTGC
  	CCGTCAAAAC
  651	ATTGGCAGGA CCTATGCCGT CTGAAAAGGC GGATTTGCCC
  	GAACAACAAT
  701	AA

• This encodes a protein having amino acid sequence <SEQ ID 744>:

• 1	MTEPAAEGGK AAKALKKYLI TGILVWLPIA VTVWVVSYIV
  	SASDQLVNLL
  51	PKQWRPQYVL GFNIPGLGVI VAIAVLFVTG LFA ANVLGRQ
  	ILAAWDSLLG
  101	RIPVVKSIYS SVKKVSXSLL SDSSRSFKTP VLVPFPQSGI
  	WTIAFVSGQV
  151	SNAVKAALPK DGDYLSVYVP TTPNPTGGYY IMVKKSDVRE
  	LDMSVDEALK
  201	YVISLGMVIP DDLPVKTLAG PMPSEKADLP EQQ*

• ORF98a and ORF98-1 show 98.7% identity in 233 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF98 shows 95.3% identity over a 233 aa overlap with a predicted ORF (ORF98ng) from N. gonorrhoeae:
• 
• The complete length ORF98ng nucleotide sequence <SEQ ID 745> is predicted to encode a protein having amino acid sequence <SEQ ID 746>:

• 1	MTEPAAEGGK AAKALKKYLI TGILVWLPIA VTVWVVSYIV
  	SASDQLVNLL
  51	PKQWRPQYVL GFNIPGLGVI VAIAVLFVTG LFA ANVLGRQ
  	ILAAWDSLLX
  101	RIPVVKSIYS SVKKVSESLL SDSSRSFKTP VLVPFPQSGI
  	WTIAFVSGQV
  151	SNAVKAALPQ DGDYLSVYVP TTPNPTGGYY IMVKKSDVRE
  	LDMSVDEALK
  201	YVISLGMVIP DDLPVKTLAG PMPPEKAELP EQQ*

• Further work revealed the complete nucleotide sequence <SEQ ID 747>:

• 1	ATGACGGAAC CTGCGGCCGA AGGCGGCAAA GCTGCCAAGG
  	CGTTAAAAAA
  51	ATATCTGATT ACAGGCATTT TGGTCTGGCT GCCGATTGCG
  	GTAACGGTTT
  101	GGGTGGTTTC CTATATCGTT TCCGCGTCCG ACCAGCTTGT
  	CAACCTGCTG
  151	CCGAAGCAAT GGCGGCCGCA ATATGTTTTG GGGTTTAATA
  	TCCCCGGGCT
  201	CGGCGTTATT GTTGCCATTG CCGTATTGTT TGTAACCGGA
  	TTATTTGCCG
  251	CAAACGTGTT GGGCCGGCAG ATTCTTGCCG CGTGGGACAG
  	CCTGTTgggg
  301	cggaTTCCGG TTGTCAAATC CATCTATTCG AGTGTGAAAA
  	AAGTATCCGA
  351	ATCGCTGCTG TCCGACAGCA GCCGTTCGTT TAAAACGCCG
  	GTACTCGTGC
  401	CGTTTCCCCA ATCGGGTATT TGGACAATCG CATTCGTGTC
  	CGGTCAGGTG
  451	TCGAATGCGG TTAAGGCCGC ATTGCCGCAG GATGGCGATT
  	ATCTTTCCGT
  501	GTATGTCCCG ACCACGCCCA ACCCGACCGG CGGTTACTAT
  	ATTATGGTAA
  551	AGAAAAGCGA TGTGCGCGAA CTCGATATGA GCGTGGACGA
  	AGCGTTGAAA
  601	TATGTGATTT CGCTGGGTAT GGTCATCCCT GACGACCTGC
  	CCGTCAAAAC
  651	ATTGGCAGGA CCTATGCCGC CTGAAAAGGC GGAGTTGCCC
  	GAACAACAAT
  701	AA

• This corresponds to the amino acid sequence <SEQ ID 748; ORF98ng-1>:

• 1	MTEPAAEGGK AAKALKKYLI TGILVWLPIA VTVWVVSYIV
  	SASDQLVNLL
  51	PKQWRPQYVL GFNIPGLGVI VAIAVLFVTG LFA ANVLGRQ
  	ILAAWDSLLG
  101	RIPVVKSIYS SVKKVSESLL SDSSRSFKTP VLVPFPQSGI
  	WTIAFVSGQV
  151	SNAVKAALPQ DGDYLSVYVP TTPNPTGGYY IMVKKSDVRE
  	LDMSVDEALK
  201	YVISLGMVIP DDLPVKTLAG PMPPEKAELP EQQ*

• ORF98ng-1 and ORF98-1 show 97.9% identity in 233 aa overlap:
• 
• Based on this analysis, including the fact that the putative transmembrane domains in the gonococcal protein are identical to the sequences in the meningococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 89
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 749>:

• 1	ATgAAAACGG TAGTCTGGAT TGTCGTCCTG TTTGCCGCCG
  	CCGTCGGACT
  51	GGCGCTGGCT TCGGGCATTT ACACCGGCGA CGTGTATATC
  	GTACTCGGAC
  101	AGACCATGCT CAGAATCAAC CTGCACGCCT TTGTGTTAGG
  	TTCGCTGATT
  151	GCCGTCGTGG TGTGGTATTT CTTGTTTAAA TTCATTATCG
  	GsGgTACTCA
  201	ATATCCCCGA AAAGATGCAG CGTTTCGGTT CGGCnCGTAA
  	AGGCCkCAAG
  251	ssCGsGCTTG CCTTGAACAA GGCGGGTTTG GCGTATTTTG
  	AAGGGCGTTT
  301	TGAAAAGGCG GAACTAGAAG CCTCACGCGT GTTGGTCAAC
  	AAAGtAGGCC
  351	GaGAGACAAC CGGACTTTGG CATTGATGCT GrGCGCGCAC
  	GCCGCCGGAC
  401	AGATGGAAAA CATCGAssTG CGCGACCGTT ATCTTGCGGA
  	AATCGCCAAA
  451	CTGCCGGAAA AACAGCAGCT TTCCCGTTAT CTTTTGTTGG
  	CGGAATCGGC
  501	GTTGAACCGG CGCGATTACG AAGCGGCGGA AGCCAATCTT
  	CATGCGGCGG
  551	CGAAGATGAA TGCCAACCTT ACGCGCCTCG TGCGTCTGCA
  	.ATTCGTTAC
  601	GCTTTCGACA GGGGCGACGC GTTGCAGGTT CTGGCAAAAA
  	CCGAAAAACT
  651	TTCCAAGGCG GGCGCGTTGG GCAAATCGGA AATGGAACGG
  	TATCAAAATT
  701	GGGCATATCC GTCGCCAGCT GGCGGATGCT GCCGATGCCG
  	CCGCTTTGAA
  751	AACCTGCCTG AAGCGGATTC CCGACAGCCT CAAAAACGGG
  	GAATTGAGCG
  801	TATCGGTTGC GGAAAAGTAC GAACGTTTGG GACTGTATGC
  	CGATGCGGTC
  851	AAATGGGTCA AACAGCATTA TCCGCAsAAC CGCCGCCCCG
  	AGCTTTTGGA
  901	AGCCTTTGTC GAAAGCGTGC GCTTTTTGGG CGAGCGCGAA
  	CAGCAGAAAG
  951	CCATCGATTT TGCCGATGCT TGGCTGAAAG AACAGCCCGA
  	TAACGCGCTT
  1001	CTGCTGATGT ATCTCGGTCG GCTCGCCTTC GGCCGCAAAC
  	TTTGGGGCAA
  1051	GGCAAAAGGC TACCTTGAAG CGAGCATTGC ATTAAAGCCG
  	AGTATTTCCG
  1101	CGCGTTTGGT TCTAACAAAG GTTTTCGACG AAATCGGAGA
  	ACCGCAGAAG
  1151	GCGGAGGCGC AC...

• This corresponds to the amino acid sequence <SEQ ID 750; ORF100>:

• 1	MKTVVWIVVL FAAAVGLALA SGIYTGDVYI VLGQTMLRIN
  	LHAFVLGSLI
  51	AVVVWYFLFK FIIGVLNIPE KMQRFGSARK GXKXXLALNK
  	AGLAYFEGRF
  101	EKAELEASRV LVNKVGRDNR TLALMLXAHA AGQMENIXXR
  	DRYLAEIAKL
  151	PEKQQLSRYL LLAESALNRR DYEAAEANLH AAAKMNANLT
  	RLVRLXIRYA
  201	FDRGDALQVL AKTEKLSKAG ALGKSEMERY QNWAYRRQLA
  	DAADAAALKT
  251	CLKRIPDSLK NGELSVSVAE KYERLGLYAD AVKWVKQHYP
  	XNRRPELLEA
  301	FVESVRFLGE REQQKAIDFA DAWLKEQPDN ALLLMYLGRL
  	AFGRKLWGKA
  351	KGYLEASIAL KPSISARLVL TKVFDEIGEP QKAEAH...

• Further work revealed the complete nucleotide sequence <SEQ ID 751>:

• 1	ATGAAAACGG TAGTCTGGAT TGTCGTCCTG TTTGCCGCCG
  	CCGTCGGACT
  51	GGCGCTGGCT TCGGGCATTT ACACCGGCGA CGTGTATATC
  	GTACTCGGAC
  101	AGACCATGCT CAGAATCAAC CTGCACGCCT TTGTGTTAGG
  	TTCGCTGATT
  151	GCCGTCGTGG TGTGGTATTT CTTGTTTAAA TTCATTATCG
  	GCGTACTCAA
  201	TATCCCCGAA AAGATGCAGC GTTTCGGTTC GGCGCGTAAA
  	GGCCGCAAGG
  251	CCGCGCTTGC CTTGAACAAG GCGGGTTTGG CGTATTTTGA
  	AGGGCGTTTT
  301	GAAAAGGCGG AACTAGAAGC CTCACGCGTG TTGGTCAACA
  	AAGAGGCCGG
  351	AGACAACCGG ACTTTGGCAT TGATGCTGGG CGCGCACGCC
  	GCCGGACAGA
  401	TGGAAAACAT CGAGCTGCGC GACCGTTATC TTGCGGAAAT
  	CGCCAAACTG
  451	CCGGAAAAAC AGCAGCTTTC CCGTTATCTT TTGTTGGCGG
  	AATCGGCGTT
  501	GAACCGGCGC GATTACGAAG CGGCGGAAGC CAATCTTCAT
  	GCGGCGGCGA
  551	AGATGAATGC CAACCTTACG CGCCTCGTGC GTCTGCAACT
  	TCGTTACGCT
  601	TTCGACAGGG GCGACGCGTT GCAGGTTCTG GCAAAAACCG
  	AAAAACTTTC
  651	CAAGGCGGGC GCGTTGGGCA AATCGGAAAT GGAACGGTAT
  	CAAAATTGGG
  701	CATACCGCCG CCAGCTGGCG GATGCTGCCG ATGCCGCCGC
  	TTTGAAAACC
  751	TGCCTGAAGC GGATTCCCGA CAGCCTCAAA AACGGGGAAT
  	TGAGCGTATC
  801	GGTTGCGGAA AAGTACGAAC GTTTGGGACT GTATGCCGAT
  	GCGGTCAAAT
  851	GGGTCAAACA GCATTATCCG CACAACCGCC GCCCCGAGCT
  	TTTGGAAGCC
  901	TTTGTCGAAA GCGTGCGCTT TTTGGGCGAG CGCGAACAGC
  	AGAAAGCCAT
  951	CGATTTTGCC GATGCTTGGC TGAAAGAACA GCCCGATAAC
  	GCGCTTCTGC
  1001	TGATGTATCT CGGTCGGCTC GCCTACGGCC GCAAACTTTG
  	GGGCAAGGCA
  1051	AAAGGCTACC TTGAAGCGAG CATTGCATTA AAGCCGAGTA
  	TTTCCGCGCG
  1101	TTTGGTTCTA GCAAAGGTTT TCGACGAAAT CGGAGAACCG
  	CAGAAGGCGG
  1151	AGGCGCAGCG CAACTTGGTT TTGGAAGCCG TCTCCGATGA
  	CGAACGTCAC
  1201	GCAGCGTTAG AGCAGCATAG CTGA

• This corresponds to the amino acid sequence <SEQ ID 752; ORF100-1>:

• 1	MKTVVWIVVL FAAAVGLALA SGIYTGDVYI VLGQTMLRIN
  	LHAFVLGSLI
  51	AVVVWYFLFK FIIGVLNIPE KMQRFGSARK GRKAALALNK
  	AGLAYFEGRF
  101	EKAELEASRV LVNKEAGDNR TLALMLGAHA AGQMENIELR
  	DRYLAEIAKL
  151	PEKQQLSRYL LLAESALNRR DYEAAEANLH AAAKMNANLT
  	RLVRLQLRYA
  201	FDRGDALQVL AKTEKLSKAG ALGKSEMERY QNWAYRRQLA
  	DAADAAALKT
  251	CLKRIPDSLK NGELSVSVAE KYERLGLYAD AVKWVKQHYP
  	HNRRPELLEA
  301	FVESVRFLGE REQQKAIDFA DAWLKEQPDN ALLLMYLGRL
  	AYGRKLWGKA
  351	KGYLEASIAL KPSISARLVL AKVFDEIGEP QKAEAQRNLV
  	LEAVSDDERH
  401	AALEQHS*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF100 shows 93.5% identity over a 386aa overlap with an ORF (ORF100a) from strain A of N. meningitidis:
• 
• The complete length ORF100a nucleotide sequence <SEQ ID 753> is:

• 1	ATGAAAACGG TAGTCTGGAT TGTCGTCCTG TTTGCCGCCG
  	CNNTCGGGCT
  51	GGCATTGGCG TCGGGCATTN ACACCGGCGA CGTGTATATC
  	GTACTCGGAC
  101	AGACCATGCT CAGAATCAAC CTGCACGCCT TTGTGTTAGG
  	TTCGCTGATT
  151	GCCGTCGTGG TGTGGTATTT CCTGTTCAAA TTCATCATCG
  	GCGTACTCAA
  201	TANCCCCGAA AAGATGCAGC GTTTCGGTTC GGCGCGTAAA
  	GGCCGCAAGG
  251	CCGCGCTTGC TTTGAACAAG GCGGGTTTGG CGTATTTTGA
  	AGGGCGTTTT
  301	GAAAAGGCGG AACTTGAAGC CTCGCGCGTA TTGGGAAACA
  	AAGAGGCGGG
  351	GGATAACCGG ACTTTGGCAT TGATGTTGGG CGCACATGCC
  	GCCGGGCAGA
  401	TGGAAAACAT CGAGCTGCGC GACCGTTATC TTGCGGAAAT
  	CGCCAAACTG
  451	CCGGAAAAGC AGCAGCTTTC CCGTTATCTT TTGTTGGCGG
  	AATCGGCGTT
  501	GAACCGGCGC GATTACGAAG CGGCGGAAGC CAATCTTCAT
  	GCGGCGGCGA
  551	AGATGAATGC CAACCTTACG CGCCTCGTGC GTCTGCAACT
  	TCGTTACGCT
  601	TTCGACAGGG GCGACGCGTT GCAGGTTCTG GCAAAAACCG
  	AAAAANTTTC
  651	CAAGGCGGGC GCGTNGGGCA AATCGGAAAT GGAACGGTAT
  	CAAAATTGGG
  701	CATACCGCCG CCAGCTGNCG GATGCTGCCG ATGCCGCCGC
  	TTTGAAAACC
  751	TGCCTGAAGC GGATTCCCGA CAGCCTCAAA AACGGGGAAT
  	TGAGCGTATC
  801	GGTTGCGGAA AAGTACGAAC GTTTGGGACT GTATGCCGAT
  	GCGGTCAAAT
  851	GGGTCAAACA GCATTATCCG CACAACCGCC GACCCGAACT
  	TTTGGAAGCN
  901	TTTGTCGAAA GCGTGCGCTT TTTGGGCGAA CGCGATCAGC
  	AGAAAGCCAT
  951	CGATTTTGCC GATGCTTGGC TGAAAGAACA GCCCGATAAT
  	GCGCTTCTGC
  1001	TGANGTATCT CGGTCGGCTC GCCTACGGCC GCAAACTTTG
  	GGGCAAGGCA
  1051	AAAGGCTACC TTGAAGCGAG CATTGCATTA AAGCCGAGTA
  	TTTCCGCGCG
  1101	TTTGGTTCTG GCAAAGGTTT TTGACGAAAC CGGAGAACCG
  	CAGAAGGCGG
  1151	AGGCGCAGCG CAACTTGGTT TTGGCAAGCG TTGCCGAGGA
  	AAACCGNCCT
  1201	TCCGCCGAAA CCCATTGA

• This encodes a protein having amino acid sequence <SEQ ID 754>:

• 	1	MKTVVWIVVL FAAAXGLALA SGIXTGDVYI VLGQTMLRIN
  		LHAFVLGSLI
  	51	AVVVWYFLFK FIIGVLNXPE KMQRFGSARK GRKAALALNK
  		AGLAYFEGRF
  	101	EKAELEASRV LGNKEAGDNR TLALMLGAHA AGQMENIELR
  		DRYLAEIAKL
  	151	PEKQQLSRYL LLAESALNRR DYEAAEANLH AAAKMNANLT
  		RLVRLQLRYA
  	201	FDRGDALQVL AKTEKXSKAG AXGKSEMERY QNWAYRRQLX
  		DAADAAALKT
  	251	CLKRIPDSLK NGELSVSVAE KYERLGLYAD AVKWVKQHYP
  		HNRRPELLEA
  	301	FVESVRFLGE RDQQKAIDFA DAWLKEQPDN ALLLXYLGRL
  		AYGRKLWGKA
  	351	KGYLEASIAL KPSISARLVL AKVFDETGEP QKAEAQRNLV
  		LASVAEENRP
  	401	SAETH*

• ORF100a and ORF100-1 show 95.1% identity in 406 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF100 shows 93.3% identity over a 386 aa overlap with a predicted ORF (ORF100ng) from N. gonorrhoeae:
• 
• The complete length ORF100ng nucleotide sequence <SEQ ID 755> is:

• 1	ATGAAAACGG TAGTCTGGAT TGTTGTCCTG TTTGCCGCCG
  	CCGTCGGACT
  51	GGCGCTGGCT TCGGGCATTT ACACCGGCGA CGTGTATATC
  	GTACTCGGAC
  101	AGACCATGCT CAGAATCAAC CTGCACGCCT TTGTGTTAGG
  	TTCGCTGATT
  151	GCCGTCGTGG TGTGGTATTT CCTGTTTAAA TTCATCATCG
  	GCGTACTCAA
  201	TATCCCCGAA AATATGCGGC GTTCCGGTTC GGCGCGGAAA
  	GGCCGCAAGG
  251	CCGCGCTTGC CTTGAATAAG GCGGGTTTGG CGTATTTCGA
  	AGGGCGTTTT
  301	GAAAAGGCGG AACTCGAAGC CTCTCGAGTG TTGGGCAACA
  	AAGAGGCCGG
  351	AGACAACCGG ACTTTGGCAT TGATGCTGGG CGCGCACGCG
  	GCAGGACAGA
  401	TGGAAAATAT CGAGCTGCGC GACCGTTATC TTGCGGAAAT
  	CGCCAAACTG
  451	CCGGAAAAAC AGCAGCTTTC CCGCTATCTT CTGCTGGCGG
  	AATCGGCGTT
  501	AAACCGGCGC GATTACGAAG CGGCGGAAGC CAATCTTCAT
  	GCGGCGGCGA
  551	AGATGAATGC CAACCTTACG CGCCTCGTGC GTCTGCAACT
  	TCGTTACGCC
  601	TTCGATCGGG GCGATGCGTT GCAGGTTCTG GCAAAAaccG
  	AAAAACTTTC
  651	CAAGGCGGGC GCGTTGGGCA AATCGGAAAT GGAACGGTAT
  	CAAAATTGGG
  701	CATACCGCCG CCAGATGGCG GATGCTGCCG ATGCCGCCGC
  	TTTGAAAACC
  751	TGCCTGAAGC GGATTCCCGA CAGCCTCAAA AACGGGGAAT
  	TGagcGTATC
  801	GGTTGCGGAA AAGTACGAAC GTTTGGGACT GTATGCCGAT
  	GCGGTCAAAT
  851	GGGTCAAACA GCATTATCCG CACAACCGCC GCCCCGAGCT
  	TTTGGAAGCC
  901	TTTGTCGAAA GCGTGCGCTT TTTGGGCGAG CGCGAACAGC
  	AGAAAGCCAT
  951	CGATTTTGCC GATTCTTGGC TGAAAGAACA GCCCGATAAC
  	GCGCTTCTGC
  1001	TGATGTATCT CGGCCGGCTC GCCTACGGCC GCAAACTTTG
  	GGGTAAGGCA
  1051	AAAGGCTACC TTGAAGCGAG TATTGCACTG AAGCCGAGTA
  	TTCCGGCGCG
  1101	TTTGGTGTTG GCAAAGGTTT TTGACGAAAC CGCACAGTCG
  	CAAAAAGCCG
  1151	AAGCACAGCG CAACTTGGTT TTGGCAAGCG TTGCCGGGGA
  	AAACCGCCCT
  1201	TCCGCCGAAA CCCGTTGA

• This encodes a protein having amino acid sequence <SEQ ID 756>:

• 	1	MKTVVWIVVL FAAAVGLALA SGIYTGDVYI VLGQTMLRIN
  		LHAFVLGSLI
  	51	AVVVWYFLFK FIIGVLNIPE NMRRSGSARK GRKAALALNK
  		AGLAYFEGRF
  	101	EKAELEASRV LGNKEAGDNR TLALMLGAHA AGQMENIELR
  		DRYLAEIAKL
  	151	PEKQQLSRYL LLAESALNRR DYEAAEANLH AAAKMNANLT
  		RLVRLQLRYA
  	201	FDRGDALQVL AKTEKLSKAG ALGKSEMERY QNWAYRRQMA
  		DAADAAALKT
  	251	CLKRIPDSLK NGELSVSVAE KYERLGLYAD AVKWVKQHYP
  		HNRRPELLEA
  	301	FVESVRFLGE REQQKAIDFA DSWLKEQPDN ALLLMYLGRL
  		AYGRKLWGKA
  	351	KGYLEASIAL KPSIPARLVL AKVFDETAQS QKAEAQRNLV
  		LASVAGENRP
  	401	SAETR*

• ORF100ng and ORF100-1 show 95.3% identity in 402 aa overlap:
• 
• Based on this analysis, including the presence of a putative leader sequence, a putative transmembrane domain, and a RGD motif, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 90
• The following DNA sequence, believed to be complete, was identified in N. meningitidis <SEQ ID 757>

• 	1	ATGATGTTTT CTTGGTTCAA GCTGTTTCAC TTGTTTTTTG
  		TCATTTCGTG
  	51	GTTTGCAGGG CTGTTTTACC TGCCGAGGAT TTTCGTCAAT
  		ATGGCGATGA
  	101	TTGATGTGCC GCGCGGCAAT CCCGAGTATG TGCGTCTGTC
  		GGGCATGGCG
  	151	GTGCGGCTGT ACCGTTTTAT GTCGCCGTTG GGCTTCGGCG
  		CGGTCGTGTT
  	201	CGGCGCGGCG ATACCGTTTG CCGCCGGCTG GTGGGGCAGC
  		GGCTGGGTAC
  	251	ACGTCAAACT GTGTTTGGGC TTGATGCTCT TGGCTTACCA
  		GTTGTATTGC
  	301	GGCGTGCTGC TGCGCCGTTT TCAGGATTAC AGCAATGCTT
  		TTTCACACCG
  	351	CTGGTACCGC GTGTTCAACG AAATCCCCGT GCTGCTGATG
  		GTTGCCGCGC
  	401	TGTATsTGGT CGTGTTCAAA CCGTTTTGA

• This corresponds to the amino acid sequence <SEQ ID 758; ORF102>:

• 	1	MMFSWFKLFH LFFVISWFAG LFYLPRIFVN MAMIDVPRGN
  		PEYVRLSGMA
  	51	VRLYRFMSPL GFGAVVFGAA IPFAAGWWGS GWVHVKLCLG
  		LMLLAYQLYC
  	101	GVLLRRFQDY SNAFSHRWYR VFNEIPVLLM VAALYXVVFK
  		PF*

• Further work revealed the complete nucleotide sequence <SEQ ID 759>:

• 	1	ATGATGTTTT CTTGGTTCAA GCTGTTTCAC TTGTTTTTTG
  		TCATTTCGTG
  	51	GTTTGCAGGG CTGTTTTACC TGCCGAGGAT TTTCGTCAAT
  		ATGGCGATGA
  	101	TTGATGTGCC GCGCGGCAAT CCCGAGTATG TGCGTCTGTC
  		GGGCATGGCG
  	151	GTGCGGCTGT ACCGTTTTAT GTCGCCGTTG GGCTTCGGCG
  		CGGTCGTGTT
  	201	CGGCGCGGCG ATACCGTTTG CCGCCGGCTG GTGGGGCAGC
  		GGCTGGGTAC
  	251	ACGTCAAACT GTGTTTGGGC TTGATGCTCT TGGCTTACCA
  		GTTGTATTGC
  	301	GGCGTGCTGC TGCGCCGTTT TCAGGATTAC AGCAATGCTT
  		TTTCACACCG
  	351	CTGGTACCGC GTGTTCAACG AAATCCCCGT GCTGCTGATG
  		GTTGCCGCGC
  	401	TGTATCTGGT CGTGTTCAAA CCGTTTTGA

• This corresponds to the amino acid sequence <SEQ ID 760; ORF102-1>:

• 1	MMFSWFKLFH LFFVISWFAG LFYLPRIFVN MAMIDVPRGN
  	PEYVRLSGMA
  51	VRLYRFMSPL GFGAVVFGAA IPFAAG WWGS GWVHVKLCLG
  	LMLLAYQLYC
  101	GVLLRRFQDY SNAFSHRWYR VFNEIPVLLM VAALYLVVFK
  	PF*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with HP1484 Hypothetical Integral Membrane Protein of H. pylori (Accession Number AE000647)
• ORF102 and HP1484 show 33% aa identity in 143aa overlap:

• orf102	3	FSWFKLFHLFFVISWFAGLFYLPRIFVNMAMIDVPRGNPEYVRLSGMAVRLYRFMSPLGF	62
  		F W K FH+  VISW A LFYLPR+FV  A     +     V++     +LY F++
  HP1484
  	8	FLWVKAFHVIAVISWMAALFYLPRLFVYHAENAHKKEFVGVVQIQEK--KLYSFIASPAM	65
  orf102	63	GAVVFGAAIPFAAG---WWGSGWVHVKLCLGLMLLAYQLYCGVLLRRFQDYSNAFSHRWY	119
  		G  +    +        +   GW+H KL L ++LLAY  YC   +R  +      + R+Y
  HP1484	66	GFTLITGILMLLIEPTLFKSGGWLHAKLALVVLLLAYHFYCKKCMRELEKDPTRRNARFY	125
  orf102	120	RVFNEIPXXXXXXXXXXXXFKPF	142
  		RVFNE P             KPF
  HP1484	126	RVFNEAPTILMILIVILVVVKPF	148

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF102 shows 99.3% identity over a 142aa overlap with an ORF (ORF102a) from strain A of N. meningitidis:
• 
• The complete length ORF102a nucleotide sequence <SEQ ID 761> is:

• 1	ATGATGTTTT CTTGGTTCAA GCTGTTTCAC TTGTTTTTTG
  	TCATTTCGTG
  51	GTTTGCAGGG CTGTTTTACC TGCCGAGGAT TTTCGTCAAT
  	ATGGCGATGA
  101	TTGATGTGCC GCGCGGCAAT CCCGAGTATG TGCGTCTGTC
  	GGGCATGGCG
  151	GTGCGGCTGT ACCGTTTTAT GTCGCCGTTG GGCTTCGGCG
  	CGGTCGTGTT
  201	CGGCGCGGCG ATACCGTTTG CCGCCGGCTG GTGGGGCAGC
  	GGCTGGGTAC
  251	ACGTCAAACT GTGTTTGGGC TTGATGCTCT TGGCTTACCA
  	GTTGTATTGC
  301	GGCGTGCTGC TGCGCCGTTT TCAGGATTAC AGCAATGCTT
  	TTTCACACCG
  351	CTGGTACCGC GTGTTCAACG AAATCCCCGT GCTGCTGATG
  	GTTGCCGCGC
  401	TGTATCTGGT CGTGTTCAAA CCGTTTTGA

• This encodes a protein having amino acid sequence <SEQ ID 762>:

• 1	MMFSWFKLFH LFFVISWFAG LFYLPRIFVN MAMIDVPRGN
  	PEYVRLSGMA
  51	VRLYRFMSPL GFGAVVFGAA IPFAAG WWGS GWVHVKLCLG
  	LMLLAYQLYC
  101	GVLLRRFQDY SNAFSHRWYR VFNEIPVLLM VAALYLVVFK
  	PF*

• ORF102a and ORF102-1 show complete identity in 142 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF102 shows 97.9% identity over a 142 aa overlap with a predicted ORF (ORF102ng) from N. gonorrhoeae:
• 
• The complete length ORF102ng nucleotide sequence <SEQ ID 763> is:

• 1	ATGATGTTTT CTTGGTTCAA GCTGTTTCAC TTGTTTTTTG
  	TCATTTCGTG
  51	GTTTGCAGGG CTGTTTTACC TGCCGAGGAT TTTCGTCAAT
  	ATGGCGATGA
  101	TTGATGCGCC GCGCGGCAAT CCCGAGTATG TGCGCCTGTC
  	GGGGATGGCG
  151	GTGCGGTTGT ACCGTTTTAT GTCGCCTTTG GGTTTCGGCG
  	CGGTCGTGTT
  201	CGGCGCGGCG ATACCGTTTG CCGCcggccg GTGGGGCagc
  	ggctggGTTC
  251	ACGTCAAACT GTGTTTGGGC TTGATGCTCT TGGCTTATCA
  	GTTGTATTGC
  301	GGCGTGCTGC TGCGCCGTTT TCAGGATTAC AGCAATGCTT
  	TTTCACACCG
  351	CTGGTACCGC GTGTTCAAcg aAATCCCCGT GCTGCTGATG
  	GTTGCCGCGC
  401	TGTATCTGGT CGTGTTCAAA CCGTTTTGA

• This encodes a protein having amino acid sequence <SEQ ID 764>:

• 1	MMFSWFKLFH LFFVISWFAG LFYLPRIFVN MAMIDAPRGN
  	PEYVRLSGMA
  51	VRLYRFMSPL GFGAVVFGAA IPFAAG RWGS GWVHVKLCLG
  	LMLLAYQLYC
  101	GVLLRRFQDY SNAFSHRWYR VFNEIPVLLM VAALYLVVFK
  	PF*

• ORF102ng and ORF102-1 show 98.6% identity in 142 aa overlap:
• 
• In addition, ORF102ng shows significant homology to a membrane protein from H. pylori:

• gi|2314656 (AE000647) conserved hypothetical integral membrane protein
  [Helicobacter pylori] Length = 148
  Score = 79.2 bits (192), Expect = 1e−14
  Identities = 50/147 (34%), Positives = 68/147 (46%), Gaps = 13/147 (8%)

  Query:	3	FSWFKLFHLFFVISWFAGLFYLPRIFVNMAMIDAPRGNPEYVRLSGMAVRLYRFMSPLGF	62
  		F W K FH+  VISW A LFYLPR+FV  A     +     V++     +LY F++
  Sbjct:	8	FLWVKAFHVIAVISWMAALFYLPRLFVYHAENAHKKEFVGVVQIQEK--KLYSFIASPAM	65
  Query:	63	GAVVFGAAIP-------FAAGRWGSGWVHVKLCLGLMLLAYQLYCGVLLRRFQDYSNAFS	115
  		G  +    +        F +G    GW+H KL L ++LLAY  YC   +R  +      +
  Sbjct:	66	GFTLITGILMLLIEPTLFKSG----GWLHAKLALVVLLLAYHFYCKKCMRELEKDPTRRN	121
  Query:	116	HRWYRVFNEIPXXXXXXXXXXXXFKPF	142
  		R+YRVFNE P             KPF
  Sbjct:	122	ARFYRVFNEAPTILMILIVILVVVKPF	148

• Based on this analysis, it is predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 91
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 765>:

• 1  ATGGCAAAAA TGATGAAATG GGCGGCTGTT GCGGCGGTCG CGGCGGCAGC
  51  GGTTTGGGGC GGATGGTCTT AACTGAAGCC CGAGCCGCAC GTGCTTGATA
  101  TTACGGAAAC GGTCAGGCGC GGC // .....
  //.. ATTTCGTTTA CGATTTTGTC CGAACCGGAT ACGCCGATTA AGGCGAAGCT
  51  CGACAGCGTC GACCCCGGGC TGACCACGAT GTCGTCGGGC GGTTACAACA
  101  GCAGTACGGA TACGGCTTCC AATGCGGTCT ACTATTATGC CCGTTCGTTT
  151  GTGCCGAATC CGGACGGCAA ACTCGCCACG GGGATGACGA CGCAGAATAC
  201  GGTTGAAATC GACGGCGTGA AAAATGTGCT GATTATTCCG TCGCTGACCG
  251  TGAAAAATCG CGGCGGCAAG GCGTTTGTGC GCGTGTTGGG TGCGGACGGC
  301  AAGGCGGCGG AACGCGAAAT CCGGACCGGT ATGAGAGACA GTATGAATAC
  351  CGAAGTAAAA AGCGGGTTGA AAGAGGGGGA CAAAGTGGTC ATCTCCGAAA
  401  TAACCGCCGC CGAGCAACAG GAAAGCGGCG AACGCGCCCT AGGCGGCCCG
  451  CCGCGCCGAT AA

• This corresponds to the amino acid sequence <SEQ ID 766; ORF85>:

• 1	MAKMMKWAAV AAVAAAAVWG GWS.LKPEPH VLDITETVRR
  	G.........
  51	.......... .......... .......... ..........
  	..........
  101	.......... .......... .......... ..........
  	..........
  151	.......... .......... .......... ..........
  	..........
  201	.......... .......... .......... .........I
  	SFTILSEPDT
  251	PIKAKLDSVD PGLTTMSSGG YNSSTDTASN AVYYYARSFV
  	PNPDGKLATG
  301	MTTQNTVEID GVKNVLIIPS LTVKNRGGKA FVRVLGADGK
  	AAEREIRTGM
  351	RDSMNTEVKS GLKEGDKVVI SEITAAEQQE SGERALGGPP
  	RR*

• Further work revealed the further partial nucleotide sequence <SEQ ID 767>:

• 1	..GTATCGGTCG GCGCGCAGGC ATCGGGGCAG ATTAAGATAC
  	TTTATGTCAA
  51	ACTCGGGCAA CAGGTTAAAA AGGGCGATTT GATTGCGGAA
  	ATCAATTCGA
  101	CCTCGCAGAC CAATACGCTC AATACGGAAA AATCCAAGTT
  	GGAAACGTAT
  151	CAGGCGAAGC TGGTGTCGGC ACAGATTGCA TTGGGCAGCG
  	CGGAGAAGAA
  201	ATATAAGCGT CAGGCGGCGT TATGGAAGGA AAACGCGACT
  	TCCAAAGAGG
  251	ATTTGGAAAG CGCGCAGGAT GCGTTTGCCG CCGCCAAAGC
  	CAATGTTGCC
  301	GAGCTGAAGG CTTTAATCAG ACAGAGCAAA ATTTCCATCA
  	ATACCGCCGA
  351	GTCGGAATTG GGCTACACGC GCATTACCGC AACGATGGAC
  	GGCACGGTGG
  401	TGGCGATTCT CGTGGAAGAG GGGCAGACTG TGAACGCGGC
  	GCAGTCTACG
  451	CCGACGATTG TCCAATTGGC GAATCTGGAT ATGATGTTGA
  	ACAAAATGCA
  501	GATTGCCGAG GGCGATATTA CCAAGGTGAA GGCGGGGCAG
  	GATATTTCGT
  551	TTACGATTTT GTCCGAACCG GATACGCCGA TTAAGGCGAA
  	GCTCGACAGC
  601	GTCGACCCCG GGCTGACCAC GATGTCGTCG GGCGGTTACA
  	ACAGCAGTAC
  651	GGATACGGCT TCCAATGCGG TCTACTATTA TGCCCGTTCG
  	TTTGTGCCGA
  701	ATCCGGACGG CAAACTCGCC ACGGGGATGA CGACGCAGAA
  	TACGGTTGAA
  751	ATCGACGGCG TGAAAAATGT GCTGATTATT CCGTCGCTGA
  	CCGTGAAAAA
  801	TCGCGGCGGC AAGGCGTTTG TGCGCGTGTT GGGTGCGGAC
  	GGCAAGGCGG
  851	CGGAACGCGA AATCCGGACC GGTATGAGAG ACAGTATGAA
  	TACCGAAGTA
  901	AAAAGCGGGT TGAAAGAGGG GGACAAAGTG GTCATCTCCG
  	AAATAACCGC
  951	CGCCGAGCAA CAGGAAAGCG GCGAACGCGC CCTAGGCGGC
  	CCGCCGCGCC
  1001	GATAA

• This corresponds to the amino acid sequence <SEQ ID 768; ORF85-1>:

• 1	..VSVGAQASGQ IKILYVKLGQ QVKKGDLIAE INSTSQTNTL
  	NTEKSKLETY
  51	QAKLVSAQIA LGSAEKKYKR QAALWKENAT SKEDLESAQD
  	AFAAAKANVA
  101	ELKALIRQSK ISINTAESEL GYTRITATMD GTVVAILVEE
  	GQTVNAAQST
  151	PTIVQLANLD MMLNKMQIAE GDITKVKAGQ DISFTILSEP
  	DTPIKAKLDS
  201	VDPGLTTMSS GGYNSSTDTA SNAVYYYARS FVPNPDGKLA
  	TGMTTQNTVE
  251	IDGVKNVLII PSLTVKNRGG KAFVRVLGAD GKAAEREIRT
  	GMRDSMNTEV
  301	KSGLKEGDKV VISEITAAEQ QESGERALGG PPRR*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF85 shows 87.8% identity over a 41 aa overlap and 99.3% identity over a 153aa overlap with an ORF (ORF85a) from strain A of N. meningitidis:
• 
• The complete length ORF85a nucleotide sequence <SEQ ID 769> is:

• 1	ATGGCAAAAA TGATGAAATG GGCGGCTGTT GCGGCGGTCG
  	CGGCGGCAGC
  51	GGTTTGGGGC GGATGGTCTT ATCTGAAGCC CGAGCCGCAG
  	GCTGCTTATA
  101	TTACGGAAAC GGTCAGGCGC GGCGACATCA GCCGGACGGT
  	TTCTGCAACA
  151	GGGGAGATTT CGCCGTCCAA CCTGGTATCG GTCGGCGCGC
  	AGGCATCGGG
  201	GCAGATTAAG AAACTTTATG TCAAACTCGG GCAACAGGTT
  	AAAAAGGGCG
  251	ATTTGATTGC GGAAATCAAT TCGACCTCGC AGACCAATAC
  	GCTCAATACG
  301	GAAAAATCCA AATTGGAAAC GTATCAGGCG AAGCTGGTGT
  	CGGCACAGAT
  351	TGCATTGGGC AGCGCGGAGA AGAAATATAA GCGTCAGGCG
  	GCGTTGTGGA
  401	AGGATGATGC GACCGCTAAA GAAGATTTGG AAAGCGCACA
  	GGATGCGCTT
  451	GCCGCCGCCA AAGCCAATGT TGCCGAGCTG AAGGCTCTAA
  	TCAGACAGAG
  501	CAAAATTTCC ATCAATACCG CCGAGTCGGA ATTGGGCTAC
  	ACGCGCATTA
  551	CCGCAACGAT GGACGGCACG GTGGTGGCGA TTCTCGTGGA
  	AGAGGGGCAG
  601	ACTGTGAACG CGGCGCAGTC TACGCCGACG ATTGTCCAAT
  	TGGCGAATCT
  651	GGATATGATG TTGAACAAAA TGCAGATTGC CGAGGGCGAT
  	ATTACCAAGG
  701	TGAAGGCGGG GCAGGATATT TCGTTTACGA TTTTGTCCGA
  	ACCGGATACG
  751	CCGATTAAGG CGAAGCTCGA CAGCGTCGAC CCCGGGCTGA
  	CCACGATGTC
  801	GTCGGGCGGC TACAACAGCA GTACGGATAC GGCTTCCAAT
  	GCGGTCTACT
  851	ATTATGCCCG TTCGTTTGTG CCGAATCCGG ACGGCAAACT
  	CGCCACGGGG
  901	ATGACGACGC AGAATACGGT TGAAATCGAC GGTGTGAAAA
  	ATGTGCTGAT
  951	TATTCCGTCG CTGACCGTGA AAAATCGCGG CGGCAGGGCG
  	TTTGTGCGCG
  1001	TGTTGGGTGC AGACGGCAAG GCGGCGGAAC GCGAAATCCG
  	GACCGGTATG
  1051	AGAGACAGTA TGAATACCGA AGTAAAAAGC GGGTTGAAAG
  	AGGGGGACAA
  1101	AGTGGTCATC TCCGAAATAA CCGCCGCCGA GCAGCAGGAA
  	AGCGGCGAAC
  1151	GCGCCCTAGG CGGCCCGCCG CGCCGATAA

• This encodes a protein having amino acid sequence <SEQ ID 770>:

• 1	MAKMMKWAAV AAVAAAAVWG GWSYLKPEPQ AAYITETVRR
  	GDISRTVSAT
  51	GEISPSNLVS VGAQASGQIK KLYVKLGQQV KKGDLIAEIN
  	STSQTNTLNT
  101	EKSKLETYQA KLVSAQIALG SAEKKYKRQA ALWKDDATAK
  	EDLESAQDAL
  151	AAAKANVAEL KALIRQSKIS INTAESELGY TRITATMDGT
  	VVAILVEEGQ
  201	TVNAAQSTPT IVQLANLDMM LNKMQIAEGD ITKVKAGQDI
  	SFTILSEPDT
  251	PIKAKLDSVD PGLTTMSSGG YNSSTDTASN AVYYYARSFV
  	PNPDGKLATG
  301	MTTQNTVEID GVKNVLIIPS LTVKNRGGRA FVRVLGADGK
  	AAEREIRTGM
  351	RDSMNTEVKS GLKEGDKVVI SEITAAEQQE SGERALGGPP
  	RR*

• ORF85a and ORF85-1 show 98.2% identity in 334 aa overlap:
• 
• FIG. 19D shows plots of hydrophilicity, antigenic index, and AMPHI regions for ORF85a.
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF85 shows a high degree of identity with a predicted ORF (ORF85ng) from N. gonorrhoeae:
• 
• The complete length ORF85ng nucleotide sequence <SEQ ID 771> is:

• 1	ATGGCAAAAA TGATGAAATG GGCGGCTGTT GCGGCGGTCG
  	CGGCGGCaac
  51	GGTTTGGGGC GGATGGTCTT ATCTGAAGCC CGAACCGCAG
  	GCTGCTTATA
  101	TTACGGAaac ggTCAGGCGC GGCGATATCA GCCGGACGGT
  	TTCCGCGACG
  151	GgcgAGATTT CGCCGTCCAA CCTGGTATCG GTCGGCGCGC
  	AGGCTTCGGG
  201	GCAGATTAAA AAGCTTTATG TCAAACTCGG GCAACAGGTC
  	AAAAAGGGCG
  251	ATTTGATTGC GGAAATCAAT TCGACCACGC AGACCAACAC
  	GATCGATATG
  301	GAAAAATCCA AATTGGAAAC GTATCAGGCG AAGCTGGTGT
  	CGGCACAGAT
  351	TGCATTGGGC AGCGCGGAGA AGAAATATAA GCGTCAGGCG
  	GCGTTGTGGA
  401	AGGATGATGC GACCTCTAAA GAAGATTTGG AAAGCGCGCA
  	GGATGCGCTT
  451	GCCGCCGCCA AAGCCAATGT TGCCGAGTTG AAGGCTTTAA
  	TCAGACAGAG
  501	CAAAATTTCC ATCAATACCG CCGAGTCGGA TTTGGGCTAC
  	ACGCGCATTA
  551	CCGCGACGAT GGACGGCACG GTGGTGGCGA TTCCCGTGGA
  	AGAGGGGCAG
  601	ACTGTGAACG CGGCGCAGTC TACGCCGACG ATTGTCCAAT
  	TGGCGAATCT
  651	GGATATGATG TTGAACAAAA TGCAGATTGC CGAGGGCGAT
  	ATTACCAAGG
  701	TGAAGGCGGG GCAGGATATT TCGTTTACGA TTTTGTCCGA
  	ACCGGATACG
  751	CCGATTAAGG CGAAGCTCGA CAGCGTCGAC CCCGGGCTGA
  	CCACGATGTC
  801	GTCGGGCGGC TACAACAGCA GTACGGATAC GGCTTCCAAT
  	GCGGTCTATT
  851	ATTATGCCCG TTCGTTTGTG CCGAATCCGG ACGGCAAACT
  	CGCCACGGGG
  901	ATGACGACGC AGAATACGGT TGAAATCGAC GGTGTGAAAA
  	ATGTGTTGCT
  951	TATTCCGTCG CTGACCGTGA AAAATCGCGG CGGCAAGGCG
  	TTCGTACGCG
  1001	TGTTGGGTGC GGACGGCAAG GCAGTGGAAC GCGAAATCCG
  	GACCGGTATG
  1051	AAAGACAGTA TGAATACCGA AGTGAAAAGC GGGTTGAAAG
  	AGGGGGACAA
  1101	AGTGGTCATC TCCGAAATAA CCGCCGCCGA GCAGCAGGAA
  	AGCGGCGAAC
  1151	GCGCCCTAGG CGGCCCGCCG CGCCGATAA

• This encodes a protein having amino acid sequence <SEQ ID 772>:

• 1	MAKMMKWAAV AAVAAAAVWG GWSYLKPEPQ AAYITEAVRR
  	GDISRTVSAT
  51	GEISPSNLVS VGAQASGQIK KLYVKLGQQV KKGDLIAEIN
  	STTQTNTIDM
  101	EKSKLETYQA KLVSAQIALG SAEKKYKRQA ALWKDDATSK
  	EDLESAQDAL
  151	AAAKANVAEL KALIRQSKIS INTAESDLGY TRITATMDGT
  	VVAIPVEEGQ
  201	TVNAAQSTPT IVQLANLDMM LNKMQIAEGD ITKVKAGQDI
  	SFTILSEPDT
  251	PIKAKLDSVD PGLTTMSSGG YNSSTDTASN AVYYYARSFV
  	PNPDGKLATG
  301	MTTQNTVEID GVKNVLLIPS LTVKNRGGKA FVRVLGADGK
  	AVEREIRTGM
  351	KDSMNTEVKS GLKEGDKVVI SEITAAEQQE SGERALGGPP
  	RR*

• ORF85ng and ORF85-1 show 96.1% identity in 334 aa overlap:
• 
• In addition, ORF85ng shows significant homology to an E. coli membrane fusion protein:

• gi|1787104 (AE000189) o380; 27% identical (27 gaps) to 332 residues from
  membrane fusion protein precursor, MTRC_NEIGO SW: P43505 (412 aa)
  [Escherichia coli] Length = 380
  Score = 193 bits (485), Expect = 2e−48
  Identities = 120/345 (34%), Positives = 182/345 (51%), Gaps = 13/345 (3%)

  Query:	29	PQAAYITETVRRGDISRTVSATGEISPSNLVSVGAQASGQIKKLYVKLGQQVKKGDLIAE	88
  		P   Y T  VR GD+ ++V ATG++     V VGAQ SGQ+K L V +G +VKK  L+
  Sbjct:	41	PVPTYQTLIVRPGDLQQSVLATGKLDALRKVDVGAQVSGQLKTLSVAIGDKVKKDQLLGV	100
  Query:	89	INSTTQTNTIDMEKSKLETYQAKLVSAQIALGSAEKKYKRQAALWKDDATSKEXXXXXXX	148
  		I+     N I   ++ L   +A+   A+  L  A   Y RQ  L +  A S++
  Sbjct:	101	IDPEQAENQIKEVEATLMELRAQRQQAEAELKLARVTYSRQQRLAQTKAVSQQDLDTAAT	160
  Query:	149	XXXXXXXXXXXXXXXIRQSKISINTAESDLGYTRITATMDGTVVAIPVEEGQTVNAAQST	208
  		I++++ S++TA+++L YTRI A M G V  I   +GQTV AAQ
  Sbjct:	161	EMAVKQAQIGTIDAQIKRNQASLDTAKTNLDYTRIVAPMAGEVTQITTLQGQTVIAAQQA	220
  Query:	209	PTIVQLANLDMMLNKMQIAEGDITKVKAGQDISFTILSEPDTPIKAKLDSVDPGLTTMSS	268
  		P I+ LA++  ML K Q++E D+  +K GQ   FT+L +P T  + ++  V P
  Sbjct:	221	PNILTLADMSAMLVKAQVSEADVIHLKPGQKAWFTVLGDPLTRYEGQIKDVLP-------	273
  Query:	269	GGYNSSTDTASNAVYYYARSFVPNPDGKLATGMTTQNTVEIDGVKNVLLIPSLTVKNRGG	328
  		+ +  ++A++YYAR  VPNP+G L   MT Q  +++  VKNVL IP   + +  G
  Sbjct:	274	-----TPEKVNDAIFYYARFEVPNPNGLLRLDMTAQVHIQLTDVKNVLTIPLSALGDPVG	328
  Query:	329	KAFVRV-LGADGKAVEREIRTGMKDSMNTEVKSGLKEGDKVVISE	372
  		+V L  +G+  ERE+  G ++  + E+  GL+ GD+VVI E
  Sbjct:	329	DNRYKVKLLRNGETREREVTIGARNDTDVEIVKGLEAGDEVVIGE	373

• Based on this analysis, it was predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF85-1 (40.4 kDa) was cloned in the pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 19A shows the results of affinity purification of the GST-fusion protein. Purified GST-fusion protein was used to immunise mice, whose sera were used for Western blot (FIG. 19B), FACS analysis (FIG. 19C), and ELISA (positive result). These experiments confirm that ORF85-1 is a surface-exposed protein, and that it is a useful immunogen.
Example 92
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 773>:

• 1	..ATTCCCGCCA CGATGACATT TGAACGCAGC GGCAATGCTT
  	ACAAAATCGT
  51	TTCGACGATT AAAGTGCCGC TATACAATAT CCGTTTCGAG
  	TCCGGCGGTA
  101	CGGTTGTCGG CAATACCCTG CACCCTACCT ACTATAGAGA
  	CATACGCAGG
  151	GGCAAACTGT ATGCGGAAgc CAAATTCGCC GACgGcAGCG
  	TAACTTACGG
  201	CAAAGCGGGC GAGAGCAAAA CCGAGCAAAG CCCCAAGGCT
  	ATGGATTTGT
  251	TCACGCTTGC CTGGCAGTTG GCGGCAAATG ACGCGAAACT
  	CCCCCCGGGG
  301	CTGAAAATCA CCAACGGCAA AAAACTTTAT TCCGTCGGCG
  	GTTTGAATAA
  351	GGCGGGTACA GGAAAATACA GCATAGGCGG CGTGGAAACC
  	GAAGTCGTCA
  401	AATATCGGGT GCGGCGCGGC GACGATGCGG TAATGTATTT
  	cTTCGCACCG
  451	TCCCTGAACA ATATTCCGGC ACAAATCGGC TATACCGACG
  	ACGGCAAAAC
  501	CTATACGCTG AAACTCAAAT CGGTGCAGAT CAACGGCCAG
  	GCAGCCAAAC
  551	CGTAA

• This corresponds to the amino acid sequence <SEQ ID 774; ORF120>:

• 1	..IPATMTFERS GNAYKIVSTI KVPLYNIRFE SGGTVVGNTL
  	HPTYYRDIRR
  51	GKLYAEAKFA DGSVTYGKAG ESKTEQSPKA MDLFTLAWQL
  	AANDAKLPPG
  101	LKITNGKKLY SVGGLNKAGT GKYSIGGVET EVVKYRVRRG
  	DDAVMYFFAP
  151	SLNNIPAQIG YTDDGKTYTL KLKSVQINGQ AAKP*

• Further work revealed the complete nucleotide sequence <SEQ ID 775>:

• 1	ATGATGAAGA CTTTTAAAAA TATATTTTCC GCCGCCATTT
  	TGTCCGCCGC
  51	CCTGCCGTGC GCGTATGCGG CAGGGCTGCC CCAATCCGCC
  	GTGCTGCACT
  101	ATTCCGGCAG CTACGGCATT CCCGCCACGA TGACATTTGA
  	ACGCAGCGGC
  151	AATGCTTACA AAATCGTTTC GACGATTAAA GTGCCGCTAT
  	ACAATATCCG
  201	TTTCGAGTCC GGCGGTACGG TTGTCGGCAA TACCCTGCAC
  	CCTACCTACT
  251	ATAGAGACAT ACGCAGGGGC AAACTGTATG CGGAAGCCAA
  	ATTCGCCGAC
  301	GGCAGCGTAA CTTACGGCAA AGCGGGCGAG AGCAAAACCG
  	AGCAAAGCCC
  351	CAAGGCTATG GATTTGTTCA CGCTTGCCTG GCAGTTGGCG
  	GCAAATGACG
  401	CGAAACTCCC CCCGGGGCTG AAAATCACCA ACGGCAAAAA
  	ACTTTATTCC
  451	GTCGGCGGTT TGAATAAGGC GGGTACAGGA AAATACAGCA
  	TAGGCGGCGT
  501	GGAAACCGAA GTCGTCAAAT ATCGGGTGCG GCGCGGCGAC
  	GATGCGGTAA
  551	TGTATTTCTT CGCACCGTCC CTGAACAATA TTCCGGCACA
  	AATCGGCTAT
  601	ACCGACGACG GCAAAACCTA TACGCTGAAA CTCAAATCGG
  	TGCAGATCAA
  651	CGGCCAGGCA GCCAAACCGT AA

• This corresponds to the amino acid sequence <SEQ ID 776; ORF120-1>:

• 1	MMKTFKNIFS AAILSAALPC AYAAGLPQSA VLHYSGSYGI
  	PATMTFERSG
  51	NAYKIVSTIK VPLYNIRFES GGTVVGNTLH PTYYRDIRRG
  	KLYAEAKFAD
  101	GSVTYGKAGE SKTEQSPKAM DLFTLAWQLA ANDAKLPPGL
  	KITNGKKLYS
  151	VGGLNKAGTG KYSIGGVETE VVKYRVRRGD DAVMYFFAPS
  	LNNIPAQIGY
  201	TDDGKTYTLK LKSVQINGQA AKP*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF120 shows 92.4% identity over a 184aa overlap with an ORF (ORF120a) from strain A of N. meningitidis:
• 
• The complete length ORF120a nucleotide sequence <SEQ ID 777> is:

• 1	ATGATGAAGA CTTTTAAAAA TATATTTTCC GCCGCCATTT
  	TGTCCGCCGC
  51	CCTGCCGTGC GCGTATGCGG CAGGGCTGCC CNAATCCGCC
  	GTGCTGCACT
  101	ATTCCGGCAG CTACGGCATT CCCGCCACNA NNANNTNNGN
  	ACNNNGNGNC
  151	AATGCTTNCA AAATCGTTTC GACGATTAAA GTGCCGCTAT
  	ACAATATCCG
  201	TTTCGAGTCC GGCGGTACGG TTGTCGGCAA TACCCTGCAC
  	CCTACCTACT
  251	ATAGAGACAT ACGCAGGGGC AAACTGTATG CGGAAGCCAA
  	ATTCGCCGAC
  301	GGCAGCGTAA CCTACGGCAA AGCGGNNNNN ANCNNNNNNG
  	NGCAAAGCCC
  351	CAAGGCTATG GATTTGTTCA CGCTTGCNTG GCAGTTGGCG
  	GCAAATGACG
  401	CGAAACTCCC CCCGGGGCTG AAAATCACCA ACGGCAAAAA
  	ACTTTATTCC
  451	GTCGGCGGTT TGAATAAGGC GGGTACAGGA AAATACAGCA
  	TAGGCGGCGT
  501	GGAAACCGAA GTCGTCAAAT ATCGGGTGCG GCGCGGCGAC
  	GATGCGGTAA
  551	TGTATTTCTT CGCACCGTCC CTGAACAATA TTCCGGCACA
  	AATCGGCTAT
  601	ACCGACGACG GCAAAACCTA TACGCTGAAA CTCAAATCGG
  	TGCAGATCAA
  651	CGGCCAGGCA GCCAAACCGT AA

• This encodes a protein having amino acid sequence <SEQ ID 778>:

• 1	MMKTFKNIFS AAILSAALPC AYAAGLPXSA VLHYSGSYGI
  	PATXXXXXXX
  51	NAXKIVSTIK VPLYNIRFES GGTVVGNTLH PTYYRDIRRG
  	KLYAEAKFAD
  101	GSVTYGKAXX XXXXQSPKAM DLFTLAWQLA ANDAKLPPGL
  	KITNGKKLYS
  151	VGGLNKAGTG KYSIGGVETE VVKYRVRRGD DAVMYFFAPS
  	LNNIPAQIGY
  201	TDDGKTYTLK LKSVQINGQA AKP*

• ORF120a and ORF120-1 show 93.3% identity in 223 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF120 shows 97.8% identity over 184 aa overlap with a predicted ORF (ORF120ng) from N. gonorrhoeae:
• 
• The complete length ORF120ng nucleotide sequence <SEQ ID 779> is:

• 1	ATGATGAAGA CTTTTAAAAA TATATTTTCC GCCGCCATTT
  	TGTCCGCCGC
  51	CCTGCCGTGC GCGTATGCGG CAAGGCTACC CCAATCCGCC
  	GTGCTGCACT
  101	ATTCCGGCAG CTACGGCATT CCCGCCACGA TGACATTTGA
  	ACGCAGCGGC
  151	AATGCTTACA AAATCGTTTC GACGATTAAA GTGCCGCTAT
  	ACAATATCCG
  201	TTTCGAATCC GGCGGTACGG TTGTCGGCAA TACCCTGCAC
  	CCTGCCTACT
  251	ATAAAGACAT ACGCAGGGGC AAACTGTATG CGGAAGCCAA
  	ATTCGCCGAC
  301	GGCAGCGTAA CCTACGGCAA AGCGGGCGAG AGCAAAACCG
  	AGCAAAGCCC
  351	CAAGGCTATG GATTTGTTCA CGCTTGCCTG GCAGTTGGCG
  	GCAAATGACG
  401	CGAAACTCCC CCCGGGTCTG AAAATCACCA ACGGCAAAAA
  	ACTTTATTCC
  451	GTCGGCGGCC TGAATAAGGC GGGTACGGGA AAATACAGCA
  	TaggCGGCGT
  501	GGAAACCGAA GTCGTCAAAT ATCGGGTGCG GCGCGGCGAC
  	GATACGGTAA
  551	CGTATTTCTT CGCACCGTCC CTGAACAATA TTCCGGCACA
  	AATCGGCTAT
  601	ACCGACGACG GCAAAACCTA TACGCTGAAG CTCAAATCGG
  	TGCAGATCAA
  651	CGGACAGGCC GCCAAACCGT AA

• This encodes a protein having amino acid sequence <SEQ ID 780>:

• 1	MMKTFKNIFS AAILSAALPC AYAARLPQSA VLHYSGSYGI
  	PATMTFERSG
  51	NAYKIVSTIK VPLYNIRFES GGTVVGNTLH PAYYKDIRRG
  	KLYAEAKFAD
  101	GSVTYGKAGE SKTEQSPKAM DLFTLAWQLA ANDAKLPPGL
  	KITNGKKLYS
  151	VGGLNKAGTG KYSIGGVETE VVKYRVRRGD DTVTYFFAPS
  	LNNIPAQIGY
  201	TDDGKTYTLK LKSVQINGQA AKP*

• In comparison with ORF120-1, ORF120ng shows 97.8% identity in 223 aa overlap:
• 
• This analysis, including the presence of a putative leader sequence in the gonococcal protein suggests that the proteins from N. meningitidis and N. gonorrhoeae, and, their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 93
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 781>:

• 1	ATGTATCGGA GGAAAGGGCG GGGCATCAAG CCGTGGATGG
  	GTGCCGGTGC
  51	.GCGTTTGCC GCCTTGGTCT GGCTGGTTTT CGCGCTCGGC
  	GATACTTTGA
  101	CTCCGTTTGC GGTTGCGGCG GTGCTGGCGT ATGTATTGGA
  	CCCTTTGGTC
  151	GAATGGTTGC AGAAAAAGGG TTTGAACCGT GCATCCGCTT
  	CGATGTCTGT
  201	GATGGTGTTT TCCTTGATTT TGTTGTTGGC ATTATTGTTG
  	ATTATCGTCC
  251	CTATGCTGGT CGGGCAGTTC AACAATTTGG CATCGCGCCT
  	GCCCCAATTA
  301	ATCGGTTTTA TGCAGAACAC GCTGCTGCCG TGGTTGAAAA
  	ATACAATCGG
  351	CGGATATGTG GAAATCGATC AGGCATCTAT TATTGCGTGG
  	CTTCAGGCGC
  401	ATACGGGAGA GTTGAGCAAC GCGCTTAAGG CGTGGTTTCC
  	CGTTTTGATG
  451	AGGCAGGGCG GCAATATT..

• This corresponds to the amino acid sequence <SEQ ID 782; ORF121>:

• 1	MYRRKGRGIK PWMGAGXAFA ALVWLVFALG DTLTPFAVAA
  	VLAYVLDPLV
  51	EWLQKKGLNR ASASMSVMVF SLILLLALLL IIVPMLVGQF
  	NNLASRLPQL
  101	IGFMQNTLLP WLKNTIGGYV EIDQASIIAW LQAHTGELSN
  	ALKAWFPVLM
  151	RQGGNI..

• Further work revealed the complete nucleotide sequence <SEQ ID 783>:

• 1	ATGTATCGGA GGAAAGGGCG GGGCATCAAG CCGTGGATGG
  	GTGCCGGTGC
  51	GGCGTTTGCC GCCTTGGTCT GGCTGGTTTT CGCGCTCGGC
  	GATACTTTGA
  101	CTCCGTTTGC GGTTGCGGCG GTGCTGGCGT ATGTATTGGA
  	CCCTTTGGTC
  151	GAATGGTTGC AGAAAAAGGG TTTGAACCGT GCATCCGCTT
  	CGATGTCTGT
  201	GATGGTGTTT TCCTTGATTT TGTTGTTGGC ATTATTGTTG
  	ATTATCGTCC
  251	CTATGCTGGT CGGGCAGTTC AACAATTTGG CATCGCGCCT
  	GCCCCAATTA
  301	ATCGGTTTTA TGCAGAACAC GCTGCTGCCG TGGTTGAAAA
  	ATACAATCGG
  351	CGGATATGTG GAAATCGATC AGGCATCTAT TATTGCGTGG
  	CTTCAGGCGC
  401	ATACGGGAGA GTTGAGCAAC GCGCTTAAGG CGTGGTTTCC
  	CGTTTTGATG
  451	AGGCAGGGCG GCAATATTGT CAGCAGTATC GGCAACCTGC
  	TGCTGCTTCC
  501	CTTGCTGCTT TACTATTTCC TGCTGGATTG GCAGCGGTGG
  	TCGTGCGGCA
  551	TTGCCAAACT GGTTCCGAgG CGTTTTGCCG GTGCTTATAC
  	GCGCATTACA
  601	GGCAATTTGA ACGAGGTATT GGGCGAATTT TTGCGCGGGC
  	AGCTTCTGGT
  651	AATGCTGATT ATGGGCTTGG TTTACGGTTT GGGATTGGTG
  	CTGGTCGGGC
  701	TGGATTCGGG GTTTGCCATC GGTATGCTTG CCGGTATTTT
  	GGTGTTTGTC
  751	CCTTATCTCG GGGCGTTTAC GGGATTGCTG CTTGCCACCG
  	TCGCCGCCTT
  801	GCTCCAGTTC GGTTCGTGGA ACGGCATCCT ATCGGTTTGG
  	GCGGTTTTTG
  851	CCGTAGGACA GTTTCTCGAA AGTTTTTTCA TTACGCCGAA
  	AATCGTGGGA
  901	GACCGTATCG GGCTGTCGCC GTTTTGGGTT ATCTTTTCGC
  	TGATGGCGTT
  951	CGGGCAGCTG ATGGGCTTTG TCGGAATGTT GGCGGGATTG
  	CCTTTGGCCG
  1001	CCGTAACCTT GGTCTTGCTT CGCGAGGGCG TGCAGAAATA
  	TTTTGCCGGC
  1051	AGTTTTTACC GGGGCAGGTA G

• This corresponds to the amino acid sequence <SEQ ID 784; ORF121-1>:

• 1	MYRRKGRGIK PWMGAGAAFA ALVWLVFALG DTLTPFAVAA
  	VLAYVLDPLV
  51	EWLQKKGLNR ASASMSVMVF SLILLLALLL IIV PMLVGQF
  	NNLASRLPQL
  101	IGFMQNTLLP WLKNTIGGYV EIDQASIIAW LQAHTGELSN
  	ALKAWFPVLM
  151	RQGGNIVSSI GNLLLLPLLL YYFLLDWQRW SCGIAKLVPR
  	RFAGAYTRIT
  201	GNLNEVLGEF LRGQLLVMLI MGLVYGLGLV LV GLDSGFAI
  	GMLAGILVFV
  251	PYLGAFTGLL LATVAALLQF GSWNGILSVW AVFAVGQFLE
  	SFFITPKIVG
  301	DRIGLSPFWV IFSLMAFGQL MGFVGMLAGL PLAAVTLVLL
  	REGVQKYFAG
  351	SFYRGR*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF121 shows 98.7% identity over a 156aa overlap with an ORF (ORF121a) from strain A of N. meningitidis:
• 
• The complete length ORF121a nucleotide sequence <SEQ ID 785> is:

• 1	ATGTATCGGA GGAAAGGGCG GGGCATCAAG CCGTGGATGG
  	ATGCCGGTGC
  51	GGCGTTTGCC GCCTTGGTCT GGCTGGTTTT CGCGCTCGGC
  	GATACTTTGA
  101	CTCCGTTTGC GGTTGCGGCG GTGCTGGCGT ATGTATTGGA
  	CCCTTTGGTC
  151	GAATGGTTGC AGAAAAAGGG TTTGAACCGT GCATCCGCTT
  	CGATGTCTGT
  201	GATGGTGTTT TCCTTGATTT TGTTGTTGGC ATTATTGTTG
  	ATTATTGTCC
  251	CTATGCTGGT CGGGCAGTTC AACAATTTGG CATCGCGCCT
  	GCCCCAATTA
  301	ATCGGTTTTA TGCAGAACAC GCTGCTGCCG TGGTTGAAAA
  	ATACAATCGG
  351	CGGATATGTG GAAATCGATC AGGCATCTAT TATTGCGTGG
  	CTTCAGGCGC
  401	ATACGGGCGA GTTGAGCAAC GCGCTTAAGG CGTGGTTTCC
  	CGTTTTGATG
  451	AGGCAGGGCG GCAATATTGT CAGCAGTATC GGCAACCTGC
  	TGCTGCTTCC
  501	CTTGCTGCTT TACTATTTCC TGCTGGATTG GCAGCGGTGG
  	TCGTGCGGCA
  551	TTGCCAAACT GGTTCCGAGG CGTTTTGCCG GTGCTTATAC
  	GCGCATTACA
  601	GGCAATTTGA ACGAGGTATT GGGCGAATTT TTGCGCGGGC
  	AGCTTCTGGT
  651	GATGCTGATT ATGGGTTTGG TTTACGGCTT GGGGTTGGTG
  	CTGGTCGGGC
  701	TGGATTCGGG GTTTGCAATC GGTATGGTTG CCGGTATTTT
  	GGTTTTTGTT
  751	CCCTATTTGG GCGCGTTTAC AGGACTGCTG CTGGCAACCG
  	TCGCCGCCTT
  801	GCTCCAGTTC GGTTCGTGGA ACGGCATCTT GGCTGTTTGG
  	GCGGTTTTTG
  851	CCGTAGGACA GTTTCTCGAA AGTTTTTTCA TTACGCCGAA
  	AATCGTGGGA
  901	GACCGTATCG GCCTGTCGCC GTTTTGGGTT ATCTTTTCGC
  	TGATGGCGTT
  951	CGGGCAGCTG ATGGGCTTTG TCGGAATGTT GGCCGGATTG
  	CCTTTGGCCG
  1001	CCGTAACCTT GGTCTTGCTT CGCGAGGGCG TGCAGAAATA
  	TTTTGCCGGC
  1051	AGTTTTTACC GGGGCAGGTA G

• This encodes a protein having amino acid sequence <SEQ ID 786>:

• 1	MYRRKGRGIK PWMDAGAAFA ALVWLVFALG DTLTPFAVAA
  	VLAYVLDPLV
  51	EWLQKKGLNR ASASMSVMVF SLILLLALLL IIV PMLVGQF
  	NNLASRLPQL
  101	IGFMQNTLLP WLKNTIGGYV EIDQASIIAW LQAHTGELSN
  	ALKAWFPVLM
  151	RQGGNIVSSI GNLLLLPLLL YYFLLDWQRW SCGIAKLVPR
  	RFAGAYTRIT
  201	GNLNEVLGEF LRGQLLVMLI MGLVYGLGLV LV GLDSGFAI
  	GMVAGILVFV
  251	PYLGAFTGLL LATVAALLQF GSWNGILAVW AVFAVGQFLE
  	SFFITPKIVG
  301	DRIGLSPFWV IFSLMAFGQL MGFVGMLAGL PLAAVTLVLL
  	REGVQKYFAG
  351	SFYRGR*

• ORF121a and ORF121-1 show 99.2% identity in 356 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF121 shows 97.4% identity over a 156 aa overlap with a predicted ORF (ORF121ng) from N. gonorrhoeae:
• 
• An ORF121ng nucleotide sequence <SEQ ID 787> was predicted to encode a protein having amino acid sequence <SEQ ID 788>:

• 1	MYRRKGRGIK PWMGAGAAFA ALVWLVYALG DTLTPFAVAA
  	VLAYVLDPLV
  51	EWLQKKGLNR ASASMSVMVF SLILLLALLL IIV PMLVGQF
  	NNLASRLPQL
  101	IGFMQNTLLP WLKNTIGGYV EIDQASIIAW FQAHTGELSN
  	ALKAWFPVLM
  151	KQGGNIVSTI GNLLLPPLLL YYFLLDWHRW SCGIPKLVPR
  	RFAGAYTRIT
  201	GNLNKVWGKF LRGQLLGETE RGAVVCRVGR ECWEGGGARS
  	RPSDDGWPRW
  251	GGG*

• Further work revealed the following gonoccocal DNA sequence <SEQ ID 789>:

• 1	ATGTATCGGA GAAAAGGACG GGGCATCAAG CCGTGGATGG
  	GTGCCGGCGC
  51	GGCGTTTGCC GCCTTGGTCT GGCTGGTTTA CGCGCTCGGC
  	GATACTTTGA
  101	CTCCGTTTGC GGTTGCGGCG GTGCTGGCGT ATGTGTTGGA
  	CCCTTTGGTC
  151	GAATGGTTGC AGAAAAAGGG TTTGAACCGT GCATCCGCTT
  	CGATGTCTGT
  201	GATGGTGTTT TCCTTGATTT TGTTGTTGGC ATTATTGTTG
  	ATTATTGTCC
  251	CTATGCTGGT CGGGCAGTTC AATAATTTGG CATCTCGCCT
  	GCCCCAATTA
  301	ATCGGTTTTA TGCAGAACAC GCTGCTGCCG TGGTTGAAAA
  	ATACAATCGG
  351	CGGATATGTG GAAATCGATC AGGCATCTAT TATTGCGTGG
  	TTTCAGGCGC
  401	ATACGGGCGA GTTGAGCAAC GCGCTTAAGG CGTGGTTTCC
  	CGTTTTGATG
  451	AAACAGGGCG GCAATATTGT CAGCAGTATC GGCAACCTGC
  	TGCTGCCGCC
  501	CTTGCTGCTT TACTATTTCC TGCTGGATTG GCAGCGGTGG
  	TCGTGCGGCA
  551	TCGCCAAACT GGTTCCGAGG CGTTTTGCCG GTGCTTATAC
  	GCGCATTACG
  601	GGTAATTTGA ACGAGGTATT GGGCGAATTT TTGCGCGGTC
  	AGCTTCTGGT
  651	GATGCTGATT ATGGGCTTGG TTTACGGTTT GGGATTGATG
  	CTAGTCGGAC
  701	TGGATTCGGG ATTTGCCATC GGTATGGTTG CCGGTATTTT
  	GGTGTTTGTC
  751	CCCTATTTGG GTGCGTTTAC GGGATTGCTG CTTGCCACTG
  	TTGCAGCCTT
  801	GCTCCAGTTC GGTTCGTGGA ACGGAATCTT GGCTGTTTGG
  	GCGGTTTTTG
  851	CCGTCGGTCA GTTTCTCGAA AGTTTTTTCA TTACGCCGAA
  	AATTGTAGGA
  901	GACCGTATCG GCCTGTCGCC GTTTTGGGTT ATCTTTTCGC
  	TGATGGCGTT
  951	CGGAGAGCTG ATGGGCTTTG TCGGAATGTT GGCCGGATTG
  	CCTTTGGCCG
  1001	CCGTAACCTT GGTCTTGCTT CGCGAGGGCG CGCAGAAATA
  	TTTTGCCGGC
  1051	AGTTTTTACC GGGGCAGGTA G

• This corresponds to the amino acid sequence <SEQ ID 790; ORF121ng-1>:

• 1	MYRRKGRGIK PWMGAGAAFA ALVWLVYALG DTLTPFAVAA
  	VLAYVLDPLV
  51	EWLQKKGLNR ASASMSVMVF SLILLLALLL IIV PMLVGQF
  	NNLASRLPQL
  101	IGFMQNTLLP WLKNTIGGYV EIDQASIIAW FQAHTGELSN
  	ALKAWFPVLM
  151	KQGGNIVSSI GNLLLPPLLL YYFLLDWQRW SCGIAKLVPR
  	RFAGAYTRIT
  201	GNLNEVLGEF LRGQLLVMLI MGLVYGLGLM LV GLDSGFAI
  	GMVAGILVFV
  251	PYLGAFTGLL LATVAALLQF GSWNGILAVW AVFAVGQFLE
  	SFFITPKIVG
  301	DRIGLSPFWV IFSLMAFGEL MGFVGMLAGL PLAAVTLVLL
  	REGAQKYFAG
  351	SFYRGR*

• ORF121ng-1 and ORF121-1 show 97.5% identity in 356 aa overlap:
• 
• In addition, ORF121ng-1 shows homology to a permease from H. influenzae:

• sp|P43969|PERM_HAEIN PUTATIVE PERMEASE PERM HOMOLOG Length = 349
  Score = 69.9 bits (168), Expect = 2e−11
  Identities = 67/317 (21%), Positives = 120/317 (37%), Gaps = 7/317 (2%)

  Query:	26	VYALGDTLTPFAVAAVLAYVLDPLVEWL-QKKGLNRASASMSVMVFSXXXXXXXXXXXVP	84
  		+Y  GD + P  +A VL+Y+L+  + +L Q     R  A++ +               VP
  Sbjct:	32	IYFFGDLIAPLLIALVLSYLLEIPINFLNQYLKCPRMLATILIFGSFIGLAAVFFLVLVP	91
  Query:	85	MLVGQFNNLASRLPQLIGFMQNTLLPWLKNTIGGYVE-IDQASIIAWFQAHTGELSNALK	143
  		ML  Q  +L S LP +     N    WL N    Y E ID + + + F +   ++    +
  Sbjct:	92	MLWNQTISLLSDLPAMF----NKSNEWLLNLPKNYPELIDYSMVDSIFNSVREKILGFGE	147
  Query:	144	AWFPVLMKQGGNIVSSIGNXXXXXXXXXXXXXDWQRWSCGIAKLVPRRFAGAYTRITGNL	203
  		+   + +    N+VS                 D      G+++ +P+    A+ R    +
  Sbjct:	148	SAVKLSLASIMNLVSLGIYAFLVPLMMFFMLKDKSELLQGVSRFLPKNRNLAFXRWK-EM	206
  Query:	204	NEVLGEFLRGQXXXXXXXXXXXXXXXXXXXXDSGFAIGMVAGILVFVPYXXXXXXXXXXX	263
  		+ +  ++ G+                    +    +    G+ V VPY
  Sbjct:	207	QQQISNYINGKLLEILIVTLITYIIFLIFGLNYPLLLAFAVGLSVLVPYIGAVIVTIPVA	266
  Query:	264	XXXXXQFGSWNGILAVWAVFAVGQFLESFFITPKIVGDRIGLSPFWVIFSLMAFGELMGF	323
  		QFG       +   FAV Q L+   + P +  + + L P  +I S++ FG L GF
  Sbjct:	267	LVALFQFGISPTFWYIIIAFAVSQLLDGNLLVPYLFSEAVNLHPLIIIISVLIFGGLWGF	326
  Query:	324	VGMLAGLPLAAVTLVLL	340
  		G+   +PLA +   ++
  Sbjct:	327	WGVFFAIPLATLVKAVI	343

• Based on this analysis, including the presence of a putative leader sequence and transmembrane domains in the two proteins, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 94
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 791>:

• 1	..ACTGCTTTTT CGGCGGCGCT GCGCTTGAGT CCATCATGAC
  	TCGTCATATT
  51	TTTGTCCTTT GGGAAACCGT ATCAACAAAC AGCCGCCATC
  	TTAACATTTT
  101	TTTGCACGTC CTGCCCGCCG CGTTCAAATG CGTACCAGCA
  	ATACCGCCGC
  151	CTGCGCCTCT ATGCCTTCCA TCCGCCCGAG ATAGCCGAGT
  	TTTTCGTTGG
  201	TTTTGCCTTT GATGTTGACG CACGAAATGT CTATGCCCAA
  	ATCGGCGGCG
  251	ATGTTGGCAC GCATTTGCGG AATGTGCGGC GCGAGTGTGG
  	GTTTCTGTGC
  301	AATCACGGTC GTATCGACAT TGACCGCCTG CCAACCCTGC
  	GCCTGAACGC
  351	TTTGATACGC CGCACGCAAA AGGACGCGGC TGTCCGCATC
  	TTTGAACTCT
  401	GCGGCGGTGT CGGGGAAATG GCTGCCGATA TCGCCCAAAC
  	CTGCCGCACC
  451	GAGCAGCGCG TCGGTAACGG CGTGCAGCAG CGCATCGGCA
  	TCGGAGTGTC
  501	CGAGCAGCCC TTTTTCAAAT GGGATTTCAA CTCCGCCAAG
  	TATCAG..

• This corresponds to the amino acid sequence <SEQ ID 792; ORF122>:

• 1	..TAFSAALRLS PSXLVIFLSF GKPYQQTAAI LTFFCTSCPP
  	RSNAYQQYRR
  51	LRLYAFHPPE IAEFFVGFAF DVDARNVYAQ IGGDVGTHLR
  	NVRRECGFLC
  101	NHGRIDIDRL PTLRLNALIR RTQKDAAVRI FELCGGVGEM
  	AADIAQTCRT
  151	EQRVGNGVQQ RIGIGVSEQP FFKWDFNSAK YQ..

• Further work revealed the complete nucleotide sequence <SEQ ID 793>:

• 1	ATATCGTACT GGGCAAGCAG TTCGCCGGAT TTTTTGGAAG
  	TAGATACCGC
  51	GCCTTTGATT TTTTTGCCGC TCTTACCCAA GGCTTCGATG
  	AAAAAGTTGA
  101	TGGTCGAGCC GGTACCGATG CCGATATATT CATTTTCGGG
  	TACGAATTCG
  151	ACTGCTTTTT CGGCGGCGAT GCGCTTGAGT TCGTCTTGTG
  	TCGTCATATT
  201	TTTGTCCTTT GGGAAACCGT ATCAACAAAC AGCCGCCATC
  	TTAACATTTT
  251	TTTGCACGTC CTGCCCGCCG CGTTCAAATG CGTACCAGCA
  	ATACCGCCGC
  301	CTGCGCCTCT ATGCCTTCCA TCCGCCCGAG ATAGCCGAGT
  	TTTTCGTTGG
  351	TTTTGCCTTT GATGTTGACG CACGAAATGT CTATGCCCAA
  	ATCGGCGGCG
  401	ATGTTGGCAC GCATTTGCGG AATGTGCGGC GCGAGTTTGG
  	GTTTCTGTGC
  451	AATCACGGTC GTATCGACAT TGACCGCCTG CCAACCCTGC
  	GCCTGAACGC
  501	TTTGATACGC CGCACGCAAA AGGACGCGGC TGTCCGCATC
  	TTTGAACTCT
  551	GCGGCGGTGT CGGGGAAATG GCTGCCGATA TCGCCCAAAC
  	CTGCCGCACC
  601	GAGCAGCGCG TCGGTAACGG CGTGCAGCAG CGCATCGGCA
  	TCGGAGTGTC
  651	CGAGCAGCCC TTTTTCAAAT GGGATTTCAA CTCCGCCAAG
  	TATCAGCTTT
  701	CTGCCTTCGG TCAGTTGGTG GACATCGTAG CCCTGTCCGA
  	TACGGATGTT
  751	CGTCATCGTT TGTGTTCCTG A

• This corresponds to the amino acid sequence <SEQ ID 794; ORF122-1>:

• 1	ISYWASSSPD FLEVDTAPLI FLPLLPKASM KKLMVEPVPM
  	PIYSFSGTNS
  51	TAFSAAMRLS SSCVVIFLSF GKPYQQTAAI LTFFCTSCPP
  	RSNAYQQYRR
  101	LRLYAFHPPE IAEFFVGFAF DVDARNVYAQ IGGDVGTHLR
  	NVRREFGFLC
  151	NHGRIDIDRL PTLRLNALIR RTQKDAAVRI FELCGGVGEM
  	AADIAQTCRT
  201	EQRVGNGVQQ RIGIGVSEQP FFKWDFNSAK YQLSAFGQLV
  	DIVALSDTDV
  251	RHALCS*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF122 shows 94.0% identity over a 182aa overlap with an ORF (ORF122a) from strain A of N. meningitidis:
• 
• The complete length ORF122a nucleotide sequence <SEQ ID 795> is:

• 1	ATATCATATT GGGCAAGCAG TTCACTGGAT TTTTTGGAAG
  	TAGATACCGC
  51	GCCTTTGATT TTTTTGCCGC TCTTACCCAA GGCTTCGATG
  	AAAAAGTTGA
  101	TGGTCGAACC GGTACCGATG CCGATGTATT CGTTTTCGGG
  	TACGAATTCG
  151	ACTGCNTTTT CGGCGGCGAT GCGCTTGAGT TCGTCTTGTG
  	TCGTCATATT
  201	TTTGTCCTTT GGGAAACCGT ATCAACAAAC AGCCGCCATC
  	TTAACATTTT
  251	TTNNNACGTC CTGCCCGCCG CGTTCAAATC CTTACCAGCA
  	ATACCGCCGC
  301	CTGCGACTCT ATGCCTTCCA TGCGCCCGAG ATAACCGAGT
  	TTTTCGTTGG
  351	TTTTGCCTTT GANGTTGACG CACGAAATGT CTATGCCCAA
  	ATCGGCGGCG
  401	ATGTTGGCAC GCATTTGCGG AATATGCGGC GCGAGTTTGG
  	GTTTCTGTGC
  451	AATCACGGTC GTATCGACAT TGACCGCCTG CCAACCCTGC
  	GCCTGAACGC
  501	TTTGATACGC CGCACGCAAA AGGACGCGGC TGTCCGCATC
  	TTTGAACTCT
  551	GCGGCGGTGT CGGGGAAATG GCTGCCGATA TCGCCCAAAC
  	CTGCCGCACC
  601	GAGCAGCGCG TCGGTAACGG CGTGCAGCAG CGCATCGGCA
  	TCGGAGTGTC
  651	CGAGCAGCCC TTTTTCAAAT GGGATTTCAA CTCCGCCAAG
  	TATCAGCTTT
  701	CTGCCTTCGG TCAGTTGGTG GACATCGTAG CCCTGTCCGA
  	TACGGATGTT
  751	CGTCATCGTT TGTGTTCCTG A

• This encodes a protein having amino acid sequence <SEQ ID 796>:

• 1	ISYWASSSLD FLEVDTAPLI FLPLLPKASM KKLMVEPVPM
  	PMYSFSGTNS
  51	TAFSAAMRLS SSCVVIFLSF GKPYQQTAAI LTFFXTSCPP
  	RSNPYQQYRR
  101	LRLYAFHAPE ITEFFVGFAF XVDARNVYAQ IGGDVGTHLR
  	NMRREFGFLC
  151	NHGRIDIDRL PTLRLNALIR RTQKDAAVRI FELCGGVGEM
  	AADIAQTCRT
  201	EQRVGNGVQQ RIGIGVSEQP FFKWDFNSAK YQLSAFGQLV
  	DIVALSDTDV
  251	RHRLCS*

• ORF122a and ORF122-1 show 96.9% identity in 256 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF122 shows 89.6% identity over a 182 aa overlap with a predicted ORF (ORF122ng) from N. gonorrhoeae:
• 
• The complete length ORF122ng nucleotide sequence <SEQ ID 797> is:

• 1	ATGTCGTACC GGGCAAGCAG TTCGCCGGAT TTTTTGGAGG
  	TTGAAACCGC
  51	GCCTTTGATT TTTTTACCGC TTTTGCCCAA GGCTTCGATG
  	AAGAAATTGa
  101	tgGTCGAACC GgtaCCGATG CCGATGTATT CGTTTTCGGG
  	TACGAATTCG
  151	ACTGCTTTTT CGGCGGCGAT GCGCttgAgt TCgtcttgcg
  	TcgTCATATT
  201	TTTAtccttt gGGAAaccct atcaAcaAAc agccgccatC
  	TTAACATTTT
  251	TTTGCACGtc ctggccgccg cgttcaAATc cgtaccaGca
  	ataccgccgc
  301	ctgcgcctCT AtgcCTTCCA TCCGCCCGAG ATAGCCGAGT
  	TTTTCGTTGG
  351	TTTTGCCTTT GATatTGACG CACGAAATAT CGatacCCAa
  	atcggcgGCG
  401	ATGTTGGCAC GCATTTGCGG AATGTGCGGT GCGAGTTTGG
  	GTTTCTGTGC
  451	AATCACGGTC GTATCGACAT TGACCACCTG CCAACCCTGC
  	GCCTGAACGC
  501	TTTGATACGC CGCACGCAAA AGGACGCGGC TGTCCGCATC
  	TTTGAACTCT
  551	GCGGCGGTGT CGGGAAAATG GCTGCCGATG TCGCCCAAAC
  	CTGCCGCACC
  601	GAGCAGCgcg tcggtaaCGG CGTGCAGCAG cgcgTcgGCA
  	TCCGAATGCC
  651	CGAGCAGCCC TTTTTCAAAT GGGATTTCAA CTCCGCCAAG
  	TATCAGCTTT
  701	CTGCCTTCGG TCAATTGGTG GACATCGTAG CCCTGTCCGA
  	TACGGATATT
  751	CGTCATCGTT TGTGTTCCTG A

• This encodes a protein having amino acid sequence <SEQ ID 798>:

• 1	MSYRASSSPD FLEVETAPLI FLPLLPKASM KKLMVEPVPM
  	PMYSFSGTNS
  51	TAFSAAMRLS SSCVVIFLSF GKPYQQTAAI LTFFCTSWPP
  	RSNPYQQYRR
  101	LRLYAFHPPE IAEFFVGFAF DIDARNIDTQ IGGDVGTHLR
  	NVRCEFGFLC
  151	NHGRIDIDHL PTLRLNALIR RTQKDAAVRI FELCGGVGKM
  	AADVAQTCRT
  201	EQRVGNGVQQ RVGIRMPEQP FFKWDFNSAK YQLSAFGQLV
  	DIVALSDTDI
  251	RHRLCS*

• ORF122ng and ORF122-1 show 92.6% identity in 256 aa overlap:
• 
• Based on this analysis, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 95
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 799>:

• 1	..GCCGGCGCGA GTGCGAACAA CATTTCCGCG CGTTTTGCGG
  	AAACACCCGT
  51	CGCTGTCAGC GTTACCCTGA TCGGCACGGT ACTTGCCGTC
  	ATGCTGCCCG
  101	TTACCGAATA TGAAAACTTC CTGCTGCTTA TCGGCTCGGT
  	ATTTGCGCCG
  151	ATGGGGCGGA TTTTGATTGC CGACTTTTTC GTCTTGAAAC
  	GGCGTGA

• This corresponds to the amino acid sequence <SEQ ID 800; ORF125>:

• 1	..AGASANNISA RFAETPVAVS VTLIGTVLAV MLPVTEYENF
  	LLLIGSVFAP
  51	MGGFDCRLFR LETA*

• Further work revealed the complete nucleotide sequence <SEQ ID 801>:

• 1	ATGTCGGGCA ATGCCTCCTC TCCTTCATCT TCCTCCGCCA
  	TCGGGCTGAT
  51	TTGGTTCGGC GCGGCGGTAT CGATTGCCGA AATCAGCACG
  	GGTACGCTGC
  101	TTGCGCCTTT GGGCTGGCAG CGCGGTCTGG CGGCTCTACT
  	TTTGGGTCAT
  151	GCCGTCGGCG GCGCGCTGTT TTTTGCGGCG GCGTATATCG
  	GCGCACTGAC
  201	CGGACGCAGC TCGATGGAAA GCGTGCGCCT GTCGTTCGGC
  	AAACGCGGTT
  251	CAGTGCTGTT TTCCGTGGCG AATATGCTGC AACTGGCCGG
  	CTGGACGGCG
  301	GTGATGATTT ACGCCGGCGC AACGGTCAGC TCCGCTTTGG
  	GCAAAGTGTT
  351	GTGGGACGGC GAATCTTTTG TCTGGTGGGC ATTGGCAAAC
  	GGCGCGCTGA
  401	TTGTGCTGTG GCTGGTTTTC GGCGCACGCA AAACAGGCGG
  	GCTGAAAACC
  451	GTTTCGATGC TGCTGATGCT GTTGGCGGTT CTGTGGCTGA
  	GTGCCGAAGT
  501	CTTTTCCACG GCAGGCAGCA CCGCCGCACA GGTTTCAGAC
  	GGCATGAGTT
  551	TCGGAACGGC AGTCGAGCTG TCCGCCGTGA TGCCGCTTTC
  	CTGGCTGCCG
  601	CTTGCCGCCG ACTACACGCG CCACGCGCGC CGCCCGTTTG
  	CGGCAACCCT
  651	GACGGCAACG CTCGCCTACA CGCTGACCGG CTGCTGGATG
  	TATGCCTTGG
  701	GTTTGGCAGC GGCGTTGTTC ACCGGAGAAA CCGACGTGGC
  	AAAAATCCTG
  751	CTGGGCGCAG GTTTGGGTGC GGCAGGCATT TTGGCGGTCG
  	TCCTCTCCAC
  801	CGTTACCACA ACGTTTCTCG ATGCCTATTC CGCCGGCGCG
  	AGTGCGAACA
  851	ACATTTCCGC GCGTTTTGCG GAAACACCCG TCGCTGTCGG
  	CGTTACCCTG
  901	ATCGGCACGG TACTTGCCGT CATGCTGCCC GTTACCGAAT
  	ATGAAAACTT
  951	CCTGCTGCTT ATCGGCTCGG TATTTGCGCC GATGGCGGCG
  	GTTTTGATTG
  1001	CCGACTTTTT CGTCTTGAAA CGGCGTGAGG AGATTGAAGG
  	CTTTGACTTT
  1051	GCCGGACTGG TTCTGTGGCT TGCGGGCTTC ATCCTCTACC
  	GCTTCCTGCT
  1101	CTCGTCCGGC TGGGAAAGCA GCATCGGTCT GACCGCCCCC
  	GTAATGTCTG
  1151	CCGTTGCCAT TGCCACCGTA TCGGTACGCC TTTTCTTTAA
  	AAAAACCCAA
  1201	TCTTTACAAA GGAACCCGTC ATGA

• This corresponds to the amino acid sequence <SEQ ID 802; ORF125-1>:

• 1	MSGNASSPSS SSAIGLIWFG AAVSIAEIST GTLLAPLGWQ
  	RGLAALLLGH
  51	AVGGALFFAA AYIGALTGRS SMESVRLSFG KRGSVLFSVA
  	NMLQLAGWTA
  101	VMIYAGATVS SALGKVLWDG ESFVWWALAN GALIVLWLVF
  	GARKTGGLKT
  151	VSMLLMLLAV LWLSAEVFST AGSTAAQVSD GMSFGTAVEL
  	SAVMPLSWLP
  201	LAADYTRHAR RPFAATLTAT LAYTLTGCWM YALGLAAALF
  	TGETDVAKIL
  251	LGAGLGAAGI LAVVLSTVTT TFLDAYSAGA SANNISARFA
  	ETPVAVGVTL
  301	IGTVLAVMLP VTEYENFLLL IGSVFAPMAA VLIADFFVLK
  	RREEIEGFDF
  351	AGLVLWLAGF ILYRFLLSSG WESSIGLTAP VMSAVAIATV
  	SVRLFFKKTQ
  401	SLQRNPS*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF125 shows 76.5% identity over a 51 as overlap with an ORF (ORF125a) from strain A of N. meningitidis:
• 
• The ORF125a partial nucleotide sequence <SEQ ID 803> is:

• 1	ATGTCGGGCA ATGCCTCCTC TCNTTCATCT TCCGCCGCCA
  	TCGGGCTGAT
  51	TTGGTTCGGC GCGGCGGTAT CGATTGCCGA AATCAGCACG
  	GGTACACTGC
  101	TTGCGCCTTT GGGCTGGCAG CGCGGTCTGG CNGCTCTGCT
  	TTTGGGTCAT
  151	GCCGTCGGCG GCGCGCTGTT TTTTGCGGCG GCGTATATCG
  	GCGCACTGAC
  201	CGGACNCANC TCGATGGAAA GCGTGCGCCT GTCGTTCGGC
  	AAACGCGGTT
  251	CAGTGCTGTT TTCCGTGGCG AATATGCTGC AACTGGCCGG
  	CTGGACGGCG
  301	GTGATGATTT ACGCCGGCGC AACGGTCAGC TCCGCTTTGG
  	GCAAAGTGTT
  351	GTGGGACGGC GAATCTTTTG TCTGGTGGGC ATTGGCAAAC
  	GGCGCGCTGA
  401	TTGTGCTGTG GCTGGTTTTC GGCGCACGCA AAACAGGCGG
  	GCTGAAAACC
  451	GTTTCGATGC TGCTGATGCT GTTGGCGGTT CTGTGGCTGA
  	GTGCCGAANT
  501	NTTTTCCACG GCAGGCAGCA CCGCCGCANN GGTNNCAGAC
  	GGCATGAGTT
  551	TCGGAACGGC AGTCGAGCTG TCCGCCGTNA TGCCGCTTTC
  	TTGGCTGCCG
  601	CTGGCCGCCG ACTACACGCG CCACGCGCGC CGCCCGTTTG
  	CGGCAACCCT
  651	GACGGCAACG CTCGCCTACA CGCTGACCGG CTGCTGGATG
  	TATGCCTTGG
  701	GTTTGGCAGC GGCGTTGTTC ACCGGAGAAA CCGACGTGGC
  	AAAAATCCTG
  751	CTGGGCGCAG GTTTGGGTGC GGCAGGCATT TTGGCGGTCG
  	TCCTGTCGAC
  801	CGTTACCACC ACTTTTCTCG ATGCNTACTC CGCCGGCGTA
  	AGTGCCAACA
  851	ATATTTCCGC CAAACTTTCG GAAATACCNA TCGCCGTTGC
  	CGTCGCCGTT
  901	GTCGGCACAC TGCTTGCCGT CCTCCTGCCC GTTACCGAAT
  	ATGAAAACTT
  951	CCTGCTGCTT ATCGGCTCGG TATTTGCGCC GATGGCGGCG
  	GTTTTGATTG
  1001	CCGACTTTTT CGTCTTGAAA CGGCGTGAGG AGATTGAAGG
  	C..

• This encodes a protein having the partial amino acid sequence <SEQ ID 804>:

• 1	MSGNASSXSS SAAIGLIWFG AAVSIAEIST GTLLAPLGWQ
  	RGLAALLLGH
  51	AVGGALFFAA AYIGALTGXX SMESVRLSFG KRGSVLFSVA
  	NMLQLAGWTA
  101	VMIYAGATVS SALGKVLWDG ESFVWWALAN GALIVLWLVF
  	GARKTGGLKT
  151	VSMLLMLLAV LWLSAEXFST AGSTAAXVXD GMSFGTAVEL
  	SAVMPLSWLP
  201	LAADYTRHAR RPFAATLTAT LAYTLTGCWM YALGLAAALF
  	TGETDVAKIL
  251	LGAGLGAAGI LAVVLSTVTT TFLDAYSAGV SANNISAKLS
  	EIPIAVAVAV
  301	VGTLLAVLLP VTEYENFLLL IGSVFAPMAA VLIADFFVLK
  	RREEIEG..

• ORF125a and ORF125-1 show 94.5% identity in 347 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF125 shows 86.2% identity over a 65aa overlap with a predicted ORF (ORF125ng) from N. gonorrhoeae:
• 
• An ORF125ng nucleotide sequence <SEQ ID 805> was predicted to encode a protein having amino acid sequence <SEQ ID 806>:

• 1	MSGNASSPSS SAAIGLVWFG AAVSIAEIST GTLLAPLGWQ
  	RGLAALLLGH
  51	AVGGALFFAA AYIGALTGRS SMESVRLSFG KCGSVLFSVA
  	NMLQLAGWTA
  101	VMIYVGATVS SALGKVLWDG ESFVWWALAN GALIVLWLVF
  	GARRTGGLKT
  151	VSMLLMLLAV LWLSVEVFAS SGTNAAPAVS DGMTFGTAVE
  	LSAVMPLSWL
  201	PLAADYTRQA RRPFAATLTA TLAYTLTGCW MYALGLAAAL
  	FTGETDVAKI
  251	LLGAGLGITG ILAVVLSTVT TTFLDTYSAG ASANNISARF
  	AEIPVAVGVT
  301	LIRTVLAVML PVTEYKNFLL LIRSVFGPMA GGFDCRLFCL
  	KTA*

• Further work revealed the following gonococcal DNA sequence <SEQ ID 807>:

• 1	ATGTCGGGCA ATGCCTCCTC TCCTTCATCT TCCGCCGCCA
  	TCGGGCTGGT
  51	TTGGTTCGGC GCGGCGGTAT CGATTGCCGA AATCAGCACG
  	GGTACGCTGC
  101	TCGCCCCCTT GGGCTGGCAG CGCGGTCTGG CGGCCCTGCT
  	TTTGGGTCAT
  151	GCCGTCGGCG GCGCGCTGTT TTTTGCGGCG GCGTATATCG
  	GCGCACTGAC
  201	CGGACGCAGC TCGATGGAAA GTGTGCGCCT GTCGTTCGGC
  	AAATGCGGTT
  251	CAGTGCTGTT TTCCGTGGCG AATATGCTGC AACTGGCCGG
  	CTGGACGGCG
  301	GTGATGATTT ACGTCGGCGC AACGGTCAGC TCCGCTTTGG
  	GCAAAGTGTT
  351	GTGGGACGGC GAATCCTTTG TCTGGTGGGC ATTGGCAAAC
  	GGCGCACTGA
  401	TCGTGCTGTG GCTGGTTTTC GGCGCACGCA GAACGGGCGG
  	GCTGAAAACC
  451	GTTTCGATGC TGCTGATGCT GCTTGCCGTG TTGTGGTTGA
  	GCGTCGAAGT
  501	GTTCGCTTCG TCCGGCACAA ACGCCGCGCC CGCCGTTTCA
  	GACGGCATGA
  551	CCTTCGGAAC GGCAGTCGAA CTGTCCGCCG TCATGCCGCT
  	TTCCTGGCTG
  601	CCGCTGGCCG CCGACTACAC GCGCCAAGCA CGCCGCCCGT
  	TTGCGGCAAC
  651	CCTGACGGCA ACGCTCGCCT ATACGCTGAC GGGCTGCTGG
  	ATGTATGCCT
  701	TGGGTTTGGC GGCGGCTCTG TTTACCGGAG AAACCGACGT
  	GGCGAAAATC
  751	CTGTTGGGCG CGGGCTTGGG CATAACGGGC ATTCTGGCAG
  	TCGTCCTCTC
  801	CACCGTTACC ACAACGTTTC TCGATACCTA TTCCGCCGGC
  	GCGAGTGCGA
  851	ACAACATTTC CGCGCGTTTT GCGGAAATAC CCGTCGCTGT
  	CGGCGTTACC
  901	CTGATCGGCA CGGTGCTTGC CGTCATGCTG CCCGTTACCG
  	AATATAAAAA
  951	CTTCCTGCTG CTTATCGGCT CGGTATTTGC GCCGATGGCG
  	GCGGTTTTGA
  1001	TTGCCGACTT TTTCGTCTTA AAACGGCGTG AGGAGATTGA
  	AGGCTTTGAC
  1051	TTTGCCGGAC TGGTTCTGTG GCTGGCAGGC TTCATCCTCT
  	ACCGCTTCCT
  1101	GCTCTCGTCC GGTTGGGAAA GCAGCATCGG TCTGACCGCC
  	CCCGTAATGT
  1151	CTGCCGTTGC CATTGCCACC GTATCGGTAC GCCTTTTCTT
  	TAAAAAAACC
  1201	CAATCTTTAC AAAGGAACCC GTCATGA

• This corresponds to the amino acid sequence <SEQ ID 808; ORF125ng-1>:

• 1	MSGNASSPSS SAAIGLVWFG AAVSIAEIST GTLLAPLGWQ
  	RGLAALLLGH
  51	AVGGALFFAA AYIGALTGRS SMESVRLSFG KCGSVLFSVA
  	NMLQLAGWTA
  101	VMIYVGATVS SALGKVLWDG ESFVWWALAN GALIVLWLVF
  	GARRTGGLKT
  151	VSMLLMLLAV LWLSVEVFAS SGTNAAPAVS DGMTFGTAVE
  	LSAVMPLSWL
  201	PLAADYTRQA RRPFAATLTA TLAYTLTGCW MYALGLAAAL
  	FTGETDVAKI
  251	LLGAGLGITG ILAVVLSTVT TTFLDTYSAG ASANNISARF
  	AEIPVAVGVT
  301	LIGTVLAVML PVTEYKNFLL LIGSVFAPMA AVLIADFFVL
  	KRREEIEGFD
  351	FAGLVLWLAG FILYRFLLSS GWESSIGLTA PVMSAVAIAT
  	VSVRLFFKKT
  401	QSLQRNPS*

• ORF125ng-1 and ORF125-1 show 95.1% identity in 408 aa overlap:
• 
• Based on this analysis, including the presence of putative leader sequence and transmembrane domains in the gonococcal protein, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 96
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 809>:

• 1	ATGACCCGTA TCGCCATCCT CGGCGGCGGC CTCTCGGGAA
  	GGCTGACCGC
  51	GTTGCAGCTT GCAGAACAAG GTTATCAGAT TGCACTTTTC
  	GATAAAAGCT
  101	GCCGCCGGGG CGAACACGCC GCCGCCTATG TAGCCGCCGC
  	CATGCTCGCG
  151	CCTGCAGCGG A.ACGGTCGA AGCCACGCCC GAAGTGGTCA
  	GGCTGGGCAG
  201	GCAGAGCATC CCGCTTTGGC GCGGCATCCG ATGCCGTCTG
  	AACACGCACA
  251	CGATGATGCA GGAAAACGGC AGCCTGATTG TATGGCACGG
  	GCAGGACAAG
  301	CCATTATCCA GCGAGTTCGT CCGCCATCTC AAACGCGGCG
  	GCGT.ACGGA
  351	TGACGAAATC GTCCGTTGGC GCGCCGACGA CATCGCCGAA
  	CGCGAACCGC
  401	AACTCGGCGG ACGTTTTTAA GACGGCATCT ACCTGCCGAC
  	CGAAGC.CAG
  451	CTCGACGGGC GGCAATTATA GTCTGCACTT GCCGACGCTT
  	TGGACGAACT
  501	GAACGTCCCC TGCCATTGGG AACACGAATG CGTCCCCGAA
  	GCCTGCAAG..

• This corresponds to the amino acid sequence <SEQ ID 810; ORF126>:

• 1	MTRIAILGGG LSGRLTALQL AEQGYQIALF DKSCRRGEHA
  	AAYVAAAMLA
  51	PAAXTVEATP EVVRLGRQSI PLWRGIRCRL NTHTMMQENG
  	SLIVWHGQDK
  101	PLSSEFVRHL KRGGXTDDEI VRWRADDIAE REPQLGGRFX
  	DGIYLPTEXQ
  151	LDGRQLXSAL ADALDELNVP CHWEHECVPE ACK...

• Further work revealed the complete nucleotide sequence <SEQ ID 811>:

• 1	ATGACCCGTA TCGCCATCCT CGGCGGCGGC CTCTCGGGAA
  	GGCTGACCGC
  51	GTTGCAGCTT GCAGAACAAG GTTATCAGAT TGCACTTTTC
  	GATAAAGGCT
  101	GCCGCCGGGG CGAACACGCC GCCGCCTATG TTGCCGCCGC
  	CATGCTCGCG
  151	CCTGCGGCGG AAGCGGTCGA AGCCACGCCC GAAGTGGTCA
  	GGCTGGGCAG
  201	GCAGAGCATC CCGCTTTGGC GCGGCATCCG ATGCCGTCTG
  	AACACGCACA
  251	CGATGATGCA GGAAAACGGC AGCCTGATTG TGTGGCACGG
  	GCAGGACAAG
  301	CCATTATCCA GCGAGTTCGT CCGCCATCTC AAACGCGGCG
  	GCGTAGCGGA
  351	TGACGAAATC GTCCGTTGGC GCGCCGACGA CATCGCCGAA
  	CGCGAACCGC
  401	AACTCGGCGG ACGTTTTTCA GACGGCATCT ACCTGCCGAC
  	CGAAGGCCAG
  451	CTCGACGGGC GGCAAATATT GTCTGCACTT GCCGACGCTT
  	TGGACGAACT
  501	GAACGTCCCC TGCCATTGGG AACACGAATG CGTCCCCGAA
  	GGCCTGCAAG
  551	CCCAATACGA CTGGCTGATC GACTGCCGCG GCTACGGCGC
  	AAAAACCGCG
  601	TGGAACCAAT CCCCCGAGCA CACCAGCACC CTGCGCGGCA
  	TACGCGGCGA
  651	AGTGGCGCGG GTTTACACAC CCGAAATCAC GCTCAACCGC
  	CCCGTGCGTC
  701	TGCTCCATCC GCGTTATCCG CTCTACATCG CCCCGAAAGA
  	AAACCACGTC
  751	TTCGTCATCG GCGCGACCCA AATCGAAAGC GAAAGCCAAG
  	CCCCCGCCAG
  801	CGTGCGTTCA GGGTTGGAAC TCTTGTCCGC ACTCTATGCC
  	ATCCACCCCG
  851	CCTTCGGCGA AGCCGACATC CTCGAAATCG CCACCGGCCT
  	GCGCCCCACG
  901	CTCAACCACC ACAACCCCGA AATCCGTTAC AACCGCGCCC
  	GACGCCTGAT
  951	TGAAATCAAC GGCCTTTTCC GCCACGGTTT CATGATCTCC
  	CCCGCCGTAA
  1001	CCGCCGCCGC CGCCAGATTG GCAGTGGCAC TGTTTGACGG
  	AAAAGACGCG
  1051	CCCGAACGCG ATAAAGAAAG CGGTTTGGCG TATATCCGAA
  	GACAAGATTA
  1101	A

• This corresponds to the amino acid sequence <SEQ ID 812; ORF126-1>:

• 1	MTRIAILGGG LSGRLTALQL AEQGYQIALF DKGCRRGEHA
  	AAYVAAAMLA
  51	PAAEAVEATP EVVRLGRQSI PLWRGIRCRL NTHTMMQENG
  	SLIVWHGQDK
  101	PLSSEFVRHL KRGGVADDEI VRWRADDIAE REPQLGGRFS
  	DGIYLPTEGQ
  151	LDGRQILSAL ADALDELNVP CHWEHECVPE GLQAQYDWLI
  	DCRGYGAKTA
  201	WNQSPEHTST LRGIRGEVAR VYTPEITLNR PVRLLHPRYP
  	LYIAPKENHV
  251	FVIGATQIES ESQAPASVRS GLELLSALYA IHPAFGEADI
  	LEIATGLRPT
  301	LNHHNPEIRY NRARRLIEIN GLFRHGFMIS PAVTAAAARL
  	AVALFDGKDA
  351	PERDKESGLA YIRRQD*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF126 shows 90.0% identity over a 180aa overlap with an ORF (ORF126a) from strain A of N. meningitidis:
• 
• The complete length ORF126a nucleotide sequence <SEQ ID 813> is:

• 1	ATGACCCGTA TCGCCATCCT CGGCGGCGGC CTCTCNGGAA
  	GGCTGACCGC
  51	ACTGCAGCTT GCAGAACAAG GTTATCAGAT TGCACTTTTC
  	GATAAAGGCT
  101	GCCGCCGGGG CGAACACGCC GCCGCCTATG TTGCCGCCGC
  	CATGCTCGCG
  151	CCTGCGGCGG AAGCGGTCGA AGCCACGCCT GAAGTGGTCA
  	GGCTGGGCAG
  201	GCAGANCATC CCGCTTTGGC GCGGCATCCG ATGCCATCTG
  	AAAACGCCTG
  251	CCATGATGCA NGAAAACGGC AGCCTGATTG TGTGGCACGG
  	GCAGGACAAA
  301	CCTTTATCCA ACGAGTTCGT CCGCCATCTC AAACGCGGCG
  	GCGTAGCGGA
  351	TGACNAAATC GTCCGTTGGC GCGCCGACGA CATCGCCGAA
  	CGCGAACCGC
  401	AACTCGGCGG ACGTTTTTCA GACGGCATCT ACCTGCCGAC
  	CGAAGGCCAG
  451	CTCGACGGGC GGCAAATATT GTCTGCACTT GCCGACGCTT
  	TGGACGAACT
  501	GAACGTCCCC TGCCATTGGG AACACGAATG TGCCCCCGAA
  	GACTTGCAAG
  551	CCCAATACGA CTGGCTGATC GACTGCCGCG GCTACGGCGC
  	AAAAACCGCG
  601	TGGAACCAAT CCCCCGANNA NACCAGCACC CTGCGCGGCA
  	TACGCGGCGA
  651	AGTGGCGCGG GTTTACACAC CCGAAATCAC GCTCAACCGC
  	CCCGTGCGCC
  701	TGCTACACCC GCGCTATCCG CTNTACATCG CCCCGAAAGA
  	AAACCNCGTC
  751	TTCGTCATCG GCGCGACCCA AATCGAAAGC GAAAGCCAAG
  	CACCTGCCAG
  801	CGTGCGTTCC GGGCTGGAAC TCTTATCCGC ACTCTATGCC
  	GTCCACCCCG
  851	CCTTCGGCGA AGCCGACATC CTCGAAATCG CCACCGGCCT
  	GCGCCCCACG
  901	CTCAATCACC ACAACCCCGA AATCCGTTAC AACCGCGCCC
  	GACGCCTGAT
  951	TGAAATCAAC GGCCTTTTCC GCCACGGTTT CATGATCTCC
  	CCCGCCGTAA
  1001	CCGCCGCCGC CGTCAGATTG GCAGTGGCAC TGTTTGACGG
  	AAAAGANGCG
  1051	CCCGAACGCG ATGAAGAAAG CGGTTTGGCG TATATCCGAA
  	GACAAGATTA
  1101	A

• This encodes a protein having amino acid sequence <SEQ ID 814>:

• 1	MTRIAILGGG LSGRLTALQL AEQGYQIALF DKGCRRGEHA
  	AAYVAAAMLA
  51	PAAEAVEATP EVVRLGRQXI PLWRGIRCHL KTPAMMXENG
  	SLIVWHGQDK
  101	PLSNEFVRHL KRGGVADDXI VRWRADDIAE REPQLGGRFS
  	DGIYLPTEGQ
  151	LDGRQILSAL ADALDELNVP CHWEHECAPE DLQAQYDWLI
  	DCRGYGAKTA
  201	WNQSPXXTST LRGIRGEVAR VYTPEITLNR PVRLLHPRYP
  	LYIAPKENXV
  251	FVIGATQIES ESQAPASVRS GLELLSALYA VHPAFGEADI
  	LEIATGLRPT
  301	LNHHNPEIRY NRARRLIEIN GLFRHGFMIS PAVTAAAVRL
  	AVALFDGKXA
  351	PERDEESGLA YIRRQD*

• ORF126a and ORF126-1 show 95.4% identity in 366 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF126 shows 90% identity over a 180 aa overlap with a predicted ORF (ORF126ng) from N. gonorrhoeae:
• 
• An ORF126ng nucleotide sequence <SEQ ID 815> was predicted to encode a protein having amino acid sequence <SEQ ID 816>:

• 1	MTRIAVLGGG LSGRLTALQL AEQGYQIELF DKGTRQGEHA
  	AAYVAAAMLA
  51	PAAEAVEATP EVIRLGRQSI PLWRGIRCRL NTLTMMQENG
  	SLIVWHGQDK
  101	PLSSEFVRHL KRGGVADDEI VRWRADEIAE REPQLGGRFS
  	DGIYLPTEGQ
  151	LDGRQILSAL ADALDELNVP CHWEHECAPQ DLQAQYDWVI
  	DCRGYGAKTA
  201	WNQSPEHTST LRGIRGEVRG FTRPKSRSTA PCACCTRAIR
  	STSPRKKTTS
  251	SSSARPKSKA KAKPPPAYVP GWNSYPRSMP STPPSAKPTS
  	SKWRPGLRPT
  301	LNHHNPEIRY SRERRLIEIN GLFRHGFMIS PAVTAAAVRL
  	AVALFDGKDA
  351	PERDEESGLA YIGRQD*

• Further work revealed the following gonococcal DNA sequence <SEQ ID 817>:

• 1	ATGACCCGTA TCGCCGTCCT CGGAGGCGGC CTTTCCGGAA
  	GGCTGACCGC
  51	ATTGCAGCTT GCAGAACAAG GTTATCAGAT TGAACTTTTC
  	GACAAGGGCA
  101	CCCGCCAAGG CGAACACGCC GCCGCCTATG TTGCCGCCGC
  	GATGCTCGCG
  151	CCTGCGGCGG AAGCGGTCGA GGCAACGCCC GAAGTCATCA
  	GGCTGGGCAG
  201	GCAGAGCATT CCGCTTTGGC GCGGCATCCG ATGCCGTCTG
  	AACACGCTCA
  251	CGATGATGCA GGAAAACGGC AGCCTGATTG TGTGGCACGG
  	GCAGGACAAG
  301	CCATTATCCA GCGAGTTCGT CCGCCATCTC AAACGCGGCG
  	GCGTAGCGGA
  351	TGACGAAATC GTCCGTTGGC GCGCCGATGA AATCGCCGAA
  	CGCGAACCGC
  401	AACTCGGCGG ACGTTTTTCA GACGGCATCT ACCTGCCGAC
  	CGAAGGCCAG
  451	CTCGACGGGC GGCAAATATT GTCTGCACTT GCCGACGCTT
  	TGGACGAACT
  501	GAACGTCCCT TGCCATTGGG AACACGAATG CGCCCCCCAA
  	GACCTGCAAG
  551	CCCAATACGA CTGGGTAATC GACTGCCGGG GCTACGGCGC
  	GAAAACCGCG
  601	TGGAACCAAT CCCCCGAGCA CACCAGCACC TTGCGCGGCA
  	TACGCGGCGA
  651	AGTGGCGCGG GTTTACACGC CCGAAATCAC GCTCAACCGC
  	CCCGTGCGCC
  701	TGCTGCACCC GCGCTATCCG CTCTACATCG CCCCGAAAGA
  	AAACCACGTC
  751	TTCGTCATCG GCGCGACCCA AATCGAAAGC GAAAGCCAAG
  	CCCCCGCCAG
  801	CGTACGTTCC GGGCTGGAAC TCTTATCCGC GCTCTATGCC
  	GTCCACCCCG
  851	CCTTCGGCGA AGCCGACATC CTCGAAATCG CCGCCGGCCT
  	GCGCCCCACG
  901	CTCAACCACC ACAACCCCGA AATCCGCTAC AGCCGCGAAC
  	GCCGCCTCAT
  951	CGAAATCAAC GGCCTTTTCC GGCACGGCTT TATGATTTCC
  	CCCGCCGTAA
  1001	CCGCCGCCGC CGTCAGATTG GCAGTGGCAC TGTTTGACGG
  	AAAAGACGCG
  1051	CCCGAACGTG ATGAAGAAAG CGGTTTGGCG TATATCGGAA
  	GACAAGATTA
  1101	A

• This corresponds to the amino acid sequence <SEQ ID 818; ORF126ng-1>:

• 1	MTRIAVLGGG LSGRLTALQL AEQGYQIELF DKGTRQGEHA
  	AAYVAAAMLA
  51	PAAEAVEATP EVIRLGRQSI PLWRGIRCRL NTLTMMQENG
  	SLIVWHGQDK
  101	PLSSEFVRHL KRGGVADDEI VRWRADEIAE REPQLGGRFS
  	DGIYLPTEGQ
  151	LDGRQILSAL ADALDELNVP CHWEHECAPQ DLQAQYDWVI
  	DCRGYGAKTA
  201	WNQSPEHTST LRGIRGEVAR VYTPEITLNR PVRLLHPRYP
  	LYIAPKENHV
  251	FVIGATQIES ESQAPASVRS GLELLSALYA VHPAFGEADI
  	LEIAAGLRPT
  301	LNHHNPEIRY SRERRLIEIN GLFRHGFMIS PAVTAAAVRL
  	AVALFDGKDA
  351	PERDEESGLA YIGRQD*

• ORF126ng-1 and ORF126-1 show 95.1% identity in 366 aa overlap:
• 
• Furthermore, ORF126ng-1 shows homology to a putative Rhizobium oxidase flavoprotein:

• gi|2627327 (AF004408) putative amino acid oxidase
  flavoprotein [Rhizobium etli]
  Length = 327
  Score = 169 bits (423), Expect = 3e−41
  Identities = 112/329 (34%), Positives = 163/329 (49%), Gaps = 25/329 (7%)

  Query:	3	RIAVLGGGLSGRLTALQLAEQGYQIELFDKGTRQGEHXXXXXXXXXXXXXXXXXXXXXXX	62
  		RI V G G++G   A QL   G+++ L ++    G
  Sbjct:	2	RILVNGAGVAGLTVAWQLYRHGFRVTLAERAGTVGA-GASGFAGGMLAPWCERESAEEPV	60
  Query:	63	IRLGRQSIPLWRGIRCRLNTLTMMQENGSLIVWHGQDKPLSSEFVRHLKRGGVADDEIVR	122
  		+ LGR +   W            +   G+L+V  G+D      F R    G    DE+
  Sbjct:	61	LTLGRLAADWWEAA-----LPGHVHRRGTLVVAGGRDTGELDRFSRRTS-GWEWLDEVA-	113
  Query:	123	WRADEIAEREPQLGGRFSDGIYLPTEGQLDGRQILSALADALDELNVPCHWEHECAPQDL	182
  		IA  EP L GRF   ++   E  LD RQ L+ALA  L++  +           +
  Sbjct:	114	-----IAALEPDLAGRFRRALFFRQEAHLDPRQALAALAAGLEDARMRLTLG---VVGES	165
  Query:	183	QAQYDWVIDCRGYGAKTAWNQSPEHTSTLRGIRGEVARVYTPEITLNRPVRLLHPRYPLY	242
  		+D V+DC G                LRG+RGE+  V T E++L+RPVRLLHPR+P+Y
  Sbjct:	166	DVDHDRVVDCTGAA-------QIGRLPGLRGVRGEMLCVETTEVSLSRPVRLLHPRHPIY	218
  Query:	243	IAPKENHVFVIGATQIESESQAPASVRSGLELLSALYAVHPAFGEADILEIAAGLRPTLN	302
  		I P++ + F++GAT IES+   P + RS +ELL+A YA+HPAFGEA + E  AG+RP
  Sbjct:	219	IVPRDKNRFMVGATMIESDDGGPITARSLMELLNAAYAMHPAFGEARVTETGAGVRPAYP	278
  Query:	303	HHNPEIRYSRERRLIEINGLFRHGFMISP	331
  		+ P  R ++E R + +NGL+RHGF+++P
  Sbjct:	279	DNLP--RVTQEGRTLHVNGLYRHGFLLAP	305

• This analysis suggests that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 97
• The following DNA sequence, believed to be complete, was identified in N. meningitidis <SEQ ID 819>:

• 1	ATGACTGATA ATCGGGGGTT TACGCTGGTT GAATTAATAT
  	CAGTGGTCTT
  51	GATATTGTCT GTACTTGCTT TAATTGTTTA TCCGAGCTAT
  	CGCAATTATG
  101	TTGAGAAAGC AAAGATAAAT GCAGTGCGGG CAGCCTTGTT
  	AGAAAATGCA
  151	CATTTTATGG AAAAGTTTTA TCTGCAGAAT GGGAGGTTTA
  	AACAAACATC
  201	TACCAAGTGG CCAAGTTTGC CGATTAAAGA GGCAGAAGGC
  	TTTTGTATCC
  251	GTTTGAATGG AATCGtCGCG CGGG..GCTT TAGACAGTAA
  	ATTCATGTTG
  301	AAGGCGGTAG CCATAGATAA AGATAAAAAT CCTTTTATTA
  	TTAAGATGAA
  351	TGAAAATCTA GTAACCTTTA aTTTGCAAGA AGTCCGCCAG
  	TTCGTGTAGT
  401	GACGGGCTGG ATTATTTTAA AGGAAATGAT AAGGACTGCA
  	AGTTACTTAA
  451	GTAG

• This corresponds to the amino acid sequence <SEQ ID 820; ORF127>:

• 1	MTDNRGFTLV ELISVVLILS VLALIVYPSY RNYVEKAKIN
  	AVRAALLENA
  51	HFMEKFYLQN GRFKQTSTKW PSLPIKEAEG FCIRLNGIVA
  	RXALDSKFML
  101	KAVAIDKDKN PFIIKMNENL VTFICKKSAS SCSDGLDYFK
  	GNDKDCKLLK
  151	*

• Further work revealed the following DNA sequence <SEQ ID 821>:

• 1	ATGACTGATA ATCGGGGGTT TACGCTGGTT GAATTAATAT
  	CAGTGGTCTT
  51	GATATTGTCT GTACTTGCTT TAATTGTTTA TCCGAGCTAT
  	CGCAATTATG
  101	TTGAGAAAGC AAAGATAAAT GCAGTGCGGG CAGCCTTGTT
  	AGAAAATGCA
  151	CATTTTATGG AAAAGTTTTA TCTGCAGAAT GGGAGGTTTA
  	AACAAACATC
  201	TACCAAGTGG CCAAGTTTGC CGATTAAAGA GGCAGAAGGC
  	TTTTGTATCC
  251	GTTTGAATGG AATCGCGCGC GGGGCTTTAG ACAGTAAATT
  	CATGTTGAAG
  301	GCGGTAGCCA TAGATAAAGA TAAAAATCCT TTTATTATTA
  	AGATGAATGA
  351	AAATCTAGTA ACCTTTATTT GCAAGAAGTC CGCCAGTTCG
  	TGTAGTGACG
  401	GGCTGGATTA TTTTAAAGGA AATGATAAGG ACTGCAAGTT
  	ACTTAAGTAG

• This corresponds to the amino acid sequence <SEQ ID 822; ORF127-1>:

• 1	MTDNRGFTLV ELISVVLILS VLALIVYPSY RNYVEKAKIN
  	AVRAALLENA
  51	HFMEKFYLQN GRFKQTSTKW PSLPIKEAEG FCIRLNGIAR
  	GALDSKFMLK
  101	AVAIDKDKNP FIIKMNENLV TFICKKSASS CSDGLDYFKG
  	NDKDCKLLK*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF127 shows 98.0% identity over a 150aa overlap with an ORF (ORF127a) from strain A of N. meningitidis:
• 
• The complete length ORF127a nucleotide sequence <SEQ ID 823> is:

• 1	ATGACTGATA ATCGGGGGTT TACGCTGGTT GAATTAATAT
  	CAGTGGTCTT
  51	GATATTGTCT GTACTTGCTT TAATTGTTTA TCCGAGCTAT
  	CGCAATTATG
  101	TTGAGAAAGC AAAGATAAAT ACAGTGCGGG CAGCCTTGTT
  	AGAAAATGCA
  151	CATTTTATGG AAAAGTTTTA TCTGCAGAAT GGGAGATTTA
  	AACAAACATC
  201	TACCAAATGG CCAAGTTTGC CGATTAAAGA GGCAGAAGGC
  	TTTTGTATCC
  251	GTTTGAATGG AATCGCGCGC GGGGCCTTAG ACAGTAAATT
  	CATGTTGAAG
  301	GCGGTAGCCA TAGATAAAGA TAAAAATCCT TTTATTATTA
  	AGATGAATGA
  351	AAATCTAGTA ACCTTTATTT GCAAGAAGTC CGCCAGTTCG
  	TGTAGTGACG
  401	GGCTGGATTA TTTTAAAGGA AATGATAAGG ACTGCAAGTT
  	ACTTAAGTAG

• This encodes a protein having amino acid sequence <SEQ ID 824>:

• 1	MTDNRGFTLV ELISVVLILS VLALIVYPSY RNYVEKAKIN
  	TVRAALLENA
  51	HFMEKFYLQN GRFKQTSTKW PSLPIKEAEG FCIRLNGIAR
  	GALDSKFMLK
  101	AVAIDKDKNP FIIKMNENLV TFICKKSASS CSDGLDYFKG
  	NDKDCKLLK*

• ORF127a and ORF127-1 show 99.3% identity in 149 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF127 shows 97.3% identity over a 150 aa overlap with a predicted ORF (ORF127ng) from N. gonorrhoeae:
• 
• The complete length ORF127ng nucleotide sequence <SEQ ID 825> is:

• 1	ATGACTGATA ATCGGGGGTT TACACTGGTT GAATTAATAT
  	CAGTGGTCTT
  51	GATATTGTCT GTACTTGCTT TAATTGTTTA TCCGAGCTAT
  	CGCAATTATG
  101	TTGAGAAAGC AAAGATAAAT GCAGTGCGGG CAGCCTTGTT
  	AGAAAATGCA
  151	CATTTTATGG AAAAGTTTTA TCTGCAGAAT GGGAGATTTA
  	AACAAACATC
  201	TACCAAATGG CCAAGTTTGC CGATTAAAGA GGCAGAAGGC
  	TTTTGTATCC
  251	GTTTGAATGG AATCGCGCGC GGGGCTTTAG ACAGTAAATT
  	CATGTTGAAG
  301	GCGGTAGCCA TAGATAAAGA TAAAAATCCT TTTATTATTA
  	AGATGAATGA
  351	AAATCTAGTA ACCTTTATTT GCAAGAAGTC CGCCAGTTCG
  	TGTAGTGACG
  401	GGCTGGATTA TTTTAAAGGA AATGATAAGG ACTGCAAGTT
  	ACTTAAGTAG

• This encodes a protein having amino acid sequence <SEQ ID 826>:

• 1	MTDNRGFTLV ELISVVLILS VLALIVYPSY RNYVEKAKIN
  	AVRAAFLENA
  51	HFMEKFYLQN GRFKQTSTKW PSLPIKEAEG FCIRLNGIAR
  	GALDSKFMLK
  101	AVAIDKDKNP FIIKMNENLV TFICKKSASS CSDRLDYFKG
  	NDKDCKLLK*

• ORF127ng and ORF127-1 show 100.0% identity in 149 aa overlap:
• 
• This analysis, including the fact that the predicted transmembrane domain is shared by the meningococcal and gonococcal proteins, suggests that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 98
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 827>

• 1	..GTGTCGCTGG CTTCGGTGAT TGCCTCTCAA ATCTTCCTTT
  	ACGAAGATTT
  51	CAACCAAATG CGGAAAACCC GTGGAGCTAT CTGCGGTTTT
  	CTTGTCCAAT
  101	ATTTATCTGG GGTTTCAGCA GGGGTATTTC GATTTGAGTG
  	CCGACGAGAA
  151	CCCCGTACTG CATATCTGGT CTTTGGCAGT AGAGGAACAG
  	TATTACCTCC
  201	TGTATCCCCT TTTGCTGATA TTTTGCTGCA AAAAAACCAA
  	ATCGCTACGG
  251	GTGCTGCGTA ACATCAGCAT CATCCTGTTT TTGATTTTGA
  	CTGCCTCATC
  301	GTTTTTGCCA AGCGGGTTTT ATACCGACAT CCTCAACCAA
  	CCCAATACTT
  351	ATTACCTTTC GACACTGAGG TTTCCCGAGC TGTTGGCAGG
  	TTCGCTGCTG
  401	GCGGTTTACG GGCAAACGCA AAACGGCAGA CGGCAAACAG
  	CAAATGGAAA
  451	ACGGCAGTTG CTTTCATCAC TCTGCTTCGG CGCATTGCTT
  	GCCTGCCTGT
  501	TCGTGATTGA CAAACACAAT CCGTTTATCC CGGGAATGAC
  	CCTGCTCCTT
  551	CCCTGCCTGC TGACGGCACT GCTTATCCGG AGTATGCAAT
  	ACGGGACACT
  601	TCCGACCCGC ATCCTGTCGG CAAGCCCCAT CGTATTTGTC
  	GGCAAAATCT
  651	CTTATTCCCT ATACCTGTAC CATTGGATTT TTATTGCTTT
  	CGCTCCGCTC
  701	ATTAGAGGCG GGAAACAGCT CGGACTGCCT GCCG..

• This corresponds to the amino acid sequence <SEQ ID 828; ORF128>:

• 1	..VSLASVIASQ IFLYEDFNQM RKTVELSAVF LSNIYLGFQQ
  	GYFDLSADEN
  51	PVLHIWSLAV EEQYYLLYPL LLIFCCKKTK SLRVLRNISI
  	ILFLILTASS
  101	FLPSGFYTDI LNQPNTYYLS TLRFPELLAG SLLAVYGQTQ
  	NGRRQTANGK
  151	RQLLSSLCFG ALLACLFVID KHNPFIPGMT LLLPCLLTAL
  	LIRSMQYGTL
  201	PTRILSASPI VFVGKISYSL YLYHWIFIAF APLIRGGKQL
  	GLPA..

• Further work revealed the complete nucleotide sequence <SEQ ID 829>:

• 1	ATGCAAGCTG TCCGATACAG ACCGGAAATT GACGGATTGC
  	GGGCCGTCGC
  51	CGTGCTATCC GTCATGATTT TCCACCTGAA TAACCGCTGG
  	CTGCCCGGAG
  101	GATTCCTGGG GGTGGACATT TTCTTTGTCA TCTCAGGATT
  	CCTCATTACC
  151	GGCATCATTC TTTCTGAAAT ACAGAACGGT TCTTTTTCTT
  	TCCGGGATTT
  201	TTATACCCGC AGGATTAAGC GGATTTATCC TGCCTTTATT
  	GCGGCCGTGT
  251	CGCTGGCTTC GGTGATTGCC TCTCAAATCT TCCTTTACGA
  	AGATTTCAAC
  301	CAAATGCGGA AAACCGTGGA GCTTTCTGCG GTTTTCTTGT
  	CCAATATTTA
  351	TCTGGGGTTT CAGCAGGGGT ATTTCGATTT GAGTGCCGAC
  	GAGAACCCCG
  401	TACTGCATAT CTGGTCTTTG GCAGTAGAGG AACAGTATTA
  	CCTCCTGTAT
  451	CCCCTTTTGC TGATATTTTG CTGCAAAAAA ACCAAATCGC
  	TACGGGTGCT
  501	GCGTAACATC AGCATCATCC TGTTTTTGAT TTTGACTGCC
  	TCATCGTTTT
  551	TGCCAAGCGG GTTTTATACC GACATCCTCA ACCAACCCAA
  	TACTTATTAC
  601	CTTTCGACAC TGAGGTTTCC CGAGCTGTTG GCAGGTTCGC
  	TGCTGGCGGT
  651	TTACGGGCAA ACGCAAAACG GCAGACGGCA AACAGCAAAT
  	GGAAAACGGC
  701	AGTTGCTTTC ATCACTCTGC TTCGGCGCAT TGCTTGCCTG
  	CCTGTTCGTG
  751	ATTGACAAAC ACAATCCGTT TATCCCGGGA ATGACCCTGC
  	TCCTTCCCTG
  801	CCTGCTGACG GCACTGCTTA TCCGGAGTAT GCAATACGGG
  	ACACTTCCGA
  851	CCCGCATCCT GTCGGCAAGC CCCATCGTAT TTGTCGGCAA
  	AATCTCTTAT
  901	TCCCTATACC TGTACCATTG GATTTTTATT GCTTTCGCCC
  	ATTACATTAC
  951	AGGCGACAAA CAGCTCGGAC TGCCTGCCGT ATCGGCGGTT
  	GCCGCGTTGA
  1001	CGGCCGGATT TTCCCTGTTG AGTTATTATT TGATTGAACA
  	GCCGCTTAGA
  1051	AAACGGAAGA TGACCTTCAA AAAGGCATTT TTCTGCCTCT
  	ATCTCGCCCC
  1101	GTCCCTGATA CTTGTCGGTT ACAACCTGTA CGCAAGGGGG
  	ATATTGAAAC
  1151	AGGAACACCT CCGCCCGTTG CCCGGCGCGC CCCTTGCTGC
  	GGAAAATCAT
  1201	TTTCCGGAAA CCGTCCTGAC CCTCGGCGAC TCGCACGCCG
  	GACACCTGAG
  1251	GGGGTTTCTG GATTATGTCG GCAGCCGGGA AGGGTGGAAA
  	GCCAAAATCC
  1301	TGTCCCTCGA TTCGGAGTGT TTGGTTTGGG TAGATGAGAA
  	GCTGGCAGAC
  1351	AACCCGTTAT GTCGAAAATA CCGGGATGAA GTTGAAAAAG
  	CCGAAGCCGT
  1401	TTTCATTGCC CAATTCTATG ATTTGAGGAT GGGCGGCCAG
  	CCTGTGCCGA
  1451	GATTTGAAGC GCAATCCTTC CTAATACCCG GGTTCCCAGC
  	CCGATTCAGG
  1501	GAAACCGTCA AAAGGATAGC CGCCGTCAAA CCCGTCTATG
  	TTTTTGCAAA
  1551	CAACACATCA ATCAGCCGTT CGCCCCTGAG GGAGGAAAAA
  	TTGAAAAGAT
  1601	TTGCCGCAAA CCAATATCTC CGCCCCATTC AGGCTATGGG
  	CGACATCGGC
  1651	AAGAGCAATC AGGCGGTCTT TGATTTGATT AAAGATATTC
  	CCAATGTGCA
  1701	TTGGGTGGAC GCACAAAAAT ACCTGCCCAA AAACACGGTC
  	GAAATATACG
  1751	GCCGCTATCT TTACGGCGAC CAAGACCACC TGACCTATTT
  	CGGTTCTTAT
  1801	TATATGGGGC GGGAATTCCA CAAACACGAA CGCCTGCTTA
  	AATCTTCCCA
  1851	CGGCGGCGCA TTGCAGTAG

• This corresponds to the amino acid sequence <SEQ ID 830; ORF128-1>:

• 1	MQAVRYRPEI DGLRAVAVLS VMIFHLNNRW LPGGFLGVDI
  	FFVISGFLIT
  51	GIILSEIQNG SFSFRDFYTR RIKRIYPAFI AAVSLASVIA
  	SQIFLYEDFN
  101	QMRKTVELSA VFLSNIYLGF QQGYFDLSAD ENPVLHIWSL
  	AVEEQYYLLY
  151	PLLLIFCCKK TKSLRVLRNI SIILFLILTA SSFLPSGFYT
  	DILNQPNTYY
  201	LSTLRFPELL AGSLLAVYGQ TQNGRRQTAN GKRQLLSSLC
  	FGALLACLFV
  251	IDKHNPFIPG MTLLLPCLLT ALLIRSMQYG TLPTRILSAS
  	PIVFVGKISY
  301	SLYLYHWIFI AFAHYITGDK QLG LPAVSAV AALTAGFSLL
  	SYYLIEQPLR
  351	KRKMTFKKAF FCLYLAPSLI LVGYNLYARG ILKQEHLRPL
  	PGAPLAAENH
  401	FPETVLTLGD SHAGHLRGFL DYVGSREGWK AKILSLDSEC
  	LVWVDEKLAD
  451	NPLCRKYRDE VEKAEAVFIA QFYDLRMGGQ PVPRFEAQSF
  	LIPGFPARFR
  501	ETVKRIAAVK PVYVFANNTS ISRSPLREEK LKRFAANQYL
  	RPIQAMGDIG
  551	KSNQAVFDLI KDIPNVHWVD AQKYLPKNTV EIYGRYLYGD
  	QDHLTYFGSY
  601	YMGREFHKHE RLLKSSHGGA LQ*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with Hypothetical Integral Membrane Protein H10392 of H. influenzae (Accession Number U32723)
• ORF128 and HI0392 show 52% aa identity in 180aa overlap:

• Orf128:	1	VSLASVIASQIFLYEDFNQMRKTVELSAVFLSNIYLGFQQGYFDLSADENPVLHIWSLAV	60
  		++L S IAS IF+Y DFN++RKT+EL+  FLSN YLG  QGYFDLSA+ENPVLHIWSLAV
  HI0392:	46	MALVSFIASAIFIYNDFNKLRKTIELAIAFLSNFYLGLTQGYFDLSANENPVLHIWSLAV	105
  Orf128:	61	EEQXXXXXXXXXIFCCKKTKSLRVLRNISIILFLILTASSFLPSGFYTDILNQPNTYYLS	120
  		E Q         I   KK + ++VL  I++ILF IL A+SF+ + FY ++L+QPN YYLS
  HI0392:	106	EGQYYLIFPLILILAYKKFREVKVLFIITLILFFILLATSFVSANFYKEVLHQPNIYYLS	165
  Orf128:	121	TLRFPELLAGSLLAVYGQTQNGRRQTANGKRQLLSSLCFGALLACLFVIDKHNPFIPGMT	180
  		LRFPELL GSLLA+Y    N + Q +     +L+ L    L +CLF+++ +  FIPG+T
  HI0392:	166	NLRFPELLVGSLLAIYHNLSN-KVQLSKQVNNILAILSTLLLFSCLFLMNNNIAFIPGIT	224

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF128 shows 98.0% identity over a 244aa overlap with an ORF (ORF128a) from strain A of N. meningitidis:
• 
• The complete length ORF128a nucleotide sequence <SEQ ID 831> is:

• 1	ATGCAAGCTG TCCGATACAG ACCGGAAATT GACGGATTGC
  	GGGCCGTCGC
  51	CGTGCTATCC GTCATGATTT TCCACCTGAA TAACCGCTGG
  	CTGCCCGGAG
  101	GATTCCTGGG GGTGGACATT TTCTTTGTCA TCTCAGGATT
  	CCTCATTACC
  151	GGCATCATTC TTTCTGAAAT ACAGAACGGT TCTTTTTCTT
  	TCCGGGATTT
  201	TTATACCCGC AGGATTAAGC GGATTTATCC TGCTTTTATT
  	GCGGCCGTGT
  251	CGCTGGCTTC GGTGATTGCC TCTCAAATCT TCCTTTACGA
  	AGATTTCAAC
  301	CAAATGCGGA AAACCGTGGA GCTTTCTGCG GTTTTCTTGT
  	CCAATATTTA
  351	TCTGGGGTTT CAGCAGGGGT ATTTCGATTT GAGTGCCGAC
  	GAGAACCCCG
  401	TACTGCATAT CTGGTCTTTG GCAGTAGAGG AACAGTATTA
  	CCTCCTGTAT
  451	CCTCTTTTGC TGATATTTTG CTGCAAAAAA ACAAAATCGC
  	TACGGGTGCT
  501	GCGTAACATC AGCATCATCC TATTTCTGAT TTTGACTGCC
  	ACATCGTTTT
  551	TGCCAAGCGG GTTTTATACC GATATTCTCA ACCAACCCAA
  	TACTTATTAC
  601	CTTTCGACAC TGAGGTTTCC CGAGCTGTTG GCAGGTTCGC
  	TGCTGGCGGT
  651	TTACGGGCAA ACGCAAAACG GCAGACGGCA AACAGCAAAT
  	GGAAAACGGC
  701	AGTTGCTTTC ATCACTCTGC TTCGGCGCAT TGCTTGCCTG
  	CCTGTTCGTG
  751	ATTGACAAAC ACAATCCGTT TATCCCGGGA ATGACCCTGC
  	TCCTTCCCTG
  801	CCTGCTGACG GCACTGCTTA TCCGGAGTAT GCAATACGGG
  	ACACTTCCGA
  851	CCCGCATCCT GTCGGCAAGC CCCATCGTAT TTGTCGGCAA
  	AATCTCTTAT
  901	TCCCTATACC TGTACCATTG GATTTTTATT GCTTTCGCCC
  	ATTACATTAC
  951	AGGCGACAAA CAGCTCGGAC TGCCTGCCGT ATCGGCGGTT
  	GCCGCGTTGA
  1001	CGGCCGGATT TTCCCTGTTG AGTTATTATT TGATTGAACA
  	GCCGCTTAGA
  1051	AAACGGAAGA TGACCTTCAA AAAGGCATTT TTCTGCCTCT
  	ATCTCGCCCC
  1101	GTCCCTGATA CTTGTCGGTT ACAACCTGTA CGCAAGGGGG
  	ATATTGAAAC
  1151	AGGAACACCT CCGCCCGTTG CCCGGCGCGC CCCTTGCTGC
  	GGAAAATCAT
  1201	TTTCCGGAAA CCGTCCTGAC CCTCGGCGAC TCGCACGCCG
  	GACACCTGCG
  1251	GGGGTTTCTG GATTATGTCG GCAGCCGGGA AGGGTGGAAA
  	GCCAAAATCC
  1301	TGTCCCTCGA TTCGGAGTGT TTGGTTTGGG TAGATGAGAA
  	GCTGGCAGAC
  1351	AACCCGTTAT GTCGAAAATA CCGGGATGAA GTTGAAAAAG
  	CCGAAGCCGT
  1401	TTTCATTGCC CAATTCTATG ATTTGAGGAT GGGCGGCCAG
  	CCCGTGCCGA
  1451	GATTTGAAGC GCAATCCTTC CTAATACCCG GGTTCCCAGC
  	CCGATTCAGG
  1501	GAAACCGTCA AAAGGATAGC CGCCGTCAAA CCCGTCTATG
  	TTTTTGCAAA
  1551	CAACACATCA ATCAGCCGTT CGCCCCTGAG GGAGGAAAAA
  	TTGAAAAGAT
  1601	TTGCCGCAAA CCAATATCTC CGCCCCATTC AGGCTATGGG
  	CGACATCGGC
  1651	AAGAGCAATC AGGCGGTCTT TGATTTGATT AAAGATATTC
  	CCAATGTGCA
  1701	TTGGGTGGAC GCACAAAAAT ACCTGCCCAA AAACACGGTC
  	GAAATATACG
  1751	GCCGCTATCT TTACGGCGAC CAAGACCACC TGACCTATTT
  	CGGTTCTTAT
  1801	TATATGGGGC GGGAATTTCA CAAACACGAA CGCCTGCTTA
  	AATCTTCTCG
  1851	CGACGGCGCA TTGCAGTAG

• This encodes a protein having amino acid sequence <SEQ ID 832>:

• 1	MQAVRYRPEI DGLRAVAVLS VMIFHLNNRW LPGGFLGVDI
  	FFVISGFLIT
  51	GIILSEIQNG SFSFRDFYTR RIKRIYPAFI AAVSLASVIA
  	SQIFLYEDFN
  101	QMRKTVELSA VFLSNIYLGF QQGYFDLSAD ENPVLHIWSL
  	AVEEQYYLLY
  151	PLLLIFCCKK TKSLRVLRNI SIILFLILTA TSFLPSGFYT
  	DILNQPNTYY
  201	LSTLRFPELL AGSLLAVYGQ TQNGRRQTAN GKRQLLSSLC
  	FGALLACLFV
  251	IDKHNPFIPG MTLLLPCLLT ALLIRSMQYG TLPTRILSAS
  	PIVFVGKISY
  301	SLYLYHWIFI AFAHYITGDK QLGLPAVSAV AALTAGFSLL
  	SYYLIEQPLR
  351	KRKMTFKKAF FCLYLAPSLI LVGYNLYARG ILKQEHLRPL
  	PGAPLAAENH
  401	FPETVLTLGD SHAGHLRGFL DYVGSREGWK AKILSLDSEC
  	LVWVDEKLAD
  451	NPLCRKYRDE VEKAEAVFIA QFYDLRMGGQ PVPRFEAQSF
  	LIPGFPARFR
  501	ETVKRIAAVK PVYVFANNTS ISRSPLREEK LKRFAANQYL
  	RPIQAMGDIG
  551	KSNQAVFDLI KDIPNVHWVD AQKYLPKNTV EIYGRYLYGD
  	QDHLTYFGSY
  601	YMGREFHKHE RLLKSSRDGA LQ*

• ORF128a and ORF128-1 show 99.5% identity in 622 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF128 shows 93.4% identity over 244 aa overlap with a predicted ORF (ORF128ng) from N. gonorrhoeae:
• 
• The complete length ORF128ng nucleotide sequence <SEQ ID 833> is:

• 1	ATGCAAGCTG TCCGATACAG GCCTGAAATT GACGGATTGC
  	GGGCCGTCGC
  51	CGTGCTATCC GTCATTATTT TCCACCTGAA TAACCGCTGG
  	CTGCCCGGAG
  101	GATTCCTGGG GGTGGACATT TTCTTTGTCA TCTCGGGATT
  	CCTCATTACC
  151	AACATCATTC TTTCTGAAAT ACAGAACGGT TCTTTTTCTT
  	TCCGGGATTT
  201	TTATACCCGC AGGATTAAGC GGATTTATCC TGCTTTTATT
  	GCGGCCGTGT
  251	CCCTGGCTTC GGTGATTGCT TCTCAAATCT TCCTTTACGA
  	AGATTTCAAC
  301	CAAATGAGGA AAACCATAGA GCTTTCTACG GTTTTTTTGT
  	CCAATATTTA
  351	TTTGGGGTTC CGATTGGGGT ATTTCGATTT GAGTGCCGAC
  	GAGAACCCCG
  401	TACTGCATAT CTGGTCTTTG GCGGTAGAGG AACAGTATTA
  	CCTCCTGTAT
  451	CCTCTTTTGC TGATATTCTG TTACAAAAAA ACCAAATCAC
  	TACGGGTGCT
  501	GCGTAATATC AGCATCATCC TGTTTCTGAT TTTGACCGCA
  	TCATCGTTTT
  551	TGCCGGCCGG GTTTTATACC GACATCCTCA ACCAACCcaa
  	TACTTATTAC
  601	CTTTCGACAC TGAGGTTTCC CGAGCTGTTG GTGGGTTCGC
  	TGTTGGCGGT
  651	TTACGGGCAA ACGCAAAACG GCAGACGGCA AACAGAAAAT
  	GGAAAACGGC
  701	AGTTGCTTTC ATTACTCTGT TTCGGCGCat tgCTTGTCTG
  	CCTGTTCGTG
  751	ATCGACAAAC ACGATCCGTT TATCCCGGGA ATAACCCTGC
  	TCCTTCCCTG
  801	CCTGCTGACG GCGCTGCTTA TCCGGAGTAT GCAATACGGG
  	ACACTTCCGA
  851	CCCGCATCCT GTCGGCAAGC CCCATCGTAT TTGTCGGCAA
  	AATCTCTTAT
  901	TCCCTATACC TGTACCATTG GATTTTTATT GCCTTCGCCC
  	ATTACATTAC
  951	AGGCGACAAA CAGCTCGGAC TGCCTGCCGT ATCGGCGGTT
  	GCCGCGTTGA
  1001	CGGCCGGATT TTCCCTGTTG AGCTATTATT TGATTGAACA
  	GCCGCTTAGA
  1051	AAACGGAAGA TGACCTTCAA AAAGGCATTT TTCTGCCTTT
  	ATCTCGCCCC
  1101	GTCCCTGATG CTTGTCGGTT ACAACCTGTA TTCAAGAGGG
  	ATATTGAAAC
  1151	AGGAACACCT CCGCCCGCTG CCCGGCACGC CCGTTGCTGC
  	GGAAAATAAT
  1201	TTTCCGGAAA CCGTCTTGAC CCTCGGCGAC TCGCACGCCG
  	GACACCTGCG
  1251	GGGGTTTCTG GATTATGTCG GCGGCAGGGA AGGGTGGAAA
  	GCTAAAATCC
  1301	TGTCCCTCGA TTCGGAGTGT TTGGTTTGGG TGGATGAGAA
  	GCTGGCAGAC
  1351	AACCCGTTGT GCCGAAAATA CCGGGATGAA GTTGAAAAAG
  	CCGAAGCTGT
  1401	TTTCATTGCC CAATTCTATG ATTTGAGGAT GGGCGGCCAG
  	CCCGTGCCGA
  1451	GATTTGAAGC GCAATCCTTC CTGATACCCG GGTTCAAAGC
  	CCGATTCAGG
  1501	GAAACCGTCA AGAGGATAGC CGCCGTCAAA CCTGTATATG
  	TTTTTGCAAA
  1551	CAATACATCA ATCAGCCGTT CTCCCTTGAG GGAGGAAAAA
  	TTGAAAAGAT
  1601	TTGCTATAAA CCAATACCTC CGGCCTATTC GGGCTATGGG
  	CGACATCGGC
  1651	AAGAGCAATC AGGCGGTCTT TGATTTGGTT AAAGATATTC
  	CCAATGTGCA
  1701	TTGGGTGGAC GCACAAAAAT ACCTGCCCAA AAACACGGTC
  	GAAATACACG
  1751	GACGCTATCT TTACGGCGAC CAAGACCACC TGACCTATTT
  	CGGTTCTTAT
  1801	TATATGGGGC GGGAATTTCA CAAACACGAA CGCCTGCTCA
  	AGCATTCCCG
  1851	AGGCGGCGCA TTGCAGTAG

• This encodes a protein having amino acid sequence <SEQ ID 834>:

• 1	MQAVRYRPEI DGLRAVAVLS VIIFHLNNRW LPGGFLGVDI
  	FFVISGFLIT
  51	NIILSEIQNG SFSFRDFYTR RIKRIYPAFI AAVSLASVIA
  	SQIFLYEDFN
  101	QMRKTIELST VFLSNIYLGF RLGYFDLSAD ENPVLHIWSL
  	AVEEQYYLLY
  151	PLLLIFCYKK TKSLRVLRNI SIILFLILTA SSFLPAGFYT
  	DILNQPNTYY
  201	LSTLRFPELL VGSLLAVYGQ TQNGRRQTEN GKRQLLSLLC
  	FGALLVCLFV
  251	IDKHDPFIPG ITLLLPCLLT ALLIRSMQYG TLPTRILSAS
  	PIVFVGKISY
  301	SLYLYHWIFI AFAHYITGDK QLGLPAVSAV AALTAGFSLL
  	SYYLIEQPLR
  351	KRKMTFKKAF FCLYLAPSLM LVGYNLYSRG ILKQEHLRPL
  	PGTPVAAENN
  401	FPETVLTLGD SHAGHLRGFL DYVGGREGWK AKILSLDSEC
  	LVWVDEKLAD
  451	NPLCRKYRDE VEKAEAVFIA QFYDLRMGGQ PVPRFEAQSF
  	LIPGFKARFR
  501	ETVKRIAAVK PVYVFANNTS ISRSPLREEK LKRFAINQYL
  	RPIRAMGDIG
  551	KSNQAVFDLV KDIPNVHWVD AQKYLPKNTV EIHGRYLYGD
  	QDHLTYFGSY
  601	YMGREFHKHE RLLKHSRGGA LQ*

• ORF128ng and ORF128-1 show 95.7% identity in 622 aa overlap:
• 
• In addition, ORF218ng shows homology to a hypothetical H. influenzae protein:

• sp|P43993|Y392_HAEIN HYPOTHETICAL PROTEIN HI0392 >gi|1074385|pir||B64007
  hypothetical protein HI0392 - Haemophilus influenzae (strain Rd KW20)
  >gi|1573364 (U32723) H. influenzae predicted coding region HI0392
  [Haemophilus influenzae] Length = 245
  Score = 239 bits (604), Expect = 3e−62
  Identities = 124/225 (55%), Positives = 152/225 (67%), Gaps = 1/225 (0%)

  Query:	38	VDIFFVISGFLITNIILSEIQNGSFSFRDFYTRRIKRIYPXXXXXXXXXXXXXXXXFLYE	97
  		+DIFFVISGFLIT II++EIQ  SFS + FYTRRIKRIYP                F+Y
  Sbjct:	1	MDIFFVISGFLITGIIITEIQQNSFSLKQFYTRRIKRIYPAFITVMALVSFIASAIFIYN	60
  Query:	98	DFNQMRKTIELSTVFLSNIYLGFRLGYFDLSADENPVLHIWSLAVEEQXXXXXXXXXIFC	157
  		DFN++RKTIEL+  FLSN YLG   GYFDLSA+ENPVLHIWSLAVE Q         I
  Sbjct:	61	DFNKLRKTIELAIAFLSNFYLGLTQGYFDLSANENPVLHIWSLAVEGQYYLIFPLILILA	120
  Query:	158	YKKTKSLRVLRNISIILFLILTASSFLPAGFYTDILNQPNTYYLSTLRFPELLVGSLLAV	217
  		YKK + ++VL  I++ILF IL A+SF+ A FY ++L+QPN YYLS LRFPELLVGSLLA+
  Sbjct:	121	YKKFREVKVLFIITLILFFILLATSFVSANFYKEVLHQPNIYYLSNLRFPELLVGSLLAI	180
  Query:	218	YGQTQNGRRQTENGKRQLLSLLCFGALLVCLFVIDKHDPFIPGIT	262
  		Y    N + Q       +L++L    L  CLF+++ +  FIPGIT
  Sbjct:	181	YHNLSN-KVQLSKQVNNILAILSTLLLFSCLFLMNNNIAFIPGIT	224

• This analysis, including the identification of several putative transmembrane domains, suggests that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 99
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 835>:

• 1	..ATTATTTACG AATACCGCTG GATGTTTCTT TACGGCGCAC
  	TGACGACCTT
  51	GGGGCTGACG GTCGTGGCAA C.GCGGGCGG TTCGGTATTG
  	GGTCTGTTGT
  101	TGGCGTTGGC GCGCCTGATT CACTTGGAAA AAGCCGGTGC
  	GCCGATGCGC
  151	GTGCTGGCGT GGGCGTTGCG TAAAGTTTCG CTGCTGTATG
  	TTACGCTGTT
  201	CCGGGGTACG CCGCTGTTTG TGCAGATTGT GATTTGGGCG
  	TATGTGTGGT
  251	TTCCGTTTTT CGTC..

• This corresponds to the amino acid sequence <SEQ ID 836; ORF129>:

• 1	..IIYEYRWMFL YGALTTLGLT VVAXAGGSVL GLLLALARLI
  	HLEKAGAPMR
  51	VLAWALRKVS LLYVTLFRGT PLFVQIVIWA YVWFPFFV..

• Further work revealed the complete nucleotide sequence <SEQ ID 837>:

• 1	ATGGATTTTC GTTTTGACAT TATTTACGAA TACCGCTGGA
  	TGTTTCTTTA
  51	CGGCGCACTG ACGACCTTGG GGCTGACGGT CGTGGCAACG
  	GCGGGCGGTT
  101	CGGTATTGGG TCTGTTGTTG GCGTTGGCGC GCCTGATTCA
  	CTTGGAAAAA
  151	GCCGGTGCGC CGATGCGCGT GCTGGCGTGG GCGTTGCGTA
  	AAGTTTCGCT
  201	GCTGTATGTT ACGCTGTTCC GGGGTACGCC GCTGTTTGTG
  	CAGATTGTGA
  251	TTTGGGCGTA TGTGTGGTTT CCGTTTTTCG TCCATCCTTC
  	AGACGGCATT
  301	TTGGTCAGCG GCGAGGCGGC AATCGCGCTG CGTCGCGGAT
  	ACGGGCCGCT
  351	GATTGCCGGT TCTTTGGCAC TGATCGCCAA CTCGGGGGCG
  	TATATCTGTG
  401	AGATTTTCCG CGCGGGCATC CAGTCTATAG ACAAAGGACA
  	GATGGAGGCG
  451	GCGCGTTCTT TGGGGCTGAC CTATCCGCAG GCGATGCGCT
  	ATGTGATTCT
  501	GCCGCAGGCA TTGCGCCGCA TGCTGCCGCC TTTGGCGAGC
  	GAGTTCATCA
  551	CGCTCTTGAA AGACAGCTCG CTGCTGTCGG TCATTGCTGT
  	GGCGGAGTTG
  601	GCGTATGTTC AGAATACGAT TACGGGCCGG TATTCGGTTT
  	ATGAAGAACC
  651	GCTTTACACC GTCGCCCTGA TTTATCTGTT GATGACGACT
  	TTCTTAGGCT
  701	GGATATTCCT GCGTTTGGAA AAACGTTACA ATCCGCAACA
  	CCGCTGA

• This corresponds to the amino acid sequence <SEQ ID 838; ORF129-1>:

• 1	MDFRFDIIYE YRWMFLYGAL TTLGLTVVAT AGGSVLGLLL
  	ALARLIHLEK
  51	AGAPMRVLAW ALRKVSLLYV TLFRGTPLFV QIVIWAYVWF
  	PFFV HPSDGI
  101	LVSGEAAIAL RRGYGPLIAG SLALIANSGA YICEIFRAGI
  	QSIDKGQMEA
  151	ARSLGLTYPQ AMRYVILPQA LRRMLPPLAS EFITLLKDSS
  	LLSVIAVAEL
  201	AYVQNTITGR YSVYEEPLYT VALIYLLMTT FLGWIFLRLE
  	KRYNPQHR*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF129 shows 98.9% identity over a 88aa overlap with an ORF (ORF129a) from strain A of N. meningitidis:
• 
• The complete length ORF129a nucleotide sequence <SEQ ID 839> is:

• 1	ATGGATTTTC GTTTTGACAT TATTTACGAA TACCGCTGGA
  	TGTTTCTTTA
  51	CGGCGCACTG ACGACCTTGG GGCTGACGGT CGTGGCGACG
  	GCGGGCGGTT
  101	CGGTATTGGG TCTGTTGTTG GCGTTGGCGC GCCTGATTCA
  	CTTGGAAAAA
  151	GCCGGTGCGC CGATGCGCGT GCTGGCGTGG GCGTTGCGTA
  	AGGTTTCGCT
  201	GCTGTATGTT ACGCTGTTCC GGGGTACGCC GCTGTTTGTG
  	CAGATTGTGA
  251	TTTGGGCGTA TGTGTGGTTT CCGTTTTTCG TCCATCCTTC
  	AGACGGCATT
  301	TTGGTTAGCG GCGAGGCGGC AATCGCGCTG CGTCGCGGAT
  	ACGGGCCGCT
  351	GATTGCCGGT TCTTTGGCAC TGATCGCCAA CTCGGGGGCG
  	TATATCTGTG
  401	AGATTTTCCG CGCGGGCATC CAGTCTATAG ACAAAGGACA
  	GATGGAGGCG
  451	GCGCGTTCTT TGGGGCTGAC CTATCCGCAG GCGATGCGCT
  	ATGTGATTCT
  501	GCCGCAGGCA TTGCGCCGTA TGCTGCCGCC TTTGGCGAGC
  	GAGTTCATCA
  551	CGCTCTTGAA AGACAGCTCG CTGCTGTCGG TCATTGCTGT
  	GGCGGAGTTG
  601	GCGTATGTTC AGAATACGAT TACGGGCCGG TATTCGGTTT
  	ATGAAGAACC
  651	GCTTTACACC GTCGCCCTGA TTTATCTGTT GATGACGACT
  	TTCTTAGGCT
  701	GGATATTCCT GCGTTTGGAA AAACGTTACA ATCCGCAACA
  	CCGCTGA

• This encodes a protein having amino acid sequence <SEQ ID 840>:

• 1	MDFRFDIIYE YRWMFLYGAL TTLGLTVVAT AGGSVLGLLL
  	ALARLIHLEK
  51	AGAPMRVLAW ALRKVSLLYV TLFRGTPLFV QIVIWAYVWF
  	PFFVHPSDGI
  101	LVSGEAAIAL RRGYGPLIAG SLALIANSGA YICEIFRAGI
  	QSIDKGQMEA
  151	ARSLGLTYPQ AMRYVILPQA LRRMLPPLAS EFITLLKDSS
  	LLSVIAVAEL
  201	AYVQNTITGR YSVYEEPLYT VALIYLLMTT FLGWIFLRLE
  	KRYNPQHR*

• ORF129a and ORF129-1 show 100.0% identity in 248 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF129 shows 98.9% identity over a 88 aa overlap with a predicted ORF (ORF129ng) from N. gonorrhoeae:
• 
• An ORF129ng nucleotide sequence <SEQ ID 841> was predicted to encode a protein having amino acid sequence <SEQ ID 842>:

• 1	MDFRFDIIYE YRWMFLYGAL TTLGLTVVAT AGGSVLGLLL
  	ALARLIHLEK
  51	AGAPMRVLAW ALRKVSLLYV TLFRGTPLFV QIVIWAYVWF
  	PFFVILHTAF
  101	LGNAMRQSRR VPDKGRWIAG SLELNCQPRG RKTRGEFPPG
  	ESNLGTEPRN
  151	PLSMGQRRFP GCENWYPPQN FIKK*

• Further work revealed the following gonococcal sequence <SEQ ID 843>:

• 1	ATGGATTTTc gtTTTGACAT TATTTAcgaA TACCGCTGGA
  	TGTTTCTTTA
  51	CGGCGCACTG Acgaccttgg ggctgacggt cgtggcgacg
  	gCGGGCGGTT
  101	CGGtattggG TCTGTTGTTG GCGTTGGCGC GCCTGATTCA
  	CTTGGAAAAA
  151	GCCGGTGCGC CGATGCGCGT GCTGGCGTGG GCGTTGCGTA
  	AGGTTTCGCT
  201	GCTGTACGTT ACCCTGTTCC GGGGTACGCC GCTGTTTGTG
  	CAGATTGTGA
  251	TTTGGGCGTA TGTGTGGTTT CCGTTTTTCG TCCATCCTTC
  	AGACGGCATT
  301	TTGGTCAGCG GCGAGGCGGC AATCGCGCTG CGTCGCGGAT
  	ACGGGCCGCT
  351	GATTGCCGGT TCTTTGGCAC TGATCGCCAA CTCGGGGGCG
  	TATATCTGTG
  401	AGATTTTCCG CGCGGGCATC CAGTCTATAG ACAAAGGACA
  	GATGGAGGCG
  451	GCGTGTTCTT TGGGACTGAC CTATCCGCAG GCGATGCGCT
  	ATGTGATTCT
  501	GCCGCAGGCA TTGCGCCGTA TGCTGCCGCC TTTGGCGAGC
  	GAGTTCATCA
  551	CGCTCTTGAA AGACAGCTCG CTGCTGTCGG TCATTGCTGT
  	GGCGGAGTTG
  601	GCGTATGTTC AGAATACGAT TACGGGCCGG TATTCGGTTT
  	ATGAAGAACC
  651	GCTTTACACC GCCGCCCTGA TTTATCTGTT GATGACGACT
  	TTCTTAGGCT
  701	GGATATTCCT GCGTTTGGAA AAACGTTACA ATCCGCAACA
  	CCGCTGA

• This corresponds to the amino acid sequence <SEQ ID 844; ORF129ng-1>:

• 1	MDFRFDIIYE YRWMFLYGAL TTLGLTVVAT AGGSVLGLLL
  	ALARLIHLEK
  51	AGAPMRVLAW ALRKVSLLYV TLFRGTPLFV QIVIWAYVWF
  	PFFVHPSDGI
  101	LVSGEAAIAL RRGYGPLIAG SLALIANSGA YICEIFRAGI
  	QSIDKGQMEA
  151	ARSLGLTYPQ AMRYVILPQA LRRMLPPLAS EFITLLKDSS
  	LLSVIAVAEL
  201	AYVQNTITGR YSVYEEPLYT VALIYLLMTT FLGWIFLRLE
  	KRYNPQHR*

• ORF129ng-1 and ORF129-1 show 99.2% identity in 248 aa overlap:
• 
• In addition, ORF129ng-1 is homologous to an ABC transporter from A. fulgidus:

• 2650409(AE001090) glutamine ABC transporter, permease protein (glnP)
  [Archaeoglobus fulgidus] Length = 224
  Score = 132 bits (329), Expect = 2e−30
  Identities = 86/178 (48%), Positives = 103/178 (57%), Gaps = 18/178 (10%)

  Query:	65	VSLLYVTLFRGTPLFVQIVIWAYVWFPFFVHPSDGILVSGEAAIALRRGYGPLIAGSLAL	124
  		+S  YV + RGTPL VQI+I       +F  P+ GI +  E A            G +AL
  Sbjct:	58	ISTAYVEVIRGTPLLVQILI------VYFGLPAIGINLQPEPA------------GIIAL	99
  Query:	125	IANSGAYICEIFRAGIQSIDKGQMEAACSLGLTYPQAMRYVILPQALRRMLPPLASEFIT	184
  		SGAYI EI RAGI+SI  GQMEAA SLG+TY QAMRYVI PQA R +LP L +EFI
  Sbjct:	100	SICSGAYIAEIVRAGIESIPIGQMEAARSLGMTYLQAMRYVIFPQAFRNILPALGNEFIA	159
  Query:	185	LLKDSSLLSVIAVAELAYVQNTITGRYSVYEEPLYTAALIYLLMTTFLGWIFLRLEKR	242
  		LLKDSSLLSVI++ EL  V   I         P    AL YL+MT  L  +    +K+
  Sbjct:	160	LLKDSSLLSVISIVELTRVGRQIVNTTFNAWTPFLGVALFYLMMTIPLSRLVAYSQKK	217

• This analysis, including the identification of transmembrane domains in the two proteins, suggests that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 100
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 845>:

• 1	..CTGAAAGAAT GCCGTCTGAA AGACCCTGTT TTTATTCCAA
  	ATATCGTTTA
  51	TAAGAACATC GCCATTACTT TCCTGCTCTT GCACGCCGCC
  	GCCGAACTTT
  101	GGCTGCCCGC GCAAACCGCC GGTTTTACCG CGCTCGCCGT
  	CGGCTTCATC
  151	CTGCTCGCCA AGCTGCGTGA gCTTCACCAT CACGAACTCT
  	TACGTAAACA
  201	cTACGTCCGC ACTTATTACy TGCTCCAACT CTTTGCCGCC
  	GCAGgcTAgT
  251	TTGTGGACAG GCGCGGCGwA ATTACAAAAC CTGCCCGCyT
  	CCGCGCCCCT
  301	GCACCTGATT ACCCTCGGCG GCATGATGGG CGGCGTGATG
  	ATGGTGTGGc
  351	TGACCGCCGG ACTGTGGCAC AGCGGCTTTA CCAAACTCGA
  	CTACCCCAAA
  401	CTCTGCCGCA TTGCCGTCCC CATCCTTTTC GCCGCCGCCG
  	TCTCGCGCGC
  451	TTTCTTGrTG AACGTGAACC CGrTATTTTT CATTACCGTT
  	CCTGCGATTC
  501	TGACCGCCGC CGTATTCGTA CTGTATCTTT TCrCGTTTAT
  	ACCGATATTT
  551	CGGGCGAATG CGTTTACAGA CGATCCGGAr TAr

• This corresponds to the amino acid sequence <SEQ ID 846; ORF130>:

• 1	..LKECRLKDPV FIPNIVYKNI AITFLLLHAA AELWLPAQTA
  	GFTALAVGFI
  51	LLAKLRELHH HELLRKHYVR TYYLLQLFAA AGSLWTGAAX
  	LQNLPASAPL
  101	HLITLGGMMG GVMMVWLTAG LWHSGFTKLD YPKLCRIAVP
  	ILFAAAVSRA
  151	FLXNVNPXFF ITVPAILTAA VFVLYLFXFI PIFRANAFTD
  	DPE*

• Further work revealed the complete nucleotide sequence <SEQ ID 847>:

• 1	ATGCGGCCGT TTTTCGTCGG CGCGGCGGTG CTTGCCATAC
  	TCGGTGCGCT
  51	GGTGTTTTTC ATCAACCCCG GTGCCATCGT CCTGCACCGC
  	CAAATTTTCT
  101	TGGAACTTAT GCTGCCGGCG GCATACGGCG GTTTTTTGAC
  	TGCGGCTTTG
  151	TTGGACTGGA CGGGTTTTTC GGGTAACCTG AAACCTGTCG
  	CGACTTTGAT
  201	GGCGGCATTA TTGCTCGCCG CATCCGCTAT ACTGCCCTTT
  	TCGCCGCAAA
  251	CTGCCTCGTT TTTCGTCGCC GCCTATTGGC TGGTGTTGCT
  	GCTGTTCTGC
  301	GCCCGGCTGA TTTGGCTAGA CCGAAACACC GACAACTTCG
  	CCCTGCTAAT
  351	GTTACTTGCC GCGTTCACTG TTTTTCAGAC GGCATATGCC
  	GTCAGCGGCG
  401	ATTTGAACCT GTTGCGCGCG CAAGTGCATC TAAATATGGC
  	GGCGGTGATG
  451	TTCGTATCCG TGCGCGTCAG TATTCTTTTG GGCGCGGAAG
  	CCCTGAAAGA
  501	ATGCCGTCTG AAAGACCCTG TTTTTATTCC AAATATCGTT
  	TATAAAAACA
  551	TCGCCATTAC TTTCCTGCTC TTGCACGCCG CCGCCGAACT
  	TTGGCTGCCC
  601	GCGCAAACCG CCGGTTTTAC CGCGCTCGCC GTCGGCTTCA
  	TCCTGCTCGC
  651	CAAGCTGCGT GAGCTTCACC ATCACGAACT CTTACGTAAA
  	CACTACGTCC
  701	GCACTTATTA CCTGCTCCAA CTCTTTGCCG CCGCAGGCTA
  	TTTGTGGACA
  751	GGCGCGGCGA AATTACAAAA CCTGCCCGCC TCCGCGCCCC
  	TGCACCTGAT
  801	TACCCTCGGC GGCATGATGG GCGGCGTGAT GATGGTGTGG
  	CTGACCGCCG
  851	GACTGTGGCA CAGCGGCTTT ACCAAACTCG ACTACCCCAA
  	ACTCTGCCGC
  901	ATTGCCGTCC CCATCCTTTT CGCCGCCGCC GTCTCGCGCG
  	CTTTCTTGAT
  951	GAACGTGAAC CCGATATTTT TCATTACCGT TCCTGCGATT
  	CTGACCGCCG
  1001	CCGTATTCGT ACTGTATCTT TTCACGTTTA TACCGATATT
  	TCGGGCGAAT
  1051	GCGTTTACAG ACGATCCGGA ATAA

• This corresponds to the amino acid sequence <SEQ ID 848; ORF130-1>:

• 1	MRPFFVGAAV LAILGALVFF INPGAIVLHR QIFLELMLPA
  	AYGGFLTAAL
  51	LDWTGFSGNL KPVATLMAAL LLAASAILP F SPQTASFFVA
  	AYWLVLLLFC
  101	ARLIWLDRNT DNFALLMLLA AFTVFQTAYA VSGDLNLLRA
  	QVHLNMAAVM
  151	FVSVRVSILL GAEALKECRL KDPVFIPNIV YKNIAITFLL
  	LHAAAELWLP
  201	AQTAGFTALA VGFILLAKLR ELHHHELLRK HYVRTYYLLQ
  	LFAAAGYLWT
  251	GAAKLQNLPA SAPLHLITLG GMMGGVMMVW LTAGLWHSGF
  	TKLDYPKLCR
  301	IAVPILFAAA VSRAFLMNVN PIFFITVPAI LTAAVFVLYL
  	FTFIPIFRAN
  351	AFTDDPE*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF130 shows 94.3% identity over a 193aa overlap with an ORF (ORF130a) from strain A of N. meningitidis:
• 
• The complete length ORF130a nucleotide sequence <SEQ ID 849> is:

• 1	ATGCGGCCGT TTTTCGTCGG CGCGGCGGTG CTTGCCATAC
  	TCGGTGCGCT
  51	GGTGTTTTTC ATCAACCCCG GTGCCATCGT CCTGCACCGC
  	CAAATTTTCT
  101	TGGAACTTAT GCTGCCGGCG GCATACGGCG GTTTTTTGAC
  	TGCGGCTTTG
  151	TTGGACTGGA CGGGTTTTTC GGGTAACCTG AAACCTGTCG
  	CGACTTTGAT
  201	GGCGGCATTA TTGCTCGCCG CATCCGCTAT ACTGCCCTTT
  	TCGCCGCAAA
  251	CTGCCTCGTT TTTCGTCGCC GCCTATTGGC TGGTGTTGCT
  	GCTGTTCTGC
  301	GCCCGGCTGA TTTGGCTAGA CCGAAACACC GACAACTTCG
  	CCCTGCTAAT
  351	GTTACTTGCC GCGTTCACTG TTTTTCAGAC GGCATATGCC
  	GTCAGCGGCG
  401	ATTTGAACCT GTTGCGCGCG CAAGTGCATC TAAATATGGC
  	GGCGGTGATG
  451	TTCGTATCCG TGCGCGTCAG TATTCTTTTG GGCGCGGAAG
  	CCCTGAAAGA
  501	ATGCCGTCTG AAAGACCCAG TATTCATCCC CAATGTCGTC
  	TATAAAAACA
  551	TCGCCATTAC CTTCCTGCTC CTGCACGCCG CCGCCGAACT
  	TTGGCTGCCT
  601	GCGCAAACCG CCGGTTTTAC CTCGCTCGCC GTCGGCTTTA
  	TCCTGCTTGC
  651	CAAGCTGCGT GAGCTTCACC ATCACGAACT CCTGCGCAAA
  	CACTACGTCC
  701	GCACTTATTA CCTGCTCCAA CTCTTTGCCG CCGCAGGCTA
  	TTTGTGGACA
  751	GGCGCGGCGA AATTACAAAA CCTGCCCGCC TCCGCGCCCC
  	TGCACCTGAT
  801	TACCCTCGGT GGCATGATGG GCAGCGTGAT GATGGTGTGG
  	CTGACTGCCG
  851	GACTGTGGCA CAGCGGCTTT ACCAAGCTCG ACTACCCGAA
  	ACTCTGCCGC
  901	ATCGCCGTCC CCATCCTNTT CGCCGCCGCC GTTTCGCGCG
  	CTGTTTTAAT
  951	GAACGTAAAC CCGATATTCT TCATCACCGT CCCCGCAATT
  	CTGACCGCCG
  1001	CCGTGTTCGT GCTTTACCTG CTGACATTCG TACCGATCTT
  	TCGGGCGAAC
  1051	GCGTTTACAG ACGATCCGGA ATAA

• This encodes a protein having amino acid sequence <SEQ ID 850>:

• 1	MRPFFVGAAV LAILGALVFF INPGAIVLHR QIFLELMLPA
  	AYGGFLTAAL
  51	LDWTGFSGNL KPVATLMAAL LLAASAILP F SPQTASFFVA
  	AYWLVLLLFC
  101	ARLIWLDRNT DNFALLMLLA AFTVFQTAYA VSGDLNLLRA
  	QVHLNMAAVM
  151	FVSVRVSILL GAEALKECRL KDPVFIPNVV YKNIAITFLL
  	LHAAAELWLP
  201	AQTAGFTSLA VGFILLAKLR ELHHHELLRK HYVRTYYLLQ
  	LFAAAGYLWT
  251	GAAKLQNLPA SAPLHLITLG GMMGSVMMVW LTAGLWHSGF
  	TKLDYPKLCR
  301	IAVPILFAAA VSRAVLMNVN PIFFITVPAI LTAAVFVLYL
  	LTFVPIFRAN
  351	AFTDDPE*

• ORF130a and ORF130-1 show 98.3% identity in 357 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF130 shows 91.7% identity over a 193 aa overlap with a predicted ORF (ORF130ng) from N. gonorrhoeae:
• 
• An ORF130ng nucleotide sequence <SEQ ID 851> was predicted to encode a protein having amino acid sequence <SEQ ID 852>:

• 1	MNKFFTHPMR PFFVGAAVLA ILGALVFFHQ PRRYHPAPPN
  	FLGTYAAGCI
  51	RRFFDYRFVG PDGFFRQPET CRYFDGGVVA CCGCFIAVFT
  	ATCRIFRRRL
  101	LAGVAAVLRL ADLARRQHRT LRSVDVTAAF TVFQTAYAVS
  	GDLNLLRAQV
  151	HLNMAAVMFV SVRVSVLLGT ETLKECRLKD PVFIPNVIYK
  	NIAITLLLHA
  201	AAELWLPAQT AGFTALAVGF ILLAKL RELH HHELLRKHYV
  	RTYYLLQLFA
  251	AAGYLWTGAA KLQNLPASAP LHLITLGGMT GGVMMVWLTA
  	GLWHSGFTKL
  301	DYPKLCRIAV SILFASAVSR AVLMNVNPIF FITVPEILTA
  	AVFMLYLLTF
  351	VPIFRANAFT DDPE*

• Further work revealed the following gonococcal DNA sequence <SEQ ID 853>:

• 1	ATGCGCCCGT TTTTCGTCGG TGCGGCAGTA CTTGCCATAC
  	TCGGTGCGTT
  51	GGTGTTTTTT ATCAACCCCG GCGCTATCAT CCTGCACCGC
  	CAAATTTTCT
  101	TGGAACTTAT GCTGCCGGCT GCATACGGCG GTTTTTTGAC
  	TACCGCTTTG
  151	TTGGACCGGA CGGGTTTTTC AGGCAACCTG AAACCTGCCG
  	CTACTTTGAT
  201	GGCGGTGTTG TTGCTTGTTG CGGCTGTTTT ATTGCCGTTT
  	TTACCGCAAC
  251	TTGCCGCATT TTTCGTCGCC GCCTATTGGC TGGTGTTGCT
  	GCTGTTCTGC
  301	GCCTGGCTGA TTTGGCTCGA CCGCAACACC GACAACTTCG
  	CTCTGTTGAT
  351	GTTACTTGCC GCATTTACCG TTTTTCAGAC GGCCTATGCC
  	GTCAGCGGCG
  401	ATTTGAACTT ACTGCGCGCG CAAGTGCATT TGAATATGGC
  	GGCGGTCATG
  451	TTCGTATCCG TCCGCGTCAG CGTCCTTTTG GGCACGGAAA
  	CCCTGAAAGA
  501	ATGCCGTCTG AAAGACCCCG TATTCATCCC CAACGTTATC
  	TATAAAAACA
  551	TCGCCATCAC CCTGCTGCTG CACGCCGCCG CCGAACTTTG
  	GCTGCCCGCG
  601	CAAACCGCCG GTTTTACTGC GCTTGCCGTC GGCTTCATCC
  	TGCTCGCCAA
  651	GCTGCGCGAA CTGCACCATC ACGAACTCTT ACGCAAACAC
  	TACGTCCGCA
  701	CTTATTACCT GCTCCAGCTC TTTGCCGCCG CAGGTTATCT
  	GTGGACAGGC
  751	GCGGCGAAAC TGCAAAACCT GCCCGCCTCC GCGCCCCTGC
  	ACCTGATTAC
  801	CCTCGGCGGC ATGACGGGTG GCGTGATGAT GGTGTGGCTG
  	ACTGCCGGAC
  851	TGTGGCACAG CGGCTTTACC AAACTCGACT ACCCGAAACT
  	CTGCCGCATC
  901	GCCGTCTCCA TCCTTTTCGC CTCCGCCGTT TCGCGCGCTG
  	TTTTAATGAA
  951	CGTGAATCCG ATATTCTTCA TCACCGTTCC CGAGATTCTG
  	ACCGCCGCCG
  1001	TGTTCATGCT TTACCTGCTG ACGTTCGTAC CGATTTTTCG
  	AGCGAACGCG
  1051	TTTACAGACG ATCCGGAATA A

• This corresponds to the amino acid sequence <SEQ ID 854; ORF130ng-1>:

• 1	MRPFFVGAAV LAILGALVFF INPGAIILHR QIFLELMLPA
  	AYGGFLTTAL
  51	LDRTGFSGNL KPAATLMAVL LLVAAVLLPF L PQLAAFFVA
  	AYWLVLLLFC
  101	AWLIWLDRNT DNFALLMLLA AFTVFQTAYA VSGDLNLLRA
  	QVHLNMAAVM
  151	FVSVRVSVLL GTETLKECRL KDPVFIPNVI YKNIAITLLL
  	HAAAELWLPA
  201	QTAGFTALAV GFILLAKLRE LHHHELLRKH YVRTYYLLQL
  	FAAAGYLWTG
  251	AAKLQNLPAS APLHLITLGG MTGGVMMVWL TAGLWHSGFT
  	KLDYPKLCRI
  301	AVSILFASAV SRAVLMNVNP IFFITVPEIL TAAVFMLYLL
  	TFVPIFRANA
  351	FTDDPE*

• ORF130ng-1 and ORF130-1 show 92.4% identity in 357 aa overlap:
• 
• Based on this analysis, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 101
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 855>:

• 1	ATGGAAATTC GGGCAATAAA ATATACGGCA ATGGCTGCGT
  	TGCTTGCATT
  51	TACGGTTGCA GGCTGCCGGC TGGCGGGGTG GTATGAGTGT
  	TCGTCCCTCA
  101	CCGGCTGGTG TAAGCCGAGA AAACCGGCTG CCATCGATTT
  	TTGGGATATT
  151	GGCGGCGAGA GTCCGCCGTC TTTAGGGGAC TACGAGATAC
  	CGCTTTCAGA
  201	CGGCAATAGT TCCGTCAGGG CAAACGAATA TGAATCCGCA
  	CAACAATCTT
  251	ACTTTTACAG GAAAATAGGG AAGTTTGAAG C.TGCGGGCT
  	GGATTGGCGT
  301	ACGCGTGACG GCAAACCTTT GATTGAGACG TTCAAACAGG
  	GAGGATTTGA
  351	CTGCTTGGAA AAG..

• This corresponds to the amino acid sequence <SEQ ID 856; ORF131>:

• 1	MEIRAIKYTA MAALLAFTVA GCRLAGWYEC SSLTGWCKPR
  	KPAAIDFWDI
  51	GGESPPSLGD YEIPLSDGNS SVRANEYESA QQSYFYRKIG
  	KFEXCGLDWR
  101	TRDGKPLIET FKQGGFDCLE K..

• Further work revealed the complete nucleotide sequence <SEQ ID 857>:

• 1	ATGGAAATTC GGGCAATAAA ATATACGGCA ATGGCTGCGT
  	TGCTTGCATT
  51	TACGGTTGCA GGCTGCCGGC TGGCGGGGTG GTATGAGTGT
  	TCGTCCCTCA
  101	CCGGCTGGTG TAAGCCGAGA AAACCGGCTG CCATCGATTT
  	TTGGGATATT
  151	GGCGGCGAGA GTCCGCCGTC TTTAGGGGAC TACGAGATAC
  	CGCTTTCAGA
  201	CGGCAATCGT TCCGTCAGGG CAAACGAATA TGAATCCGCA
  	CAACAATCTT
  251	ACTTTTACAG GAAAATAGGG AAGTTTGAAG CCTGCGGGCT
  	GGATTGGCGT
  301	ACGCGTGACG GCAAACCTTT GATTGAGACG TTCAAACAGG
  	GAGGATTTGA
  351	CTGCTTGGAA AAGCAGGGGT TGCGGCGCAA CGGTCTGTCC
  	GAGCGCGTCC
  401	GATGGTAA

• This corresponds to the amino acid sequence <SEQ ID 858; ORF131-1>:

• 1	MEIRAIKYTA MAALLAFTVA GCRLAGWYEC SSLTGWCKPR
  	KPAAIDFWDI
  51	GGESPPSLGD YEIPLSDGNR SVRANEYESA QQSYFYRKIG
  	KFEACGLDWR
  101	TRDGKPLIET FKQGGFDCLE KQGLRRNGLS ERVRW*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF131 shows 95.0% identity over a 121 aa overlap with an ORF (ORF131a) from strain A of N. meningitidis:
• 
• The complete length ORF131a nucleotide sequence <SEQ ID 859> is:

• 1	ATGGAAATTC GGGCAATAAA ATATACGGCA ATGGCTGCGT
  	TGCTTGCATT
  51	TACGGTTGCA GGCTGCCGGT TGGCAGGTTG GTATGAGTGT
  	TCGTCCCTGT
  101	CCGGCTGGTG TAAGCCGAGA AAACCTGCCG CCATCGATTT
  	TTGGGATATT
  151	GGCGGCGAGA GTCCTCCGTC TTTAGAGGAC TACGAGATAC
  	CGCTTTCAGA
  201	CGGCAATCGT TCCGTCAGGG CAAACGAATA TGAATCCGCA
  	CAACAATCTT
  251	ACTTTTACAG GAAAATAGGG AAGTTTGAAG CCTGCGGGTT
  	GGATTGGCGT
  301	ACGCGTGACG GCAAACCTTT GATTGAGACG TTCAAACAGG
  	AAGGTTTTGA
  351	TTGTTTGAAA AAGCAGGGGT TGCGGCGCAA CGGTCTGTCC
  	GAGCGCGTCC
  401	GATGGTAA

• This encodes a protein having amino acid sequence <SEQ ID 860>:

• 1	MEIRAIKYTA MAALLAFTVA GCRLAGWYEC SSLSGWCKPR
  	KPAAIDFWDI
  51	GGESPPSLED YEIPLSDGNR SVRANEYESA QQSYFYRKIG
  	KFEACGLDWR
  101	TRDGKPLIET FKQEGFDCLK KQGLRRNGLS ERVRW*

• ORF131a and ORF131-1 show 97.0% identity in 135 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF131 shows 89.3% identity over 121 aa overlap with a predicted ORF (ORF131ng) from N. gonorrhoeae:
• 
• A complete length ORF131ng nucleotide sequence <SEQ ID 861> was predicted to encode a protein having amino acid sequence <SEQ ID 862>:

• 1	MEIRVIKYTA TAALFAFTVA GCRLAGWYEC LSLSGWCKPR
  	KPAAIDFWDI
  51	GGESPLSLED YEIPLSDGNR SVRANEYESA QKSYFYRKIG
  	KFEACGLDWR
  101	TRDGKPLVER FKQEGFDCLE KQGLRRNGLS ERVRW*

• Further work revealed the following gonococcal DNA sequence <SEQ ID 863>:

• 1	ATGGAAATTC GGGTAATAAA ATATACGGCA ACGGCTGCGT
  	TGTTTGCATT
  51	TACGGTTGCA GGCTGCCGGC TGGCGGGGTG GTATGAGTGT
  	TCGTCCTTGT
  101	CCGGCTGGTG TAAGCCGAGA AAACCTGCCG CCATCGATTT
  	TTGGGATATT
  151	GGCGGCGAGA GtccgctGTC TTTAGAGGAC TACGAGATAC
  	CGCTTTCAGA
  201	CGGCAATCGT TCCGTCAGGG CAAACGAATA TGAATCCGCG
  	CAAAAATCTT
  251	ACTTTTATAG GAAAATAGGG AAGTTTGAAG CCTGCGGGTT
  	GGATTGGCGT
  301	ACGCGTGACG GCAAACCTTT GGTTGAGAGG TTCAAACAGG
  	AAGGTTTCGA
  351	CTGTTTGGAA AAGCAGGGGT TGCGGCGCAA CGGCCTGTCC
  	GAGCGCGTCC
  401	GATGGTAA

• This corresponds to the amino acid sequence <SEQ ID 864; ORF131ng-1>:

• 1	MEIRVIKYTA TAALFAFTVA GCRLAGWYEC SSLSGWCKPR
  	KPAAIDFWDI
  51	GGESPLSLED YEIPLSDGNR SVRANEYESA QKSYFYRKIG
  	KFEACGLDWR
  101	TRDGKPLVER FKQEGFDCLE KQGLRRNGLS ERVRW*

• ORF131ng-1 and ORF131-1 show 92.6% identity in 135 aa overlap:
• 
• Based on the presence of a predicted prokaryotic membrane lipoprotein lipid attachment site, it is predicted that the proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 102
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 865>

• 1	ATGAAACACA TCCATATTAT CGGTATCGGC GGCACGTTTA
  	TGGGCGGGCT
  51	TGCCGCCATT GCCAAAGAAG CGGGGTTTGA AGTCAGCGGT
  	TGCGACGCGA
  101	AGATGTATCC GCCGATGAGC ACCCAGCTCG AAGCCTTGGG
  	TATAGACGTG
  151	TATGAAGGCT TCGATGCCGC TCAGTTGGAC GAATTTAAAG
  	CCGACGTTTA
  201	CGTTATCGGC AATGTCGCCA AGCGCGGGAT GGATGTGGTT
  	GAAGCGATTT
  251	TGAACCTCGG CCTGCCtTAT ATtTcCGGCC CGCAATGGCT
  	GTCGGAAAAC
  301	GTGCTGCACC ATCATTGGGT ACTCGGTGTG GCGGGGACgC
  	ACGGCAAAAC
  351	GACCACCGCC TCCATGCTCG CATGGGTCTT GGAATATgCC
  	GGCCTCGCGC
  401	CGGGCTTCCT TATtGGCGGC GTACC.GGAA AATttCGGCG
  	TTTCCGCCCG
  451	CCTGCCGCAA ACGCCGCGCC AAGACCCGAA CAGCCAATCG
  	CCGTTTTTcG
  501	TCATCGAAGC CGACGAATAC GACACCGCCT TTtTCGACAA
  	ACGTTCTAAA
  551	TtCGTGCATT ACCGTCCGCG TACCGCCGTG TTGAACAATC
  	TGGAATTCGA
  601	CCACGCCGAC ATCTTTGCCG ACTTGGGCGC GATACAGACc
  	CAGTTCCACT
  651	ACCTCGTGCG TACCGTGCCG TCTGAAGGCT TAATCGTCTG
  	CAACGGACGG
  701	CAGCAAAGCC TGCAAGATAC TTTGGACAAA GGCTGCTGGA
  	CGCCGGTGGA
  751	AAAATTCGGC ACGGAACACG GCTGGCA..

• This corresponds to the amino acid sequence <SEQ ID 866; ORF132>:

• 1	MKHIHIIGIG GTFMGGLAAI AKEAGFEVSG CDAKMYPPMS
  	TQLEALGIDV
  51	YEGFDAAQLD EFKADVYVIG NVAKRGMDVV EAILNLGLPY
  	ISGPQWLSEN
  101	VLHHHWVLGV AGTHGKTTTA SMLAWVLEYA GLAPGFLIGG
  	VXGKFRRFRP
  151	PAANAAPRPE QPIAVFRHRS RRIRHRLFRQ TFXIRALPSA
  	YRRVEQSGIR
  201	PRRHLCRLGR DTDPVPLPRA YRAVXRLNRL QRTAAKPARY
  	FGQRLLDAGG
  251	KIRHGTRLA..

• Further work revealed the complete nucleotide sequence <SEQ ID 867>:

• 1	ATGAAACACA TCCATATTAT CGGTATCGGC GGCACGTTTA
  	TGGGCGGGCT
  51	TGCCGCCATT GCCAAAGAAG CGGGGTTTGA AGTCAGCGGT
  	TGCGACGCGA
  101	AGATGTATCC GCCGATGAGC ACCCAGCTCG AAGCCTTGGG
  	TATAGACGTG
  151	TATGAAGGCT TCGATGCCGC TCAGTTGGAC GAATTTAAAG
  	CCGACGTTTA
  201	CGTTATCGGC AATGTCGCCA AGCGCGGGAT GGATGTGGTT
  	GAAGCGATTT
  251	TGAACCTCGG CCTGCCTTAT ATTTCCGGCC CGCAATGGCT
  	GTCGGAAAAC
  301	GTGCTGCACC ATCATTGGGT ACTCGGTGTG GCGGGGACGC
  	ACGGCAAAAC
  351	GACCACCGCC TCCATGCTCG CATGGGTCTT GGAATATGCC
  	GGCCTCGCGC
  401	CGGGCTTCCT TATTGGCGGC GTACCGGAAA ATTTCGGCGT
  	TTCCGCCCGC
  451	CTGCCGCAAA CGCCGCGCCA AGACCCGAAC AGCCAATCGC
  	CGTTTTTCGT
  501	CATCGAAGCC GACGAATACG ACACCGCCTT TTTCGACAAA
  	CGTTCTAAAT
  551	TCGTGCATTA CCGTCCGCGT ACCGCCGTGT TGAACAATCT
  	GGAATTCGAC
  601	CACGCCGACA TCTTTGCCGA CTTGGGCGCG ATACAGACCC
  	AGTTCCACTA
  651	CCTCGTGCGT ACCGTGCCGT CTGAAGGCTT AATCGTCTGC
  	AACGGACGGC
  701	AGCAAAGCCT GCAAGATACT TTGGACAAAG GCTGCTGGAC
  	GCCGGTGGAA
  751	AAATTCGGCA CGGAACACGG CTGGCAGGCC GGCGAAGCCA
  	ATGCCGACGG
  801	CTCGTTCGAC GTGTTGCTCG ACGGCAAAAC CGCCGGACGC
  	GTCAAATGGG
  851	ATTTGATGGG CAGGCACAAC CGCATGAACG CGCTCGCCGT
  	CATTGCCGCC
  901	GCGCGTCATG TCGGTGTCGA TATTCAGACC GCCTGCGAAG
  	CCTTGGGCGC
  951	GTTTAAAAAC GTCAAACGCC GGATGGAAAT CAAAGGCACG
  	GCAAACGGCA
  1001	TCACCGTTTA CGACGACTTC GCCCACCACC CGACCGCCAT
  	CGAAACCACG
  1051	ATTCAAGGTT TGCGCCAACG CGTCGGCGGC GCGCGCATCC
  	TCGCCGTCCT
  1101	CGAACCGCGT TCCAACACGA TGAAGCTGGG CACGATGAAG
  	TCCGCCCTGC
  1151	CTGTAAGCCT CAAAGAAGCC GACCAAGTGT TCTGCTACGC
  	CGGCGGCGTG
  1201	GACTGGGACG TCGCCGAAGC CCTCGCGCCT TTGGGCGGCA
  	GGCTGAACGT
  1251	CGGCAAAGAC TTCGATGCCT TCGTTGCCGA AATCGTGAAA
  	AACGCCGAAG
  1301	TAGGCGACCA TATTTTGGTG ATGAGCAACG GCGGTTTCGG
  	CGGAATACAC
  1351	GGAAAGCTGC TGGAAGCTTT GAGATAG

• This corresponds to the amino acid sequence <SEQ ID 868; ORF132-1>:

• 1	MKHIHIIGIG GTFMGGLAAI AKEAGFEVSG CDAKMYPPMS
  	TQLEALGIDV
  51	YEGFDAAQLD EFKADVYVIG NVAKRGMDVV EAILNLGLPY
  	ISGPQWLSEN
  101	VLHHHWVLGV AGTHGKTTTA SMLAWVLEYA GLAPGFLIGG
  	VPENFGVSAR
  151	LPQTPRQDPN SQSPFFVIEA DEYDTAFFDK RSKFVHYRPR
  	TAVLNNLEFD
  201	HADIFADLGA IQTQFHYLVR TVPSEGLIVC NGRQQSLQDT
  	LDKGCWTPVE
  251	KFGTEHGWQA GEANADGSFD VLLDGKTAGR VKWDLMGRHN
  	RMNALAVIAA
  301	ARHVGVDIQT ACEALGAFKN VKRRMEIKGT ANGITVYDDF
  	AHHPTAIETT
  351	IQGLRQRVGG ARILAVLEPR SNTMKLGTMK SALPVSLKEA
  	DQVFCYAGGV
  401	DWDVAEALAP LGGRLNVGKD FDAFVAEIVK NAEVGDHILV
  	MSNGGFGGIH
  451	GKLLEALR*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with the Hypothetical o457 Protein of E. coli (Accession Number U14003)
• ORF132 and o457 show 58% aa identity in 140 aa overlap:

• Orf132:	4	IHIIGIGGTFMGGLAAIAKEAGFEVSGCDAKMYPPMSTQLEALGIDVYEGFDAAQLDEFK	63
  		IHI+GI GTFMGGLA +A++ G EV+G DA +YPPMST LE  GI++ +G+DA+QL+  +
  o457:	3	IHILGICGTFMGGLAMLARQLGHEVTGSDANVYPPMSTLLEKQGIELIQGYDASQLEP-Q	61
  Orf132:	64	ADVYVIGNVAKRGMDVVEAILNLGLPYISGPQWLSENVLHHHWVLGVAGTHGKTTTASML	123
  		D+ +IGN   RG   VEA+L   +PY+SGPQWL + VL   WVL VAGTHGKTTTA M
  o457:	62	PDLVIIGNAMTRGNPCVEAVLEKNIPYMSGPQWLHDFVLRDRWVLAVAGTHGKTTTAGMA	121
  Orf132:	124	AWVLEYAGLAPGFLIGGVXG	143
  		W+LE  G  PGF+IGGV G
  o457:	122	TWILEQCGYKPGFVIGGVPG	141

  
  Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF132 shows 74.6% identity over a 189aa overlap with an ORF (ORF132a) from strain A of N. meningitidis:
• 
• The complete length ORF132a nucleotide sequence <SEQ ID 869> is:

• 1 ATGAAACACA TCCACATTAT CGGTATCGGC GGCACGTTTA TGGGTGGGAT
  51 TGCCGCCATT GCCAAAGAAG CAGGGTTTGA ANTCAGCGGT TGCGATGCGA
  101 AGATGTATCC GCCGATGAGC ACCCAGCTCG AAGCCTTGGG CATAGGCGTG
  151 TATGAAGGCT TCGACACCGC GCAGTTGGAC GAATTTAAAG CCGACGTTTA
  201 CGTTATCGGC AATGTCGCCA AGCGCGGGAT GGATGTGGTT GAAGCGATTT
  251 TGAACCGTGG GCTGCCTTAT ATTTCCGGCC CGCAATGGCT GGCTGAAAAC
  301 NTGCTGCACC ATCATTGGNN ACTCGGCGTG GCGGNGACGC ACGGCAAAAC
  351 GACCACCGCG TCTATGCTCG CGTGGGTTTT GGAATATGCC GGACTCGCAC
  401 CGGGCTTCNT TATCGGCGGC GTACCGGAAA ACTTCAGCGT TTCCGCCCGC
  451 CTGCCGCAAA CGCCGCGCCA AGACCCGAAC AGCCAATCGC CGTTTTTCGT
  501 CATTGAAGCC GACGAATACG ACACCGCGTT TTTCGACAAA CGCTCCAAAT
  551 TCGTGCATTA CCGTCCGCGT ACCGCCGTGT TGAACAATCT GGAATTCGAC
  601 CACGCCGACA TCTTCGCCGA TTTGGGCGCG ATACAGACCC AGTTCCACCA
  651 CCTCGTGCGT ACCGTGCCGT CTGAAGGCCT CATCGTCTGC AACGGACGGC
  701 AGCAAAGCCT GCAAGACACT TTGGACAAAG GCTGCTGGAC GCCGGTGGAA
  751 AAATTCGGCA CGGAACACGG CTGGCAGGCC GGCGAAGCCA ATGCCGATGG
  801 CTCGTTCGAC GTGTTGCTTG ACGGCAAAAA AGCCGGACAC GTCGCTTGGA
  851 GTTTGATGGG CGGACACAAC CGCATGAACG CGCTCGCNGT CATCGCCGCC
  901 GCGCGTCATG CCGGAGTNGA CATTCAGACG GCCTGCGAAG CCTTGAGCAC
  951 GTTTAAAAAC GTCAAACGCC GCATGGAAAT CAAAGGCACG GCAAACGGTA
  1001 TCACCGTTTA CGACGACTTC GCCCACCATC CGACCGCTAT CGAAACCACG
  1051 ATTCAAGGTT TGCGCCAGCG CGTCGGCGGC GCGCGCATCC TCGCCGTCCT
  1101 CGAACCGCGT TCCAATACGA TGAAGCTGGG TACGATGAAA GCCGCCCTGC
  1151 CCGCAAGCCT CAAAGAAGCC GACCAAGTGT TCTGNTACGC CGGCGGCGCG
  1201 GACTGGGACG TTGCCGAAGC CCTCGCGCCT TTGGGCGGCA GGCTGCACGT
  1251 CGGCAAAGAC TTCGATGCCT TCGTTGCCGA AATCGTGAAA AACGCCGAAG
  1301 CAGGCGACCA TATTTTGGTG ATGAGCAACG GCGGTTTCGG CGGAATACAC
  1351 ACCAAACTGC TGGACGCTTT GAGATAG

• This encodes a protein having-amino acid sequence <SEQ ID 870>:

• 1 MKHIHIIGIG GTFMGGIAAI AKEAGFEXSG CDAKMYPPMS TQLEALGIGV
  51 YEGFDTAQLD EFKADVYVIG NVAKRGMDVV EAILNRGLPY ISGPQWLAEN
  101 XLHHHWXLGV AXTHGKTTTA SMLAWVLEYA GLAPGFXIGG VPENFSVSAR
  151 LPQTPRQDPN SQSPFFVIEA DEYDTAFFDK RSKFVHYRPR TAVLNNLEFD
  201 HADIFADLGA IQTQFHHLVR TVPSEGLIVC NGRQQSLQDT LDKGCWTPVE
  251 KFGTEHGWQA GEANADGSFD VLLDGKKAGH VAWSLMGGHN RMNALAVIAA
  301 ARHAGVDIQT ACEALSTFKN VKRRMEIKGT ANGITVYDDF AHHPTAIETT
  351 IQGLRQRVGG ARILAVLEPR SNTMKLGTMK AALPASLKEA DQVFXYAGGA
  401 DWDVAEALAP LGGRLHVGKD FDAFVAEIVK NAEAGDHILV MSNGGFGGIH
  451 TKLLDALR*

• ORF132a and ORF132-1 show 93.9% identity in 458 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF132 shows 89.6% identity over 259 aa overlap with a predicted ORF (ORF132ng) from N. gonorrhoeae:
• 
• An ORF132ng nucleotide sequence <SEQ ID 871> was predicted to encode a protein having amino acid sequence <SEQ ID 872>:

• 1	MKHIHIIGIG GTFMGGIAAI AKEAGFKVSG CDAKMYPPMS
  	TQLEALGIGV
  51	HEGFDAAQLE EFQADIYVIG NVARRGMDVV EAILNRGLPY
  	ISGPQWLAEN
  101	VLHHHWVLGV AGTHGKTTTA SMLAWVLEYA GLAPGFLIGG
  	VPGKFRRFRP
  151	PTANAASRPE QQIAVFRHRS RRIRHRLFRQ TLQIRALSPA
  	YRRVEQSGIR
  201	PRRHLRRLGR DTDPVPPPRA HRTIRRPHRL QRTAAKPARY
  	FGQRLLDAGG
  251	KIRHRTRLAD W*

• Further work revealed the following gonococcal DNA sequence <SEQ ID 873>:

• 1	ATGAAACACA TCCACATTAT CGGTATCGGC GGCACGTTTA
  	TGGGCGGGAT
  51	TGCCGCCATT GCCAAAGAAG CCGGGTTCAA AGTCAGCGGT
  	TGCGACGCGA
  101	AGATGTATCC GCCGATGAGC ACCCAGCTCG AAGCCTTGGG
  	CATAGGCGTA
  151	CACGAAGGCT TCGATGCCGC GCAGTTGGAA GAATTTCAAG
  	CCGATATTTA
  201	CGTCATCGGC AATGTCGCCA GGCGCGGGAT GGATGTGGTC
  	GAGGCGATTT
  251	TGAACCGTGG GCTGCCTTAT ATTTCCGGCC CGCAATGGCT
  	GGCTGAAAac
  301	GTGCtgcacc atcaTTGGgt ACTCGGCGTG GcagggaCGC
  	ACGGcaaAac
  351	gaccaCcGcg tCCATGCTCG CCTGGGTCTT GGAATATGCC
  	GGACTCGCGC
  401	CGGGCTTCCT CATCGGCGGt gtaccggaAA ATTTCGGCGT
  	TTCCGCCCGC
  451	CTACCGCAAA CGCCGCGTCA AGACCCGAAC AGCAAATCGC
  	CGTTTTTCGT
  501	CATCGAAGCC GACGAATACG ACACCGCCTT TTTCGACAAA
  	CGCTCCAAAT
  551	TCGTGCATTA TCGCCCGCGT ACCGCCGTGT TGAACAATCT
  	GGAATTCGAC
  601	CACGCCGACA TCTTCGCCGA CTTGGGCGCG ATACAGACCC
  	AGTTCCACCA
  651	CCTCGTGCGC ACCGTACCAT CCGAAGGCCT CATCGTCTGC
  	AACGGACAGC
  701	AGCAAAGCCT GCAAGATACT TTGGACAAAG GCTGCTGGAC
  	GCCGGTGGAA
  751	AAATTCGGCA CCGGACACGG CTGGCAGATT GGTGAAGTCA
  	ATGCCGACGG
  801	CTCGTTCGAC GTATTGCTTG ACGGCAAAAA AGCCGGACAC
  	GTCGCATGGG
  851	ATTTGATGGG CGGACACAAC CGCATGAACG CGCTCGCCGT
  	CATCGCTGCC
  901	GCACGCCATG CCGGAGTCGA TGTTCAGACG GCCTGCGAAG
  	CCTTGGGTGC
  951	GTTTAAAAAC GTCAAACGCC GCATGGAAAT CAAAGGCACG
  	GCAAACGGCA
  1001	TCACCGTTTA CGACGATTTC GCCCACCACC CGACCGCCAT
  	CGAAACCACG
  1051	ATTCAAGGTT TGCGCCAACG TGTCGGCGGC GCGCGCATCC
  	TCGCCGTCCT
  1101	CGAGCCGCGT TCCAACACCA TGAAACTCGG CACGATGAAG
  	TCCGCCCTGC
  1151	CCGCAAGCCT CAAAGAAGCC GACCAAGTGT TCTGCTACGC
  	CGGCGGCGCG
  1201	GACTGGGACG TTGCCGAAGC CCTCGCGCCT TTGGGCTGCA
  	GGCTGCGCGT
  1251	CGGTAAAGAT TTCGATACCT TCGTTGCCGA AATTGTGAAA
  	AACGCCCGAA
  1301	CCGGCGACCA TATTTTGGTG ATGAGCAACG GCGGTTTCGG
  	CGGAATACAC
  1351	ACCAAACTGC TGGACGCTTT GAGATAG

• This corresponds to the amino acid sequence <SEQ ID 874; ORF132ng-1>:

• 1	MKHIHIIGIG GTFMGGIAAI AKEAGFKVSG CDAKMYPPMS
  	TQLEALGIGV
  51	HEGFDAAQLE EFQADIYVIG NVARRGMDVV EAILNRGLPY
  	ISGPQWLAEN
  101	VLHHHWVLGV AGTHGKTTTA SMLAWVLEYA GLAPGFLIGG
  	VPENFGVSAR
  151	LPQTPRQDPN SKSPFFVIEA DEYDTAFFDK RSKFVHYRPR
  	TAVLNNLEFD
  201	HADIFADLGA IQTQFHHLVR TVPSEGLIVC NGQQQSLQDT
  	LDKGCWTPVE
  251	KFGTGHGWQI GEVNADGSFD VLLDGKKAGH VAWDLMGGHN
  	RMNALAVIAA
  301	ARHAGVDVQT ACEALGAFKN VKRRMEIKGT ANGITVYDDF
  	AHHPTAIETT
  351	IQGLRQRVGG ARILAVLEPR SNTMKLGTMK SALPASLKEA
  	DQVFCYAGGA
  401	DWDVAEALAP LGCRLRVGKD FDTFVAEIVK NARTGDHILV
  	MSNGGFGGIH
  451	TKLLDALR*

• ORF132ng-1 and ORF132-1 show 93.2% identity in 458 aa overlap:
• 
• In addition, ORF132ng-1 is homologous to a hypothetical E. coli protein:

• pir||S556459 hypothetical protein o457 - Escherichia coli >gi|537075 (U14003)
  ORF_o457 [Escherichia coli] >gi|1790660 (AE000494). hypothetical 48.5 kD protein
  in fbp-pmba intergenic region [Escherichia coli] Length = 457
  Score = 474 bits (1207), Expect = e−133
  Identities = 249/439 (56%), Positives = 294/439 (66%), Gaps = 13/439 (2%)
  Query:  22 KEAGFKVSGCDAKMYPPMSTQLEALGIGVHEGFDAAQLEEFQADIYVIGNVARRGMDVVE  81
  ++ G +V+G DA +YPPMST LE  GI + +G+DA+QLE  Q D+ +IGN   RG    VE
  Sbjct:  21 RQLGHEVTGSDANVYPPMSTLLEKQGIELIQGYDASQLEP-OPDLVIIGNAMTRGNPCVE  79
  Query:  82 AILNRGLPYISGPQWLAENVLHHHWVLGVAGTHGKTTTASMLAWVLEYAGLAPGFLIGGV 141

  A+L  ++PY+SGPQWL  +VL   WVL VAGTHGKTTTA M  W+LE  G   PGF+IGGV

  Sbjct:  80 AVLEKNIPYMSGPQWLHDFVLADRWVLAVAGTHGKTTTAGMATWILEQCGYKPGFVIGGV 139
  Query: 142 PENFGVSARLPQTPRQDPNSKSPFFVIEADEYDTAFFDKRSKFVHYRPRTAVLNNLEFDH 201

  P NF VSA L          +S FFVIEADEYD AFFDKRSKFVHY PRT +LNNLEFDH

  Sbjct: 140 PGNFEVSAHL---------GESDFFVIEADEYDCAFFDKRSKFVHYCPRTLILNNLEFDH 190
  Query: 202 ADIFADLGAIQTQFHHLVRTVPSEGLIVCNGQQQSLQDTLDKGCWTPVEKFGTGHGWQIG 261
  ADIF DL AIQ QFHHLVR VP +G I+      +L+ T+  GCW+  E  G     WQ
  Sbjct: 191 ADIFDDLKAIQKQFHHLVRIVPGOGRIIWPENDINLKQTMAMGCWSEQELVGEQGHWQAK 250
  Query: 262 EVNADGS-FDVLLDGKKAGHVAWDLMGGHNRMNALAVIAAARHAGVDVQTACEALGAFKN 320
  ++  D S ++VLLDG+K G V W L+G HN  N L  IAAARH GV    A   ALG+FN
  Sbjct: 251 KLTTDASEWEVLLDGEKVGEVKWSLVGEHNMHNGLMAIAAARHVGVAPADAANALGSFIN 310
  Query: 321 VKRRMEIKGTANGITVYDDFAHHPTAIETTIQGLRQRVGG-ARILAVLEPRSNTMKLGTM 379
  +RR+E++G ANG+TVYDDFAHHPTAI  T+  LR +VGG  ARI+AVLEPRSNTMK+G
  Sbjct: 311 ARRRLELRGEANGVTVYDDFAHHPTAILATLAALRGKVGGTARIIAVLEPRSNTMKMGIC 370
  Query: 380 KSALPASLKEADQVF-CYAGGADWDVAEALAPLGCRLRVGKDFDTFVAEIVKNARTGDHI 438
  K  L  SL  AD+VF        W VAE             D DT    +VK A+ GDHI
  Sbjct: 371 KDDLAPSLGRADEVFLLQPAHIPWQVAEVAEACVQPAHWSGDVDTLADMVVKTAQPGDHI 430
  Query: 439 LVMSNGGFGGIHTKLLDAL                                          457
  LVMSNGGFGGIH KLLD L
  Sbjct: 931 LVMSNGGFGGIHQKLLDGL                                          999

• Based on this analysis, it was predicted that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• ORF132-1 (26.4 kDa) was cloned in pET and pGex vectors and expressed in E. coli, as described above. The products of protein expression and purification were analyzed by SDS-PAGE. FIG. 20A shows the results of affinity purification of the His-fusion protein, and FIG. 20B shows the results of expression of the GST-fusion in E. coli. Purified His-fusion protein was used to immunise mice, whose sera were used for FACS analysis (FIG. 20C) and ELISA (positive result). These experiments confirm that ORF132 is a surface-exposed protein, and that it is a useful immunogen.
Example 103
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 875>

• 1 ..CCGGGCTATT ACGGCTCGGA TGACGAATTT AAGCGGGCAT TCGGAGAAAA
  51   CTCGCCGACA TmCAAGAAAC ATTGCAACCG GAGCTGCGGG ATTTATGAAC
  101   CCGTATTGAA AAAATACGGC AAAAAGCGCG CCAACAACCA TTCGGTCAGC
  151   ATTAGTGCGG ACTTCGGCGA TTATTTCATG CCGTTCGCCA GCTATTCGCG
  201   CACACACCGT ATGCCCAACA TCCAAGAAAT GTATTTTTCC CAAATCGGCG
  251   ACTCCGGCGT TCACACCGCC TTAAAACCAG AGCGCGCAAA CACTTGGCAA
  301   TTTGGCTTCr ATACCTATAA AAAAGGATTG TTAAAACAAG ATGATACATT
  351   AGGATTAAAA CTGGTCGGCT ACCGCAGCCG CATCGACAAC TACATCCACA
  401   ACGTTTACGG GAAATGGTGG GATTTGAACG GGGATATTCC GAGCTGGGTC
  451   AGCAGCACCG GGCTTGCCTA CACCATCCAA CATCGCrATT TCAwAGACAA
  501   AGTGCATCAA nnnnnnnnnn nnnnnnnnnn nnnnTACGAT TATGGGCGTT
  551   TTTTCACCAA CCTTTCTTAC GCCTATCAAA AAAGCACGCA ACCGACCAAC
  601   TTCAGCGATG CGAGCGAATC GCCCAACAAT GCGTCCAAAG AAGACCAACT
  651   CAAACAAGGT TATGGGTTGA GCAGGGTTTC CGCCCTGCCG CGAGATTACG
  701   GACGTTTGGA AGTCGGTACG CGCTGGTTGG GCAACAAACT GACTTTGGGC
  751   GGCGCGATGC GCTATTTCGG CAAGAGCATC CGCGCGACGG CTGAAGAACG
  801   CTATATCGAC GGCACCAACG GGGGAAATAC CAGCAATTTC CGGCAACTGG
  851   GCAAGCGTTC CATCAAACAA ACCGAAACTC TTGCCCGCCA GCCTTTGATT
  901   TTwGATTTTa ACGCCGCTTA CGAGCCGAAG AAAAACCTTA TTTTCCGCGC
  951   CGAAGTCAAA AATCTGTTCG ACAGGCGTTA TATCGATCCG CTCGATGCGG
  1001   GCAATGATGC GGCAAC.GAG CGTTATTACA GCTCGTTCGA CCCGAAAGAC
  1051   AAGGACrrAG ACGTAACGTG TAATGCTGAT AAAACGTTGT GCaACGGCAA
  1101   ATACGGCGGC ACAAGCAAAA GCGTATTGAC CAATTTTGCA CGCGGACGCA
  1151   CCTTTTTgAT GACGATGAGC TACAAGTTTT AA

• This corresponds to the amino acid sequence <SEQ ID 876; ORF133>:

• 1	..PGYYGSDDEF KRAFGENSPT XKKHCNRSCG IYEPVLKKYG
  	KKRANNHSVS
  51	ISADFGDYFM PFASYSRTHR MPNIQEMYFS QIGDSGVHTA
  	LKPERANTWQ
  101	FGFXTYKKGL LKQDDTLGLK LVGYRSRIDN YIHNVYGKWW
  	DLNGDIPSWV
  151	SSTGLAYTIQ HRXFXDKVHQ XXXXXXXXYD YGRFFTNLSY
  	AYQKSTQPTN
  201	FSDASESPNN ASKEDQLKQG YGLSRVSALP RDYGRLEVGT
  	RWLGNKLTLG
  251	GAMRYFGKSI RATAEERYID GTNGGNTSNF RQLGKRSIKQ
  	TETLARQPLI
  301	XDFNAAYEPK KNLIFRAEVK NLFDRRYIDP LDAGNDAAXE
  	RYYSSFDPKD
  351	KDXDVTCNAD KTLCNGKYGG TSKSVLTNFA RGRTFLMTMS
  	YKF*

• Further work revealed the further partial DNA sequence <SEQ ID 877>:

• 1	GAGGCGCAGA TACAGGTTTT GGAAGATGTG CACGTCAAGG
  	CGAAGCGCGT
  51	ACCGAAAGAC AAAAAAGTGT TTACCGATGC GCGTGCCGTA
  	TCGACCCGTC
  101	AGGATATATT CAAATCCAGC GAAAACCTCG ACAACATCGT
  	ACGCAGCATC
  151	CCCGGTGCGT TTACACAGCA AGATAAAAGC TCGGGCATTG
  	TGTCTTTGAA
  201	TATTCGCGGC GACAGCGGGT TCGGGCGGGT CAATACGATG
  	GTGGACGGCA
  251	TCACGCAGAC CTTTTATTCG ACTTCTACCG ATGCGGGCAG
  	GGCAGGCGGT
  301	TCATCTCAAT TCGGTGCATC TGTCGACAGC AATTTTATTG
  	CCGGACTGGA
  351	TGTCGTCAAA GGCAGCTTCA GCGGCTCGGC AGGCATCAAC
  	AGCCTTGCCG
  401	GTTCGGCGAA TCTGCGGACT TTAGGCGTGG ATGACGTCGT
  	TCAGGGCAAT
  451	AATACCTACG GCCTGCTGCT AAAAGGTCTG ACCGGCACCA
  	ATTCAACCAA
  501	AGGTAATGCG ATGGCGGCGA TAGGTGCGCG CAAATGGCTG
  	GAAAGCGGAG
  551	CATCTGTCGG TGTGCTTTAC GGGCACAGCA GGCGCAGCGT
  	GGCGCAAAAT
  601	TACCGCGTGG GCGGCGGCGG GCAGCACATC GGAAATTTTG
  	GCGCGGAATA
  651	TTTGGAACGG CGCAAGCAGC GATATTTTGT ACAAGAGGGT
  	GCTTTGAAAT
  701	TCAATTCCGA CAGCGGAAAA TGGGAGCGGG ATTTACAAAG
  	GCAACAGTGG
  751	AAATACAAGC CGTATAAAAA TTACAACAAC CAAGAACTAC
  	AaAAATACAT
  801	CGAAGAGCAT GACAAAAGCT GGCGGGAAAA CCTg.CaCCG
  	CAATACGACA
  851	TTACCCCCAT CGATCCGTCC AGCCTGAAGC AGCAGTCGGC
  	AGGCAATCTG
  901	TTTAAATTGG AATACGACGG CGTATTCAAT AAATACACGG
  	CGCAATTTCG
  951	CGATTTAAAC ACCAAAATCG GCAGCCGCAA AATCATCAAC
  	CGCAATTATC
  1001	AGTTCAATTA CGGTTTGTCT TTGAACCCGT ATACCAACCT
  	CAATCTGACC
  1051	GCAGCCTACA ATTCGGGCAG GCAGAAATAT CCGAAAGGGT
  	CGAAGTTTAC
  1101	AGGCTGGGGG CTTTTAAAGG ATTTTGAAAC CTACAACAAC
  	GCGAAAATCC
  1151	TCGACCTCAA CAACACCGCC ACCTTCCGGC TGCCCCGCGA
  	AACCGAGTTG
  1201	CAAACCACTT TGGGCTTCAA TTATTTCCAC AACGAATACG
  	GCAAAAACCG
  1251	CTTTCCTGAA GAATTGGGGC TGTTTTTCGA CGGTCCTGAT
  	CAGGACAACG
  1301	GGCTTTATTC CTATTTGGGG CGGTTTAAGG GCGATAAAGG
  	GCTGCTGCCC
  1351	CAAAAATCAA CCATTGTCCA ACCGGCCGGC AGCCAATATT
  	TCAACACGTT
  1401	CTACTTCGAT GCCGCGCTCA AAAAAGACAT TTACCGCTTA
  	AACTACAGCA
  1451	CCAATACCGT CGGCTACCGT TTCGGCGGCG AATATACGGG
  	CTATTACGGC
  1501	TCGGATGACG AATTTAAGCG GGCATTCGGA GAAAACTCGC
  	CGACATACAA
  1551	GAAACATTGC AACCGGAGCT GCGGGATTTA TGAACCCGTA
  	TTGAAAAAAT
  1601	ACGGCAAAAA GCGCGCCAAC AACCATTCGG TCAGCATTAG
  	TGCGGACTTC
  1651	GGCGATTATT TCATGCCGTT CGCCAGCTAT TCGCGCACAC
  	ACCGTATGCC
  1701	CAACATCCAA GAAATGTATT TTTCCCAAAT CGGCGACTCC
  	GGCGTTCACA
  1751	CCGCCTTAAA ACCAGAGCGC GCAAACACTT GGCAATTTGG
  	CTTCAATACC
  1801	TATAAAAAAG GATTGTTAAA ACAAGATGAT ACATTAGGAT
  	TAAAACTGGT
  1851	CGGCTACCGC AGCCGCATCG ACAACTACAT CCACAACGTT
  	TACGGGAAAT
  1901	GGTGGGATTT GAACGGGGAT ATTCCGAGCT GGGTCAGCAG
  	CACCGGGCTT
  1951	GCCTACACCA TCCAACATCG CAATTTCAAA GACAAAGTGC
  	ACAAACACGG
  2001	TTTTGAGTTG GAGCTGAATT ACGATTATGG GCGTTTTTTC
  	ACCAACCTTT
  2051	CTTACGCCTA TCAAAAAAGC ACGCAACCGA CCAACTTCAG
  	CGATGCGAGC
  2101	GAATCGCCCA ACAATGCGTC CAAAGAAGAC CAACTCAAAC
  	AAGGTTATGG
  2151	GTTGAGCAGG GTTTCCGCCC TGCCGCGAGA TTACGGACGT
  	TTGGAAGTCG
  2201	GTACGCGCTG GTTGGGCAAC AAACTGACTT TGGGCGGCGC
  	GATGCGCTAT
  2251	TTCGGCAAGA GCATCCGCGC GACGGCTGAA GAACGCTATA
  	TCGACGGCAC
  2301	CAACGGGGGA AATACCAGCA ATTTCCGGCA ACTGGGCAAG
  	CGTTCCATCA
  2351	AACAAACCGA AACTCTTGCC CGCCAGCCTT TGATTTTTGA
  	TTTTTACGCC
  2401	GCTTACGAGC CGAAGAAAAA CCTTATTTTC CGCGCCGAAG
  	TCAAAAATCT
  2451	GTTCGACAGG CGTTATATCG ATCCGCTCGA TGCGGGCAAT
  	GATGCGGCAA
  2501	CGCAGCGTTA TTACAGCTCG TTCGACCCGA AAGACAAGGA
  	CGAAGACGTA
  2551	ACGTGTAATG CTGATAAAAC GTTGTGCAAC GGCAAATACG
  	GCGGCACAAG
  2601	CAAAAGCGTA TTGACCAATT TTGCACGCGG ACGCACCTTT
  	TTGATGACGA
  2651	TGAGCTACAA GTTTTAA

• This corresponds to the amino acid sequence <SEQ ID 878; ORF133-1>:

• 1	EAQIQVLEDV HVKAKRVPKD KKVFTDARAV STRQDIFKSS
  	ENLDNIVRSI
  51	PGAFTQQDKS SGIVSLNIRG DSGFGRVNTM VDGITQTFYS
  	TSTDAGRAGG
  101	SSQFGASVDS NFIAGLDVVK GSFSGSAGIN SLAGSANLRT
  	LGVDDVVQGN
  151	NTYGLLLKGL TGTNSTKGNA MAAIGARKWL ESGASVGVLY
  	GHSRRSVAQN
  201	YRVGGGGQHI GNFGAEYLER RKQRYFVQEG ALKFNSDSGK
  	WERDLQRQQW
  251	KYKPYKNYNN QELQKYIEEH DKSWRENLXP QYDITPIDPS
  	SLKQQSAGNL
  301	FKLEYDGVFN KYTAQFRDLN TKIGSRKIIN RNYQFNYGLS
  	LNPYTNLNLT
  351	AAYNSGRQKY PKGSKFTGWG LLKDFETYNN AKILDLNNTA
  	TFRLPRETEL
  401	QTTLGFNYFH NEYGKNRFPE ELGLFFDGPD QDNGLYSYLG
  	RFKGDKGLLP
  451	QKSTIVQPAG SQYFNTFYFD AALKKDIYRL NYSTNTVGYR
  	FGGEYTGYYG
  501	SDDEFKRAFG ENSPTYKKHC NRSCGIYEPV LKKYGKKRAN
  	NHSVSISADF
  551	GDYFMPFASY SRTHRMPNIQ EMYFSQIGDS GVHTALKPER
  	ANTWQFGFNT
  601	YKKGLLKQDD TLGLKLVGYR SRIDNYIHNV YGKWWDLNGD
  	IPSWVSSTGL
  651	AYTIQHRNFK DKVHKHGFEL ELNYDYGRFF TNLSYAYQKS
  	TQPTNFSDAS
  701	ESPNNASKED QLKQGYGLSR VSALPRDYGR LEVGTRWLGN
  	KLTLGGAMRY
  751	FGKSIRATAE ERYIDGTNGG NTSNFRQLGK RSIKQTETLA
  	RQPLIFDFYA
  801	AYEPKKNLIF RAEVKNLFDR RYIDPLDAGN DAATQRYYSS
  	FDPKDKDEDV
  851	TCNADKTLCN GKYGGTSKSV LTNFARGRTF LMTMSYKF*

• Computer analysis of this amino acid sequence gave the following results:
• Homology with the Probable TonB-Dependent Receptor HI121 of H. influenzae (Accession Number U32801)
• ORF133 and HI121 show 57% aa identity in 363aa overlap:
• 
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF133 shows 90.8% identity over a 392aa overlap with an ORF (ORF133a) from strain A of N. meningitidis:
• 
• A partial ORF133a nucleotide sequence <SEQ ID 879> is:

• 1	AAAGACAAAA AAGTGTTTAC CGATGCGCGT GCCGTATCGA
  	CCCGTCAGGA
  51	TATATTCAAA TCCANCGAAA ACCTCGACAA CATCGTACGC
  	ANCATCCCCG
  101	GTGCGTTTAC ACANCAANAT AAAAGCTCGG GCNTTGTGTC
  	TTTGAATATT
  151	CGCNGCGACA GCGGGTTCGG GCGGGTCAAT ACNATGGTNG
  	ACGGCATCAC
  201	NCANACCTTT TATTCGACTT CTACCGATGC GGGCAGGGCA
  	GGCGGTTCAT
  251	CTCAATTCGG TGCATCTGTC GACAGCAATT TTATNGCCGG
  	ACTGGATGTC
  301	GTCAAAGGCA GCTTCAGCGG CTCGGCAGGC ATCAACAGCC
  	TTGCCGGTTC
  351	GGCGAATCTG CGGACTTTAN GCGTGGATGA TGTCGTTCAG
  	GGCAATANTA
  401	CNTACGGCCT GCTGCTAAAA GGTCTGACCG GCACCAATTC
  	AACCAAAGGT
  451	AATGCGATGG CGGCGATAGG TGCGCGCAAA TGGCTGGAAA
  	GCGGAGCATC
  501	TGTCGGTGTG CTTTACGGGC ACAGCAGGCG CAGCGTGGCG
  	CAAAATTACC
  551	GCGTGGGCGG CGGCGGGCAG CACATCGGAA ATTTTGGCGC
  	GGAATATCTG
  601	GAACGACGCA AGCAACGATA TTTTGAGCAA GAAGGCGGGT
  	TGAAATTCAA
  651	TTCCAACAGC GGAAAATGGG AGCGGGATTT CCAAAAGTCG
  	TACTGGAAAA
  701	CCAAGTGGTA TCAAAAATAC GATGCCCCCC AAGAACTGCA
  	AAAATACATC
  751	GAAGGTCATG ATAAAAGCTG GCGGGAAAAC CTGGCGCCGC
  	AATACGACAT
  801	CACCCCCATC GATCCGTCCA GCCTGAAGCN GCAGTCGGCA
  	GGCAACCTGT
  851	TTAAATTGGA ATACGACGGC GTATTCAATA AATACACGGC
  	GCAATTTCGC
  901	GATTTAAACA CCAAAATCGG CAGCCGCAAA ATCATCAACC
  	GCAATTATCA
  951	ATTCAATTAC GGTTTGTCTT TGAACCCGTA TACCAACCTC
  	AATCTGACCG
  1001	CAGCCTACAA TTCGGGCAGG CAGAAATATC CGAAAGGGTC
  	GAAGTTTACA
  1051	GGCTGGGGGC TTTTNAAAGA TTTTGAAACC TACAACAACG
  	CAAAAATCCT
  1101	CGACCTCANC AACACCTCCA CCTTCCGGCT GCCCCGTGAA
  	ACCGAGTTGC
  1151	AAACCACTTT GGGCTTCAAT TATTTCCACA ACGAATACGG
  	CAAAAACCGC
  1201	TTTCCTGAAG AATTGGGGCT GTTTTTCGAC GGTCCGGATC
  	ANGACAACGG
  1251	GCTTTATTCC TATTTGGGGC GGTTTAAGGG CGATAAAGGG
  	CTGCTGCCCC
  1301	AAAAATCAAC CATTGTCCAA CCGGCCGGCA GCCAATATTT
  	CAACACGTTC
  1351	TACTTCGATG CCGCGCTCAA AAAAGACATT TACCGCTTAA
  	ACTACAGCAC
  1401	CAATACCGTC GGCTACCGTT TCGGCGGCNA ATATACGGGC
  	TATTACNGCT
  1451	CGGATGACGA ATTTAAGCGG GCATTCGGAG AAAACTCGCC
  	GACATACANG
  1501	AAACATTGCA ACCAGAGCTG CGGAATTTAT GAACCCGTAT
  	TGAAAAAATA
  1551	CGGCAAAAAG CGCGCCAACA ACCATTCGGT CAGCATTAGT
  	GCGGACTTCG
  1601	GCGATTATTT CATGCCGTTC GCCAGCTATT CGCGCACACA
  	CCGTATGCCC
  1651	AACATCCAAG AAATGTATTT TTCCCAAATC GGCGACTCCG
  	GCGTTCACAC
  1701	CGCCTTAAAA CCAGAGCGCG CAAACACTTG GCAATTTGGC
  	TTCAATACCT
  1751	ATAAAAAAGG ATTGTTAAAA CAAGATGATA TATTAGGATT
  	AAAACTGGTC
  1801	GGCTACCGCA GCCGCATCGA CNACTACATC CACAACGTTT
  	ACGGGAAATG
  1851	GTGGGATTTG AACGGGAATA TTCCGAGCTG GGTCAGCAGC
  	ACCGGGCTTG
  1901	CCTACACCAT CCAACACCGC AATTTCAAAG ACAAAGTGCA
  	CAAACACGGT
  1951	TTTGAGTTGG AGCTGAATTA CGATTATNGG CGTTTTTTCA
  	CCAACCTTTC
  2001	TTACGCCTAT CAAAAAAGCA CGCAACCGAC CAACTTCAGC
  	GATGCGAGCG
  2051	AATCGCCCAA CAATGCGTCC AAAGAAGACC AACTCAAACA
  	AGGTTATGGG
  2101	TTGAGCAGGG TTTCCGCCCT GCCGCGAGAT TACGGACGTT
  	TGGAAGTCGG
  2151	TACGCGCTGG TTGGGCAACA AACTGACTTT GGGCGGCGCG
  	ATGCGCTATT
  2201	TCGGCAAGAG CATCCGCGCG ACGGCTGAAG AACGCTATAT
  	CGACGNCACC
  2251	AATGGGGNAN NTACCAGCAA TTTCCGGCAA CTGGGCAAGC
  	GTTCCATCAN
  2301	ACAAACCGAA ACCCTTGCCC GCCAGCCTTT GATTTTTGAT
  	TTNTACGCCG
  2351	CTTACGAGCC GAAGAAAAAN CTTATTTTCC GCGCCGAAGT
  	CAAAAATCTG
  2401	TTCGACAGGC GTTATATCGA TCCGCTCGAT GCGGGCAATG
  	ATGCGGCAAC
  2451	GCAGCGTTAT TACAGTTCGT TCGACCCGAA AGACAAGGAC
  	GAAGAAGTAA
  2501	CGTGTAATGA TGATAACACG TTATGCAACG GCAAATACGG
  	CGGCACAAGC
  2551	AAAAGCGTAT TGACCAATTT TGCACGCGGA CNCACCTTTT
  	TGATAACGAT
  2601	GAGCTACAAG TTTTAA

• This encodes a protein having (partial) amino acid sequence <SEQ ID 880>:

• 1	KDKKVFTDAR AVSTRQDIFK SXENLDNIVR XIPGAFTXQX
  	KSSGXVSLNI
  51	RXDSGFGRVN TMVDGITXTF YSTSTDAGRA GGSSQFGASV
  	DSNFXAGLDV
  101	VKGSFSGSAG INSLAGSANL RTLXVDDVVQ GNXTYGLLLK
  	GLTGTNSTKG
  151	NAMAAIGARK WLESGASVGV LYGHSRRSVA QNYRVGGGGQ
  	HIGNFGAEYL
  201	ERRKQRYFEQ EGGLKFNSNS GKWERDFQKS YWKTKWYQKY
  	DAPQELQKYI
  251	EGHDKSWREN LAPQYDITPI DPSSLKXQSA GNLFKLEYDG
  	VFNKYTAQFR
  301	DLNTKIGSRK IINRNYQFNY GLSLNPYTNL NLTAAYNSGR
  	QKYPKGSKFT
  351	GWGLXKDFET YNNAKILDLX NTSTFRLPRE TELQTTLGFN
  	YFHNEYGKNR
  401	FPEELGLFFD GPDXDNGLYS YLGRFKGDKG LLPQKSTIVQ
  	PAGSQYFNTF
  451	YFDAALKKDI YRLNYSTNTV GYRFGGXYTG YYXSDDEFKR
  	AFGENSPTYX
  501	KHCNQSCGIY EPVLKKYGKK RANNHSVSIS ADFGDYFMPF
  	ASYSRTHRMP
  551	NIQEMYFSQI GDSGVHTALK PERANTWQFG FNTYKKGLLK
  	QDDILGLKLV
  601	GYRSRIDXYI HNVYGKWWDL NGNIPSWVSS TGLAYTIQHR
  	NFKDKVHKHG
  651	FELELNYDYX RFFTNLSYAY QKSTQPTNFS DASESPNNAS
  	KEDQLKQGYG
  701	LSRVSALPRD YGRLEVGTRW LGNKLTLGGA MRYFGKSIRA
  	TAEERYIDXT
  751	NGXXTSNFRQ LGKRSIXQTE TLARQPLIFD XYAAYEPKKX
  	LIFRAEVKNL
  801	FDRRYIDPLD AGNDAATQRY YSSFDPKDKD EEVTCNDDNT
  	LCNGKYGGTS
  851	KSVLTNFARG XTFLITMSYK F*

• ORF133a and ORF133-1 show 94.3% identity in 871 aa overlap:
• 

  
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF133 shows 92.3% identity over 392 aa overlap with a predicted ORF (ORF133ng) from N. gonorrhoeae:
• 
• The complete length ORF133ng nucleotide sequence <SEQ ID 881> is predicted to encode a protein having amino acid sequence <SEQ ID 882>:

• 1	MRSSFRLKPI CFYLMGVMLY HHSYAEDAGR AGSEAQIQVL
  	EDVHVKAKRV
  51	PKDKKVFTDA RAVSTRQDVF KSGENLDNIV RSIPGAFTQQ
  	DKSSGIVSLN
  101	IRGDSGFGRV NTMVDGITQT FYSTSTDAGR AGGSSQFGAS
  	VDSNFIAGLD
  151	VVKGSFSGSA GINSLAGSAN LRTLGVDDVV QGNNTYGLLL
  	KGLTGTNSTK
  201	GNAMAAIGAR KWLESGASVG VLYGHSRRGV AQNYRVGGGG
  	QHIGNFGEEY
  251	LERRKQQYFV QEGGLKFNAG SGKWERDLQR QYWKTKWYKK
  	YEDPQELQKY
  301	IEEHDKSWRE NLAPQYDITP IDPSGLKQQS AGNLLNLEYD
  	GVFNKYTAQF
  351	RDLNTRIGSR KIINRNYQFN YGLSLNPYTN LNLTAAYNSG
  	RQKYPKGAKF
  401	TGWGLLKDFE TYNNAKILDL NNTATFRLPR ETELQTTLGF
  	NYFHNEYGKN
  451	RFPEELGLFF DGPDQDNGLY SYLGRFKGDK GLLPQKSTIV
  	QPAGSQYFNT
  501	FYFDAALKKD IYRLNYSTNA INYRFGGEYT GYYGSENEFK
  	RAFGENSPAY
  551	KEHCDPSCGL YEPVLKKYGK KRANNHSVSI SADFGDYFMP
  	FAGYSRTHRM
  601	PNIQEMYFSQ IGDSGVHTAL KPERANTWQF GFNTYKKGLL
  	KQDDILGLKL
  651	VGYRSRIDNY IHNVYGKWWD LNGDIPSWVG STGLAYTIRH
  	RNFKDKVHKH
  701	GFELELNYDY GRFFTNLSYA YQKSTQPTNF SDASESPNNA
  	SKEDQLKQGY
  751	GLSRVSALPR DYGRLEVGTR WLGNKLTLGG AMRYFGKSIR
  	ATAEERYIDG
  801	TNGGNTSNVR QLGKRSIKQT ETLARQPLIF DFYAAYEPKK
  	NLIFRAEVKN
  851	LFDRRYIDPL DAGNDAATQR YYSSFDPKDK DEDVTCNADK
  	TLCNGKYGGT
  901	SKSVLTNFAR GRTFLMTMSY KF*

• A variant was also identified, being encoded by the gonococcal DNA sequence <SEQ ID 883>:

• 1 ATGAGATCTT CTTTCCGGTT GAAGCCGATT TGTTTTTATC TTATGGGTGT
  51 TATGCTATAT CATCATAGTT ATGCCGAAGA TGCAGGGCGC GCGGGCAGCG
  101 AGGCGCAGAT ACAGGTTTTG GAAGATGTGC ACGTCAAGGC GAAGCGCGTA
  151 CCGAAAGACA AAAAAGTGTT TACCGATGCG CGTGCCGTAT CGACCCGTca
  201 gGATGTGTTC AAATCCGGCG AAAACCTCGA CAACATCGTA CGCAGCATAC
  251 CCGGTGCGTT TACACAGCAA GATAAAAGCT CGGGCATTGT GTCTTTGAAT
  301 ATTCCCGGCG ACAGCGGGTT CGGGCGGGTC AATACGATGG TGGACGGCAT
  351 CACGCAGACC TTTTATTCGA CTTCTACCGA TGCGGGCAGG GCAGGCGGTT
  401 CATCTCAATT CGGTGCATCT GTCGACAGCA ATTTTATTGC CGGACTGGAT
  451 GTCGTCAAAG GCAGCTTCAG CGGCTCGGCA GGCATCAACA GCCTTGCCGG
  501 TTCGGCGAAT CTGCGGACTT TAGGCGTGGA TGACGTCGTT CAGGGCAATA
  551 ATACCTACGG CCTGCTGCTA AAAGGTCTGA CCGGCACCAA TTCAACCAAA
  601 GGTAATGCGA TGGCGGCGAT AGGTGCGCGC AAATGGCTGG AAAGCGGAGC
  651 GTCTGTCGGT GTGCTTTACG GGCACAGCAG GCGCGGCGTG GCGCAAAATT
  701 ACCGCGTGGG CGGCGGCGGG CAGCACATCG GAAATTTTGG TGAAGAATAT
  751 CTGGAACGGC GCAAACAGCA ATATTTTGTA CAAGAGGGTG GTTTGAAATT
  801 CAATGCCGGC AGCGGAAAAT GGGAACGGGA TTTGCAAAGG CAATACTGGA
  851 AAACAAAGTG GTATAAAAAA TACGAAGACC CCCAAGAACT GCAAAAATAC
  901 ATCGAAGAGC ATGATAAAAG CTGGCGGGAA AACCTGGCGC CGCAATACGA
  951 CATCACCCCC ATCGATCCGT CCGGCCTGAA GCAGCAGTCG GCAGGCAATC
  1001 TGTTTAAATT GGAATACGAC GGCGTATTCA ATAAATACAC GGCGCAATTT
  1051 CGCGATTTAA ACACCAGAAT CGGCAGCCGC AAAATCATCA ACCGCAATTA
  1101 TCAATTCAAT TACGGTTTGT CTTTGAACCC GTATACCAAC CTCAATCTGA
  1151 CCGCAGCCTA CAATTCGGGC AGGCAGAAAT ATCCGAAAGG GGCGAAGTTT
  1201 ACAGGCTGGG GGCTTTTAAA AGATTTTGAA ACCTACAACA ACGCGAAAAT
  1251 CCTCGACCTC AACAACACCG CCACCTTCCG GCTGCCCCGC GAAACCGAGT
  1301 TGCAAACCAC TTTGGGCTTC AATTATTTCC ACAACGAATA CGGCAAAAAC
  1351 CGCTTTCCTG AAGAATTGGG GCTGTTTTTC GACGGTCCTG ATCAGGACAA
  1401 CGGGCTTTAT TCCTATTTGG GGCGGTTTAA GGGCGATAAA GGGCTGTTGC
  1451 CTCAAAAATC AACCATTGTC CAACCGGCCG GCAGCCAATA TTTCAACACG
  1501 TTCTACTTCG ATGCCGCGCT CAAAAAAGAC ATTTACCGCT TAAACTACAG
  1551 CACCAATGCA ATCAACTACC GTTTCGGCGG CGAATATACG GGCTATTACG
  1601 GCTCGGAAAA CGAATTTAAG CGGGCATTCG GAGAAAACTC GCCGGCATAC
  1651 AAGGAACATT GCGACCCGAG CTGCGGGCTT TATGAACCCG TATTGAAAAA
  1701 ATACGGCAAA AAGCGCGCCA ACAACCATTC GGTCAGCATT AGTGCGGACT
  1751 TCGGCGATTA TTTCATGCCG TTCGCCGGCT ATTCGCGCAC ACACCGTATG
  1801 CCCAACATCC AAGAAATGTA TTTTTCCCAA ATCGGCGACT CCGGCGTTCA
  1851 CACCGCCTTA AAACCAGAGC GCGCAAACAC TTGGCAATTT GGCTTCAATA
  1901 CCTATAAAAA AGGATTGTTA AAACAAGATG ATATATTAGG ATTGAAACTG
  1951 GTCGGCTACC GCAGCCGCAT TGACAACTAC ATCCACAACG TTTACGGGAA
  2001 ATGGTGGGAT TTGAACGGGG ATATTCCGAG CTGGGTCGGC AGCACCGGGC
  2051 TTGCCTACAC CATCCGACAC CGCAATTTCA AAGACAAAGT GCACAAACAC
  2101 GGTTTTGAGC TGGAGCTGAA TTACGATTAT GGGCGTTTTT TCACCAACCT
  2151 TTCTTACGCC TATCAAAAAA GCACGCAACC GACCAATTTC AGCGATGCGA
  2201 GCGAATCGCC CAACAATGCC tccaaAGAAG ACCAACTCAA ACAAGGTTAT
  2251 GGGCTGAGCA GGGTTTCCGC CCTGCCGCGA GATTACGGAC GTTTGGAAGT
  2301 CGGTACGCGC TGGTTGGGCA ACAAACTGAC TTTGGGCGGC GCGAtgCGCT
  2351 ATTTCGGCAA GAGCATCCGC GCGACGGCTG AAGAACGCTA TATCGACGGC
  2401 ACCAACGGGG GAAATACCAG CAATGTCCGG CAACTGGGCA AGCGTTCCAT
  2451 CAAACAAACC GAAACCCTTG CCCGACAGCC TTTGATTTTT GATTTTTACG
  2501 CCGCTTACGA GCCGAAGAAA AACCTTATTT TCCGCGCCGA AGTCAAAAAC
  2551 CTGTTCGACA GGCGTTATAT CGATCCGCTC GATGCGGGCA ATGATGCGGC
  2601 AACGCAGCGT TATTACAGCT CGTTCGACCC GAAAGACAAG GACGAAGACG
  2651 TAACGTGTAA TGCTGATAAA ACGTTGTGCA ACGGCAAATA CGGCGGCACA
  2701 AGCAAAAGCG TATTGACCAA TTTCGCACGC GGACGCACCT TCTTGATGAC
  2751 GATGAGCTAC AAGTTTTAA

• This corresponds to the amino acid sequence <SEQ ID 884; ORF133ng-1>:

• 1 MRSSFRLKPI CFYLMGVMLY HHSYAEDAGR AGSEAQIQVL EDVHVKAKRV
  51 PKDKKVFTDA RAVSTRQDVF KSGENLDNIV RSIPGAFTQQ DKSSGIVSLN
  101 IRGDSGFGRV NTMVDGITQT FYSTSTDAGR AGGSSQFGAS VDSNFIAGLD
  151 VVKGSFSGSA GINSLAGSAN LRTLGVDDVV QGNNTYGLLL KGLTGTNSTK
  201 GNAMAAIGAR KWLESGASVG VLYGHSRRGV AQNYRVGGGG QHIGNFGEEY
  251 LERRKQQYFV QEGGLKFNAG SGKWERDLQR QYWKTKWYKK YEDPQELQKY
  301 IEEHDKSWRE NLAPQYDITP IDPSGLKQQS AGNLFKLEYD GVFNKYTAQF
  351 RDLNTRIGSR KIINRNYQFN YGLSLNPYTN LNLTAAYNSG RQKYPKGAKF
  401 TGWGLLKDFE TYNNAKILDL NNTATFRLPR ETELQTTLGF NYFHNEYGKN
  451 RFPEELGLFF DGPDQDNGLY SYLGRFKGDK GLLPQKSTIV QPAGSQYFNT
  501 FYFDAALKKD IYRLNYSTNA INYRFGGEYT GYYGSENEFK RAFGENSPAY
  551 KEHCDPSCGL YEPVLKKYGK KRANNHSVSI SADFGDYFMP FAGYSRTHRM
  601 PNIQEMYFSQ IGDSGVHTAL KPERANTWQF GFNTYKKGLL KQDDILGLKL
  651 VGYRSRIDNY IHNVYGKWWD LNGDIPSWVG STGLAYTIRH RNFKDKVHKH
  701 GFELELNYDY GRFFTNLSYA YQKSTQPTNF SDASESPNNA SKEDQLKQGY
  751 GLSRVSALPR DYGRLEVGTR WLGNKLTLGG AMRYFGKSIR ATAEERYIDG
  801 TNGGNTSNVR QLGKRSIKQT ETLARQPLIF DFYAAYEPKK NLIFRAEVKN
  851 LFDRRYIDPL DAGNDAATQR YYSSFDPKDK DEDVTCNADK TLCNGKYGGT
  901 SKSVLTNFAR GRTFLMTMSY KF*

• ORF133ng-1 and ORF133-1 show 96.2% identity in 889 aa overlap:
• 

  
• In addition, ORF133ng-1 is homologous to a TonB-dependent receptor in H. influenzae:

• sp|P45114|YC17_HAEIN PROBABLE TONB-DEPENDENT RECEPTOR HI1217 PRECURSOR
  >gi|1075372|pir||G64110 transferrin binding protein 1 precursor (tbp1)
  homolog - Haemophilus influenzae (strain Rd KW20) >gi|1574147 (U32801)
  transferrin binding protein 1 precursor (tbp1) [Haemophilus influenzae]
  Length = 913
  Score = 930 bits (2377), Expect = 0.0
  Identities = 476/921 (51%), Positives = 619/921 (66%), Gaps = 72/921 (7%)

  Query:	38	QVLEDVHVKAKRVPKDKKVFTDARAVSTRQDVFKSGENLDNIVRSIPGAFTQQDKSSGIV	97
  		+ L  + V  K +  DKK FT+A+A STR++VFK  + +D ++RSIPGAFTQQDK SG+V
  Sbjct:	29	ETLGQIDVVEKVISNDKKPFTEAKAKSTRENVFKETQTIDQVIRSIPGAFTQQDKGSGVV	88
  Query:	98	SLNIRGDSGFGRVNTMVDGITQTFYSTSTDAGRAGGSSQFGASVDSNFIAGLDVVKGSFS	157
  		S+NIRG++G GRVNTMVDG+TQTFYST+ D+G++GGSSQFGA++D NFIAG+DV K +FS
  Sbjct:	89	SVNIRGENGLGRVNTMVDGVTQTFYSTALDSGQSGGSSQFGAAIDPNFIAGVDVNKSNFS	148
  Query:	158	GSAGINSLAGSANLRTLGVDDVVQXXXXXXXXXXXXXXXXXXXXXAMAAIGARKWLESGA	217
  		G++GIN+LAGSAN RTLGV+DV+                       M     RKWL++G
  Sbjct:	149	GASGINALAGSANFRTLGVNDVITDDKPFGIILKGMTGSNATKSNFMTMAAGRKWLDNGG	208
  Query:	218	SVGVLYGHSRRGVAQNYRVGGGGQHIGNFGEEYLERRKQQYFVQEGGLKFNAGSGKWERD	277
  		VGV+YG+S+R V+Q+YR+ GGG+ + + G++ L + K+ YF +  G   N   G+W  D
  Sbjct:	209	YVGVVYGYSQREVSQDYRI-GGGERLASLGQDILAKEKEAYF-RNAGYILNP-EGQWTPD	265
  Query:	278	LQRQYWK-----------TKWY--------------------KKYEDPQELQK---YIEE	303
  		L +++W              +Y                    KK +D ++LQK    IEE
  Sbjct:	266	LSKKHWSCNKPDYQKNGDCSYYRIGSAAKTRREILQELLTNGKKPKDIEKLQKGNDGIEE	325
  Query:	304	HDKSWRENLAPQYDITPIDPSGLKQQSAGNLFKLEYDGVFNKYTAQFRDLNTRIGSRKII	363
  		DKS+  N   QY + PI+P  L+ +S  +L K EY        AQ R L+ +IGSRKI
  Sbjct:	326	TDKSFERN-KDQYSVAPIEPGSLQSRSRSHLLKFEYGDDHQNLGAQLRTLDNKIGSRKIE	384
  Query:	364	NRNYQFNYGLSLNPYTNLNLTAAYNSGRQKYPKGAKFTGWGLLKDFETYNNAKILDLNNT	423
  		NRNYQ NY  + N Y +LNL AA+N G+  YPKG  F GW +     T N A I+D+NN+
  Sbjct:	385	NRNYQVNYNFNNNSYLDLNLMAAHNIGKTIYPKGGFFAGWQVADKLITKNVANIVDINNS	444
  Query:	424	ATFRLPRETELQTTLGFNYFHNEYGKNRFPEELGLFFDGPDQDNGLYSY--LGRFKGDKG	481
  		TF LP+E +L+TTLGFNYF NEY KNRFPEEL LF++    D GLYS+   GR+ G K
  Sbjct:	445	HTFLLPKEIDLKTTLGFNYFTNEYSKNRFPEELSLFYNDASHDQGLYSHSKRGRYSGTKS	504
  Query:	482	LLPQKSTIVQPAGSQYFNTFYFDAALKKDIYRLNYSTNAINYRFGGEYTGYYGSENEFKR	541
  		LLPQ+S I+QP+G Q F T YFD AL K IY LNYS N  +Y F GEY GY
  Sbjct:	505	LLPQRSVILQPSGKQKFKTVYFDTALSKGIYHLNYSVNFTHYAFNGEYVGY---------	555
  Query:	542	AFGENSPAYKEHCDPSCGLYEPVLKKYGKKRANNHSVSISADFGDYFMPFAGYSRTHRMP	601
  		EN+   +        + EP+L K G K+A NHS ++SA+  DYFMPF  YSRTHRMP
  Sbjct:	556	---ENTAGQQ--------INEPILHKSGHKKAFNHSATLSAELSDYFMPFFTYSRTHRMP	604
  Query:	602	NIQEMYFSQIGDSGVHTALKPERANTWQFGFNTYKKGLLKQDDILGLKLVGYRSRIDNYI	661
  		NIQEM+FSQ+ ++GV+TALKPE+++T+Q GFNTYKKGL  QDD+LG+KLVGYRS I NYI
  Sbjct:	605	NIQEMFFSQVSNAGVNTALKPEQSDTYQLGFNTYKKGLFTQDDVLGVKLVGYRSFIKNYI	664
  Query:	662	HNVYGKWWDLNGDIPSWVGSTGLAYTIRHRNFKDKVHKHGFELELNYDYGRFFTNLSYAY	721
  		HNVYG WW     +P+W  S G  YTI H+N+K  V K G ELE+NYD GRFF N+SYAY
  Sbjct:	665	HNVYGVWW--RDGMPTWAESNGFKYTIAHQNYKPIVKKSGVELEINYDMGRFFANVSYAY	722
  Query:	722	QKSTQPTNFSDASESPNNASKEDQLKQGYGLSRVSALPRDYGRLEVGTRWLGNKLTLGGA	781
  		Q++ QPTN++DAS  PNNAS+ED LKQGYGLSRVS LP+DYGRLE+GTRW   KLTLG A
  Sbjct:	723	QRTNQPTNYADASPRPNNASQEDILKQGYGLSRVSMLPKDYGRLELGTRWFDQKLTLGLA	782
  Query:	782	MRYFGKSIRATAEERYIDGTNGGNTSNVRQLGKRSIKQTETLARQPLIFDFYAAYEPKKN	841
  		RY+GKS RAT EE YI+G+     + +R+    ++K+TE + +QP+I D + +YEP K+
  Sbjct:	783	ARYYGKSKRATIEEEYINGSR-FKKNTLRRENYYAVKKTEDIKKQPIILDLHVSYEPIKD	841
  Query:	842	LIFRAEVKNLFDRRYIDPLDAGNDAATQRYYSSFDPKDKDEDVTCNADKTLCNGKYGGTS	901
  		LI +AEV+NL D+RY+DPLDAGNDAA+QRYYSS      +  + C  D + C    GG+
  Sbjct:	842	LIIKAEVQNLLDKRYVDPLDAGNDAASQRYYSSL-----NNSIECAQDSSAC----GGSD	892
  Query:	902	KSVLTNFARGRTFLMTMSYKF	922
  		K+VL NFARGRT++++++YKF
  Sbjct:	893	KTVLYNFARGRTYILSLNYKF	913

• The underlined motif in the gonococcal protein (also present in the meningococcal protein) is predicted to be an ATP/GTP-binding site motif A (P-loop), and the analysis suggests that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
Example 104
• The following partial DNA sequence was identified in N. meningitidis <SEQ ID 885>

• 1	ATGAACCTGA TTTCACGTTA CATCATCCGT CAAATGGCGG
  	TTATGGCGGT
  51	TTACGCGCTC CTTGCCTTCC TCGCTTTGTA CAGCTTTTTT
  	GAAATCCTGT
  101	ACGAAACCGG CAACCTCGGC AAAGGCAGTT ACGGCATATG
  	GGAAATGCTG
  151	GGCTACACCG CCCTCAAAAT GCCCGCCCGC GCCTACGAAC
  	TGATTCCCCT
  201	CGCCGTCCTT ATCGGCGGAC TGGTCTCCCT CAGCCAGCTT
  	GCCGCCGGCA
  251	GCGAACTGAC CGTCATCAAA GCCAGCGGCA TGAGCACCAA
  	AAAGCTGCTG
  301	TTGATTCTGT CGCAGTTCGG TTTTATTTTT GCTATTGCCA
  	CCGTCGCGCT
  351	CGGCGAATGG GTTGCGCCCA CACTGAGCCA AAAAGCCGAA
  	AACATCAAAG
  401	CCGCCGCCAT CAACGGCAAA ATCAGCACCG GCAATACCGG
  	CCTTTGGCTG
  451	AAAGAAAAAA ACAGCGTGAT CAATGTGCGC GAAATGTTGC
  	CCGACCAT..

• This corresponds to the amino acid sequence <SEQ ID 886; ORF112>:

• 1	MNLISRYIIR QMAVMAVYAL LAFLALYSFF EILYETGNLG
  	KGSYGIWEML
  51	GYTALKMPAR AYELIPLAVL IGGLVSLSQL  AAGSELTVIK
  	ASGMSTKKLL
  101	LILSQFGFIF AIATVALGEW VAPTLSQKAE NIKAAAINGK
  	ISTGNTGLWL
  151	KEKNSVINVR EMLPDH...

• Further work revealed further partial nucleotide sequence <SEQ ID 887>:

• 1	ATGAACCTGA TTTCACGTTA CATCATCCGT CAAATGGCGG
  	TTATGGCGGT
  51	TTACGCGCTC CTTGCCTTCC TCGCTTTGTA CAGCTTTTTT
  	GAAATCCTGT
  101	ACGAAACCGG CAACCTCGGC AAAGGCAGTT ACGGCATATG
  	GGAAATGCTG
  151	gGCTACACCG CCCTCAAAAT GCCCGCCCGC GCCTACGAAC
  	TGATTCCCCT
  201	CGCCGTCCTT ATCGGCGGAC TGGTCTCCCT CAGCCAGCTT
  	GCCGCCGGCA
  251	GCGAACTGAC CGTCATCAAA GCCAGCGGCA TGAGCACCAA
  	AAAGCTGCTG
  301	TTGATTCTGT CGCAGTTCGG TTTTATTTTT GCTATTGCCA
  	CCGTCGCGCT
  351	CGGCGAATGG GTTGCGCCCA CACTGAGCCA AAAAGCCGAA
  	AACATCAAAG
  401	CCGCCGCCAT CAACGGCAAA ATCAGCACCG GCAATACCGG
  	CCTTTGGCTG
  451	AAAGAAAAAA ACAGCrTkAT CAATGTGCGC GAAATGTTGC
  	CCGACCATAC
  501	GCTTTTGGGC ATCAAAATTT GGGCGCGCAA CGATAAAAAC
  	GAATTGGCAG
  551	AGGCAGTGGA AGCCGATTCC GCCGTTTTGA ACAGCGACGG
  	CAGTTGGCAG
  601	TTGAAAAACA TCCGCCGCAG CACGCTTGGC GAAGACAAAG
  	TCGAGGTCTC
  651	TATTGCGGCT GAAGAAAACT GGCCGATTTC CGTCAAACGC
  	AACCTGATGG
  701	ACGTATTGCT CGTCAAACCC GACCAAATGT CCGTCGGCGA
  	ACTGACCACC
  751	TACATCCGCC ACCTCCAAAA CAACAGCCAA AACACCCGAA
  	TCTACGCCAT
  801	CGCATGGTGG CGCAAATTGG TTTACCCCGC CGCAGCCTGG
  	GTGATGGCGC
  851	TCGTCGCCTT TGCCTTTACC CCGCAAACCA CCCGCCACGG
  	CAATATGGGC
  901	TTAAAACTCT TCGGCGGCAT CTGTsTCGGA TTGCTGTTCC
  	ACCTTGCCGG
  951	ACGGCTCTTT GGGTTTACCA GCCAACTCGG...

• This corresponds to the amino acid sequence <SEQ ID 888; ORF112-1>:

• 1	MNLISRYIIR QMAVMAVYAL LAFLALYSFF EILYETGNLG
  	KGSYGIWEML
  51	GYTALKMPAR AYELIPLAVL IGGLVSLSQL  AAGSELTVIK
  	ASGMSTKKLL
  101	LILSQFGFIF AIATVALGEW VAPTLSQKAE NIKAAAINGK
  	ISTGNTGLWL
  151	KEKNSXINVR EMLPDHTLLG IKIWARNDKN ELAEAVEADS
  	AVLNSDGSWQ
  201	LKNIRRSTLG EDKVEVSIAA EENWPISVKR NLMDVLLVKP
  	DQMSVGELTT
  251	YIRHLQNNSQ NTRIYAIAWW RKLVYPAAAW VMALVAFAFT
  	PQTTRHGNMG
  301	LKLFGGICXG LLFHLAGRLF GFTSQL...

• Computer analysis of this amino acid sequence predicts two transmembrane domains and gave the following results:
• Homology with a Predicted ORF from N. meningitidis (Strain A)
• ORF112 shows 96.4% identity over a 166aa overlap with an ORF (ORF112a) from strain A of N. meningitidis:
• 
• The ORF112a nucleotide sequence <SEQ ID 889> is:

• 1	ATGAACCTGA TTTCACGTTA CATCATCCGT CAAATGGCGG
  	TTATGGCGGT
  51	TTACGCGCTC CTTGCCTTCC TCGCTTTGTA CAGCTTTTTT
  	GAAATCCTGT
  101	ACGAAACCGG CAACCTCGGC AAAGGCAGTT ACGGCATATG
  	GGAAATGNTG
  151	GGNTACACCG CCCTCAAAAT GNCCGCCCGC GCCTACGAAC
  	TGATGCCCCT
  201	CGCCGTCCTT ATCGGCGGAC TGGTCTCTNT CAGCCAGCTT
  	GCCGCCGGCA
  251	GCGAACTGAN CGTCATCAAA GCCAGCGGCA TGAGCACCAA
  	AAAGCTGCTG
  301	TTGATTCTGT CGCAGTTCGG TTTTATTTTT GCTATTGCCA
  	CCGTCGCGCT
  351	CGGCGAATGG GTTGCGCCCA CACTGAGCCA AAAAGCCGAA
  	AACATCAAAG
  401	CCGCGGCCAT CAACGGCAAA ATCAGTACCG GCAATACCGG
  	CCTTTGGCTG
  451	AAAGAAAAAA ACAGCATTAT CAATGTGCGC GAAATGTTGC
  	CCGACCATAC
  501	CCTGCTGGGC ATTAAAATCT GGGCCCGCAA CGATAAAAAC
  	GAACTGGCAG
  551	AGGCAGTGGA AGCCGATTCC GCCGTTTTGA ACAGCGACGG
  	CAGTTGGCAG
  601	TTGAAAAACA TCCGCCGCAG CACGCTTGGC GAAGACAAAG
  	TCGAGGTCTC
  651	TATTGCGGCT GAAGAAAANT GGCCGATTTC CGTCAAACGC
  	AACCTGATGG
  701	ACGTATTGCT CGTCAAACCC GACCAAATGT CCGTCGGCGA
  	ACTGACCACC
  751	TACATCCGCC ACCTCCAAAN NNACAGCCAA AACACCCGAA
  	TCTACGCCAT
  801	CGCATGGTGG CGCAAATTGG TTTACCCCGC CGCAGCCTGG
  	GTGATGGCGC
  851	TCGTCGCCTT TGCCTTTACC CCGCAAACCA CCCGCCACGG
  	CAATATGGGC
  901	TTAAAANTCT TCGGCGGCAT CTGTCTCGGA TTGCTGTTCC
  	ACCTTGCCGG
  951	NCGGCTCTTC NGGTTTACCA GCCAACTCTA CGGCATCCCG
  	CCCTTCCTCG
  1001	NCGGCGCACT ACCTACCATA GCCTTCGCCT TGCTCGCCGT
  	TTGGCTGATA
  1051	CGCAAACAGG AAAAACGCTA A

• This encodes a protein having the amino acid sequence <SEQ ID 890>:

• 1	MNLISRYIIR QMAVMAVYAL LAFLALYSFF EILYETGNLG
  	KGSYGIWEMX
  51	GYTALKMXAR AYELMPLAVL IGGLVSXSQ L AAGSELXVIK
  	ASGMSTKKLL
  101	LILSQFGFIF AIATVALGEW VAPTLSQKAE NIKAAAINGK
  	ISTGNTGLWL
  151	KEKNSIINVR EMLPDHTLLG IKIWARNDKN ELAEAVEADS
  	AVLNSDGSWQ
  201	LKNIRRSTLG EDKVEVSIAA EEXWPISVKR NLMDVLLVKP
  	DQMSVGELTT
  251	YIRHLQXXSQ NTRIYAIAWW RKLVYPAAAW VMALVAFAFT
  	PQTTRHGNMG
  301	LKXFGGICLG LLFHLAGRLF XFTSQLYGIP PFLXGALPTI
  	AFALLAVWLI
  351	RKQEKR*

• ORF112a and ORF112-1 show 96.3% identity in 326 aa overlap:
• 
• Homology with a Predicted ORF from N. gonorrhoeae
• ORF112 shows 95.8% identity over 166aa overlap with a predicted ORF (ORF112ng) from N. gonorrhoeae:
• 
• The complete length ORF112ng nucleotide sequence <SEQ ID 891> is:

• 1	ATGAACCTGA TTTCACGTTA CATCATCCGC CAAATGGCGG
  	TTATGGCGGT
  51	TTACGCGCTC CTTGCCTTCC TCGCTTTGTA CAGCTTTTTT
  	GAAATCCTGT
  101	ACGAAACCGG CAACCTCGGC AAAGGCAGTT ACGGCATATG
  	GGAAATGCTG
  151	GGCTACACCG CCCTCAAAAT GCCCGCCCGC GCCTACGAAC
  	TCATGCCCCT
  201	CGCCGTCCTC ATCGGCGGAC TGGCCTCTCT CAGCCAGCTT
  	GCCGCCGGCA
  251	GCGAACTGGC CGTCATCAAA GCCAGCGGCA TGAGCACCAA
  	AAAGCTGCTG
  301	TTGATTCTGT CTCAGTTCGG TTTTATTTTT GCTATTGCCG
  	CCGTCGCGCT
  351	CGGCGAATGG GTTGCGCCCA CGCTGAGCCA AAAAGCCGAA
  	AACATCAAag
  401	cCGCCGCCAt taacggCAAA ATCAGCAccg gcAATACCGG
  	CCTTTggcTG
  451	AAAGAAAAAa ccAGCATTAT CAATGTGcGc GGAATGTTGC
  	CCGACCATAC
  501	GCTTTTGGGC ATCAAAATTT GGGCGCGCAA CGATAAAAAC
  	GAATTGGCAG
  551	AGGCAGTGGA AGCCGATTCC GCCGTTTTGA ACAGCGACGG
  	CAGCTGGCAG
  601	TTGAAAAACA TCCGCCGCAG CATCATGGGT ACAGACAAAA
  	TCGAAACATC
  651	cgCCGCCGCC GAAGAAACTT gGCCGATTGC CGTCAGACGC
  	AACCTGATGG
  701	ACGTATTGCT CGTCAAGCCC GACCAAATGT CCGTCGGCGA
  	GCTGACCACC
  751	TACATCCGCC ACCTCCAAAA CAACAGCCAA AACACCCAAA
  	TCTACGCCAT
  801	CGCATGGTGG CGTAAACTCG TTTACCCCGT CGCCGCATGG
  	GTCATGGCGC
  851	TCGTTGCCTT CGCCTTTACG CCGCAAACCA CGCGCCACGG
  	CAATATGGGC
  901	TTAAAACTCT TCGGCGGCAT CTGTCTCGGA TTGCTGTTCC
  	ACCTTGCCGG
  951	CAGGCTCTTC GGGTTTACCA GCCAACTCTA CGGCACCCCA
  	CCCTTCCTCG
  1001	CCGGCGCACT GCCTACCATA GCCTTCGCCT TGCTCGCTGT
  	TTGGCTGATA
  1051	CGCAAACAGG AAAAACGTTG A

• This encodes a protein having amino acid sequence <SEQ ID 892>:

• 1	MNLISRYIIR QMAVMAVYAL LAFLALYSFF EILYETGNLG
  	KGSYGIWEML
  51	GYTALKMPAR AYELMPLAVL IGGLASLSQL  AAGSELAVIK
  	ASGMSTKKLL
  101	LILSQFGFIF AIAAVALGEW VAPTLSQKAE NIKAAAINGK
  	ISTGNTGLWL
  151	KEKTSIINVR GMLPDHTLLG IKIWARNDKN ELAEAVEADS
  	AVLNSDGSWQ
  201	LKNIRRSIMG TDKIETSAAA EETWPIAVRR NLMDVLLVKP
  	DQMSVGELTT
  251	YIRHLQNNSQ NTQIYAIAWW RKLVYPVAAW VMALVAFAFT
  	PQTTRHGNMG
  301	LKLFGGICLG LLFHLAGRLF GFTSQLYGTP PFLAGALPTI
  	AFALLAVWLI

• • 351 RKQEKR*
• ORF112ng and ORF112-1 show 94.2% identity in 326 aa overlap:
• 
• This analysis suggests that these proteins from N. meningitidis and N. gonorrhoeae, and their epitopes, could be useful antigens for vaccines or diagnostics, or for raising antibodies.
• It will be appreciated that the invention has been described by means of example only, and that modifications may be made whilst remaining within the spirit and scope of the invention.

• TABLE I
  PCR primers

  ORF	Primer	Sequence	Restriction sites
  ORF 1	Forward	CGCGGATCCGCTAGC-GGACACACTTATTTCGG	BamHI-NheI
  	Reverse	CCCGCTCGAG-CCAGCGGTAGCCTAATT	XhoI
  ORF 2	Forward	GCGGATCCCATATG-TTTGATTTCGGTTTGGG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-GACGGCATAACGGCG	XhoI
  ORF 2-1	Forward	GCGGATCCCATATG-TTTGATTTCGGTTTGGG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TGATTTACGGACGCGCA	XhoI
  ORF 4	Forward	GCGGATCCCATATG-TGCGGAGGTCAAAAAGAC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TTTGGCTGCGCCTTC	XhoI
  ORF 5	Forward	GGAATTCCATATGGCCATGG-TGGAAGGCGCACAACC	NdeI-NcoI
  	Forward	CGGGATCC-ATGGAAGGCGCACAAC	BamHI
  	Reverse	CCCGCTCGAG-GACTGTGCAAAAACGG	XhoI
  ORF 6	Forward	CGCGGATCCCATATG-ACCCGTCAATCTCTGCA	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TGCGCCGAACACTTTC	XhoI
  ORF 7	Forward	CGCGGATCCGCTAGC-GCGCTGCTTTTTGTTCC	BamHI-NheI
  	Reverse	CCCGCTCGAG-TTTCAAAATATATTTGCGGA	XhoI
  ORF 8	Forward	GCGGATCCCATATG-GCTCAACTGCTTCGTAC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-AGCAGGCTTTGGCGC	XhoI
  ORF 9	Forward	CGCGGATCCCATATG-CCGAAGGAAGTCGGAAA	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TTTCCGAGGTTTTCGGG	XhoI
  ORF 10	Forward	GCGGATCCCATATG-GACACAAAAGAAATCCTC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TAATGGGAAACCTTGTTTT	XhoI
  ORF 11	Forward	GCGGATCCCATATG-GCGGTCAACCTCTACG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-GGAAACGACTTCGCC	XhoI
  ORF 13	Forward	CGCGGATCCCATATG-GCTCTGCTTTCCGCGC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-AGGGTGTGTGATAATAAG	XhoI
  ORF 15	Forward	GGAATTCCATATGGCCATGG-GCGGGACACTGACAG	NdeI-NcoI
  	Forward	CGGGATCC-TGCGGGACACTGACAGG	BamHI
  	Reverse	CCCGCTCGAG-AGGTTGGCCTTGTCTATG	XhoI
  ORF 17	Forward	GGAATTCCATATGGCCATGG-TTGCCGGCCTGTTCG	NdeI-NcoI
  	Forward	CGGGATCC-ATTGCCGGCCTGTTCG	BamHI
  	Reverse	CCCGCTCGAG-AAGCAGGTTGTACAGC	XhoI
  ORF 18	Forward	GCGGATCCCATATG-ATTTTGCTGCATTTGGAT	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TCTTCCAATTTCTGAAAGC	XhoI
  ORF 19	Forward	GGAATTCCATATGGCCATGG -TCGCCAGTGTTTTTACC	NdeI-NcoI
  	Forward	CGGGATCC-TTCGCCAGTGTTTTTACCG	BamHI
  	Reverse	CCCGCTCGAG-GGTGTTTTTGAAGCTGCC	XhoI
  ORF 20	Forward	GGAATTCCATATGGCCATGG -TCGGCGCGGGTATG	NdeI-NcoI
  	Forward	CGGGATCC-TTCGGCGCGGGTATG	BamHI
  	Reverse	CCCGCTCGAG-CGGCGAGCGAGAGCA	XhoI
  ORF 22	Forward	GGAATTCCATATGGCCATGG-TGATTAAAATCAAAAAAGGTCT	NdeI-NcoI
  	Forward	CGGGATCC-ATGATTAAAATCAAAAAAGGTCTAAACC	BamHI
  	Reverse	CCCGCTCGAG-ATTATGATAGCGGCCC	XhoI
  ORF 23	Forward	CGCGGATCCCATATG-GATGTTTCTGTTTCAGAC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TTTAAACCGATAGGTAAACG	XhoI
  ORF 24	Forward	GGAATTCCATATGGCCATGG- TGATGCCGGAAATGGTG	NdeI-NcoI
  	Forward	CGGGATCC-ATGATGCCGGAAATGGTG	BamHI
  	Reverse	CCCGCTCGAG-TGTCAGCGTGGCGCA	XhoI
  ORF 25	Forward	GCGGATCCCATATG-TATCGCAAACTGATTGC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-ATCGATGGAATAGCCG	XhoI
  ORF 26	Forward	GCGGATCCCATATG -CAGCTGATCGACTATTC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-GACATCGGCGCGTTTT	XhoI
  ORF 27	Forward	GGAATTCCATATGGCCATGG-AGACCTATTCTGTTTA	NdeI-NcoI
  	Forward	CGGGATCC- CAGACCTATTCTGTTTATTTTAATC	BamHI
  	Reverse	CCCGCTCGAG-GGGTTCGATTAAATAACCAT	XhoI
  ORF 28	Forward	GGAATTCCATATGGCCATGG-ACGGCTGTACGTTGATGT	NdeI-NcoI
  	Forward	CGGGATCC-AACGGCTGTACGTTGATG	BamHI
  	Reverse	CCCGCTCGAG-TTTGTCAGAGGAATTCGCG	XhoI
  ORF 29	Forward	GCGGATCCCATATG -AACGGTTTGGATGCCCG	BamHI-NdeI
  	Forward	CGCGGATCCGCTAGC-AACGGTTTGGATGCCCG	BamHI-NheI
  	Reverse	CCCGCTCGAG-TTTGTCTAAGTTCCTGATATG	XhoI
  ORF 32	Forward	CGCGGATCCCATATG-AATACTCCTCCTTTTG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-GCGTATTTTTTGATGCTTTG	XhoI
  ORF 33	Forward	GCGGATCCCATATG -ATTGATAGGGATCGTATG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TTGATCTTTCAAACGGCC	XhoI
  ORF 35	Forward	GCGGATCCCATATG-TTCAGAGCTCAGCTT	BamHI-NdeI
  	Forward	CGCGGATCCGCTAGC-TTCAGAGCTCAGCTT	BamHI-NheI
  	Reverse	CCCGCTCGAG-AAACAGCCATTTGAGCGA	XhoI
  ORF 37	Forward	GCGGATCCCATATG-GATGACGTATCGGATTTT	BamHI-NdeI
  	Reverse	CCCGCTCGAG-ATAGCCCGCTTTCAGG	XhoI
  ORF 58	Forward	CGCGGATCCGCTAGC-TCCGAACGCGAGTGGAT	BamHI-NheI
  	Reverse	CCCGCTCGAG-AGCATTGTCCAAGGGGAC	XhoI
  ORF 65	Forward	GGAATTCCATATGGCCATGG -TGCTGTATCTGAATCAAG	NdeI-NcoI
  	Forward	CGGGATCC-TTGCTGTATCTGAATCAAGG	BamHI
  	Reverse	CCCGCTCGAG-CCGCATCGGCAGACA	XhoI
  ORF 66	Forward	GCGGATCCCATATG-TACGCATTTACCGCCG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TGGATTTTGCAGAGATGG	XhoI
  ORF 72	Forward	CGCGGATCCCATATG- AATGCAGTAAAAATATCTGA	BamHI-NdeI
  	Reverse	CCCGCTCGAG-GCCTGAGACCTTTGCAA	XhoI
  ORF 73	Forward	GCGGATCCCATATG-AGATTTTTCGGTATCGG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TTCATCTTTTTCATGTTCG	XhoI
  ORF 75	Forward	GCGGATCCCATATG- TCTGTCTTTCAAACGGC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TTTGTTTTTGCAAGACAG	XhoI
  ORF 76	Forward	GATCAGCTAGCCATATG-AAACAGAAAAAAACCGC	NheI-NdeI
  	Reverse	CGGGATCC-TTACGGTTTGACACCGTT	BamHI
  ORF 79	Forward	CGCGGATCCCATATG-GTTTCCGCCGCCG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-GTGCTGATGCGCTTCG	XhoI
  ORF 83	Forward	GCGGATCCCATATG-AAAACCCTGCTGCTGC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-GCCGCCTTTGCGGC	XhoI
  ORF 84	Forward	GCGGATCCCATATG-GCAGAGATCTGTTTG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-GTTTGCCGATCCGACCA	XhoI
  ORF 85	Forward	CGCGGATCCCATATG- GCGGTTTGGGGCGGA	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TCGGCGCGGCGGGC	XhoI
  ORF 89	Forward	GGAATTCCATATGGCCATGG-CCATACCTTCTTATCA	NdeI-NcoI
  	Forward	CGGGATCC-GCCATACCTTCTTATCAGAG	BamHI
  	Reverse	CCCGCTCGAG-TTTTTTGCGATTAGAAAAAGC	XhoI
  ORF 97	Forward	GCGGATCCCATATG-CATCCTGCCAGCGAAC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TTCGCCTACGGTTTTTTG	XhoI
  ORF 98	Forward	GCGGATCCCATATG-ACGGTAACTGCGG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TTGTTGTTCGGGCAAATC	XhoI
  ORF 100	Forward	GCGGATCCCATATG-TCGGGCATTTACACCG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-ACGGGTTTCGGCGGAA	XhoI
  ORF 101	Forward	GCGGATCCCATATG-ATTTATCAAAGAAACCTC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TTTTCCGCCTTTCAATGT	XhoI
  ORF 102	Forward	GCGGATCCCATATG-GCAGGGCTGTTTTACC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-AAACGGTTTGAACACGAC	XhoI
  ORF 103	Forward	GCGGATCCCATATG-AACCACGACATCAC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-CAGCCACAGGACGGC	XhoI
  ORF 104	Forward	GCGGATCCCATATG-ACGTGGGGAACGC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-GCGGCGTTTGAACGGC	XhoI
  ORF 105	Forward	GCGGATCCCATATG-ACCAAATTTCAAACCCCTC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TAAACGAATGCCGTCCAG	XhoI
  ORF 106	Forward	GCGGATCCCATATG-AGGATAACCGACGGCG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TTTGTTCCCGATGATGTT	XhoI
  ORF 109	Forward	GCGGATCCCATATG-GAAGATTTATATATAATACTCG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-ATCAGCTTCGAACCGAAG	XhoI
  ORF 110	Forward	AAAGAATTC-ATGAGTAAATCCCGTAGATCTCCC	EcoRI
  	Reverse	AAACTGCAG-GGAAAACCACATCCGCACTCTGCC	PstI
  ORF 111	Forward	AAAGAATTC-GCACCGCAAAAGGCAAAAACCGCA	EcoRI
  	Reverse	AAACTGCAG-TCTGCGCGTTTTCGGGCAGGGTGG	PstI
  ORF 113	Forward	AAAGAATTC-ATGAACAAAACCCTCTATCGTGTGATTTTCAACCG	EcoRI
  	Reverse	AAACTGCAG-TTACGAATGCCTGCTTGCTCGACCGTACTG	PstI
  ORF 115	Forward	AAAGAATTC-TTGCTTGTGCAAACAGAAAAAGACGG	EcoRI
  	Reverse	AAAAAAGTCGAC-CTATTTTTTAGGGGCTTTTGCTTGTTTGAAAAGCCTGCC	SalI
  ORF 119	Forward	AAAGAATTC-TACAACATGTATCAGGAAAACCAATACCG	EcoRI
  	Reverse	AAACTGCAG-TTATGAAAACAGGCGCAGGGCGGTTTTGCC	PstI
  ORF 120	Forward	AAAGAATTC-GCAAGGCTACCCCAATCCGCCGTG	EcoRI
  	Reverse	AAACTGCAG-CGGTTTGGCTGCCTGGCCGTTGAT	PstI
  ORF 121	Forward	AAAGAATTC-GCCTTGGTCTGGCTGGTTTTCGC	EcoRI
  	Reverse	AAACTGCAG-TCATCCGCCACCCCACCTCGGCCATCCATC	PstI
  ORF 122	Forward	AAAAAAGTCGAC-ATGTCTTACCGCGCAAGCAGTTCTCC	SalI
  	Reverse	AAACTGCAG-TCAGGAACACAAACGATGACGAATATCCGTATC	PstI
  ORF 125	Forward	AAAGAATTC-GCGCTGTTTTTTGCGGCGGCGTAT	EcoRI
  	Reverse	AAACTGCAG-CGCCGTTTCAAGACGAAAAAGTCG	PstI
  ORF 126	Forward	AAAGAATTC-GCGGAAACGGTCGAAG	EcoRI
  	Reverse	AAACTGCAG-TTAATCTTGTCTTCCGATATAC	PstI
  ORF 127	Forward	AAAGAATTC-ATGACTGATAATCGGGGGTTTACG	EcoRI
  	Reverse	AAAAAAGTCGAC-CTTAAGTAACTTGCAGTCCTTATC	SalI
  ORF 128	Forward	AAAGAATTC-ATGCAAGCTGTCCGCTACAGGCC	EcoRI
  	Reverse	AAACTGCAG-CTATTGCAATGCGCCGCCGCGGGAATGTTTGAGCAGGCG	PstI
  ORF 129	Forward	AAAGAATTC-ATGGATTTTCGTTTTGACATTATTTACGAATACCG	EcoRI
  	Reverse	AAACTGCAG-TTATTTTTTGATGAAATTTTGGGGCGG	PstI
  ORF 130	Forward	AAAGAATTC-GCAGTACTTGCCATTCTCGGTGCG	EcoRI
  	Reverse	AAACTGCAG-CTCCGGATCGTCTGTAAACGCATT	PstI
  ORF 131	Forward	GCGGATCCCATATG-GAAATTCGGGCAATAAAAT	BamHI-NdeI
  	Reverse	CCCGCTCGAG-CCAGCGGACGCGTTC	XhoI
  ORF 132	Forward	GCGGATCCCATATG-AAAGAAGCGGGGTTTG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-CCAATCTGCCAGCCGT	XhoI
  ORF 133	Forward	CGCGGATCCCATATG-GAAGATGCAGGGCGCG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-AAACTTGTAGCTCATCGT	XhoI
  ORF 134	Forward	GCGGATCCCATATG-TCTGTGCAAGCAGTATTG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-ATCCTGTGCCAATGCG	XhoI
  ORF 135	Forward	GCGGATCCCATATG-CCGTCTGAAAAAGCTTT	BamHI-NdeI
  	Reverse	CCCGCTCGAG-AAATACCGCTGAGGATG	XhoI
  ORF 136	Forward	CGCGGATCCGCTAGC-ATGAAGCGGCGTATAGCC	BamHI-NheI
  	Reverse	CCCGCTCGAG-TTCCGAATATTTGGAACTTTT	XhoI
  ORF 137	Forward	CGCGGATCCCATATG-GGCACGGCGGGAAATA	BamHI-NdeI
  	Reverse	CCCGCTCGAG-ATAACGGTATGCCGCC	XhoI
  ORF 138	Forward	GCGGATCCCATATG-TTTCGTTTACAATTCAGGC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-CGGCGTTTTATAGCGG	XhoI
  ORF 139	Forward	GCGGATCCCATATG-GCTTTTTTGGCGGTAATG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TAACGTTTCCGTGCGTTT	XhoI
  ORF 140	Forward	GCGGATCCCATATG-TTGCCCACAGGCAGC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-GACGATGGCAAACAGC	XhoI
  ORF 141	Forward	GCGGATCCCATATG-CCGTCTGAAGCAGTCT	BamHI-NdeI
  	Reverse	CCCGCTCGAG-ATCTGTTGTTTTTAAAATATT	XhoI
  ORF 142	Forward	GCGGATCCCATATG-GATAATTCTGGTAGTGAAG	BamHI-NdeI
  	Reverse	CCCGCTCGAG-AAACGTATAGCCTACCT	XhoI
  ORF 143	Forward	GCGGATCCCATATG-GATACCGCTTTGAACCT	BamHI-NdeI
  	Reverse	CCCGCTCGAG-AATGGCTTCCGCAATATG	XhoI
  ORF 144	Forward	GCGGATCCCATATG-ACCTTTTTACAACGTTTGC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-AGATTGTTGTTGTTTTTTCG	XhoI
  ORF 147	Forward	GCGGATCCCATATG-TCTGTCTTTCAAACGGC	BamHI-NdeI
  	Reverse	CCCGCTCGAG-TTTGTTTTTGCAAGACAG	XhoI
  NB:
  restriction sites are underlined
  for ORFs 110-130, where the ORF itself carries an EcoRI site (eg. ORF 122), a SalI site was used in the forward primer instead. Similarly, where the ORF carries a PstI site (eg. ORFs 115 and 127), a SalI site was used in the reverse primer.

• TABLE II
  Summary of cloning, expression and purification

  		PCR/	His-fusion	GST-fusion
  	ORF	cloning	expression	expression	Purification
  	orf 1	+	+	+	His-fusion
  	orf 2	+	+	+	GST-fusion
  	orf 2.1	+	n.d.	+	GST-fusion
  	orf 4	+	+	+	His-fusion
  	orf 5	+	n.d.	+	GST-fusion
  	orf 6	+	+	+	GST-fusion
  	orf 7	+	+	+	GST-fusion
  	orf
  8	+	n.d.	n.d.
  	orf 9	+	+	+	GST-fusion
  	orf
  10	+	n.d.	n.d.
  	orf 11	+	n.d.	n.d.
  	orf 13	+	n.d.	+	GST-fusion
  	orf 15	+	+	+	GST-fusion
  	orf 17	+	n.d.	n.d.
  	orf 18	+	n.d.	n.d.
  	orf 19	+	n.d.	n.d.
  	orf 20	+	n.d.	n.d.
  	orf 22	+	+	+	GST-fusion
  	orf 23	+	+	+	His-fusion
  	orf 24	+	n.d.	n.d.
  	orf 25	+	+	+	His-fusion
  	orf 26	+	n.d.	n.d.
  	orf 27	+	+	+	GST-fusion
  	orf 28	+	+	+	GST-fusion
  	orf 29	+	n.d.	n.d.
  	orf 32	+	+	+	His-fusion
  	orf 33	+	n.d.	n.d.
  	orf 35	+	n.d.	n.d.
  	orf 37	+	+	+	GST-fusion
  	orf 58	+	n.d.	n.d.
  	orf 65	+	n.d.	n.d.
  	orf 66	+	n.d.	n.d.
  	orf 72	+	+	n.d.	His-fusion
  	orf 73	+	n.d.	+	n.d.
  	orf 75	+	n.d.	n.d.
  	orf 76	+	+	n.d.	His-fusion
  	orf 79	+	+	n.d.	His-fusion
  	orf 83	+	n.d.	+	n.d.
  	orf 84	+	n.d.	n.d.
  	orf 85	+	n.d.	+	GST-fusion
  	orf 89	+	n.d.	+	GST-fusion
  	orf 97	+	+	+	GST-fusion
  	orf 98	+	n.d.	n.d.
  	orf 100	+	n.d.	n.d.
  	orf 101	+	n.d.	n.d.
  	orf 102	+	n.d.	n.d.
  	orf 103	+	n.d.	n.d.
  	orf 104	+	n.d.	n.d.
  	orf 105	+	n.d.	n.d.
  	orf 106	+	+	+	His-fusion
  	orf 109	+	n.d.	n.d.
  	orf 110	+	n.d.	n.d.
  	orf 111	+	+	n.d.	His-fusion
  	orf 113	+	+	n.d.	His-fusion
  	orf 115	n.d.	n.d.	n.d.
  	orf 119	+	+	n.d.	His-fusion
  	orf
  120	+	+	n.d.	His-fusion
  	orf 121	+	n.d.	n.d.
  	orf 122	+	+	n.d.	His-fusion
  	orf
  125	+	+	n.d.	His-fusion
  	orf 126	+	+	n.d.	His-fusion
  	orf 127	+	+	n.d.	His-fusion
  	orf 128	+	n.d.	n.d.
  	orf 129	+	+	n.d.	His-fusion
  	orf
  130	+	n.d.	n.d.
  	orf 131	+	+	+	n.d.
  	orf 132	+	+	+	His-fusion
  	orf 133	+	n.d.	+	GST-fusion
  	orf 134	+	n.d.	n.d.
  	orf 135	+	n.d.	n.d.
  	orf 136	+	n.d.	n.d.
  	orf 137	+	n.d.	+	GST-fusion
  	orf 138	+	n.d.	+	GST-fusion
  	orf 139	+	n.d.	n.d.
  	orf 140	+	n.d.	n.d.
  	orf 141	+	n.d.	n.d.
  	orf 142	+	n.d.	n.d.
  	orf 143	+	n.d.	n.d.
  	orf 144	+	n.d.	+	n.d.
  	orf 147	+	n.d.	n.d.

Claims (7)

1: An isolated protein comprising:

(a) the amino acid sequence of SEQ ID NO: 654; or

(b) an amino acid sequence having 80% or greater sequence identity to the amino acid sequence of SEQ ID NO: 654; or

(b) a fragment of SEQ ID NO: 654 of at least 10 contiguous amino acids in length.

2: The isolated protein of claim 1 comprising (b).

3: The isolated protein of claim 2, wherein the amino acid sequence has 90% or greater sequence identity to the amino acid sequence of SEQ ID NO: 654.

4: The isolated protein of claim 2, wherein the amino acid sequence has 95% or greater sequence identity to the amino acid sequence of SEQ ID NO: 654.

5: The isolated protein of claim 1 comprising (c).

6: A composition comprising the protein of any one of claims 1-5 and an adjuvant.

7: The composition of claim 6 further comprising a pharmaceutically acceptable carrier.

Images