NZ533932A - MS11 gene product encoding for a phosphoglycerate kinase, and a purine nucleoside phosphatase and glucose-6-phosphate isomerase proteins from Group B streprococcus and their use in treating bacterial infection - Google Patents

Abstract

A peptide encoded by the nucleotide sequence of SEQ ID NO: 3, or a homologue having greater than 70% sequence similarity is described. The peptide sequence can be the amino acid sequence of SEQ ID NO: 4 and can be obtained from Group B streptococcus. The peptide or polynucleotide can be used in a medicament for the treatment of bacterial infections, preferably when the infection is a focal or urinary tract infection. Also described are antibodies and hosts transformed with the peptide.

Description

New Zealand Paient Spedficaiion for Paient Number 533932 53 39 3 2 NEW ZEALAND PATENTS ACT, 1953 No: Date: Divided out of New Zealand Specification No. 525538 dated 22 December 1999 COMPLETE SPECIFICATION OUTER SURFACE PROTEINS, THEIR GENES, AND THEIR USE We, MICROSCIENCE LIMITED, 545 Eskdale Road, Winnersh Triangle, Wokingham, Berkshire RG41 5TU, United Kingdom, do hereby declare the invention for which we pray that a patent may be granted to us, and the method by which it is to be performed, to be particularly described in and by the following statement: INTELLECTUAL PROPERTY OFFICE OF N.Z. - 6 jul m receiver (followed by page 1 a) la OUTER SURFACE PROTEINS. THEIR GENES, AND THEIR USE Fjeld of the Invention This invention relates to the identification of outer 5 surface proteins, their genes, and their use. More particularly, it relates to their use in therapy, for immunisation and in screening for drugs.

Background to the Invention Group B Streptococcus (GBS), also known as 10 Streptococcus agalactiae, is the causative agent of various conditions. In particular, GBS causes: Early onset neonatal infection.

This infection usually begins in utero and causes severe septicaemia and pneumonia in infants, which is 15 lethal if untreated and even with treatment is associated with a 10-20% mortality rate.

Late onset neonatal infection.

This infection occurs in the period shortly after birth until about 3 months of age. It causes a 20 septicaemia, which is complicated by meningitis in 90% of cases. Other focal infections also occur including osteomyelitis, septic arthritis, abscesses and endopthalmitis.

Mult infections.

These appear to be increasingly common and occur most frequently in women who have just delivered a baby, the elderly and the immunocompromised. They are characterised by septicaemia and focal infections including osteomyelitis, septic arthritis, abscesses and 30 endopthalmitis.

Urinary tract infections.

GBS is a cause of urinary tract infections and in pregnancy accounts for about 10% of all infections. Veterinary infections.

GBS causes chronic mastitis in cows. This, in turn, leads to reduced milk production and is therefore of considerable economic importance. / l- / / GBS infections can be treated with antibiotics. However, immunisation is preferable. It is therefore desirable to develop an immunogen that could be used in a therapeutically-effective vaccine.

Summary of the Invention The present invention is based on the identification of a series of genes in GBS, and also related organisms, the products of which may be associated with the outer surface of the organism, and may therefore be useful as 10 targets for immunotherapy.

According to one aspect of the invention, a peptide^ encoded by an operon comprising a nucleotide sequence of SEQ ID NO: 3 including the gene identified herein as MS 11 obtainable from Group B Streptococcus, or a homologue having greater than 70% sequence similarity thereto, or functional fragment thereof is provided. 15 Such a peptide is suitable for therapeutic use, e.g. when isolated.

The term "functional fragments" is used herein to define a part of the gene or peptide which retains the activity of the whole gene or peptide. For example, a 20 functional fragment of the peptide may be used as an antigenic determinant, useful in a vaccine or in the production of antibodies.

A gene fragment may be used to encode the active peptide. Alternatively, the gene fragment may have utility 25 in gene therapy, targetting the wild-type gene in vivo to exert a therapeutic effect.

A peptide according to the present invention may comprise the amino acid sequence identified herein as SEQ ID NO, 4, or a functional fragment thereof.

Because of the extracellular or cell surface location, the peptides of the present invention may be suitable candidates for the production of therapeutically-effective vaccines against GBS. The term "therapeutically-effective" 35 is intended to include the prophylactic effect of vaccines. For example, a vaccine may comprise a peptide according to p INTELLECTUAL PROPERTY OFFICE 1 OF NX - 1 mar 2005 ei\/cn the invention, or the means for its expression, for the treatment of infection.

This vaccine may be administered to females either prior to or during pregnancy to protect mother and neonate against infection by GBS.

According to another aspect of the invention, the peptides or genes may be used for screening potential antimicrobial drugs or for the detection of virulence.

A further aspect of this invention is the use of any of the products identified herein, for the treatment or prevention of a condition associated with infection by a Group B Streptococcal strain.

Although the protein has been described for use in the treatment of patients, veterinary uses of the products of the invention are also considered to be within the scope of the present invention. In particular, the peptides or the vaccines may be used in the treatment of chronic mastitis, especially in cows.

Description of the Invention The present invention is described with reference to Group B Streptococcal strain M732. However, all the GBS strains and many other bacterial strains are likely to include related peptides or proteins having amino acid sequence homology with the peptide of M732. Organisms likely to contain the peptides include, but are not limited to, S. pneumoniae, S. pyogenes, S. suis, S. milleri, Group C and Group G streptococci and Enterococci. Vaccines to each of these may be developed in the same way as described for GBS.

Preferably, the peptides that may be useful for the production of vaccines have greater than 70% sequence similarity with the peptides identified herein. More preferably, the peptides have greater than 75% sequence similarity. Most preferably, the peptides have greater than 80% sequence similarity, e.g. 95% similarity.

Having characterised a gene according to the invention, it is possible to use the gene sequence to establish homologies in other microorganisms. In this way it is possible to determine whether other microorganisms have similar outer surface products. Sequence homologies may be established by searching in existing databases, e.g. EMBL or Genbank.

Peptides or proteins according to the invention may be purified and isolated by methods known in the art. In particular, having identified the gene sequence, it will be possible to use recombinant techniques to express the genes in a suitable host. Active fragments and homologues can be identified and may be useful in therapy. For example, the peptides or their active fragments may be used as antigenic determinants in a vaccine, to elicit an immune response. They may also be used in the preparation of antibodies, for passive immunisation, or diagnostic applications. Suitable antibodies include monoclonal antibodies, or fragments thereof, including single chain fv fragments. Methods for the preparation of antibodies will be apparent to those skilled in the art.

The preparation of vaccines based on attenuated microorganisms is known to those skilled in the art. Vaccine compositions can be formulated with suitable carriers or adjuvants, e.g. alum, as necessary or desired, and used in therapy, to provide effective immunisation against Group B Streptococci or other related microorganisms. The preparation of vaccine formulations will be apparent to the skilled person.

More generally, and as is well known to those skilled in the art, a suitable amount of an active component of the invention can be selected, for therapeutic use, as can suitable carriers or excipients, and routes of administration. These factors will be chosen or determined according to known criteria such as the nature/severity of the condition to be treated, the type or health of the subject etc.

The products of the present invention were identified as follows: Todd-Hewitt broth was inoculated with GBS and allowed to grow overnight at 37°C. The cells were harvested by centrifugation and washed with Phosphate Buffered Saline (PBS). The cells were resuspended in an osmotic buffer 5 (20% (w/v) Sucrose, 20mM Tris-HCl pH 7.0, lOmM MgCl2) containing protease inhibitors (1 mM PMSF, 10 fM Iodoeacetic Acid, 10 mM 1,10-Phenanthroline, l /xM Pepstatin A) and Mutanolysin at a final concentration of 4 Units per microlitre. This was incubated (shaking) at 37°C for 2 10 hours.

Cells and debris were removed first by high speed centrifugation, then ultra-centrifugation for 1 hour. The resultant supernatant containing cell wall proteins was concentrated under pressure using an ultrafiltration device 15 (10,000 molecular weight cut-off).

The sample was dialysed against ultra high quality water and lyophilised. After resuspension in loading buffer, the proteins were separated by preparative 2-Dimensional-Gel Electrophoresis. Following electrophoresis 20 an individual spot was chosen for study. The spot was subjected to in-gel tryptic digestion. The resulting peptides were extracted from the gel and purified using microbore RP-HPLC. Fractions were collected every 45 seconds and a portion of these consistent with the regions 25 of OV absorbance were analysed by Delayed Extraction-Matrix Assisted Laser Desorption-Time of Flight Mass Spectrometry (DE-MALDI-TOF-MS). Peptides not observed in a blank preparation were then subjected to sequencing using Nanospray-MS/MS Using this peptide sequence information, degenerate oligonucleotides were designed to be used in a polymerase chain reaction (PCR) to amplify the DNA segment lying between the peptide sequences identified.

PCR amplification resulted in the production of 35 several polynucleotide fragments, each of which was cloned into the pCR 2.1-T0P0 vector (Invitrogen BV, Netherlands) according to manufacturers protocol. 6 The DNA fragment in each plasmid was identified by sequencing and then used to obtain the full-length gene sequence, as follows.

Using the identified DNA fragment, oligonucleotide 5 primers were designed for genomic DNA sequencing. These primers were designed so as to sequence in an "outward* direction from the obtained sequence. Once read, the sequence obtained was checked to see if the 5' and 3 * termini of the gene had. been reached. The presence of 10 these features was identified by checking against homologous sequences, and for the 5' end the presence of an AUG start codon (or accepted alternative) preceded by a Shine-Dalgarno consensus sequence, and for the 3' end, the presence of a translation termination (Stop) codon. 15 Upon identification of the full-length gene, primers were designed for amplification of full-length product from GBS genomic DNA. Primers used included restriction enzyme recognition sites (Ncol at the 5*end and Eco0109I at the 3' end) to allow subsequent cloning of the product into the 20 Lactococcal expression system used.

PCR was carried out using the primers, and the products cloned into a pCR 2.1 cloning vector (In Vitrogen). Following confirmation of the presence of the cloned fragment, the DNA was'excised using the restriction 25 enzymes Ncol and Eco0109I.

The vector into which this fragment was inserted was a modified version of pNZ8048 (Kuipers, O. P. et al. (1998) J. Biotech 64: 15-21). This vector, harbouring a lactococcal origin of replication, a chloramphenicol 30 resistance marker, an inducible nisin promoter and a multicloning site was altered by the replacement of the multicloning site with two 10X His tags, flanked on the 5-most end with an Ncol site, split in the middle with a multicloning site (including an Eco0l091 site), and a Stop 35 (termination) codon at the 3*end of the His tags.

The gene of interest was inserted so that a 10X His tag was in the 3* position relative to the coding region. 7 Following transformation of the recombinant plasmid into L.lactis (strain NZ9000 - Kuipers, 0. P. et al. (1998) supra), a 400 ml liquid culture was set up and translation of the protein was induced by the addition of nisin to the 5 culture. After a 2 hour incubation, the cells were harvested and lysed by bead beating. The resultant lysate was cleared by centrifugation, then passed over a metal affinity (Talon, Clonetech) column. The column was washed repeatedly before bound proteins were eluted with 10 Imidazole.

To identify fractions containing the His-tagged recombinant protein, an aliquot from each fraction was analysed by SDS-PAGE, Western blotted and probed with anti-His antibodies. -15 The recombinant protein obtained was then used to immunise New Zealand white rabbits, with pre-immune sera being harvested prior to immunisation. Following a boost, the rabbits were sacrificed and sera collected. This sera was used in Western blots, ELISA and animal protection 20 models.

Using the sera obtained from the animal studies, immunosorption studies were carried out.

Group B Streptococcus was grown in 20ml Todd Hewitt broth (THB) for 8 hours, harvested and resuspended in 5ml 25 PBS. 50/zl aliquots of this were used to coat wells in a 96 well plate (Nunc Immuno-Sorb) . This was left at 4°C overnight to allow for absorbance of the bacteria onto the plate. Plates were washed twice with PBS, then blocked with 3%BSA in PBS for lhr at 37°C. Plates were again 30 washed. Serial 10 fold dilutions of the sera were made in PBS and 50/tl of these dilutions were added to the wells of the plate, in duplicate. The plate was covered and incubated for 1 hr at 37°C. The plate was washed, then 50jul anti-rabbit alkaline phosphatase conjugated secondary 35 antibody at a concentration of 1:5000 was added to each well. Following incubation at 37°C for an hour, the plate was washed again. 50/zl substrate (PNPP) was added to each 8 well, and the reaction allowed to proceed for 30min before the absorbance was read at 405 ni.

Animal protection studies were also carried out to test the effectiveness of protection on the immunised rabbits.

GBS M732 was grown up in THB until mid-log phase was reached - approximately 5 hours. Cells were counted in a counting chamber, and bacteria were diluted to give a concentration of 2xl07 bacteria per ml in pre-immune or test sera. 50/Ltl of this was injected via the intraperitoneal route into 0-1 day old mice. The mice were observed for survival over 48 hours.

The following Examples illustrate the invention. Example 1 A first plasmid was termed MS4. The cloned DNA fragment was sequenced and the nucleotide and deduced amino acid sequence (SEQ ID NO. 1 and 2) was used to search protein databases.

Homologues to the GBS MS4 gene product can be identified in Clostridium perfingens, Haemophilus influenzae, Neisseria flavescens and Thermatoga maritima. In all cases the homologues are the genes for Ornithine Carbamoyltransferase (OCT). In eukaryotic systems this enzyme catalyses the second step in the Urea cycle, the conversion of ornithine to citruliine, a reaction requiring carbomyl phosphate. In prokaryotes, ODC is one of the three enzymes involved in Arginine Deaminase activity - a system which protects bacteria from acid damage. In particular, ODC is responsible for the conversion of citruliine to ornithine and carbamoyl phosphate (the opposite role to that in eukaryotes) (Casiano-Colon, A and Marquis, R. E. 1988. Appl. Environ. Microbiol. 54: 1318-1324, Cunin, R. et al. 1986. Microbiol. Rev. 50: 314-352).

Animal protection studies were carried out as described above. The results are as follows: 9 Treatment # pups # pups surviving at time (hrs) 24 48 PBS 6 0 Pre-immune 41 18 1 Test 41 33 14 0 25 Example 2 A second plasmid was termed MS11. The nucleotide and deduced amino acid sequence (SEQ ID NOS. 3 and 4) were used to search protein databases.

Homologues to the GBS MS11 gene product can be identified in Lactobacillus delbrueckii, Thermotoga maritima, Clostridium acetobulylicum, Bacillus megaterium, Triticum aestivium and Synechocystis PCC6803.

In all cases the homologues are the genes for the protein Phosphoglycerate Kinase (PGK) . PGK is a major enzyme in the glycolytic pathway, being involved in the conversion of Glyceraldehyde-3-phosphate to Phosphoenolpyruvate. In particular, it is involved in the catalysis of the reaction between Glycerate-l,3-diphosphate and 3 —Phospho—Gly cerate, releasing a phosphate in the forward reaction.

Example 3 A third plasmid was termed pMS16. The 5' and 3* cloned DNA fragments were sequenced and the nucleotide and deduced amino acid sequences for each are shown as SEQ ID NOS. 5 and 6 for the 5' fragment and SEQ ID NOS. 7 and 8 for the 3' fragment.

Homologues to the GBS MS 16 gene product can be identified in Bacillus stearothermophilus, Bacillus subtilis and Mycoplasma genitalium.

In all cases the homologues are the genes for the protein Glucose-6-Phosphate Isomerase (GPI).

The enzyme Glucose-6-Phospate Isomerase catalyses the reaction between Glucose-6-phosphate and Fructose-6-Phosphate in both glycolysis (G6P to F6P) and gluconeogenesis (F6P to G6P) . Mutations in the gpi gene have been shown to confer purine analogue sensitivity to organisms.

Example 4 A fourth plasmid was termed pMS14. The cloned DNA fragment was sequenced and the nucleotide and deduced amino acid sequence (SEQ ID NOS. 9 and 10) was used to search protein databases.

Homologues to the. GBS MS14 gene product can be identified in Bacillus subtilis, Bacillus stearothermophilus, Mus musculus, Bos taurus and Zea mays. In all cases the homologues are the genes for the protein Purine Nucleoside Phosphatase (PNP). The function of this enzyme is to cleave the nucleosides guanosine or inosine to their respective basis and sugar-l-phosphate molecules in the presence of orthophosphate. ffrapple 5 A fifth plasmid was termed pMSlO. The cloned DNA fragment was sequenced. The nucleotide and deduced amino acid sequence (SEQ ID NOS. 11 and 12) was used to search protein databases.

Homologues to the GBS MS10 gene product can be identified in Streptococcus mutans, Nicotiana plumb, Pisum sativum and Zea mays. In all cases the homologues are the genes for the protein Nonphosphorylating, NADP-Dependent Glyceraldehyde-3-Phosphate Dehydrogenase (NPGAP-3-DH). NPGAP-3-DH has been reported as being an important means of generating NADPH for biosynthetic reactions in S. mutans (as opposed to NAD-specific GAP—3-DH which satisfies the requirements of the glycolytic pathway) (Boyd, D.A., Cvitkovitch, D. G. and Hamilton, I. R 1995 J. Bacterid. 177: 2622-2727).

SEQUENCE LISTING <110> Microscience Limited <120> OUTER SURFACE PROTEINS, THEIR GENES, AND THEIR USE <130> REP05969WO <140> <141> <160> 12 <170> Patentln Ver. 2.1 <210> 1 <211> 1014 <212> DNA <213> group B streptococcus <220> <221> CDS <222> (1)..(1014) <400> 1 atg aca caa gta ttt caa gga cgt agt ttc tta gca gaa aaa gat ttt 48 Met Thr Gin Val Phe Gin Gly Arg Ser Phe Leu Ala Glu Lys Asp Phe 15 10 15 tct cgt gag gaa ttt gaa tat ctt att gat ttt tea get cat tta aaa 96 Ser Arg Glu Glu Phe Glu Tyr Leu lie Asp Phe Ser Ala His Leu Lys 20 25 30 gac ctt aaa aaa cgt ggt gtt cct cat cat tat ctt gaa ggt aaa aat 144 Asp Leu Lys Lys Arg Gly Val Pro His His Tyr Leu Glu Gly Lys Asn 35 40 45 att get ctc tta ttt gaa aaa aca tct act cgt act cgc gca gcc ttt 192 lie Ala Leu Leu Phe Glu Lys Thr Ser Thr Arg Thr Arg Ala Ma Phe 50 55 60 aca act gca gca att gac eta ggc get cat ccg gaa tac ctt ggt gca 240 Thr Thr Ala Ala lie Asp Leu Gly Ala His Pro Glu Tyr Leu Gly Ala 65 70 75 80 aat gat att caa ctt ggt aaa aaa gaa tea aca gaa gat act get aag 288 Asn Asp lie Gin Leu Gly Lys Lys Glu Ser Thr Glu Asp Thr Ala Lys 85 90 95 11 gtt tta gga cgt atg ttt gat ggt att gaa ttc cgt ggt ttt age caa 336 Val Leu Gly Arg Met Phe Asp Gly lie Glu Phe Arg Gly Phe Ser Gin 100 105 110 aga atg gtt gaa gag ctt get gaa ttt tct gga gta cct gtc tgg aat 384 Arg Met Val Glu Glu Leu Ala Glu Phe Ser Gly Val Pro Val Trp Asn 115 120 125 ggt tta aca gat gaa tgg cat cca aca caa atg eta get gac tac ctt 432 Gly Leu Thr Asp Glu Trp His Pro Thr Gin Met Leu Ala Asp Tyr Leu 130 135 140 act ate aaa gaa aac ttc ggt aaa ctt gaa ggt att act ctt gtt tac 480 Thr lie Lys Glu Asn Phe Gly Lys Leu Glu Gly lie Thr Leu Val Tyr 145 150 155 160 tgt ggt gac gga cgt aac aat gtt gcc aac teg ctt tta gtg get ggg 528 Cys Gly Asp Gly Arg Asn Asn Val Ala Asn Ser Leu Leu Val Ala Gly 165 170 175 act ttg atg ggg gtc aat gta cac ate ttt tct cca aaa gaa ctt tty 576 Thr Leu Met Gly Val Asn Val His lie Phe Ser Pro Lys Glu Leu Phe 180 185 190 ccw get gaa gag att gtt aaa ttg get gaa gga tat gcc aaa gaa tct 624 Xaa Ala Glu Glu lie Val Lys Leu Ala Glu Gly Tyr Ala Lys Glu Ser 19S 200 205 ggg get cac gtt etc gtt act gat aat gta gac gaa get gta aag gga 672 Gly Ala His Val Leu Val Thr Asp Asn Val Asp Glu Ala Val Lys Gly 210 215 220 gca gac gtc ttt tac act gat gtc tgg gta teg atg gga gaa gaa gat 720 Ala Asp Val Phe Tyr Thr Asp Val Trp Val Ser Met Gly Glu Glu Asp 225 230 235 240 aag ttc aaa gaa cgc gtt gaa ctt ctt caa cca tat caa gta aac atg 768 Lys Phe Lys Glu Arg Val Glu Leu Leu Gin Pro Tyr Gin Val Asn Met 245 250 255 gaa ctg att aaa aaa get aat aat gat aat ctt ate ttc tta cac tgc 816 Glu Leu lie Lys Lys Ala Asn Asn Asp Asn Leu lie Phe Leu His Cys 260 265 270 tta cct gca ttc cat gat aca aat ace gtt tat ggc aaa gac gtc get 864 Leu Pro Ala Phe His Asp Thr Asn Thr Val Tyr Gly Lys Asp Val Ala 275 280 285 12 gaa aaa ttt ggg gtc aag gaa atg gaa gtt act gat gaa gtc ttc cgt 912 Glu Lys Phe Gly Val Lys Glu Met Glu Val Thr Asp Glu Val Phe Arg 290 295 300 age aaa tat get cgt cat ttc gac caa get gaa aat cgt atg cac act 960 Ser Lys Tyr Ala Arg His Phe Asp Gin Ala Glu Asn Arg Met His Thr 305 310 315 320 att aaa get gta atg get gca acc ctt gga aat ctt ttc att cca aaa 1008 He Lys Ala Val Met Ala Ala Thr Leu Gly Asn Leu Phe He Pro Lys 325 330 335 gtt taa 1014 Val <210> 2 <211> 337 <212> PRT <213> group B streptococcus <400> 2 Met Thr Gin Val Phe Gin Gly Arg Ser Phe Leu Ala Glu Lys Asp Phe 15 10 15 Ser Arg Glu Glu Phe Glu Tyr Leu lie Asp Phe Ser Ala His Leu Lys 20 25 30 Asp Leu Lys Lys Arg Gly Val Pro His His Tyr Leu Glu Gly Lys Asn 35 40 45 lie Ala Leu Leu Phe Glu Lys Thr Ser Thr Arg Thr Arg Ala Ala Phe 50 55 60 Thr Thr Ala Ala lie Asp Leu Gly Ala His Pro Glu Tyr Leu Gly Ala 65 70 75 80 Asn Asp He Gin Leu Gly Lys Lys Glu Ser Thr Glu Asp Thr Ala Lys 85 90 95 Val Leu Gly Arg Met Phe Asp Gly lie Glu Phe Arg Gly Phe Ser Gin 100 105 110 Arg Met Val Glu Glu Leu Ala Glu Phe Ser Gly Val Pro Val Trp Asn 115 120 125 Gly Leu Thr Asp Glu Trp His Pro Thr Gin Met Leu Ala Asp Tyr Leu 13 130 135 140 Thr lie Lys Glu Asn Phe Gly Lys Leu Glu Gly lie Thr Leu Val Tyr 145 150 155 160 Cys Gly Asp Gly Arg Asn Asn Val Ala Asn Ser Leu Leu Val Ala Gly 165 170 175 Thr Leu Met Gly Val Asn Val His lie Phe Ser Pro Lys Glu Leu Phe 180 185 190 Xaa Ala Glu Glu lie Val Lys Leu Ala Glu Gly Tyr Ala Lys Glu Ser 195 200 205 Gly Ala His Val Leu Val Thr Asp Asn Val Asp Glu Ala Val Lys Gly 210 215 220 Ala Asp Val Phe Tyr Thr Asp Val Trp Val Ser Met Gly Glu Glu Asp 225 230 235 240 Lys Phe Lys Glu Arg Val Glu Leu Leu Gin Pro Tyr Gin Val Asn Met 245 250 255 Glu Leu lie Lys Lys Ala Asn Asn Asp Asn Leu lie Phe Leu His Cys 260 265 270 Leu Pro Ala Phe His Asp Thr Asn Thr Val Tyr Gly Lys Asp Val Ala 275 280 285 Glu Lys Phe Gly Val Lys Glu Met Glu Val Thr Asp Glu Val Phe Arg 290 295 300 Ser Lys Tyr Ala Arg His Phe Asp Gin Ala Glu Asn Arg Met His Thr 305 310 315 320 lie Lys Ala Val Met Ala Ala Thr Leu Gly Asn Leu Phe lie Pro Lys 325 330 335 Val <210> 3 <211> 1197 <212> DNA <213> group B streptococcus <220> 14 <221> CDS <222> (1)..(1197) <400> 3 atg get aaa ttg act gtt aaa gac gtt gat ttg aag gta aaa aaa gtc 48 Met Ala Lys Leu Thr Val Lys Asp Val Asp Leu Lys Val Lys Lys Val 15 10 15 etc gtt cgt gtt gac ttt aat gtg cct ttg aaa gac ggc gtt ate act 96 Leu Val Arg Val Asp Phe Asn Val Pro Leu Lys Asp Gly Val lie Thr 20 25 30 aac gac aac cgt ate act gcg get ctt cca aca ate aag tat ate ate 144 Asn Asp Asn Arg lie Thr Ala Ala Leu Pro Thr lie Lys Tyr lie lie 35 40 45 gaa caa ggt ggt cgt get ate ctc ttc tct cac ctt gga cgt gtt aaa 192 Glu Gin Gly Gly Arg Ala lie Leu Phe Ser His Leu Gly Arg Val Lys 50 55 60 gaa gaa get gac aaa gaa gga aaa tea ctt gca ccg gta get get gat 240 Glu Glu Ala Asp Lys Glu Gly Lys Ser Leu Ala Pro Val Ala Ala Asp 65 70 75 80 tta get get aaa ctt ggt caa gat gtt Leu Ala Ala Lys Leu Gly Gin Asp Val gta ttc cca ggt gtt act cgt 288 Val Phe Pro Gly Val Thr Arg 90 - 95 ggt gca aaa tta gaa gaa gca ate aat get ttg gaa gat gga caa gtt 336 Gly Ala Lys Leu Glu Glu Ala lie Asn Ala Leu Glu Asp Gly Gin Val 100 105 110 ctt ttg gtt gaa aac act cgt ttt gaa gat gtt gac ggt aag aaa gaa 384 Leu Leu Val Glu Asn Thr Arg Phe Glu Asp Val Asp Gly Lys Lys Glu 115 120 125 tct aag aat gac gaa gaa ctt ggt aaa tac tgg get tea ctt gga gat 432 Ser Lys Asn Asp Glu Glu Leu Gly Lys Tyr Trp Ala Ser Leu Gly Asp 130 135 140 gga ate ttc gtt aac gat gca ttt ggt aca gca cac cgt get cat gca 480 Gly lie Phe Val Asn Asp Ala Phe Gly Thr Ala His Arg Ala His Ala 145 150 155 160 tea aac gta ggt att tea gca aac gtt gaa aaa get gta get ggt ttc 528 Ser Asn Val Gly lie Ser Ala Asn Val Glu Lys Ala Val Ala Gly Phe 165 170 175 ctt ctt gaa aac gaa att get tac ate caa gaa gca gtt gaa act cca Leu Leu Glu Asn Glu lie Ala Tyr lie Gin Glu Ala Val Glu Thr Pro 180 185 190 576 gaa egc cca ttc gta get att ctt ggt gge tea aaa gtt tct gat aag 624 Glu Arg Pro Phe Val Ala lie Leu Gly Gly Ser Lys Val Ser Asp Lys 195 200 205 att ggt gtt ate gaa aac ctt ctt gaa aaa get gat aaa gtt ctt ate 672 lie Gly Val lie Glu Asn Leu Leu Glu Lys Ala Asp Lys Val Leu lie 210 215 220 ggt ggt ggt atg act tac aca ttc tac- aaa get caa ggt ate gaa ate 720 Gly Gly Gly Met Thr Tyr Thr Phe Tyr Lys Ala Gin Gly lie Glu He 225 230 235 240 ggt aac tea ctt gta gaa gaa gac aaa ttg gat gtt get aaa gac ctc 768 Gly Asn Ser Leu Val Glu Glu Asp Lys Leu Asp Val Ala Lys Asp Leu 245 250 255 ctt gaa aaa tea aac ggt aaa ttg ate ttg cca gtt gac tea aaa gaa 816 Leu Glu Lys Ser Asn Gly Lys Leu lie Leu Pro Val Asp Ser Lys Glu 260 265 270 gca aac gca ttt get ggt tat act gaa gtt egc gac act gaa ggt gaa 864 Ala Asn Ala Phe Ala Gly Tyr Thr Glu Val Arg Asp Thr Glu Gly Glu 275 280 285 gca gtt tea gaa ggg ttc ctt ggt ctt gac ate ggt cct aaa tea ate 912 Ala Val Ser Glu Gly Phe Leu Gly Leu Asp lie Gly Pro Lys Ser lie 290 295 300 get aaa ttt gat gaa gca ctt act ggt get aaa aca gtt gta tgg aac 960 Ala Lys Phe Asp Glu Ala Leu Thr Gly Ala Lys Thr Val Val Trp Asn 305 310 315 320 gga cct atg ggt gtc ttt gaa aac cct gac ttc caa get ggt aca ate 1008 Gly Pro Met Gly Val Phe Glu Asn Pro Asp Phe Gin Ala Gly Thr lie 325 330 335 ggt gta atg gac get ate gtt aaa caa cca gge gtt aaa tea ate ate 1056 Gly Val Met Asp Ala lie Val Lys Gin Pro Gly Val Lys Ser lie lie 340 345 350 ggt ggt ggt gat tea gca gca get get ate aac ctt ggt cgt get gac 1104 Gly Gly Gly Asp Ser Ala Ala Ala Ala lie Asn Leu Gly Arg Ala Asp 355 360 365 16 aaa ttc tea tgg ate tct act ggt ggt gga gca age atg gaa ttg ctc 1152 Lys Phe Ser Trp lie Ser Thr Gly Gly Gly Ala Ser Met Glu Leu Leu 370 375 380 gaa ggt aaa gta tta cca ggt ttg gca gca ttg act gaa aaa taa 1197 Glu Gly Lys Val Leu Pro Gly Leu Ala Ala Leu Thr Glu Lys 385 390 395 <210> 4 <211> 398 <212> PRT <213> group B streptococcus <400> 4 Met Ala Lys Leu Thr Val Lys Asp Val Asp Leu Lys Val Lys Lys Val 15 10 15 Leu Val Arg Val Asp Phe Asn Val Pro Leu Lys Asp Gly Val lie Thr 20 25 30 Asn Asp Asn Arg lie Thr Ala Ala Leu Pro Thr lie Lys Tyr lie lie 35 40 45 Glu Gin Gly Gly Arg Ala lie Leu Phe Ser His Leu Gly Arg Val Lys 50 55 60 Glu Glu Ala Asp Lys Glu Gly Lys Ser Leu Ala Pro Val Ala Ala Asp 65 70 75 80 Leu Ala Ala Lys Leu Gly Gin Asp Val Val Phe Pro Gly Val Thr Arg 85 90 95 Gly Ala Lys Leu Glu Glu Ala lie Asn Ala Leu Glu Asp Gly Gin Val 100 105 110 Leu Leu Val Glu Asn Thr Arg Phe Glu Asp Val Asp Gly Lys Lys Glu 115 120 125 Ser Lys Asn Asp Glu Glu Leu Gly Lys Tyr Trp Ala Ser Leu Gly Asp 130 135 140 Gly lie Phe Val Asn Asp Ala Phe Gly Thr Ala His Arg Ala His Ala 145 150 155 160 Ser Asn Val Gly lie Ser Ala Asn Val Glu Lys Ala Val Ala Gly Phe 165 170 175 17 Leu Leu Glu Asn Glu lie Ala Tyr lie Gin Glu Ala Val Glu Thr Pro 180 185 190 Glu Arg Pro Phe Val Ala lie Leu Gly Gly Ser Lys Val Ser Asp Lys 195 200 205 lie Gly Val lie Glu Asn Leu Leu Glu Lys Ala Asp Lys Val Leu lie 210 215 220 Gly Gly Gly Met Thr Tyr Thr Phe Tyr Lys Ala Gin Gly lie Glu lie 225 230 235 240 Gly Asn Ser Leu Val Glu Glu Asp Lys Leu Asp Val Ala Lys Asp Leu 245 250 255 Leu Glu Lys Ser Asn Gly Lys Leu 260 Ala Asn Ala Phe Ala Gly Tyr Thr 275 280 lie Leu Pro Val Asp Ser Lys Glu 265 270 Glu Val Arg Asp Thr Glu Gly Glu 285 Ala Val Ser Glu Gly Phe Leu Gly Leu Asp lie Gly Pro Lys Ser lie 290 295 300 Ala Lys Phe Asp Glu Ala Leu Thr Gly Ala Lys Thr Val Val Trp Asn 305 310 315 320 Gly Pro Met Gly Val Phe Glu Asn Pro Asp Phe Gin Ala Gly Thr lie 325 330 335 Gly Val Met Asp Ala lie Val Lys Gin Pro Gly Val Lys Ser lie lie 340 345 350 Gly Gly Gly Asp Ser Ala Ala Ala Ala lie Asn Leu Gly Arg Ala Asp 355 360 365 Lys Phe Ser Trp lie Ser Thr Gly Gly Gly Ala Ser Met Glu Leu Leu 370 375 380 Glu Gly Lys Val Leu Pro Gly Leu Ala Ala Leu Thr Glu Lys 385 390 395 <210> 5 <211> 516 <212> DNA <213> group B streptococcus 18 <220> <221> CDS <222> (1)..(516) <400> 5 atg aca cat att aca ttt gac tta ttc aaa gtc ttg ggt caa ttt gta 48 Met Thr His lie Thr Phe Asp Leu Phe Lys Val Leu Gly Gin Phe Val 15 10 15 gge gaa cac gag tta gac tac eta cca cca caa gta agt gca gca gat 96 Gly Glu His Glu Leu Asp Tyr Leu Pro Pro Gin Val Ser Ala Ala Asp 20 25 30 get ttc ctt cgt caa ggg act ggt cct gga tea gat ttt ctc gga tgg 144 Ala Phe Leu Arg Gin Gly Thr Gly Pro Gly Ser Asp Phe Leu Gly Trp 35 40 45 atg gaa cct cca gaa aac tat gac aaa gaa gaa ttt tct cgc att caa 192 Met Glu Pro Pro Glu Asn Tyr Asp Lys Glu Glu Phe Ser Arg lie Gin 50 55 60 aaa gcc get gaa aag att aaa tea gat age gaa gta ctc gtg gtt att 240 Lys Ala Ala Glu Lys lie Lys Ser Asp Ser Glu Val Leu Val Val lie 65 70 75 80 ggt att ggt ggt teg tac ctt ggt gca aaa gca gca att gac ttt ttg 288 Gly lie Gly Gly Ser Tyr Leu Gly Ala Lys Ala Ala lie Asp Phe Leu 85 90 95 aat aat cat ttt get aat ttg caa acc gca gaa gaa cgt aaa gcg cct 336 Asn Asn His Phe Ala Asn Leu Gin Thr Ala Glu Glu Arg Lys Ala Pro 100 105 110 cag att ctt tat get gga aat tct att tea tct act tac ctt gcc gat 384 Gin lie Leu Tyr Ala Gly Asn Ser lie Ser Ser Thr Tyr Leu Ala Asp 115 120 125 tta gtt gaa tac gtc caa gat aaa gaa ttc tea gta aat gtc att tea 432 Leu Val Glu Tyr Val Gin Asp Lys Glu Phe Ser Val Asn Val lie Ser 130 135 140 aaa tea ggt aca aca act gaa cca gcg att get ttc cgt gta ttt aaa 480 Lys Ser Gly Thr Thr Thr Glu Pro Ala lie Ala Phe Arg Val Phe Lys 145 150 155 160 gaa ctt eta gtt aaa aag tac egg tea aga aga age 516 Glu Leu Leu Val Lys Lys Tyr Arg Ser Arg Arg Ser 19 165 170 <210> 6 <211> 172 <212> PRT <213> group B streptococcus <400> 6 Met Thr His lie Thr Phe Asp Leu Phe Lys Val Leu Gly Gin Phe Val 15 10 15 Gly Glu His Glu Leu Asp Tyr Leu Pro Pro Gin Val Ser Ala Ala Asp 20 25 30 Ala Phe Leu Arg Gin Gly Thr Gly Pro Gly Ser Asp Phe Leu Gly Trp 35 40 45 Met. Glu Pro Pro Glu Asn Tyr Asp Lys Glu Glu Phe Ser Arg He Gin 50 55 60 Lys Ala Ala Glu Lys lie Lys Ser Asp Ser Glu Val Leu Val Val lie 65 70 75 80 Gly lie Gly Gly Ser Tyr Leu Gly Ala Lys Ala Ala He Asp Phe Leu 85 90 95 Asn Asn His Phe Ala Asn Leu Gin Thr Ala Glu Glu Arg Lys Ala Pro 100 105 110 Gin lie Leu Tyr Ala Gly Asn Ser lie Ser Ser Thr Tyr Leu Ala Asp 115 120 125 Leu Val Glu Tyr Val Gin Asp Lys Glu Phe Ser Val Asn Val lie Ser 130 135 140 Lys Ser Gly Thr Thr Thr Glu Pro Ala lie Ala Phe Arg Val Phe Lys 145 150 155 160 Glu Leu Leu Val Lys Lys Tyr Arg Ser Arg Arg ser 165 170 <210> 7 <211> 318 <212> DNA <213> group B streptococcus <220> <221> CDS <222> (1)..(318) <400> 7 att aac cga aga ttt aga tgg tct tgg tta tct tea aga aaa gat gta 48 lie Asn Arg Arg Phe Arg Trp Ser Trp Leu Ser Ser Arg Lys Asp Val 10 15 gat ttt gtt aat aaa aaa gca aca gat ggt gtg ctt ctt get cat aca 96 Asp Phe Val Asn Lys Lys Ala Thr Asp Gly Val Leu Leu Ala His Thr 20 25 30 gat ggt ggg gtt cca aat atg ttt gta acg ctt cct aca caa gac get 144 Asp Gly Gly Val Pro Asn Met Phe Val Thr Leu Pro Thr Gin Asp Ala 35 40 45 tac act ctt ggt tac act att tac ttc ttt gag tta gca att gge ctt 192 Tyr Thr Leu Gly Tyr Thr lie Tyr Phe Phe Glu Leu Ala lie Gly Leu 50 55 60 tea ggt tat ctt aac tea gta aat cca ttt gat caa ceg ggg gta gaa 240 Ser Gly Tyr Leu Asn Ser Val Asn Pro Phe Asp Gin Pro Gly Val Glu 65 70 75 80 gca tat aaa cgt aat atg ttc gca ttt ggt aaa cct gga ttc gaa gag 288 Ala Tyr Lys Arg Asn Met Phe Ala Phe Gly Lys Pro Gly Phe Glu Glu 85 90 95 ctt age get gaa ttg aat gca cgt ctt taa 318 Leu Ser Ala Glu Leu Asn Ala Arg Leu 100 105 <210> 8 <211> 105 <212> PRT <213> group B streptococcus <400> 8 lie Asn Arg Arg Phe Arg Trp Ser Trp Leu Ser Ser Arg Lys Asp Val 1 5 10 15 Asp Phe Val Asn Lys Lys Ala Thr Asp Gly Val Leu Leu Ala His Thr 20 25 30 Asp Gly Gly Val Pro Asn Met Phe Val Thr Leu Pro Thr Gin Asp Ala 21 40 45 Tyr Thr Leu Gly Tyr Thr lie Tyr Phe Phe Glu Leu Ala lie Gly Leu 50 55 60 Ser Gly Tyr Leu Asn Ser Val Asn Pro Phe Asp Gin Pro Gly Val Glu 65 70 75 80 Ala Tyr Lys Arg Asn Met Phe Ala Phe Gly Lys Pro Gly Phe Glu Glu 85 90 95 Leu Ser Ala Glu Leu Asn Ala Arg Leu 100 105 <210> 9 <211> 804 <212> DNA <213> group B streptococcus <220> <221> CDS <222> (1)..(804) <400> 9 atg aca tta tta gaa aaa att aat gag act aga gac ttt ttg caa gca 48 Met Thr Leu Leu Glu Lys lie Asn Glu Thr Arg Asp Phe Leu Gin Ala 1 5 10 15 aaa gge gtc aca gca cca gaa ttt ggy ctt att tta gge tct ggt tta 96 Lys Gly Val Thr Ala Pro Glu Phe Xaa Leu lie Leu Gly Ser Gly Leu 20 25 30 gga gaa ttg get gaa gaa ate gaa aat cct att gtt gtg gat tat gca 144 Gly Glu Leu Ala Glu Glu lie Glu Asn Pro lie Val Val Asp Tyr Ala 35 40 45 gac ate ccm aat tgg gga cag tea aca gta gtt ggt cat get gga aaa 192 Asp He Xaa Asn Trp Gly Gin Ser Thr Val Val Gly His Ala Gly Lys 50 55 60 ttt agt gta tgg gat tta tea gge cgt aag gta tta gcg ctt caa ggt 240 Phe Ser Val Trp Asp Leu Ser Gly Arg Lys Val Leu Ala Leu Gin Gly 65 70 75 80 cgt ttt cat ttt tay gaa ggw aat aca atg gaa gtc gtt act ttc cca 288 Arg Phe His Phe Tyr Glu Xaa Asn Thr Met Glu Val Val Thr Phe Pro 22 85 90 95 gta cgt ate atg aga gca ttg get tgc cac agt gtg ctt gtg act aat 336 Val Arg lie Met Arg Ala Leu Ala Cys His Ser Val Leu Val Thr Asn 100 105 110 gca gcg ggt ggg att gga tac gga cca gga act tta atg ctg ate aaa 384 Ala Ala Gly Gly lie Gly Tyr Gly Pro Gly Thr Leu Met Leu lie Lys 115 120 125 gac cac ate aat atg att ggg act aac cct ctc ata ggt gag aac ctt 432 Asp His lie Asn Met lie Gly Thr Asn Pro Leu lie Gly Glu Asn Leu 130 135 140 gaa. gaa ttt gga cca cgt ttc cca gac atg teg gat get tay aca gca 480 Glu Glu Phe Gly Pro Arg Phe Pro Asp Met Ser Asp Ala Tyr Thr Ala 145 150 155 160 aca tat cga caa aaa get cac caa att get gaa aac gat ate aaa ctc 528 Thr Tyr Arg Gin Lys Ala His Gin lie Ala Glu Asn Asp lie Lys Leu 165 170 175 gaa gaa ggt gtg tac ttg ggt gta tea gga ccc act tat gaa aca cct 576 Glu Glu Gly Val Tyr Leu Gly Val Ser Gly Pro Thr Tyr Glu Thr Pro 180 185 190 gca gaa att cgt gca ttc caa aca atg gge gca caa gcg gta ggt atg 624 Ala Glu lie Arg Ala Phe Gin Thr Met Gly Ala Gin Ala Val Gly Met 195 200 205 tec acg gtt cca gag gtg ate gtt gca get cac tea ggg ctt aaa gtg 672 Ser Thr Val Pro Glu Val lie Val Ala Ala His Ser Gly Leu Lys Val 210 215 220 tta gga att tea gca att act aac ctt gcc get gge ttc caa tea gag 720 Leu Gly lie Ser Ala lie Thr Asn Leu Ala Ala Gly Phe Gin Ser Glu 225 230 235 240 ctc aat cat gag gag gtc gtt gaa gtt act cag cgt att aaa gaa gat 768 Leu Asn His Glu Glu Val Val Glu Val Thr Gin Arg lie Lys Glu Asp 245 250 255 ttc aag gga tta ggt aaa tea tta gtt Phe Lys Gly Leu Gly Lys Ser Leu Val 260 265 get gaa ctc 804 Ala Glu Leu <210> 10 23 <Z11> 268 <212> PRT <213> group B streptococcus <400> 10 Met Thr Leu Leu Glu Lys lie Asn Glu Thr Arg Asp Phe Leu Gin Ala 15 10 15 Lys Gly Val Thr Ala Pro Glu Phe Xaa Leu lie Leu Gly Ser Gly Leu 20 25 30 Gly Glu Leu Ala Glu Glu lie Glu Asn Pro lie Val Val Asp Tyr Ala 35 40 45 Asp lie Xaa Asn Trp Gly Gin Ser Thr Val Val Gly His Ala Gly Lys 50 55 60 Phe Ser Val Trp Asp Leu Ser Gly Arg Lys Val Leu Ala Leu Gin Gly 65 70 75 80 Arg Phe His Phe Tyr Glu Xaa Asn Thr Met Glu Val Val Thr Phe Pro 85 90 95 Val Arg lie Met Arg Ala Leu Ala Cys His Ser Val Leu Val Thr Asn 100 105 110 Ala Ala Gly Gly lie Gly Tyr Gly Pro Gly Thr Leu Met Leu lie Lys 115 120 125 Asp His lie Asn Met lie Gly Thr Asn Pro Leu lie Gly Glu Asn Leu 130 135 140 Glu Glu Phe Gly Pro Arg Phe Pro Asp Met Ser Asp Ala Tyr Thr Ala 145 150 155 160 Thr Tyr Arg Gin Lys Ala His Gin He Ala Glu Asn Asp lie Lys Leu 165 170 175 Glu Glu Gly Val Tyr Leu Gly Val Ser Gly Pro Thr Tyr Glu Thr Pro 180 185 190 Ala Glu lie Arg Ala Phe Gin Thr Met Gly Ala Gin Ala Val Gly Met 195 200 205 Ser Thr Val Pro Glu Val lie Val Ala Ala His Ser Gly Leu Lys Val 210 215 220 Leu Gly He Ser Ala lie Thr Asn Leu Ala Ala Gly Phe Gin Ser Glu 24 225 230 235 240 Leu Asn His Glu Glu Val Val Glu Val Thr Gin Arg lie Lys Glu Asp 245 250 255 Phe Lys Gly Leu Gly Lys Ser Leu Val Ala Glu Leu 260 265 <210> 11 <211> 1428 <212> DNA <213> group B streptococcus <220> <221> CDS <222> (1) .. (1428) <400> 11 ttg aca aaa gaa tat caa aat tat gtc aat gge gaa tgg aaa tea tct 48 Leu Thr Lys Glu Tyr Gin Asn Tyr Val Asn Gly Glu Trp Lys Ser Ser 15 10 15 gtt aat cag att gag att ttg tea cca att gat gat tct tea ttg gga 96 Val Asn Gin lie Glu He Leu Ser Pro lie Asp Asp Ser Ser Leu Gly 20 25 30 ttc gtg cca gcg atg act cga gaa gaa gtt gat cat get atg aaa gcg 144 Phe Val Pro Ala Met Thr Arg Glu Glu Val Asp His Ala Met Lys Ala 35 40 45 ggt cgt gag get tta cca get tgg get get tta aca gta tat gaa cgt 192 Gly Arg Glu Ala Leu Pro Ala Trp Ala Ala Leu Thr Val Tyr Glu Arg 50 55 60 gca caa tac ctt cat aaa gcc gca gac att att gaa cgt gat aaa gaa 240 Ala Gin Tyr Leu His Lys Ala Ala Asp lie lie Glu Arg Asp Lys Glu 65 70 75 80 gaa att get act gtt tta gca aaa gaa att tct aaa get tac aat get 288 Glu lie Ala Thr Val Leu Ala Lys Glu lie Ser Lys Ala Tyr Asn Ala 85 90 95 tea gta act gag gtt gta agg aca get gat ctt att cgt tat gca gca 336 Ser Val Thr Glu Val Val Arg Thr Ala Asp Leu lie Arg Tyr Ala Ala 100 105 110 gaa gaa gga att cgt tta tea act tea get gac gaa ggt gga aaa atg 384 Glu Glu Gly lie Arg Leu Ser Thr Ser Ala Asp Glu Gly Gly Lys Met 115 120 125 gat get tea aca ggt cat aag ttg get gtt att cgt cgt caa cca gta 432 Asp Ala Ser Thr Gly His Lys Leu Ala Val lie Arg Arg Gin Pro Val 130 135 140 ggt ate gtt tta gca ate gca cct tat aat tac cct gtt aac ctc tea 480 Gly lie Val Leu Ala lie Ala Pro Tyr Asn Tyr Pro Val Asn Leu Ser 145 150 155 160 gga tea aaa att gcg cca get eta att ggt gga aac gtt gtg atg ttt 528 Gly Ser Lys lie Ala Pro Ala Leu lie Gly Gly Asn Val Val Met Phe 165 170 175 aaa cca cca aca caa ggt tea gtc tea gga ctt gtt tta gca aaa get 576 Lys Pro Pro Thr Gin Gly Ser Val Ser Gly Leu Val Leu Ala Lys Ala 180 185 190 ttt gca gaa gca ggt ctt cca gca ggt gtc ttt aat act att aca gga 624 Phe Ala Glu Ala Gly Leu Pro Ala Gly Val Phe Asn Thr lie Thr Gly 195 200 205 cgc ggt tct gag att gga gat tac att gtt gag cat gaa gaa gtt aat 672 Arg Gly Ser Glu lie Gly Asp Tyr lie Val Glu His Glu Glu Val Asn 210 215 220 ttt att aac ttt aca gga tea acg cca gtt gga caa cgt att ggt aag 720 Phe lie Asn Phe Thr Gly Ser Thr Pro Val Gly Gin Arg lie Gly Lys 225 230 235 240 ttg gca gga atg cgt cca att atg ctt gag ttg gge ggt aag gat gca 768 Leu Ala Gly Met Arg Pro lie Met Leu Glu Leu Gly Gly Lys Asp Ala 245 250 255 ggt ate gtc tta get gat get gac ctt gat aac get get aaa caa ate 816 Gly lie Val Leu Ala Asp Ala Asp Leu Asp Asn Ala Ala Lys Gin lie 260 265 270 gtt gca ggt get tat gat tac tct gga caa cgc tgt acg gca att aag 864 Val Ala Gly Ala Tyr Asp Tyr Ser Gly Gin Arg cys Thr Ala lie Lys 275 280 285 cgt gtg ctt gtc gtt gaa gaa gtt gew gat gaa ttg gca gaa aaa ata 912 Arg Val Leu Val Val Glu Glu Val Xaa Asp Glu Leu Ala Glu Lys lie 290 295 300 26 tct gaa aat gta gca aaa tta tea gta ggt gat cca ttt gat aat gca 960 Ser Glu Asn Val Ala Lys Leu Ser Val Gly Asp Pro Phe Asp Asn Ala 305 310 315 320 acg gtg aca ceg gtt att gat gat aat tea get gac ttt att gaa age 1008 Thr Val Thr Pro Val lie Asp Asp Asn Ser Ala Asp Phe lie Glu Ser 325 330 335 tta gta gta gat gca cgt caa aaa ggt gcg aaa gaa ttg aat gaa ttt 1056 Leu Val Val Asp Ala Arg Gin Lys Gly Ala Lys Glu Leu Asn Glu Phe 340 345 350 aaa cgt gat ggt cgt eta tta act cea gga ttg ttt gat cat gtt act 1104 Lys Arg Asp Gly Arg Leu Leu Thr Pro Gly Leu Phe Asp His Val Thr 355 360 365 tta gat atg aaa eta get tgg gaa gag cct ttt gga cca att ctc cca 1152 Leu Asp Met Lys Leu Ala Trp Glu Glu Pro Phe Gly Pro lie Leu Pro 370 375 380 att att cgt gtc aag gat gca gaa gaa get gtt get att gcc aac aaa 1200 lie lie Arg Val Lys Asp Ala Glu Glu Ala Val Ala lie Ala Asn Lys 385 390 395 400 tct gat ttt gga tta caa tea tea gtc ttt aca cgt gat ttc caa aaa 1248 Ser Asp Phe Gly Leu Gin Ser Ser Val Phe Thr Arg Asp Phe Gin Lys 405 410 415 gca ttt gat ata gca aat aaa ctt gaa gtt ggt aca gtt cac att aac 1296 Ala Phe Asp lie Ala Asn Lys Leu Glu Val Gly Thr Val His lie Asn 420 425 430 aat aag act gga cgt ggt ccw gat aat ttc cca ttc tta gga ctc aaa 1344 Asn Lys Thr Gly Arg Gly Xaa Asp Asn Phe Pro Phe Leu Gly Leu Lys 435 440 445 gga tct ggt gca ggt gtt caa ggt ate aga tat tea att gaa gca atg 1392 Gly Ser Gly Ala Gly Val Gin Gly lie Arg Tyr Ser lie Glu Ala Met 450 455 460 aca aat gta aaa teg att gtt etc gat atg aaa tag 1428 Thr Asn Val Lys Ser lie Val Leu Asp Met Lys 465 470 475 <210> 12 <211> 475 <212> PRT 27 <213> group B streptococcus <400> 12 Leu Thr Lys Glu Tyr Gin Asn Tyr Val Asn Gly Glu Trp Lys Ser Ser 1 5 10 15 Val Asn Gin lie Glu lie Leu Ser Pro lie Asp Asp Ser Ser Leu Gly 20 25 30 Phe Val Pro Ala Met Thr Arg Glu Glu Val Asp His Ala Met Lys Ala 35 40 45 Gly Arg Glu Ala Leu Pro Ala Trp Ala Ala Leu Thr Val Tyr Glu Arg 50 55 60 Ala Gin Tyr Leu His Lys Ala Ala Asp lie lie Glu Arg Asp Lys Glu 65 70 75 80 Glu lie Ala Thr Val Leu Ala Lys Glu lie Ser Lys Ala Tyr Asn Ala 85 90 95 Ser Val Thr Glu Val Val Arg Thr Ala Asp Leu lie Arg Tyr Ala Ala 100 105 110 Glu Glu Gly lie Arg Leu Ser Thr Ser Ala Asp Glu Gly Gly Lys Met 115 120 125 Asp Ala Ser Thr Gly His Lys Leu Ala Val lie Arg Arg Gin Pro Val 130 135 140 Gly lie Val Leu Ala lie Ala Pro Tyr Asn Tyr Pro Val Asn Leu Ser 145 150 155 160 Gly Ser Lys lie Ala Pro Ala Leu lie Gly Gly Asn Val Val Met Phe 165 170 17S Lys Pro Pro Thr Gin Gly Ser val Ser Gly Leu Val Leu Ala Lys Ala 180 185 190 Phe Ala Glu Ala Gly Leu Pro Ala Gly Val Phe Asn Thr lie Thr Gly 195 200 205 Arg Gly Ser Glu lie Gly Asp Tyr lie Val Glu His Glu Glu Val Asn 210 215 220 Phe lie Asn Phe Thr Gly Ser Thr Pro Val Gly Gin Arg lie Gly Lys 225 230 235 240 28

Claims (18)

Leu Ala Gly Met Arg Pro lie Met Leu Glu Leu Gly Gly Lys Asp Ala 245 250 255 Gly lie Val Leu Ala Asp Ala Asp Leu Asp Asn Ala Ala Lys Gin lie 260 265 270 Val Ala Gly Ala Tyr Asp Tyr Ser Gly Gin Arg Cys Thr Ala lie Lys 275 280 285 Arg Val Leu Val Val Glu Glu Val Xaa Asp Glu Leu Ala Glu Lys lie 290 295 300 Ser Glu Asn Val Ala Lys Leu Ser Val Gly Asp Pro Phe Asp Asn Ala 305 310 315 320 Thr Val Thr Pro Val lie Asp Asp Asn Ser Ala Asp Phe lie Glu Ser 325 330 335 Leu Val Val Asp Ala Arg Gin Lys Gly Ala Lys Glu Leu Asn Glu Phe 340 345 350 Lys Arg Asp Gly Arg Leu Leu Thr Pro Gly Leu Phe Asp His Val Thr 355 360 365 Leu Asp Met Lys Leu Ala Trp Glu Glu Pro Phe Gly Pro lie Leu Pro 370 375 380 He lie Arg Val Lys Asp Ala Glu Glu Ala Val Ala He Ala Asn Lys 385 390 395 400 Ser Asp Phe Gly Leu Gin Ser Ser Val Phe Thr Arg Asp Phe Gin Lys 405 410 415 Ala Phe Asp lie Ala Asn Lys Leu Glu Val Gly Thr Val His lie Asn 420 425 430 Asn Lys Thr Gly Arg Gly Xaa Asp Asn Phe Pro Phe Leu Gly Leu Lys 435 440 445 Gly Ser Gly Ala Gly Val Gin Gly lie Arg Tyr Ser lie Glu Ala Met 450 455 460 Thr Asn Val Lys Ser lie Val Leu Asp Met Lys 465 470 475 29 30 CLAIMS

1. A peptide encoded by an operon comprising the nucleotide sequence of SEQ ID NO: 3 obtainable from Group B streptococcus, or a homologue thereof having greater than 70% sequence similarity thereto or a functional fragment thereof.

2. A peptide according to claim 1, comprising the amino acid sequence identified herein as SEQ ID NO: 4.

3. A peptide according to claim 1 or claim 2, for therapeutic use.

4. A polynucleotide encoding a peptide according to claim 1 or claim 2, for therapeutic use.

5. A host transformed to express a peptide according to claim 1 or claim 2 with the proviso that said host is not present in a human body.

6. A vaccine comprising a peptide according to claim 1 or claim 2, or the means for its expression.

7. Use of a product according to any of claims 1 to 5, for screening potential drugs or for the detection of virulence.

8. Use of a product according to any of claims 1 to 5, for the manufacture of a medicament for use in the treatment or prevention of a condition associated with bacterial infection.

9. Use according to claim 8, wherein the infection is a Group B streptococcal infection.

10. Use according to claim 8 or claim 9, wherein the infection is a focal infection.

11. Use according to claim 8 or claim 9, wherein the infection is a urinary tract infection.

12. An antibody raised against a peptide according to claim 1 or claim 2.

13. A peptide as claimed in claim 1 substantially as herein described with reference to any example thereof.

14. A polynucleotide as claimed in claim 4 substantially as herein described with reference to any example thereof.

15. A host as claimed in claim 5 substantially as herein described with reference to any example thereof.

16. A vaccine as claimed in claim 6 substantially as herein described with reference to any example thereof. iNltOtCTUAL PKOPERFY OFFICE I OF HI. | - t mar 2085 j 338143_1.DOC 31

17. A use as claimed in claims 7 or 8 substantially as herein described with reference to any example thereof.

18. An antibody as claimed in claim 12 substantially as herein described with reference to any example thereof. INTELLECTUAL PROPERTY OFFICE OF N.Z. - 3 OCT 2005 RECEIVED 338143_1.DOC