SA113340369B1 - Neisseria meningitidis compositions and methods thereof - Google Patents

Abstract

In one aspect, the invention relates to an isolated polypeptide comprising an amino acid sequence that is at least 95% identical to SEQ ID NO: 71. In another aspect, the invention relates to an immunogenic composition including an isolated non-lipidated, non-pyruvylated ORF2086 polypeptide from Neisseria meningitidis serogroup B, and at least one conjugated capsular saccharide from a meningococcal serogroup. Figure 9A-9B.

Description

— \ — Neisseria meningitidis compositions and methods thereof FULL DESCRIPTION BACKGROUND The present invention relates to Neisseria meningitidis compositions and methods for their preparation. Neisseria meningitides is a Gram-negative encapsulated bacterium © that can cause sepsis (515m58); Meningitis and death (Sa (death) Classification of N. meningitides into about 1 serogroups (including serogroups A s; D 29 A ol K A (ZY X W=-135 depending on the chemically distinct polysaccharide capsules and antigenicity. Five serogroups (W-1 35, «CB A]) are responsible for most of the disease. The bacterial meningococcus meningococcal (meningococcal Neisseria meningitides) is a devastating disease that can kill children and adults within hours despite the availability of antibiotics. Improved immune combinations are needed against meningococcal serogroups Y C B A and 35 W1 and/or X Description The general invention of the invention (Vo) to satisfy these needs and the needs of (gyal) the present invention relates to formulations of Neisseria meningitides and methods for their preparation. In one aspect, the invention relates to an isolated polypeptide comprising acid order 800100 having a homology of at least 795 with order identification no. : (VY) where the first twenty acid 801100 residues of the order do not contain .cysteine (yoy).

Ladd in one of the embodiments; The isolated polypeptide includes an amino acid arrangement at positions 1854-1 of arrangement identification number: YY in one embodiment; Isolated polypeptide includes amino acid arrangement at positions 185-198 of arrangement identification No.: VY YY in one embodiment” The isolated polypeptide includes at least 1 neighboring amino acids of acid order 80100 at positions 7554-1185 of arrangement identification no.: .71 in one embodiment ; The isolated polypeptide is .pyruvylated in one embodiment.” The isolated polypeptide is non-lipidated .001 in one embodiment; The isolated polypeptide is immunogenic in one embodiment; The isolated polypeptide includes an amino acid arrangement consisting of the arrangement mentioned later in the definition of arrangement number: except in one respect; The invention relates to an isolated polypeptide comprising at least arrangement 800100 ila 0 795 with arrangement identification number: VT wherein the first twenty acid residues 10 80100 of the arrangement do not contain .cysteine in one embodiment; The isolated polypeptide includes acid order 80100 of order definition AA tad) in one embodiment; Isolate polypeptide includes acid order 80100 from arrangement definition (VT: where the cysteine at position A is omitted in another embodiment). Isolate polypeptide | 00 includes acid order 8007100 from arrangement definition: VT where 6 is substituted at position 1 with an amino acid that is not a Cys residue One embodiment of the isolated polypeptide includes acid order 800100 from order identification number: YY L

— ¢ — in one embodiment; The isolated polypeptide is not .pyruvylated in one embodiment; The isolated polypeptide is non-lipidic. In one embodiment the isolated polypeptide is an immunogen. In another aspect the invention relates to an immunogenic composition comprising a polypeptide as in o any of the aforementioned embodiments. In terms of Al » the invention relates to an immunogenic composition comprising a polypeptide as in any of the embodiments described herein. in the best aspects; The invention relates to an isolated nucleic acid arrangement encoding an isolated polypeptide consisting of the amino acid arrangement mentioned later in the definition of arrangement No.: YY in an embodiment; The isolated arrangement of Nucleic acid includes arrangement identification number: 77. Va on one side; The invention relates to an immunogenic composition comprising ORF2086 a non-lipidic polypeptide; Non-pyruvylated isolated from serogroup Neisseria B 1©01090©5; and at least one conjugate selected from: 0 encapsulated saccharide conjugate of serogroup A meningitides 38:1 (b) conjugate from 06 encapsulated serogroup ¢Neisseria meningitides C (c) conjugate Vo of encapsulated saccharide of serogroup Neisseria meningitides W135 5 (3) 30k conjugate of encapsulated saccharide of serogroup Neisseria meningitides Y in one embodiment; the immunogenic composition includes at least two conjugates that select From: 0 Encapsulated Neisseria A saccharide conjugate (b) Encapsulated Serum C saccharide conjugate (Neisseria 00601091065 | 0 (c) Encapsulated saccharide conjugate Serogroup W135 ¢ (Neisseria meningitides 5 (3) encapsulated saccharide conjugate of Serogroup Y meningitides Y 61556118 L. L.

El in one embodiment; The immunogenic composition includes at least three selected conjugates

Neisseria A saccharide encapsulated from: 0 conjugate from

C encapsulated serum saccharide conjugate (B) encapsulated serum saccharide conjugate 5 encapsulated serum saccharide conjugate (C) encapsulated Neisseria meningitides conjugate and (D) conjugate of ¢ (Neisseria 06010910065 W135 © 61556118L. meningitides serogroup Y saccharide encapsulated In one embodiment; the immunogenic composition saccharide encapsulated includes a conjugate from Neisseria meningitides A bad Encapsulated from group saccharide conjugate from 1L 61556118 meningitides C encapsulated from group saccharide and conjugate from (Neisseria meningitides 135 // A 0 61556118 L. meningitides serotype A from Subfamily polypeptide is a polypeptide in one embodiment; is Embodiments: (non-lipidated). 0 is pyruvylated. It is 812 non-polypeptide. A pyruvylated lipid other than A22 is a polypeptide in one embodiment; .(non-lipidated) is pyruvylated .501 is a non-polypeptide in one embodiment; is 00 (non-lipidated). pyruvylated is 509 non-polypeptide in one embodiment; .(non-lipidated) (yoy

— a — in one embodiment; The polypeptide is non-pyruvylated B44 (non-lipidated) in one embodiment; The polypeptide is non-pyruvylated B22.(non-lipidated) 0 in one embodiment; The polypeptide is non-pyruvylated B24 (non-lipidated) in one embodiment; The polypeptide is 862 non-pyruvylated (non-lipidated) in one embodiment; The polypeptide includes acid order 801100 selected from group A consisting of order identification number A 2 order identification number : 59 order identification number : ©5); CI definition: Ie CI: CUA: CI: VY and CI: Dla in aad embodiments; The polypeptide amino acid arrangement includes the identification number: YY on one side; The invention relates to a method for inducing an immune response against meningitides 10 398 in a mammal. The method involves administration of the reptile organism with an effective amount of an immunogenic formulation comprising non-lipidic polypeptide 0472086; Other than 71177/18160/a0 isolated from serogroup “Neisseria meningitides B” and at least one conjugate extracted from: (1) a conjugate of encapsulated saccharide from serogroup Neisseria meningitides A (b) conjugate (c) Encapsulated serogroup 0 saccharide conjugate C 61556118; (c) Encapsulated serogroup saccharide conjugate ¢Neisseria meningitides W135 5 (3) serogroup encapsulated saccharide conjugate .Neisseria meningitides Y L

—y— on one side; The invention relates to a method for eliciting a bactericidal antibody against a mammalian organism. The method involves administering C. Neisseria meningitides serogroup non-ORF2086 polypeptide to the mammalian organism an effective amount of the immunogenic formulation containing 8. Neisseria meningitides isolated from the pyruvylated lipidic serogroup; not specified in the definition of an amino acid of the polypeptide arrangement in an embodiment; The selected o consists of the set consisting of arrangement definition no: 807100 acid or arrangement 71 arrangement number: arrangement identification number: 40; order identification number: 0; Arrangement Identification VY Arrangement Identification No.: 0; VA Arrangement Identification No.: OA Arrangement Identification No.: OY Arrangement Identification No.: OT No.: IN Embodiment 1. At position cysteine and arrangement definition no: 17; Where 01 deletes the order number:

YT mentioned in order identification number: amino acid other polypeptide arrangement including Vo in one additional embodiment; polypeptide of 1 at position cysteine another embodiment also omitted

YY stated in order identification number: an amino acid polypeptide arrangement containing in one embodiment; The immunogenic composition additionally includes at least one conjugate enzyme of the serum saccharide group selected from: 0 conjugate of the saccharide group (B) conjugate of Neisseria meningitides A Yo precipitated from (c) conjugate from ¢Neisseria 0060109111065 serogroup C saccharide 61556118L; and (d) conjugate from serogroup W135 meningitides

Neisseria meningitides Y encapsulated from the serogroup on one side; The invention relates to a method for eliciting a bactericidal antibody against a mammalian organism. The method involves administration of Neisseria meningitides 1 serogroup 00 non-ORF2086 polypeptide mammalian organism an effective amount of the included immunogenic composition

Neisseria meningitides B isolated from serogroup pyruvylated lipids; not specified in the definition of an amino acid of the polypeptide arrangement in an embodiment; The selector consists of the group consisting of the definition of the order No.: 807100 acid or the order of VY the order No.: L

—A— arrangement definition V0 arrangement definition number: 4 0; Arrangement Identification Number: OY Arrangement Identification Number: NY VA Arrangement Identification Number: OA Arrangement Identification Number: OY Arrangement Identification Number: OT No.: IN Embodiment 1. At position cysteine and arrangement definition no: 17; Where 01 deletes arrangement number: mentioned in the definition of arrangement number: 77. In amino acid another polypeptide arrangement; includes in additional embodiment; polypeptide of 1 at position cysteine also omit AT embodiment 0

YY stated in order identification number: an amino acid polypeptide arrangement containing in one embodiment; The immunogenic composition additionally includes at least one encapsulated conjugate of the serum saccharide group selected from: 0 encapsulated conjugate of saccharide group (B) conjugate of Neisseria meningitides A encapsulated from the saccharide (c) conjugate from Neisseria serogroup 06010910065 0 saccharide 61556118L and (d) conjugate from serogroup W135 meningitides Y serogroup 17 encapsulated. The invention is a method for eliciting a bactericidal antibody against a mammalian organism, comprising the administration of a mammal with an effective amount of Neisseria meningitides pyruvylated non-ORF2086 polypeptide immunogenic composition containing 1 8; and at least one conjugate Neisseria meningitides isolated from the serogroup

HEY selects Neisseria A encapsulated serum group saccharide conjugate from 0

C encapsulated saccharide conjugate (b) conjugate from ¢meningitides encapsulated saccharide conjugate (C) conjugate (Neisseria 00601091066 00

W135 Encapsulated Conjugated Saccharide Group of 32k (3) ¢ Neisseria meningitides

Neisseria meningitides Y serotypes (yoy

q — — Brief Explanation of Drawings Figure 1: Jill Nucleic Acid Contrast Form P2086 Figure ?: Nucleic Acid Configuration of Variant Form 02086. The Gly/Ser stem is in the terminal tail-no for each form contrast underlined. o Figure ?: construct of .ORF2086 Protein Figure 4: Terminal Cys removal - does not lead to loss of expression in E.coli form ro Effect of stem length Gly/Ser on the expression of heteromorph 0852086 Non-lipidic. The order associated with the variant isoform for protein marker 801 is given in the definition of order number: 5?. The arrangement associated with the variant isoform for suspension 00 844 is given in 0 arrangement definition no. protein suspensor 2 is given in arrangement definition no.: FA. The conjugate arrangement for variant isoform for protein suspensor 822 is given in arrangement identification no.: 34. The concomitant arrangement for variant isoform for 07 suspensor 819 is given in arrangement identification no.: 460 Figure 6: High levels of non-lipidic Express 809 despite the presence of a short Gly/Ser stalk. The two boxes on the left side show a variation of 809 eluted Cys-terminal L! before and after induction. The third digits dally show the positive variation 809 Cys N- bk before and after induction. The first digit from the right is the standard value for the molecular weight. The arrangement of the amino acid shown under the image is mentioned in the definition of arrangement number: .41 The arrangement of nucleotide 0 representing the variant 822 isoform Cys-terminal 8; Referred to as 8222_0017" in Figure 1 is shown in Arrangement Definition No.: EF shown under Arrangement Definition No.: 41 in Figure. Arrangement 106 is representative of variant 822 with a Cys-terminal omitted; Referred to as 822_17" in the figure; it is mentioned in the definition of arrangement number: (OY) The nucleotide arrangement represented by

-1 “= for the variation 809 elimed Cys terminal-l8; indicated B09_004" in the figure; shown in the definition of order number: OF Fig. 7: The presence of an optimal codon increases the display of non-lipidic variants 822 and 0822. The left panel shows the manifestation of the Cys-deleted variant 822 terminal-no before © (box (YF, 1) and after (block 1 f;) urg 1016. The right panel shows the showing of the variation 2 from which Cys terminal-l! is omitted before (box (V) and after ( Digit A (inductance of IPTG, bits o and “ are standard values for molecular weight. Figure tA Amino Acid Nucleic Acid arrangements of anisotropy P2086 Figure d(T) ( b): It is an orderly parallelism of variants of the selected wild-type subfamily fHBP BSA described in (abd) 15-19. Note that the !terminus of 862 is 1% homologous to 809 and its C-end is symmetric by 1% 1% with A222 the arrangements shown are (arrangement identifier #: 11); 812 (arrangement identifier #: 1 4); 822 (arrangement identifier #: 0); 862 (arrangement identifier #: 007) BOO (Arrangement Definition #: 81); 824 (Arrangement Definition #: 10); Compatibility Arrangement (Arrangement Definition #: VA Vo) Detailed Description 1: Arrangement Definitions Seq# identification: 1 states the DNA sequence for meningitidis a.a serogroup 8; Gene 804 isoform 2086 which includes a codon with a terminal Cys code no. 0 Sequence identification number: Y states the DNA sequence for meningitidis. aa serogroup 8; The AOS gene is a variant of 2086 which includes a codon that encodes a terminal Cys no. L

Sequence identification number: YF states the DNA sequence for <N. meningitidis serogroup 8; Gene 812 isoform 2086 which includes a codon with a terminal Cys code no. Definition of order number: ; List the DNA sequence for Meningitidis. aa serogroup 8; Gen 812-2 Variant form 2086; which includes a codon that carries a terminal Cys code no. o Definition of sequence number: © The DNA sequence for <N. meningitidis serogroup 8; Gene 822 variant 2086 which includes a codon carrying an N-terminal Cys code Sequence identification number: states the DNA sequence for meningitidis. aa serogroup 8; gene 802 variant 2086; which includes a codon that carries a terminal Cys code no. Sequence identification number: 1 states DNA sequence for meningitidis a.a serogroup 0 8 gen 803 variant 2086; which includes a codon that carries a terminal Cys code no. Definition of sequence number: A states the DNA sequence for <N. meningitidis serogroup 8; gene 809 variant 2086; which includes a codon that carries a terminal Cys code no. Sequence identification number: 4 mentions the DNA sequence for meningitidis. a serogroup 8; Gene 822 isoform 2086 which includes a codon with a terminal Cys code no. Vo sequence identification number: 01 DNA sequence for “N. meningitidis” mentions serogroup B gene B24 variant 2086” which includes a codon carrying a 545 N-terminal Cys Sequence identification number: 11 DNA sequence for (N. meningitidis) mentions serogroup 8; gen 844 variant 2086; which includes a codon carrying Cys-terminal code no. Sequence identification number: 11 State the amino acid order for meningitidis. No 0 serogroup 8; 804 variant 2086; which includes an N-terminal Cys at the .1 amino acid site (yoy).

Sequence # identification: 1 states the amino acid order for meningitidis. No serogroup 8; AOS Variation 2086; which includes the N-terminal Cys at the amino acid site 1. Arrangement identification number: 14 mentions the amino acid arrangement for meningitidis. no © serogroup B 812 Variant 2086; which includes Cys terminal la at amino acid site 1. Arrangement identification number: 11 mentions acid arrangement 8010100 for meningitidis. No serogroup 8; A22 contrast 2086; Which includes Cys terminal N at amino acid 1 site Va. Arrangement identification number: 16 mentions the amino acid arrangement for meningitidis. No serogroup 8; 802 Variation 2086” which includes Cys terminal N at amino acid 1 site. Arrangement identification number: 117 mentions the amino acid arrangement for meningitidis. No serogroup 8; 803 variations 2086; which includes the N-terminal Cys at amino site 1. 800 10 Arrangement identification number: VA states the amino acid arrangement for meningitidis. no serogroup 8; 809 variations 2086; Which includes the N-terminal Cys at the amino acid site 1. Arrangement identification number: 19 mentions the amino acid arrangement for meningitidis. No 0 serogroup 8; 822 variations 2086; Which includes Cys terminal N at amino acid 1 site. Ed

Arrangement identification number: Yo states the amino acid arrangement for meningitidis. No serogroup 8; 824 variations 2086; which includes the N-terminal Cys at the 1 amino acid site. Arrangement identification number: 71 mentions the amino acid arrangement for meningitidis. No © serogroup (B 844 Variant 2086 ; which includes a Cys terminal LA at the amino acid site 1. Sequence identification number: YY Lists the DNA sequence for a pre-precursor sale shown in Example 2. Sequence identification number: YY List the DNA sequence for an inverted precursor shown in Example 0 2. List the sequence number: 4 7 List the DNA sequence for the precursor sale precursor shown in Example 2 Table 1. List the sequence number : Yo reports the DNA sequence for an inverted precursor; shown in Example 2 Table 1. Yo represents the sequence number: YT reports the DNA sequence for a precursor sale; shown in Example 2 Table 1. Sequence identification: YY states the DNA sequence for an inverted precursor shown in Example 2 Table 1. Arrangement definition: YA states the DNA sequence for a stem +© 5//A6; shown in JE) 00 4 Arrangement Identification No: 79 states the amino acid order for the stem (Gly/Ser shown in Example 4 which bears its code; for example For example, the definition of the order (YA tad) Amad

Identification of sequence number: To states the DNA order for the (Gly/Ser) leg shown in example 4 Definition of sequence number: 1 states the amino acid order for the (Gly/Ser) leg shown in example 4 that carries its code; eg sequence definition (Yo tad) > sequence identification number: TY lists the DNA sequence for the stem (Gly/Ser shown in Example 4 sequence definition number: YY lists the sequence acid 801100 for the stem of Gly/Ser that encodes it; eg sequence identification number:77 and sequence identification number Yeo sequence definition number: T states the DNA sequence for the stem (Gly/ Ser shown in example 00 4 sequence identification number: Yo mentions acid order 800100 for terminal no in <N. meningitidis serogroup 8;801 variant 2086; shown in figure 0 sequence definition number: 1 states acid order 800100 for terminal no in (N. meningitidis 10 serogroup 8; 844 variant 2086; shown in Fig. 0 arrangement identification number: YY states acid order 800100 for terminal no in 5 . No Serogroup 8 805 Variant Figure 2086 shown in Figure 0 0 Definition of order number: YA acid order 800100 is listed for peripheral no in 5. no serogroup 8 JEN A22 variant 2086; Shown in Figure 0 is nad

a J \ -

Arrangement identification number: Ye mentions acid order 800100 for terminal no in 5. no serogroup 8 822 variant 2086; shown in Figure .O

Arrangement identification number: 406 mentions acid arrangement 800100 for terminal not in

© meningitidis No; serogroup 8; 819 Variant Fig. 2086; shown in fig

.O

Arrangement identification number: 41 mentions acid arrangement 800100 for peripheral not in “(N. meningitidis serogroup 8; variant 2086; shown in Figure 6.

Definition of sequence number: “?; DNA sequence for N-terminal is mentioned in “N. meningitidis”

.6 serogroup 8; 822 contrast Fig. 2086; shown in Figure 0

Define order number: ?; Referred DNA sequence 00000 mentions for N. 5 serogroup 8; gene 844; variant 2086; where 00017 encodes a terminal cysteine (N) as the comparator with sequence identification number: 11. Plasmid 7 includes sequence definition number: 4 .

Vo Define the number order: ££ State the amino acid order of N. meningitidis is non-lipidic; Serogroup B44 BB Variant Figure 2086 Arrangement Identification No: £8 corresponds to Arrangement Identification YY where the N-terminal cysteine is canceled out at position 1 in Arrangement Identification No: 1 ?. The collation definition code $e: is carried in e.g. collation definition: .

N. For 00000 as DNA Arrangement Identification Number: 80 mentions the arrangement of 0. .; serogroup 8; gene 809; variant 2086; where 5 terminal A is canceled and where the order includes 00001765 carrying cysteine region code bearing 07 Plasmid identification pEBO63 includes additional A; As a comparator with the definition of order number: Gly [Ser . 45 Arrangement No.:

Lad

Arrangement Identification No: 67 mentions Arrangement as DNA 00000 for N. 5 .; serogroup 8; gene 809; variant 2086; where is canceled 0017 carries a terminal cysteine code GN as the comparator with sequence identification number: A Plasmid 04 includes sequence definition number: LET ee sequence definition number: 7; The order as DNA 00000 is listed for N. 5).; serogroup 8; gene 809; variant 2086; where 0017 is canceled carrying a terminal cysteine GN as the comparator with sequence identification number: A Plasmid pEB 5 includes sequence identification number: 497 . Arrangement identification number: 8 SY DNA sequence for L. meningitidis; group 0 serotype 8; gene 809; variant 2086; where a codon encoding a terminal cysteine is canceled out; As a comparator with sequence identification number: A Plasmid pLAL34 includes sequence identification number: LEA sequence definition number: 4 mentions the amino acid sequence for N. non-lipidic meningitides serogroup 8 809 variant 2086. Configuration identification No: £9 Matches Vo to arrangement identification VA where the terminal cysteine is omitted at position 1 in arrangement identification OA coded Arrangement Identification Number Q For example, DNA Qi selected from the set formed by Arrangement Identification Number: ET Arrangement Identification Number: EY and Arrangement Identification Number: EA Arrangement Identification Number: 0 © mentions the amino arrangement acid for meningitidis No. 0 serogroup 8; 809 Variant Fig. 2086; where a codon encoding a 6-terminal no is omitted and where the order includes codons encoding an additional Gly/Ser region; Comparative with the definition of order number: 0. Configuration identification code “On” is carried in eg JE Configuration identification number: 2 char

l \ — sequence identification number: SY 51 DNA sequence for ‎meningitidis .l; serogroup 8; gene 844; variant 2086; where a codon encoding a terminal cysteine is canceled out; As comparator with sequence definition #: .11 01-8056 Plasmid includes sequence definition #: 0 o ) sequence definition #: of states the HLA order for the N-terminal in “N. meningitidis serogroup 8 B22 Variant Figure 2086 as shown in JRE Sequence identification number: of The DNA sequence for the N-terminal is stated in “N. meningitidis serogroup 8 5809 variant 2086; As described in JRE Sequence Identification #: 4 states the DNA sequence for L. meningitidis; group 0 serotype 8; gene 805; variant 2086; Where a codon encoding a terminal cysteine “N” is canceled as the comparator with sequence definition yo: sequence definition number: 00 mentions the amino acid order for N. meningitidis non-lipidic serogroup 8; 805 Variation Fig. 2086. Configuration identification number: 00 corresponds to arrangement identification number 11 where the N-terminal cysteine is canceled out at position 1 in arrangement definition Yo number 7: 1. The sequence identification code is number :55 carried in eg sequence definition number: 0¢ sequence definition number: 07 states the amino acid order of the repeating arrangement “serine—glycine” shown in example J sequence definition number: 517 states the order of amino acid for N. meningitidis other than Ye lipidic serogroup 8; 801 Variation Fig. 2086. Configuration definition #: OV corresponds to configuration identification #0A where the N-terminal cysteine is canceled out at position 1 in configuration identification #: OA char

Arrangement identification number: 0A mentions acid arrangement 801100 for meningitidis. No serogroup 8; 801 variations 2086; which includes the N-terminal Cys at the 1 amino acid site. Arrangement identification number: 09 mentions acid arrangement 8010100 for meningitidis. no © serogroup 8 815 Variant 2086; Which includes a terminal Cys no at the amino acid site 1. Arrangement identification number: Te mentions the amino acid order for meningitidis. No serogroup 8; 816 variations 2086; which includes Cys terminal N at the site of amino acid 1 Va. Arrangement identification number: 11 mentions the DNA sequence for (N. meningitidis) serogroup 8 B22 variant form 2086 where the codon for Cys replaces the N-terminal at the 1 amino acid position of sequence identification number: 09 with 000 for .Glycine. arrangement identification number: TY mentions acid order 8010100 for meningitidis.no serogroup 8 822 variant 2086; where Cys replaces the N-terminal at amino site 1 860 Vo of sequence identification number: 0 with Glycine sequence identification number : TY mentions the DNA sequence for (N. meningitidis) serogroup 8; A22 variant 2086; where the codon for Cys terminal replaces NO at the 1 amino acid position of the definition of sequence no. : V0 with 000 for Glycine. Arrangement identification number: 14 mentions acid arrangement 8010100 for meningitidis. No Yo serogroup 8; 822 variant 2086; where Cys replaces the N-terminal At amino site 1 0 of sequence definition number: Vo with 6176la6. sequence definition number: 65 mentions an enhanced DNA sequence (PEBO42) codon encoding a polypeptide other than Land Non pyruvylated 05. Lad

Arrangement definition #17: The amino acid arrangement is stated for N. non-cipal meningitides serogroup 8; 812 Variant 2086. The definition of order number: TT is the same as the definition of order number: ; 1 While cysteine omits the N terminus at position 1 of sequence definition number: Ne encodes sequence definition number: 11 dail gs 3 for example sequence definition number LAV ° sequence definition number : 1 mentions the order of the DNA enhancer polypeptide Ja¥ codon other than (ial is not \Al2 pyruvylated identifier of the order number: TA mentions the order of the amino acid for N. meningitides other than cipal Serogroup 8; 822 variant 2086. Sequence identification #: TA is identical to Sequence identification #: 11 while the cysteine omits the N-terminal at position 1 of sequence identification #: No Yo encodes the identification Arrangement No.: 186 dail gs 3 For example, Arrangement Definition No.: 14 Arrangement Definition No.: 4 mentions an improved DNA arrangement, polypeptide Ja¥ codon, other than (dal, non-pyruvylated) 22. Arrangement identification, No.: 0 mentions the amino acid order for meningitides. No serogroup 8; 862 variant 2086; lain includes Cys Jie terminal Aa at the 1 amino acid site 0 identifying order number: 1 does not mention the order amino acid for N. meningitides non-cipal serogroup 8; 862 variants 2086. Stripping is P arrangement number: 1 is identical to the definition of arrangement number: Ve, while cysteine omits the N terminus at position 1 of the definition of arrangement number: Ya Ye definition of arrangement number: YY mentions arrangement Enhanced DNA codon for sequence identification number: aA (yoy

=« \ — sequence identification number: VF mentions the DNA sequence optimizer (pDK086) codon for meningitidis ...for serogroup 8; ADS gene variant 2086 in which a deletion of 0007 encodes a terminal cysteine for; comparison with sequence definition (Yad) sequence definition number: VE lists the amino acid sequence for 008010910065. Lal © serogroup 8; 829 variations 2086; Where Cys Jie includes a terminal A at the 1 amino acid position identifying the order number: Vo states the amino acid order of N. non-lipidic meningitides; serogroup 8; 822 Variation 2086. Arrangement definition #: VO is identical to Arrangement definition #: 19 while cysteine omits the N terminus at position 1 of Arrangement Definition #: VO is the same as Arrangement Definition #: 776 mentions Arrangement amino acid for meningitides. A serogroup B 805 variant 2086. Arrangement identification number: VV states the amino acid arrangement for N. non-lipidic meningitides; Serogroup B ¢ blood variant 2086. NAS configuration identification number is the same as Vo for configuration definition: 19 where there is no terminal cysteine L at position 1 of arrangement identification number: AA configuration identification number: YA mentions arrangement acid 801100 for the order of the sequence positioned in Figure 4(a)-)=( sequence definition #: 79 is the same as sequence definition #: YA except that there is no Cys at yo position 1 of sequence definition # no sequence definition No.: 0 mentions the order of amino acid for “N. meningitidis serogroup B 84 variant 2086 0. The definition of order No. Avs is the same as the definition of order No.: 01 where the terminal cysteine is omitted No at position 1 of Arrangement Identification No.: .01 (yoy yy no. meningitidis for amino acid mentions Arrangement A) Arrangement Identification No.: Arrangement Identification Slay AY Group Serum 8 824 Variation 2086. Arrangement Definition No.: .01 where residuals are omitted at position 1-? From Arrangement Definition No.: 01 No.: If not specified otherwise, all technical and scientific terms used herein have the same Ordinary skill in the art to which this invention belongs A 5 Although methods and materials similar or equivalent to those described herein may be used in practice or testing of the present invention; Suitable methods and materials are described below. The materials, methods and examples are illustrative only and are not intended to be specific. All publications; Patents and other documents cited herein are incorporated by reference in full. Generally refers to a biomolecule; Usually (antigen) the term 'antigen' or a conjugate containing at least one 011006 lipid «polysaccharide» peptide «protein or in some cases a ¢ (cognate antibody) with which an antibody can selectively bind A partner or both of them in TR is an immunogenic substance that can trigger antibody production or animal responses, including formulations injected or absorbed into an animal. The immune response may be triggered by the molecule (hapten or epitope) can be used. The whole or of one or more different sections of a molecule (homogeneous) 1 Term to refer to a single molecule or to a homologous or heterologous group of antigenic molecules. The antigen is recognized by antibodies; 1 cell receptors or other elements of adaptive immunity Significant antigenic and/or cellular-specific epitopes. The term 'antigen (8019860)' includes each antigen identified using any number of antigen-specific epitopes mapping techniques. Well known in the art may be identified. See; eg epitope Ye

Epitope Mapping Protocols in Methods in Molecular Biology, Vol. 66 (Glenn E. Morris, Ed., 1996) Humana Press, Totowa, N. J. Linearity can be determined by, for example, the correlative formation of large numbers of epitopes; The corresponding sections of the peptides molecule (50) supports) on solid supports peptides of L yy are still bound with the peptides while they are salad) with the bodies of the peptides and interact scaffolding protein. These techniques are known in the art and are described in eg US Patent No. 1 N1

Geysen et al. (1984) Proc. Natl. Acad. Sci. USA 81:3998-4002; Geysen et al. (1986) Molec. Immunol. 23:709-715, © All are incorporated herein by reference to their contents. similarly; 60,110,065 structures can be determined by X magnetic resonance identification, for example, by; Crystallography of the amino acids spatial configuration for the above. In addition; Epitope Mapping Protocols «ie dimensions. See; ALS nuclear protein for purposes of the present invention; 'antigen (8011960)' may also be used to denote modifications, such as removals, additions and substitutions (generally protective in nature, but which may retain the ability to induce a nonprotective protein response) of the original arrangement; As long as immunogenicity, these modifications may be dependent, for example, through locus-directed mutagenesis;

Sia special formative procedures; or through a genetic engineering treatment method; or may be accidental; Or (BEE lA can be produced through mutations of hosts” that produce antigens. Add a whole live. in a GIS image or it may be an anti-CLS from a microbe, for example; alse isolate 1 (alae Shows a similar polynucleotide or oligonucleotide antigen; definition also includes Synthetic antigens also include .00016:6 acid for example in side-branch immunization applications; and other synthetic or derived epitopes for example synthetic antigens (Bergmann et al. (1993) Eur. J. Immunol. 23:2777 2781; Bergmann 0 etal. (1996) J. Immunol. 157:3242 3249; Suhrbier, A. (1997). Immunol. and Cell Biol. 75:402 408; Gardner et al. (1998) 12th World AIDS.

Conference, Geneva, Switzerland, Jun. 28 - Jul. 3, 1998). Lad

SN The term amino acid substitutions "protectant (©6005607/817)" may be according to the similarity in polarity; the shipment; solubility » water repellency; absorption (el) and/or the duality nature of the included residues. For example (JBL includes nonpolar (hydrophobic) acids 800100 “tryptophan (proline (valine < isoleucine (leucine) 830106) and ¢methionine© includes [iad neutral amino acids” cysteine (threonine (serine (glycine) (glutamine “asparagine” tyrosine) includes amino acids with a positive charge (basic) ¢ histidine dysine “arginine” and includes amino acids with a negative charge (acid) aspartic acid glutamic acid in some embodiments; the acid 800100 protective changes modify the initial order for cORF2086 polypeptides but do not modify the function of the molecule.When these 0 mutations are generated, we can consider the hydropathy index for -amino acids. aqueous for acid 800100 in providing a bio-reactive function for an Ale-sensitive polypeptide in art (Kyte 8 Doolittle, 1982, J.

Mol.

Biol., 157(1):105-32). It has a similar biological activity. Each amino acid is assigned a Sle pathogenicity index based on its charge and water repellency properties. These indicators are: (4.5+) leucine ¢ valine (+4.2) isoleucine (3.8+); (2.8+) ¢ cysteine/cystine (+2.5) «phenylalanine (1.9+) 0191100106; tglutamate (-3.5) ¢histidine (-3.2) ¢proline (-1,6) styrosine (-1.3) ¢(-0.9) 0 dysine (-3.9) «asparagine (-3.5) aspartate (-3,5) «glutamine (- 3.5) and (4.5-) .arginine It is believed that the special hydropathic characteristic of the amino acid residue determines the secondary and tertiary structure of the resulting polypeptide; which in turn determines the interaction of the polypeptide with other Yo molecules such as . substrates receptors ide bodies can be replaced by 80100 acid enzymes antigens etc. It is known in the art that a similar polypeptide equivalent with obtaining Sle Another that has an indicator of amino acid impairment has indicators of hydropathy within the limits of amino acids functionally. In those changes, it is preferable to replace and also preferably more particularly those within the limits of OF and it is especially preferred those within the limits of oY b with b

Lead can make amino acid protective substitutions or inserts on a water repellent basis. As described in the US Patent 0 (£008) incorporated herein by reference, the larger topical average water repellency for a polypeptide which is governed by the water repellency for neighboring amino acids; corresponds to its immunogenicity and antigenicity; i.e., with a biological property of polypeptide 0 US Patent 0 (£006) indicates that the following water repellency values are attributed to amino residues 0 as follows: (3.0+) ¢lysine (+3.0) ¢arginine (3.0+1+) 8508:1816; ) ¢threonine (-0.4) ¢proline (-0.5+1) tleucine (-1.8) ¢ valine (1.5) tmethionine (-1.3) ¢ cysteine (-1.0) tryptophan ¢ phenylalanine (-2.5) dyrosine (- 2.3) ¢isoleucine (-1.8) 0 (3.4-). In Wd bee ellen the amino said 5 water repellency values within +?; specifically prefer those within VE and YN particularly very much prefer those within .0.*+ The representational substitutions which take account of many of the foregoing distinguishing features are well known to those skilled in the art and include; Unspecified: arginine caspartate glutamate ¢ lysine, 561108; glutamine ¢ threonine 35081891068;

leucine, valine, and isoleucine. © The term “effective immunogenic amount” as used herein refers to an amount of a polypeptide or a combination including an effective immunogenic amount in

supported

E01 “Inducing an immune response in a vertebrate host For example, an immunogenic effective amount of TLP2086 protein of this invention is an effective quantity in inducing an immune response in a vertebrate host. The dose or effective quantity will depend The specific "immunogenic" depends on the age; weight and medical condition of the host; as well as the method of administration. Appropriate doses are readily determined by © skilled in the art. The term 'Gly [Ser stalk] Gly/Ser' as used herein refers to the residue series ‎ Ser; Gly is in the direct downstream of a Cys-terminal residue - not in a protein encoded by ORF2086. These residues may be between © and 1 residue of Gly and 5613 in the .Gly/Ser stem accordingly; The Gly/Ser stalk consists of 2 amino acids to 7 and 13 of a protein encoded by ORF2086 | 0 preferably; The Gly/Ser stem consists of 2 acids 801100 and up to between 7 and 13 of the protein encoded by 0572086. The Gly [Ser] stems of variants 6 of the present invention are represented by the underlined arrangements in Figure 1 (Arrangement Identification No.: 711-1). As shown here, the +58//A6 stalk length could affect stability levels or exhibit a non-lipidic variant of P2086. In a representative embodiment, the effects of stalk length of Gly/SerVO are comparable with those of the corresponding wild-type isoform. The term “immunogenic” refers to the ability of an antigen or a vaccine to induce an immune response; either adaptive or by cell; or both. '40 immunogenic amount' or 'immunologically effective amount' or 'dcp (0056)' are each used herein 0 interchangeably; generally referring to the amount of antigen or An immunogenic composition sufficient to induce a response ae le either a cellular (cell-) or adaptive (8-cell or antibody) response; or all lagi as measured by standard tests known to those skilled in the art. The term 'immunogenic composition' ( immunogenic composition) of any drug combination containing an antigen; Sle is a microorganism; or made up of it; That combination can be used for L

-'1?-

To induce an immune response in Cathn. The immunogenic compositions of the present invention may be used to treat = susceptible to L. 0601791015 infection; By administering immunogenic formulations by a general transdermal or mucosal route of administration. Such administrations may include injections into the (Jamal) in the peritoneum; in the dermis of the skin or under the skin; application with a patch or other transdermal delivery device; or by mucosal administration into the mouth/gut; respiratory tract or genitourinary tract. in one embodiment; An immunogenic composition can be used to manufacture a vaccine or generate polyclonal or monoclonal antibodies that can be used to protect or treat an organism. after an initial vaccination; Objects can be given one

or more reinforcements separated from each other to an appropriate degree. The term 'isolated' means that a substance is removed from its original environment (eg the natural environment if it exists in nature or from its host organism if it is of a synthetic type or taken from one environment to a different environment). For example (Jil) a 'isolated' protein or peptide is Vo that is substantially free of cellular material or other contaminating proteins from the cell or tissue source from which the cprotein or cell is derived substantially from chemical primary sources or other chemicals when chemically synthesized” or otherwise present in a mixture as part of a chemical reaction. In the present invention, Je proteins can be extracted from the bacterial cell or cellular residue so that they are available in a form useful in the manufacture of an immunogenic composition. The term “isolated” includes “(isolated) or Je” (15018109)” purification includes for example Ye (JE) the methods for purifying proteins as described herein. The term “substantially free of cellular material” includes ) Polypeptide or protein preparations in which the polypeptide or protein is separated from cellular components of cells isolated or produced synthetically. Therefore, “protein or peptide is essentially devoid of cellular substance” includes polysaccharide preparations “The capsule” protein or peptide has less than about 710 Jo 20 170 Yo 27.5 or ZY (of dry weight) of protein 0017586008106 contaminated

L

or other cellular material. Laie synthetically produced polypeptide protein is also preferably substantially free from culture medium; That is, the culture medium represents less than about 7700 700 or 75 of the volume of the protein preparation when producing a polypeptide or 7 with a chemical composition; It is preferable that it be essentially free of chemical primary sources or chemicals (opal) that is, it is © separated from chemical primary sources or other chemicals included in the formation of protein or polysaccharide accordingly; those preparations are of polypeptide or protein with a Jil of approximately 6776 0770 20 725 (dry weight) from chemical primary sources or compounds other than a polypeptide [protein] or significant polysaccharide fraction. The term 'L-terminal tail!' (N) denotes -terminal tail) as used here to the N= Lil 0 section of a protein encoded by 01472086; which attaches the protein to the cell membrane. A tail (N= 8) is shown at the bottom of the side view build in Figure oF The terminal-LA tail typically includes the six secret terminal-LA amino acids in the protein encoded by [4]2086a0. in some embodiments; The N-terminal tail is 1-16 amino acids of any one of Order Identification Number: 1-17 7. Vo The term “ORF2086” as used herein refers to Open Reading Frame 2086 of a bacterium of spp. Neisseria 082086 proteins (Neisseria encoding terminator parts of these and immunogenic assemblies comprising these proteins are known in the art and described Mis in WO02003/063766, and in U.S.

Patent Application Publication Nos.

US and US 20090202593, Y. 20060257413 are each incorporated herein by reference. The term “P2086” generally refers to the protein 0472086 encodes. The “P” before “2086” is an abbreviation for “1016:0p”. The P2086 proteins of the invention may be lipid or non-lipidic. “P2086” 5 LP2086 typically refers to the lipid and non-lipid forms of LW 2086.

Ya

TLP2086" of the invention is synthesized. P2086 protein denotes, respectively. "protein at" (313.2086 protein) and "2020867" may typically be the lipid and non-lipid forms for the order. wll means adh " Diluent; excipient and/or Lili pied lola ‎"(pharmaceutically acceptable diluent, excipient and/or carrier) © Depending on the usage herein, it may include any or all of the solvents; dispersing media; coatings; antibacterial and antifungal agents; frequency agents to delay absorption, etc. compatible with administration to humans and other vertebrate hosts Typically, a Pharmaceutical Acceptable Carrier is a diluted; excipient and/or carrier authorized by a federal regulatory agency, a state government, or other regulatory agency, or listed in the Encyclopedia of Pharmacopoeia In the US or other pharmacological encyclopedia generally recognized for use in animals, including humans as well as non-human mammals, the term 'diluent, excipient and/or 'lela' refers to a diluent, adjuvant, excipient, or carrier given together with the medicinal composition.These substances may be diluents; the excipients and/or carriers of the drug are sterile liquids; Jie water and oils; including those from petroleum origin; animal; vegetable or industrial. 0 can use water; Salt solutions and aqueous glycerol 5 dextrose solutions as diluents; excipient and/or liquid carrier; Specifically for injectable solutions. Suitable pharmaceutical diluents and/or excipients include starch, malt (gelatin, sucrose, lactose (glucose), corn flour, chalk, silica, sodium chloride, talc (glycerol monostearate (sodium stearate), skimmed milk powder, glycol) propylene “glycerol” water; “ethanol” etc. Composition 00 may also contain on request minor amounts of wetting, bulking, emulsifying, or pH-stabilizing agents These formulations may be in the form of solutions; suspensions; emulsifier; extended-remaining preparations (DULY) etc. Examples of diluents; excipients and/or suitable drug carriers are described in 'Remington's Pharmaceutical Sciences' by ay.E.W.Martin. The justification and/or appropriate carrier for YO will be obvious to those skilled in the art and will depend in large part on the mode of administration.

—vq-

'protective' immune response refers to the ability of an immunogenic formulation to induce an immune response; either adaptive or by cell; They serve to protect the organism from infection. The prevention provided need not be absolute; any; that there is no indication of preventing or eradicating LIS infection if there is a statistically beneficial improvement compared to comparison subjects; Sia infected animals that have not been given a vaccine or an immunogenic combination. Protection may be limited by diminishing the severity or speed of onset of infection symptoms. in general; includes "protective immune response" inducing an increase in antibody levels specific to a given antigen in at least 075 organisms; including some level of functional antibody responses that can be measured for each antigen. In special cases Including “protective immune response (16500058) 0 Inducing a 2-fold increase in antibody levels or a 2-fold increase in antigen-specific antibody levels in at least 0 75 organisms including a certain level of antibody responses Functionality that can be measured for each antigen.In particular embodiments, antibodies related to the anticoagulant in the blood correspond to a protective immune response.Therefore, the protective immune response can be tested by measuring deficiency of bacterial number 1 in a serum bactericidal activity (SBA) test ) or a cath phagocytosis test for example those described below.

Bacterial number in the absence of immunogenic composition. The terms “protein” polypeptide and “peptide” 00 refer to a polymer of amino acid residues and are not restricted to a minimum product length. So; The definition includes emultimers “dimers (oligopeptides peptides etc.) The definition includes both full-length proteins and parts thereof. Terms also include modifications; jie removals; additions and substitutions (generally protective in nature; but may be non-protective) For a primary arrangement; preferably so that the protein retains the ability to induce an immune response in an animal to which it is given

L

for 0017 . The definition also includes modifications after expression “acetylation (glycosylation (fic ¢phosphorylation” lipidation) etc.

Active isoforms and parts of polypeptides are also described herein; Declared polynucleotides. “Variants” refer to substantially similar arrangements. As used here © “polypeptide” (variant polypeptide) refers to a polypeptide that is derived from a parent protein with one or more amino modifications acids at the N and/or © terminus of the parent protein. The modification may include the deletion (so-called amputation) of one or more amino 5's at the la and/or © terminus of the original protein; Deletion and/or addition of one or more lsss + o_-_-----++6«6«6+6+6+6«----“---p amino acids) at an internal site one or more in the parent protein; or substitution of one or more amino acids at one or more sites in the original protein. Variegated polypeptides still possess the desired biological activity original polypeptide (sal; i.e., they are immunogenic compounds. Typically a variant of the polypeptide or polynucleotide arrangement declared herein contains (i.e. arrangement number definitions: Yoo or ( YF on Bla to rank at least by about 7715.

(9A 294 or greater for the reference sequence. The term “fragment) denotes a section of a § amino acid nucleotide arrangement comprising a specified number of contiguous amino acid or nucleotide residues. In special embodiments; retained Part of the polypeptide declared herein has biological activity as a full-length polypeptide (Sal) Yo and is thus immunogenic. Parts of the polynucleotide may encode 07 fragments that retain the biological activity of a protein and thus be immunogenic. Alternatively, parts of the polynucleotide Polynucleotides that are useful as PCR raw materials do not generally retain biological activity. Therefore, the portions of the nucleotide arrangement declared here may range from about at least 100 to 1000 to 1000 (Vo). Raqqa Da Pak Fazakt Foe “YOu Yo Yo Branch of Bairaq Pebel Baal Yabil Barak Takhil EIR IES for Kalfa or Youu

(yoy

“yy full length. Portions of a polynucleotide arrangement contiguous to or up to nucleotides of your he Ye declared here comprise at least 0 sq of polypeptide

YOu Mataf “Yer BL YA Kalaf D F ANAF D Alf Graf AY BL Y Yes contiguous; Or it may be up to amino acids 800 or EVO (£04 full length nic nic nic nic. polypeptide contained in amino acids total number of “protein” depending on use here to any ( recombinant) The term 'recombinant

SHAG or a cell that displays an important gene produced by genetic engineering methods. The term “polypeptide” means “a polypeptide or protein as used herein in relation to” (266000510801) of the present invention Jie synthetic proteins. The resulting polynucleotide can display 0/6 according to (recombinant) From a natural source or produced by means of genetic engineering. The “‘creature’ which, in light of its source or its processing, does not use 01001816 acid here, additionally describing a molecule

Glad associated with it in nature. The term polynucleotide from aud is accompanied by each or as used here in relation to a family cell means a family cell that includes “(266000510801) an organism. (subject) The term "object Yo

SS also includes humans. The term (subject) is another animal. The term organism also includes domesticated animals. Examples of domesticated animals include: hamsters (subject) guinea pigs ferrets; oi muscles; (oli) dogs; cats; pigs; rabbits; rats; Also (subject) birds; snakes; lizards; fish; turtles; and frogs. The term “organism” includes animals and cattle. Non-restricted examples of animals and livestock include: alpacas; bison; camels; cattle; goats; rabbits; reindeer. yaks, poultry, cabal, deer, pigs, horses; animas mules monkeys geese and turkeys. Mammal (01807071815) as used herein refers to any mammal; For example; primates. in a preferred embodiment; The mammal being is human

DJ

The terms “vaccine” or “vaccine composition” used interchangeably herein refer to drug formulations comprising at least one immunogenic formulation

Induces an immune response in CAS The present invention has also identified previously unidentified difficulties of the non-lipidic P2086 variant D and provided methods to overcome these difficulties and novel combinations thereof. Although plasmid constructs encoding the non-lipidic P2086 variant provided strong leads for the non-lipidic variant; these heteromorphs were pyruvylated on the Cys-terminal no. Pyruvylation prevents or reduces the possibility of consistent manufacturing of aay. polypeptides The authors further add that deletion of the Cys-terminal non-lipidic P2086 heteroform arrangement avoids pyruvylation of the non-lipidic P2086 heteroform. Attempts to overcome pyruvylation by deleting the codon for the terminal Cys-No either nullified the manifestation or resulted in the manifestation of insoluble heteromorphs. in an alternative way The removal of the Cys-terminal! The non-lipidic P2086 variant has reduced expression in some variants. surprisingly sy fie; anyway; ose sid found that the variants are 805 812 822 862; 801; 809; 822 and 844 non-0 non-pyruvylated lipids can at least be shown despite the omission of the terminal Cys-aa residue. in general; These polypeptides can be shown without further modifications other than deletion of (Cys) compared to the corresponding non-lipidic wild-type rearrangement later. plas eg; JU) 2 and Example 4. Additionally; Inventors discover that non-pyruvylated non-lipid variants are surprisingly immunogenic and unexpectedly produce killer antibodies

0 a for electria. accordingly; The present invention provides two methods to overcome or to reduce the possibility of these difficulties in exhibiting non-lipidic heteromorphs. However la the present invention foresees additional methods. The first method is to vary the length of the Gly/Ser stalk in the C-terminal tail directly into the downstream of the Cys-terminal residue. The second way is to optimize 000017 in Yo tail terminal - no. anyway; The present invention foresees optimization of additional codons. L

These methods provide overexpression of soluble non-lipidic heterostructures©2086. Jd) in one embodiment; The overexpression of the soluble P2086 non-lipidic ALE variant is compared with that of the wild-type non-lipidic variant. Isolated Polypeptides ° The inventors surprisingly discovered ORF2086 isolated non-pyruvylated non-lipidic polypeptides. The inventors further discovered that polypeptides are unexpectedly de idl-generating and capable of inducing an immune response that kills bacteria. By usage cba the term non (non-pyruvylated) pyruvylated refers to a polypeptide that does not have a pyruvate content. The polypeptides 01472086 other than Land Ve that do have a pyruvylated content typically show transition ABS +70 compared to the corresponding wild-type polypeptide. in one embodiment; The invented polypeptide does not show a +0 mass transfer compared to the corresponding wild-type non-lipidic polypeptide when measured with a Cada mass assay. See Das example 10. In another embodiment including non-pyruvylated polypeptide 0472086 isolate 1 non-lipidic cada of retarded N= 4k cysteine compared with the corresponding wild-type non-pyruvylated polypeptide 01472086. The term “cyste ine” (N-terminal cysteine) refers to cysteine (Cys) at the L-terminal or N-terminal tail for polypeptide more specifically “the term “cysteine” denotes terminal "(N-terminal cysteine) as used herein refers to the L-terminal cysteine at which the lipidic proteins 0 LP2086 form a lipid with a lipid tail except 0110a0108; as known in art. eg ( JB When referring to any one of order definition #: 71-11 as a reference order, the terminal cysteine- does not lie at position -1. As another example when referring to order definition #: Ve as a reference order N= djl cysteine at the site. 1 hand

duet

The term “polypeptide 46 wild-type” (wild-type non-lipidated ORF2086 polypeptide) or “polypeptide 2086 wild-type” (wild~type non-lipidated 2086 polypeptide) or '(wild-type non-lipidated polypeptide)' © as used herein to polypeptide 0142086 having an amino acid arrangement homologous to the acid order 8060 MLN ORF2086 fatty polypeptide The corresponding mutation is found in the natural one. The only difference between lipid and non-lipid molecules is that polypeptide 05472086 is a wild-type non-lipid

Non-lipidic with a palmitoyl triple lipid tail at terminal cysteine-no. Ya as known in dll produces a non-lipidic form 2086 because the protein lacks the original leader arrangement or because the leader arrangement is replaced with a section of an arrangement that does not specify the Ja¥ acylation site of the lipidic acid in a host cell. See; Sis 02003/063766// ; merged here for reference

entire. Examples of non-lipidic ORF2086 include not only the wild-type ORF2086 polypeptide NO non-lipidic wild type described, but also polypeptides having an amino acid order according to any one of the sequence definitions: 71-1 where 0 is omitted. A terminal Cys that does not have polypeptides has a sp aa amino acid according to any one of the definition of the arrangement numbers: 71-11 where the N-terminal Cys replaces with acid 801100 is not retarded.Cys Other examples of an ORF2086 polypeptide other than a Yo lipid include a polypeptide having an amino acid arrangement according to the definition of the arrangement number: Vo in which the Cys terminal is omitted and polypeptides having an amino acid arrangement according to For any one of the definition of order number: 110 where the N-terminal Cys replaces with an amino acid that is not retarded 5 not 0. An additional lid on the ORF2086 non-lipidic polypeptide includes acid order 807100 selected from order identification number: ;; (B44) Arrangement identification number: £9 (809); Arrangement ID: 00 Yo (805); Arrangement identification number: (BOL) ©1 Arrangement identification number: (BOT) OA Arrangement identification number: L

—yvo- Examples are Vo (822) . Arrangement identification number: o(A22) 14 identifier ai ill (B22) TY selected from 807100 non-lipidic acid containing arrangement ORF2086 polypeptide additional on

VY (822); order definition: TA (812); Arrangement identification number: TT Arrangement identification number:

AY and sequence identification number: (B24) 80 more includes sequence identification number: AR (862) is a non-lipidic amino containing the ORF2086 polypeptide arrangement. Additional examples are (824) ©.

OB in an embodiment; 177. and arrangement identification number: VT mentioned in arrangement identification number: 0

To 750 shall be at least about a non-lipidic amino acid including a polypeptide arrangement per JAY JAY JAY Kamal Mukkal TAN TAY without a single non-lipidic amino acid JY an isomorphic polypeptide with an encoding arrangement 2001 NO 244 OR JAY A non-lipid probiotic containing 862 corresponding polypeptide. For example; in an analog embodiment; The Ye Val Tall TA is at least about 50 pH.

YY symmetric to order identification number: 0 N 18d on polypeptide 6 non-lipidic wild type include dpolypeptides Yo | The amino acid order of any one of the number order definition: 1-11 shown in Fig. ?; Arrangement Identification No.: OA Arrangement Identification No.: 09 and Arrangement Identification Number: Ve The AT example of polypeptide 05472086 is a non-wild type lipid that includes a polypeptide having an amino acid arrangement according to Arrangement Identification No.: . 7010 The polypeptide 05472086 is a non-lipidide representative wild type. This includes Cys terminal no. Ye as used for clin for example; Polypeptide 844 is "non-lipidated" includes a polypeptide with an arrangement of @mino acid picked from arrangement definition: 10; arrangement definition: VY where the terminal Cys is omitted! A at position 1; sequence identification number: 4 4. Polypeptide 844 "wild-type non-lipidated" includes acid-order polypeptide 800100 sequence identification number: . 1 The B44 polypeptide Yo is “(non-pyruvylated non-lipidated) pyruvylated (yoy pe where YY is omitted) Selected from arrangement identification number: 800170 acid has a polypeptide arrangement Includes .4 4 and arrangement identification number: (NO terminal Cys

As another example, depending on usage cla includes polypeptide 809 "non-lipidated (non— polypeptide 'lipidated) of acid order 800100 picked from definition of order number: VA 8 and oo' - order number : VA where Cys is an N-terminal cancel at position 1; order ID#: £9 and order ID#: 0 5. The term polypeptide 809 includes “a wild-type non-lipid polypeptide” (wild -type non-lipidated) has the order amino acid Arrangement identification number: VA includes 5809 "non-pyruvylated non- (non-pyruvylated non— aul polypeptide "6108180) 0 has the amino acid order Selected from the definition of the order number: VA where

terminal Cys omits no at position 1; identifies order number: £9 and identifies order number: On as an additional example; By use (lia) includes polypeptide 805 "non-lipidated polypeptide "(non-lipidated) of acid order 800170 picked from order identification number: 11 where the terminal Cys is omitted not at position 1 And the definition of arrangement number: 00 includes an example of AT on the polypeptide Yo 805 “non-lipidated polypeptide” (lipidated) having an amino acid arrangement picked from the definition of arrangement number: VY where the Cys-terminal A is replaced at Position 1 with an amino acid that is not retarded (Cys includes an additional example of a polypeptide 05m 'non-lipid (0108180a)' polypeptide having the amino acid arrangement mentioned in the Yad arrangement definition) Example AT also includes an AOS polypeptide "(lipidated) polypeptide having a Ye amino acid order mentioned in order definition #: YY ADS includes a "non-lipidic EE" Polypeptide "(wild-type non-lipidated) has an amino acid arrangement Definition of arrangement number: VY AOS includes "non-pyruvylated non-lipidated" (non—-pyruvylated non-lipidated) polypeptide 4 amino acid arrangement picked from the definition of arrangement number: 11 where the Cys Y terminal of L is omitted A at position 1 and order identification number: 00 Additional examples of 805 include "non

supported

The pyruvylated non-lipidic polypeptide "(non-pyruvylated non-lipidated) has the @amino acid arrangement picked from the arrangement definition number: 11 where the terminal Cys is replaced not at position 1 with an amino acid that is not retarded (Cys sequence definition: 1/6 where Cys at position 1 is omitted; sequence definition: VT where Cys at position 1 is replaced with amino acid Which © is not retarded (Cys and order definition number: YY

By usage clia includes polypeptide 262 "non-lipidated" polypeptide having an amino acid arrangement picked from arrangement definition number: Ve definition arrangement number: Vo where Cys is omitted terminal N at position 0) and configuration identification number: VY includes another example of polypeptide 862 other than polypeptide sand with arrangement identification number: 11 where Cys 0 replaces the N-terminal at position 1 with an amino acid that is not retarded (Cys 62 includes a "wild-type non-lipidated" polypeptide having an amino acid arrangement identification number: .171 Includes 862 "non-pyruvylated non-lipidated polypeptide "(non-pyruvylated non-lipidated) having an amino acid arrangement picked from the definition of arrangement number: Vo where the terminal Cys!a is omitted at position 1 and the definition of Order number: YY Vo Another example of a non-pyruvylated non-lipidic AG2 polypeptide includes polypeptide 41 Define order number: Ve where the terminal Cys replaces !a at position 1 with 0 800100 which not be retarded 5/a0. de "(non—pyruvylated non-lipidated) has the amino acid order

Mentioned in the definition of arrangement number: YY Y. According to the use cla Al2 polypeptide includes a “non-lipidated” polypeptide having an amino acid arrangement selected from the definition of arrangement number: VE sequence identification number: 4 where the Cys terminal is omitted at position 1; sequence identification number: AT includes an Al2 polypeptide that has a “wild-type non-lipidated” acid order 800100 Arrangement identification number: VE 812 includes a "non-pyruvylated non-Yo lipid polypeptide "(non—pyruvylated non-lipidated) having an amino acid order selected

supported

M from the definition of arrangement number: dua 0 the Cys terminal ae N omit position 0 and the definition of arrangement number: a by use clia includes A22 polypeptide "non-lipidated" (non-lipidated) A polypeptide has an arrangement @mino acid picked from arrangement identification number: (V0 arrangement identification number: dua 0 2 omits the N-terminal Cys at position 6 arrangement identification number cit: and arrangement identification number: : TA includes A22 polypeptide "wild-type non— polypeptide "01081©0) having acid order 800100 Arrangement identification number: V0 includes 22 "non-pyruvylated A non-lipidic polypeptide "(non-pyruvylated non-lipidated) of acid order 800100 picked from sequence identification number: 10 where the terminal Cys is omitted at Ye position 1; sequence identification number: TE and the definition of arrangement No. 5. Preferably”; 822 includes “a non-pyruvylated non-lipidated polypeptide” (non-pyruvylated non-lipidated) having the amino acid arrangement mentioned in the arrangement definition: SUA The term “includes omitted Cys (deletion) terminal-no as used here Mutation deleting Cys-terminal 8; compared to p-order A non-lipidic olypeptide of wild type No. For example; “Delete (61100a06)” Cys-terminal denotes the removal of amino acid Cys from the reference order; Sle from the corresponding wild type order; This results in a lower acid lag of 60 compared to the reference arrangement. Unless otherwise specified eel, terms Cys "(N-terminal Cys) N-terminal not at position Cys" (N-terminal Cys at position 1) 1 at "(Cys at position 1) 1 0 is interchangeable. in another embodiment; Cys replaces the N terminal with acid 80100 which is not retarded by eg Cys; in an analog embodiment; includes the terminal Cys aa at position 1 of sequence definitions: 71-11 substituting CoG at position 1 eg (JE) arrangement identification number: TY compared with arrangement identification number: : 19 (Wild Type 822); Configuration ID No.: 1140 YO compared to Configuration Identification No.: Vo (Wild Type (A22) containing Amino Acids Amino Acids

And the*

To replace the N-terminal Cys with any amino acid other than “Cys”, an uncharged acid 80100 is preferred

polar (Jie 9170106. In a preferred embodiment; substituted with a successive residue with Cys The inventors surprisingly discovered that showing ORF2086 non-lipidic polypeptides has cada for a residue N= djl Cys no gas to pyruvylation Determined upon measurement© by mass spectrometry; comparison with corresponding wild-type non-pyruvylated polypeptide ORF2086 Examples of 01472086 non-pyruvylated non-lipidic polypeptides include those of acid order 807100 selected from the group consisting of Arrangement Identification #: VY (804); Arrangement Identification #: (ADS) 11 Arrangement Identification #: VE (812); Arrangement Identification #: 11 (22m); Arrangement Identification #: 116 (802); CI#: VY (803); CI#: VA 0 (809); CI#: 19 (822); CI#: 01 (824) ; arrangement identification number: (B44) YY and arrangement identification number: 1700 (862) where cysteine is omitted at position 1. Example ORF2086 includes a non-pyruvylated non-lipidic polypeptide Jal jal with an arrangement amino acid Arrangement identification number: 0A (801), where cysteine is omitted at position 1. Additional examples of ORF2086 non-pyruvylated non-lipidic polypeptides include isolated VO polypeptides having an amino acid arrangement picked from the group consisting of arrangement identification number: 44; Arrangement Definition No.: 49; order identification number: 0 ; CI#: 00) CI#: TT CI#: 14; CI#: 17; CI#: 5/A. Additional example includes polypeptide 05472086 non-pyruvylated non-lipidic polypeptide Has an amino acid order ID: (BOL) oY Includes example Aly. on polypeptide 01472086 Non-pyruvylated non-lipidic isolated Polypeptide with order ID: YY (805) ; polypeptide has sequence definition number: (ADS) 1776 where Cys is omitted at position 1; 5 polypeptide has sequence identification number: (ADS) 1776 where it replaces Cys at position 1 with acid 801100 which is not retarded 5/O0. Additional examples of ORF2086 non-pyruvylated non-lipidic polypeptides include those with an amino acid Yo arrangement plucked from the group consisting of order identification number: 11 (804); definition of tad).

no

PR

Arrangement Identification ((A22) 11 (812); Arrangement Identification No: VE Arrangement Identification No: (AOS) VY (803); VV Identification (802); Arrangement Identification No: 11 Arrangement Identification No.: (BOL) 0A Arrangement Identification No.: “(B24) 01 (822) Arrangement Identification No.: 19 (809) Arrangement Identification No.: VA Arrangement Identification No.: at cysteine where (AG2) replaces 171 (844); arrangement definition number: YY arrangement definition number: 2086 to include (Jandy which is not retarded 8l7a0.80100 acid with 1 in position ©

YT. Or (Yoo (Yo). At least a non-lipidic about pyruvylated non-polypeptide ahhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhhtih You (YoV (YOoA (Yod) (YT.) on the polypeptide has a minimum value with no maximum value to specify a range. More preferably, it is from § SUA IL 1-X Honey 0 767 It has at most consecutive polypeptide. In some embodiments the amino acids are amino acids Yos have a consecutive polypeptide. In other embodiments it is a pyruvylated non-lipidic amino acid other than a consecutive polypeptide. In one embodiment ; includes

JAY next TNS work Ne according to .al Je which has at least symmetry amino acid 1001 as a whole or JAA JAY said a0 (at JAY JAY JAY Jar JAS JAA 01 the opposite non-lipidic. On A pyruvylated non-polypeptide pathway with the arrangement encoding a non-lipidic arrangement of a pyruvylated non-A62 polypeptide in a representative embodiment; includes (JE)

JAY TAL TNS WORKERS NE JY TOW J THAT HAS AT LEAST AMINO ACIDS 1001 KHK OR TAN AY AT MEKL JAE JAY JAY (JAY NAKAL A TAA. L1 WITH IDENTIFICATION Arrangement #: .0 in one embodiment; the isolated non-pyruvylated polypeptide 0872086 is a non-lipidic polypeptide encoded by an operationally connected nucleotide arrangement with the led system) in which the display system is capable of being expressed in a bacterial cell. » The order of the NUcleotide is connected with a regulatory order that controls the expression of the nucleotide order, i.e. gy

The display systems; organizational arrangements; The appropriate bacterial cells are known in the art. For example JUAN any expression vector “Jie plasmid PET™ (Novogen, Madison Wis.) or PMAL™ (New England Biolabs, Beverly, Mass.) can be used as long as the polypeptide has The ability to appear in a bacterial cell. © preferred; The PET™ vector is used to copy and express synthetic proteins in coli.. In the PET™ system, the transcribed gene may appear under the control of a phage 17 promoter.

The WIA includes representative bacteria (Pseudomonas fluorescens, preferably E. coli). In one aspect; ORF2086 relates to a non-pe pyruvylated lipid polypeptide obtained by a process. The polypeptide is preferably isolated. 0 The invention (Lilia) relates to compositions comprising ORF2086 a non-pyruvylated non-lipid polypeptide obtainable by a process. Preferably, the combination should be an immunogenic formula. The process involves showing an arrangement 0110160116 encoding a polypeptide having an arrangement of an amino acid selected from the set formed by arrangement identification number: VY arrangement identification number: OY arrangement identification number: VE arrangement identification number: (V0 identification Arrangement ID#: OT Arrangement ID#: VY Arrangement ID#: OA 1 Arrangement ID#: 90; Arrangement ID#: 10; Arrangement ID#: FY Arrangement ID#: 4; And sequence identification number: 70 omits Cysteine at position 1. In another embodiment, the process involves showing a nucleotide arrangement encoding a polypeptide having an arrangement of @mino acid from arrangement identification number: 776 where Cysteine is omitting at position . 1. In another embodiment, the process involves showing a nucleotide arrangement encoding a polypeptide having an amino acid arrangement selected from group Y consisting of arrangement identification number: OY arrangement identification number: VY arrangement identification number: VE sequence identification number: (V0 arrangement identification number: OT arrangement identification number: VY arrangement identification number: OA arrangement identification number: 90; arrangement identification number: 10; arrangement identification number: : oF) Arrangement definition No: OA and Arrangement definition t Do: Ve, where Cysteine is replaced at position 1 with an amino acid It is not retarded Cys The arrangement of the nucleotide is operationally connected with a display system capable of showing

Yo in a bacterial cell. L

and in one embodiment; The process involves showing a nucleotide arrangement encoding a polypeptide of acid order 8001700 picked from the set formed by sequence identification number: 44; Arrangement Identification No.: 29) Arrangement Identification No.: 0; Arrangement Identification No.: 00) Arrangement Identification No.: VT Arrangement Identification No.: CTA Arrangement Identification No.: (VY) Arrangement Identification No.: OV and Arrangement Identification No.: Arrangement number: VO© In another embodiment, the process involves showing a nucleotide arrangement encoding a polypeptide having an acid order of 200100 Defining arrangement number: 7/7 In the HAT embodiment “The nucleotide arrangement is selected from the constituent group From Configuration Identification No.: (EY) Configuration Identification No.: (0) Arrangement Identification No.: 7 Arrangement Identification No.: 66 Arrangement Identification No.: 44; Arrangement Identification No.: 5; Arrangement Identification No.: 02 Arrangement Identification No.: (TO) Arrangement identification number: TY Arrangement identification number: VA and 0 arrangement identification number: VY The bacterial cell is preferably E.coli (B44 (B09) 205: In one aspect; the invention relates in composition including first isolate polypeptide; includes acid order 80100 given in arrangement definition No.: £9 (809); second isolate polypeptide” includes amino acid order given in arrangement definition No.: 4 (844). In a preferred embodiment the polypeptides are immunogenic. In an embodiment (Junin HAT 1) 1 The formulation additionally includes polypeptide 0472086 of subfamily A of group EE OPN .N. meningitidis preferably; The ORF2086 polypeptide of the M8 subfamily is the non-pyruvylated non-lipid ORF2086 polypeptide of family A del. in an analog embodiment; Polypeptide 0872086 of subfamily A is ADS and examples include Yo for example; Arrangement Identification No.: VV omitting terminal cysteine-No at position 1; Arrangement Identification No.: 8.00 Other representative embodiment The composition includes a non-pyruvylated non-lipidic AOS polypeptide with an acid arrangement 800100 Arrangement Identification No. : YT where Cys is omitted at position 1; definition of arrangement number: VT where Cys at position 1 is replaced with amino 0 which is not retarded 5 no; And the definition of arrangement number: YY YO domains of control Polypeptide char

In another aspect the invention relates to a method for producing an isolated polypeptide. The method involves showing the polypeptide in a cell (AS includes an arrangement that has symmetry greater than 790 with the arrangement definition number: oF) The said arrangement includes at least one domain of domain selected from the group of 13-18 @amino acids from Definition of Arrangement No.: Amino Acids 21-34 71 © .From Arrangement Identification No.: 71 and 70-80 acids 801100 of Arrangement Identification No.: 91 or a combination thereof; where the cysteine polypeptide is missing a -terminus. Method Walia) involves purifying a .polypeptide The resulting polypeptide includes a non-pyruvylated non-lipidic polypeptide 01472086. Preferably an immunogenic polypeptide. in a preferred embodiment; ah aad lye —

E.coli 0 includes at least one domain of control selected from polypeptides for atid The amino acids 71 of the identification order number: 80100 acids 13-18 the group consisting of

YY from Configuration Identification No.: @mino acids and 70-80 “YY from Configuration Identification No.: 21-4 (AOS) VY (804); Arrangement Definition No.: VY or a combination thereof” includes Arrangement Definition No.: 116 (822); Order identification number: VO (812); Arrangement Identification Number: VE Arrangement Identification Number: 0 (809); Arrangement Identification Number: VA (803); Arrangement Identification Number: VY (802); Sequence identification number: Preferably (844) YY (824); Sequence identification number: Yo. Sequence identification number: (B22) 4 cysteine in another embodiment substitutes .1 at site polypeptides This (cysteine omitting an additional analog polypeptides .Cys that is not retarded 800100 acid with 1 at position 5; definition 0 includes arrangement identification number: 4 4; arrangement identification TY: 54; sequence identification number: 0 polypeptides and arrangement identification number: 64. TY sequence identification number: 00 (ty) polypeptides N. 71 and arrangement identification number: Vo (An additional exemplar includes Arrangement Definition Number: Additional examples include Arrangement Definition Number: YT An additional exemplar includes Arrangement Definition Number:

AY and arrangement definition: (B24) Av. Additional examples include arrangement definition: (B24) vo (yoy).

and in a representative embodiment” the further isolated polypeptide arrangement includes at least one domain of control selected from the group consisting of 96-116 acids 811100 of arrangement definition no. -185 71 801100 From Configuration Definition No.: “VY 187-199 80005 80100 From Configuration Identification No.: YY @mino acids 213-224 © From Configuration Identification No.: (amino acids 226-237 71) Configuration Identification No.: acids 239-248 71 807100 from Configuration Identification No.: 1 or a combination thereof. Examples for polypeptides include at least one domain selected from the group of 13-18 @amino acids From Configuration Definition No.: amino acids 21-34 71 from Configuration Definition No.: YY and 70-80 acid 800100 from Configuration Definition No.: 1 or a combination of 00 thereof; additionally including one domain of control over Least is selected from the set consisting of acid 96-6 807110 of order definition number: acid 158-170 “YY 817170 of order definition number: acid 172-185 71 807100 of order definition number: amino 80105 71 187-199 of Arrangement Definition No.: ‎@mino acids 213-224 “VY from Amino acids 226-237 71 Amino acids 239-248 71 801100 from Vo Amino acids 226-237 71 or a combination thereof; Including arrangement identification number: 16 (802)); Arrangement identification number: VV (803); Arrangement Identification No: (BOO) VA Arrangement Identification No: 09 (822); Arrangement ID #: (B24) 710 and Arrangement ID #: YY (844). Preferably omit polypeptides (this cysteine at position 1. includes a polypeptide of amino order 0 selected from order identification number: ; 4; order identification number: £9) (order identification number: Or 0 Arrangement identification No.: 00 and Arrangement identification No.: OY In one respect; the invention relates to an isolated polypeptide produced by a process described herein. In one embodiment the non-pyruvylated polypeptide is non-lipidic. In another respect it relates The invention relates to an immunogenic composition produced by a process described herein. Nucleotide arrangements encoding polypeptides (yoy

Ha

9: on one side; The invention relates to an isolated Polypeptide compound comprising the amino acid arrangement mentioned in arrangement identification No.: VA with a Cys-terminal deletion of L at position 1 or arrangement identification No.: 49. It includes representative nucleotide arrangements encoding identification Arrangement #: 4 includes arrangements derived from Arrangement Definition #: (ET) Arrangement Definition #: 497; and Arrangement Definition #: 497;

© Arrangement No: LEA preferred; The nucleotide order is the definition of order number: ET on one side; The invention relates to an isolated NUCleotide arrangement that includes the definition of arrangement number: GET on one side; The invention relates to an isolated NUCleotide arrangement incorporating arrangement identification number: “SEY one of the sides”; The invention relates to an isolated NUCleotide arrangement that includes arrangement identification number: EA on one side; The invention relates to a plasmid comprising a nucleotide arrangement selected from Ve arrangement identification number: fT arrangement identification number: 497; Arrangement identification number: 8A and Arrangement identification number: £0 where the plasmid is able to appear in a bacterial cell. Olea systems are organizational arrangements; Bacterial cells are well known in (pall) as described above. Preferably; the bacterial cell is E.coli In another aspect; the invention relates to an isolated Polypeptide compound incorporating the arrangement Vo 8010 800 100 mentioned in arrangement definition No.: 0 5. In an analog embodiment, the order definition number: 0 is encoded by the order definition number: 45 .

4: In another aspect, clad, the invention relates to an isolated Polypeptide compound that includes the amino acid arrangement mentioned in the definition of arrangement No.: YY omitting (N= dhl) Cys at position 1 or the definition of arrangement No.: ; ;. Includes representative nucleotide arrangements that encode the definition of the arrangement

Yo:£2 includes arrangements picked from sequence definition: EV and sequence definition: 0) Preferably the nucleotide arrangement is sequence definition: ?4. In one respect » the invention relates to an isolated arrangement 71101601106 which includes the definition of arrangement No.: 67.

5 on one side; The invention relates to an isolated Polypeptide compound that includes the amino acid arrangement mentioned in Arrangement Definition No.: (ADS) VY where the N-terminal Cys is omitted at

Ed

Pr representative encoding nucleotide identification or sequence identification number: 100 Arrangements of 1 position V0 arrangement number: 00 include arrangements selected from sequence identification number: 4 5; Defining sequence number: is defining sequence number: nucleotide is preferred; The YY arrangement and arrangement definition No. 5.4 on one side; The invention relates to arrangement 7110160006 isolated containing arrangement identification number: 10 isolated containing arrangement identification number: NUCleOtide on one side; The invention relates to the order of 0

YY isolated Configuration definition includes number: NUCleOtide on one side; The invention relates to an isolated arrangement that includes a polypeptide arrangement in another aspect. The invention relates to a compound: 2

SN terminal Cys where (812) omits 1 mentioned in Sequence Definition No.: 800100 acid 17 exemplar that encodes Sequence Identification No.: nucleotide The LTT Arrangements Sequence Identification No.: Isolated nucleotide Arrangement definition No. 3.7 includes an aspect; the invention relates to arranging 0

SY The arrangement definition includes No.: Isolated includes Polypeptide The invention relates to a clad compound in another aspect : 2

Cys where (A22) Vo is omitted. Mentioned later in sequence definition No.: 800100 acid Chaplet arrangement that encodes nucleotide identification The terminal STA rearrangements do not or sequence identification number: include sequence identification number: 14. On one side; The invention relates to Arrangement TA Arrangement No: Vo .14 Isolated The definition of Arrangement No: 16 includes an isolate having a polypeptide arrangement on one side; The invention relates to a compound: 2 wherein the first 71 isomer of at least 795 percent with the definition of arrangement number: amino acid does not contain a polypeptide preferably cysteine of the arrangement contains 801100 retarded acid 00 preferably That the VY from the definition of the arrangement No.: 1854-1 as shown at the sites 800100 acid of the Yo arrangement is “AT a non-lipidic compound other than 18160/a/717/a0. In the polypeptide embodiment Le lis polypeptide isolated is a part of 862. The polypeptide representative fragments include « AT in the embodiment or definition of arrangement 71 of 862 include any number of contiguous residues of the definition of arrangement No.: Ed

— 7 ¢ — No.: 1 no. in one embodiment”; The isolated polypeptide includes the amino acid arrangement at positions 185-1978 of arrangement identification number: 1 No. In another embodiment the isolated polypeptide includes the amino acid arrangement at positions 1871-1949 of arrangement identification Arrangement No.: .91 in one embodiment » The polypeptide comprises at least 1 contiguous amino acids of amino acids of arrangement © 80610 at positions 1954-1785 of Arrangement Definition No.: YY Elsewhere the invention relates With an isolated Nucleic acid arrangement that encodes an isolated polypeptide having an amino acid arrangement of at least 795 symmetry with the definition of arrangement number: VY where the Yo J amino acid residue does not contain cysteine it is preferable that A polypeptide consists of the amino acid arrangement given in Arrangement Definition No. 1: No. In one embodiment; The isolated nucleic 8610 Ye includes the arrangement definition No.: IY In another aspect baal The invention relates to an isolated Polypeptide compound that includes the amino acid arrangement mentioned in the arrangement definition No.: 171 (862) where it is omitted Cys terminal SN sequence definition: WV The representative nucleotide arrangements encoding sequence identification number: VY include sequence identification number: 77. On one side; The invention relates to an isolated Nucleotide Arrangement VO the definition of Arrangement No. IY includes Immunogenic Compositions in a preferred embodiment; The constructs described herein containing an isolated non-pyruvylated non-lipidic polypeptide ORF2086 are immunogenic. Immunogenic constructs that include a protein encoded by the nucleotide sequence of Neisseria meningitidis Yo. 0472086 are known in the art. Include complementary immunogenic formulations those described in WO2003/063766 and RAY (US Patent Application Publications Nos. 20060257413 US and US 20090202593 (their respective contents are merged herein). These immunogenic formulations described herein include a protein that exhibits activity Bactericidal defined as portions 0872086 protein of gh protein to ORF2086 protein thereof immunogenic; and/or bioequivalents thereof.

Neisseria of type Yo AT encoded by the open reading frame is original. Neisseria isolate of protein type 0101810 isolate or protein may be

Jie from bacterial strains; Neisseria 01472086 proteins, for example, can be isolated

A (Neiisseria meningitidis serogroups, including those of Neisseria serogroups) ©

Neisseria Neisseria gonorrhoeae ((29E;.Z.Y X W-135:D C) immunogenic proteins Ja¥ and/or bioequivalents portions in addition to the mentioned 38. B proteins 5 «2086 For subfamily A proteins include ORF2086 01016105 The A proteins are immunogenic terminators and/or bioequivalents thereof. 5 proteins of subfamily; 0 of subfamily 2086 are known in the art; see for example 8 proteins 5 2086; ): 379-89 proteins associated with an amino acid arrangement of Jia, the arrangement numbers 770-7750 in which, in addition to this, 02003/063766//a reveals the definition of LA subfamily 2086 Ve 2086 proteins Accompanying with amino acid arrangements Jia Arrangement of numbers 44-7174 ?in which 01472086 01016175 Al-Farhiyya family 8. Merge here 02003/063766/a/a as a full reference.

Neisseria ORF2086 or equivalents may be lipid or non-lipidic. Preferably non-lipidic. alternatively; The immunogenic constructs may be combinations of 007 lipid and non-lipidic. 01472086 proteins 0 isolate having homology to the immunogenic protein (sal sal) in one embodiment; Formula included

Neisseria encoded by the 0101601606 order of protein at least with 740 amino acid isolated on protein for immunogenicity on sal gd) The composition includes “AT in embodiment ORF2086. TAY TAN TAY IAT AS” said Money INO .the Jo Te with the lowest percentage ila (yoy

EVAR VAR SIVA said impossible CIT TR POA IN VAS EVA VIVA EVAR 0 homologous to a protein encoding by the nucleotide sequence of Neisseria ORF2086 in one embodiment; The immunogen sal (sal) construct includes an isolated protein having at least 7955 amino acid sequence homology to a subfamily protein A encoded by arrangement 1711001601606 of Neisseria ORF2086 preferably ©; The generated construct includes protein dc all an isolated dcp family A encoding the Neisseria ORF2086 nucleotide arrangement in some embodiments; ORF2086 subfamily A polypeptide is a variant (ADS 04H12 of AG2 or 2). In some embodiments, subfamily A polypeptide 0472086 is a variant (ADS 812 or A22 Ya conjugate of polypeptides subfamily 8/: In one embodiment the combination includes any conjugate of ORF2086 polypeptides of family JA Representative conjugates of polypeptides 042086 of family A due dll include; eg Example » 805 and 12m; 05m ADS ¢A22 t2mm; ORF2086 polypeptides 1 subfamily A is non-lipidic and non-pyruvylated in another embodiment The immunogenic composition includes an isolated protein having homology of at least 7955 amino acid sequence with protein subfamily 8 encoded by sequence 171101601606 of Preferably Neisseria 046; the immunogenic formulation includes protein DC family 8 isolate encoded by order 101161601008 of 0142086 Neisseria in some embodiments; protein 0 0852086 subfamily 8 is a variant 844 B24 (B22 B03 B02 or 809. in some embodiments; protein 0472086 de dll family 8 is a variant of B22 B44 or B09. Consortium of polypeptides subfamily 8: in one embodiment; The composition includes any conjugate of polypeptides 0472086 Faris family 8. Representative conjugates of (yoy)

ORF2086 polypeptides dell family 8 includes; For example, “Jiu 809 and 822; B22, 844; 844 and 809; BO1 and 809; 5801 and 822; 801 (B22 B09 “B44, &544; 9 &824;”B24; BO1 “B44; B24” B24, B22 802 &824; 802 & “B01 502 B24 B01” B24 5 B09 B01 “B44 ; 802 “B09 and B44 o. in the HAT embodiment ¢ The composition (sal gd) of the immunogen includes an isolated protein having at least 7955 amino acid sequence homology to the B subfamily protein it encodes Order 171101601606 from Neisseria 046 for example; in some embodiments; ORF2086 007 subfamily 8 is arrangements selected from 844 having acid order 800100 as given in arrangement definition no: 17; 8022 has acid order 807100 as given in arrangement definition 0 no: 761 803 has the order amino acid as described in the arrangement definition: VY 822 has the acid order 8771700 as shown in the arrangement definition: 14 824 has the order amino WS acid shown in the definition order number: 10; BOL has an acid order of 800100 as shown in the order identification number: 48©; Or a variant form having the arrangement of amino acid as shown in the definition of arrangement number: VA where the terminal Cys is omitted or any conjugates thereof. yo it is most preferable that the immunogenic construct include non-pyruvylated and non-lipidic polypeptide 809” B44 non-pyruvylated and non-lipidic polypeptide; or unions thereof. in one embodiment; The composition includes a non-pyruvylated, non-lipidic variant 809 with an acid arrangement of 800100 as indicated in the definition of the arrangement number: VA, where the terminal (N 844 other than 18160/A7117/A0 and a non-lipidic with an amino acid arrangement) is omitted. As indicated in sequence definition: ua (YY 0 omits the terminal Cys no or a union thereof. In the OAT embodiment the immunoconstruct includes 809 non-pyruvylated, non-lipidic having sequence definition: gd 844 is a non-pyruvylated and non-lipidic having order definition No.: 4 4; or a combination thereof. In af aspects the invention relates to an immunocomplex comprising a polypeptide of subfamily B 046 of the serogroup “B N. meningitidis where polypeptide is Yo of 844 other than 18160/a/7117/a0 and non-lipidic. B44 may have acid order 800100 as (yoy

— \ g shown in the order definition oY) where the terminal Cys does not or the Getty order definition omits one of the embodiments; The formulation additionally includes a second polypeptide of subfamily 8 ORF2086 of the meningitidis serogroup Aa 8; where the second polypeptide is an 809 non-ney pyruvylated lipid. 809 may have an acid arrangement 800170 as indicated in © Arrangement Definition No: OA where terminal Cys omits no; or definition of order number: 3.89 one of the embodiments; The formulation is a vaccine. In another embodiment the composition includes not more than three Farahia family polypeptides ORF2086 8 In an additional embodiment; The combination includes not more than two polypeptides of ORF2086 B family .dcp "0 in an additional embodiment; the combination includes at most (0 ? or © species of the heteromorph subfamily ORF2086 B .complexes including the polypeptide Subfamily 8 and polypeptide subfamily A: in one embodiment; the formulation additionally includes one or more polypeptides of subfamily ORF2086 A in a preferred embodiment; the formulation includes a polypeptide of family Vo Subfamily 8/8 AOS Most preferred; ADS subfamily polypeptide A is a non-lipidic compound and is 18160N/1111/A0.In a preferred embodiment AT the composition includes a subfamily A polypeptide 862. More preferably Alike) polypeptide subunit A 862 is a non-lipidic, non-pyruvylated compound. In another embodiment the immunogenic composition also includes an isolated protein having a Bla of an amino acid order of at least 740 0 with protein subfamily A encoded by the nucleotide arrangement of (Neisseria 006) and an isolated protein having at least 740 amino acid sequence homology with protein subfamily 8 encoded by the nucleotide arrangement of 01472086 Neisseria Preferably »; That the immune construct includes a subfamily protein A isolate encoded by the nucleotide arrangement of Neisseria ORF2086 and a subfamily protein 8 isolate encoded by the yoy order

1e nucleotide from Neisseria ORF2086 “iT prefers that the immunogenic construct include a non-pyruvylated, non-lipidic ORF2086 subfamily polypeptide and a non-pyruvylated ORF2086 A subfamily polypeptide and non-libido. COMPOSITIONS: Any combination of 4.0RF2086 polypeptides can be visualized in one of the © embodiments; The composition includes at least one family A polypeptide in the absence of subfamily 8 polypeptides. For example; The formulation includes iil subunit polypeptides/ only. in another embodiment; The composition includes at least one ejb family 8 polypeptide in the absence of e.g. WA subfamily polypeptides; The formulation includes only subfamily A polypeptides. Ya The immunogenic construct includes polypeptide subfamily 8 or a combination thereof. in some embodiments”; The polypeptide is subfamily ORF2086 A in a variant form (ADS h2 or 2022. In another embodiment the polypeptide of family ORF2086 A ie has 2. In a preferred embodiment the polypeptide is subfamily A 05472086 is AOS has an acid order of 800100 as shown in the order definition: VY 804 has an acid order of 800100 as VO -_shown in the order definition: OY 812 has an acid order 800100 as indicated in arrangement definition No.: 1; or a variant 822 having an acid arrangement 801100 as indicated in arrangement definition No.: V0 where the terminal Cys omits no; or any union thereof. Another representative immunoassay also contains a consortium of polypeptides 0872086 subfamily A625 ADS A non-lipidic, non-pyruvylated isolate.For example, the immunoassay may include polypeptide |0 having order identification number: 00 and 106M708A00 having identification Arrangement No.: LV A representative immunocomplex comprising a conjugate of polypeptides 0472086 family A 05m and 12 non-pyruvylated and non-lipidic isolated A representative immunocomplex gal comprising a conjugate of polypeptides 01472 086 subfamily A 712 and 8.62 non-nespyruvylated lipid isolates. no

Saddle (Sal Sal) Immunogen The composition includes any polypeptide of subfamily 8 or a combination thereof. in some embodiments; protein of subfamily 8 0472086 is a variant (B22 (B03 B02 B44 824; or 809). In the June embodiment protein of subfamily B 0472086 is 844 with an acid order of 807100 According to the explanation in the arrangement definition © No.: 1; 502 has the arrangement definition 8100100 according to the explanation in the arrangement definition No.: V7 803 has the sp a acid 800100 according to the explanation in the arrangement definition No: 711; 822 has The amino acid arrangement as described in the arrangement definition: 91; 824 has the amino acid arrangement as indicated in the arrangement definition: 10; or a variant form 809 has the amino acid arrangement as indicated in the Yo arrangement definition No.: VA where Cys-terminal no is omitted or a combination thereof. Another representative immunogenic formulation also includes a conjugate of polypeptides 0872086 of subfamily 8 809 B44 non-pyruvylated non-lipidic isolated. Includes a representative immunogenic formulation Additional conjugate of ORF2086 polypeptides of family B dc all 809 and isolated non-pyruvylated non-lipidic B22. Another representative immunogenic formulation includes conjugate of polypeptides 0872086 subfamily 1 B44 822 B Non-pyruvylated non-lipidic isolated. An additional representative immunogenic formulation includes a conjugate of ORF2086 polypeptides from the family B22, B09 B44, B09 B, non-pyruvylated, non-lipidic isolated.

in one embodiment; The formulation includes non-lipidic polypeptide 0872086 in the absence of lipidic polypeptide ORF2086. in another embodiment; The formula includes ORF2086

polypeptide 0 is non-lipidic and at least one ORF2086 polypeptide. in one embodiment; The formulation includes non-pyruvylated non-lipidic polypeptide 05472086 in the absence of lipidic polypeptide 01472086. in another embodiment; The formulation includes lipidic polypeptide 0472086 and non-pyruvylated non-lipidic polypeptide 01472086. For example; The formulation may include a lipidic AOS polypeptide having the sequence definition yo: AOS VT non-pyruvylated non-lipidic AOS polypeptide having the sequence definition yo: 797. The formulation includes

(yoy

Another representative lipid 5-lipidic polypeptide having sequence identification number: 7/76 and 862 non-pyruvylated non-lipidic sequence identification number: WV) Additional representative composition includes 801 gal polypeptide having sequence definition number: 0A and 2862 non pyruvylated non-lipidic having order identification number: YY © Representative consortia including one representative immunogenic combination conjugate of ORF2086 polypeptides (B09 (AOS 822; 5 B44) isolated non-lipidic. For example; may The immunogenic formulation includes ORF2086 non-pyruvylated non-lapidated 05/non-pyruvylated non-lapidated (identification order number: 00) ORF2086 polypeptide from All subtype A and 8509 non-pyruvylated non-lapidated isolated (identification 0 order number : o(£9 822 (order identification no.: V0) and 844 (order identification number: £4) polypeptides 0872086 of subfamily 8. Another representative immunogenic formulation includes a consortium of subfamily ORF2086 polypeptides A125 AOS A non-pyruvylated non-lipidic isolated and ORF2086 Ahk polypeptides sub-B 822 and 5844 non-pyruvylated non-lipidic isolated. additive (Al12 (AOS polypeptides) 8509; and 844 non-pyruvylated non-lipidic isolates. Another example also includes Al2 polypeptides 62H9, and 844 non-pyruvylated non-lipidic isolates. An additional example also includes A62 12 (AOS polypeptides 809) and 5844 non-pyruvylated non-lipidic isolates. Includes another representative immunogenic formulation A62 polypeptides and 809 non-pyruvylated Yo non-lipid isolates. Another representative immunogenic formulation includes B09 (A62 polypeptides) 862 and B44 non-pyruvylated non-lipidic isolates. Another representative immunogenic formulation includes B09 (A62 polypeptides and 844 non-pyruvylated non-lipidic isolates. Another representative immunogenic formulation includes B44 5 A62 (ADS polypeptides) non-pyruvylated non-lipidic isolates. Another representative immunogenic formulation includes ADS polypeptides 62m 809 Yo and 5844 non Pyruvylated non-lipidic isolated

ee in one embodiment; The immunogenic combination includes a 0:1 ratio of subfamily protein A to subfamily protein 8. In the AT embodiment ¢ the immunogenic combination includes any one of the following ratios of subfamily A polypeptide to polypeptide subfamily 1 8 hi-net in yes sunna (Ar) nc or .01 in other embodiments © the immunogenic composition includes any one of the following ratios of polypeptide subfamily 8 to ALLY subfamily polypeptide GA made in Netflix EADY VEY 1:1; or He) bactericidal immune responses in one aspect”; The isolated polypeptides and constructs described here show a response of 0 . Bactericidal immunity in a mammal against infection with any serogroup of Meningitidis. The isolated polypeptides and constructs described herein show a bactericidal immune response in a mammalian serotype against infection with a serogroup of W=CB (A Y “135 and/or X yo). On the other hand, the isolated polypeptides and constructs are shown Described herein is a bactericidal immune response in a mammal against the ORF2086 polypeptide of WN serogroup 8. meningitides formulations have the ability to induce antibodies against bactericidal meningococcus after administration to a mammal; Strains with specific subfamilies. Additional information on bactericidal responses is given below. See for example (Sta) Examples 11 (6) 12 and 13. In one embodiment the constructs show a bactericidal immune response against the heterotrophic family of Serogroup 8 WN. meningitides for example may show the structure A bactericidal immune response containing a polypeptide from the non-lipid subfamily A against a variant of the family (A deal) of serogroup B meningitides.

-4e- and/or against a variant of subfamily 8 of serogroup N. meningitides B “hail for example; Examples 18 - 19. In an additional aspect; The isolated polypeptides and constructs described herein exhibit a bactericidal immune response against at least one strain of L. meningitidis. Serogroup © A Serogroup 8; serogroup ©; serogroup W135 and/or serogroup 7. In a preferred embodiment; The constructs show a bactericidal immune response against at least serogroup 8; serogroup ©; and serogroup oe 17. no meningitides. See, for example, JE eg 21. Bacteriicidal antibodies are an indicator of protection in humans and preclinical studies serve as a surrogate.” Any new immunogenic combination candidate should These elicit functional antibodies. 9 ;: in one of the sides »; elicited polypeptide 809 non-lipidic isolated; immunogenic formulations of it; Bacteriicidal antibodies against (eg that can bind to) ORF2086 polypeptide of serogroup 8 (N. meningitidis) or subfamily 8.Vo in a representative embodiment; eliciting a non-pyruvylated polypeptide 809 An isolate has an order number definition: VA where the terminal Cys is omitted at position 1 or an order number definition: £9 (LT) that is an immunogenic antibody that elicits bactericidal antibodies against (for example; that can be be associated with ORF2086 polypeptide of serogroup 8 meningitidis. not of subfamily E or 0 preferred subfamily 8. Non-pyruvylated polypeptide 809 and immunogenic formulations of it are preferred; Elicit bactericidal antibodies against the ADS variant (contrast identification number: VF (contrast identification number: 844) (contrast identification number: 17); variant 6 (contrast identification number: Te) variant 824 ( arrangement number: 10); variant form 809 (definition arrangement number: 81); or a combination of them. Nabat Amad bodies

—oy— 816 antibacterial against variant form 844 (order identification no: 17); Variation Figure Variation Figure 824 (Arrangement Identification Number: 10); Variational form Te (Order Definition No.: 12 (Order Definition No.: 81); or a combination thereof. See for example; Example 11; Example 13.9 and Example

4°: on one side; elicited polypeptide 844 non-lipidic isolated; immunogenic formulations of it; Bacteriicidal antibodies against (eg that can bind to) ORF2086 polypeptide of serogroup 8 (N. meningitidis or subfamily 8). In another representative embodiment, B44 polypeptide is non-lipidic; non-pyruvylated The isolate has order definition no: dua YY) Cys-terminal N omitted at position 1 or order definition no: 4 4;

0 and combinations of immunogenic terminators eliciting bactericidal antibodies against (eg; that can bind with) polypeptide 05472086 of serogroup 8 meningitidis family B44 polypeptide (Jag 8 de dl) non-lipidic non pyruvylated and immunogenic formulations die to elicit bactericidal antibodies against variant 844 (order identification no: 17); variant 816 (order identification no: Te); 10); Variant Figure 809 (Arrangement Definition No.: (VA) or a combination thereof. hail eg; Example No. 11. In addition to dia the polypeptide 844 is a non-pyruvylated non-lipidic and immunogenic formulation From it you may also derive bactericidal antibodies that bind to isoform 802 (order identification number: 176). See, for example, Example 12 and Example 13. Furthermore, the B44 polypeptide is non-gal. pyruvylated 0 and immunogenic formulations thereof may also elicit bactericidal antibodies that bind to variant 3 (order identification number: VY) and variant variant BIS (Definition of Arrangement No. 09) See; on

Example “Example 6.2: On one side”; elicited polypeptide 822 non-lipidic isolated; immunogenic formulations thereof; Bacteriicidal antibodies against (eg, that can bind to) polypeptide Yo 01472086 of serogroup 8 (N. meningitidis of subfamily 8. in

Insomnia

—oA-

Additional representative embodiment » The B22 polypeptide is non-lipidic; The non-cpyruvylated isolate has an order identification number: 19 where a terminal Cys deletion is not at position 1; and a terminator immunogenic construct elicits a bactericidal antibody against (eg; that can bind to ORF2086 (a= polypeptide from serogroup meningitidis B0a; of subfamily 8. A non-lipidic, non-cpyruvylated B22 polypeptide© preferably elicits bactericidal antibodies against variant 844 (seq identification number: YY) variant 816 (Arrangement Identification No.: 0) Variation 824 (Arrangement Identification No.: 10); Variation BOO (Arrangement Identification No.: 10)

No.: (((VA) or a combination thereof. Sle hail Example 13.: In one aspect’; elicits the isolated non-lipidic AOS polypeptide; and immunogenic 0 constructs thereof; May be associated with) polypeptide 01472086 of serogroup 8 meningitidis. No subfamily A is one embodiment; the AOS polypeptide is non-lipidic; non-cpyruvylated isolate has order identification number: 1 where omitted Cys N-terminal or order identification number: 00 and immunogenic combinations Lge elicit bactericidal antibodies against (eg that can bind to) polypeptide 1 01472086 of serogroup 8 meningitidis subfamily L. A One embodiment; includes isolated polypeptide 805 of the acid order 807170 identification of arrangement number: (YT where cystein is omitted at position 1). In another embodiment the isolated AOS polypeptide includes the amino acid order Arrangement identification number: VT where cystein substitutes at position 1 with an amino acid that does not have a -Cys retard in one embodiment”; isolate polypeptide 05 0 includes the amino acid arrangement definition of arrangement t Qom: (Jandy WVY) ADS is non-lipidic; non-pyruvylated and immunogenic formulations thereof; To devise bactericidal antibodies against the ADS variant (VY variant 822 (Vo variant 812) (fof VE) a consortium thereof. Sle hl Example 6 and 13. :AG2 3 one side”; elicits isolated non-lipidic polypeptide 862; and immunogenic Yo constructs thereof; bactericidal antibodies against (eg, that can bind with)

no

-and 4e-

A of the subfamily (N. meningitidis 8) of serogroup 01472086 polypeptide sequence identification number: amino acid. Arrangement 862 polypeptide includes one of the embodiments.

Cys that is not an amino acid residue with 1 at the cysteine position where the isolated non-lipidic Ve substitutes for the pyruvylated non-A62 polypeptide in another embodiment the VY-terminal structures neither or Seq definition: Cys where Vo omits Seq definition: © Elicits bactericidal antibodies against (eg that can bind to) immunogenic cate and/or A.No of the family . of bacteria vs. variant Variant Figure 812 (order identification number: 1); Variant Figure 822 (order definition (VY) other elicit 862 other than JUS (Vo number: 11); and Variant 862 (order definition number: 11). : non-lipidic and immunogenic formulations thereof; bactericidal antibodies against pyruvylated formulation 829 variant 809; and variant form 824. See for example; examples non-lipidic and pyruvylated formulations ylated other than AG2 in another embodiment elicits .19 - 8

B16 bactericidal antibody against the isoform (is immunoglobulin 10 non-pyruvylated non-pyruvylated 812 polypeptide in one embodiment); elicit : 2

SN-terminal Cys where VE deletes a lipid isolate with SID: SID: 16 and immunogenic constructs thereof; Bactericidal antibodies against

A subfamily (N. meningitidis 8) of the serogroup ORF2086 non-lipid polypeptide and pyruvylated structures other than A12 subfamily 8. It is preferable to elicit ff, 0 Figure (VY) (order definition no. : ADS immunogenic from it; antibodies against the VE variant (Vo variant 812 (configuration identification number: Vo) (contrast identification number: A22 variant (JE) as lash .809) Variant form Vee Variant Fig. 862 (order identification no. 19 - 18 andy (yoy qm non-lipidic pyruvylated non-A22 polypeptide in one embodiment); elicit terminal no or arrangement identification no. Cys where 11 isolates with defining order number: and immunogenic combinations deleted from it; bactericidal antibodies against (which can bind with) 14

A subfamily (N. meningitidis 8 of serogroup ORF2086 non-lipidic polypeptide and pyruvylated constructs other than A22 and/or subfamily 8. preferably elicited © tad) (order definition AOS Variant form 862 (order definition) Vo variant 822 (order identification number: VY .19 - 18 ABN (JE) as hail .829) Variant (Ve no.: method of elicitation of bactericidal antibodies) in one aspect; the invention relates to a method for eliciting bactericidal antibodies specific for Ya no ... in a mammal. in one aspect; relates to meningitides A towards the serogroup

C The invention is a method to generate specific bactericidal antibodies against the serogroup in a mammal. on one side”; The invention relates to a method for culture of the bodies of N. meningitides an additive organism. AN. meningitides W135 is antibacterial specific to the serogroup in one respect; The invention relates to a method for eliciting bactericidal antibodies against group 0 no in a mammal. on one side; The invention relates to a method of meningitides X serotyped in N. meningitides Y to generate specific bactericidal antibodies against the serogroup of a mammal. on one side; The invention relates to a method for eliciting specific bactericidal antibodies. Bactericidal antibodies specific for 0 no in a mammal In a representative embodiment; the method includes meningitides 8 liad (group 8 L. meningitides) elicitation of bactericidal antibodies specific for 0.

A ... for serogroup 8 subfamily meningitides (ORF2086 8 subfamily ); or union thereof. 086 Amad

-? 1-

The method involves administering to the mammal an isolated amount of non-pyruvylated non-lipid polypeptide 2086 or an immunogenic formulation cate as described above. Look

For (Jad) Examples 18 - 19 and 22. in a preferred embodiment; The method includes elicitation of specific bactericidal antibodies against N. meningitides© serogroup 8 subfamily LORF2086 B containing the isolated polypeptide formulation or the immunogenic formulation B44 non-pyruvylated non-lipidic polypeptide. In another preferred embodiment; The formulation additionally includes 809 non-0 non-lipidic polypeptide. in an analog embodiment; Isolated polypeptide or immunogenic combination includes order identification number: 49; Arrangement Definition No.: 44; or a union thereof. In another representative 0 embodiment the isolated polypeptide or immunogenic composition thereof includes order identification number: VA where terminal Cys omits !a at position 1; order identification number: oF) where Cys omits terminal A ! at position 0) or a union from it. In another representative embodiment (Lay) the isolated polypeptide or immunogenic combination includes order identification number: 99 where the N-terminal Cys at position 1 is omitted. 15- Bacteria have specificity towards meningitides. L serogroup 8 family ORF2086 B ic sll at least one of (AOS polypeptide 812 and 862 is non-pyruvylated non-lipidic.

See for example; Example 19. In a preferred embodiment; The method includes elicitation of specific bactericidal antibodies against N. meningitides serogroup 8 subfamily ORF2086 A containing polypeptide 0 isolate or immunogenic formulation AOS non-pyruvylated non-lipidic polypeptide. in a preferred embodiment; The isolated polypeptide or immunogenic formulation includes order identification number: OY where the N-terminal Cys is omitting at position 1. In a preferred embodiment “SAT” the formulation additionally includes a non-pyruvylated polypeptide B44 Non-lipidic. See eg JE Example 6 and Example 3. In a representative embodiment, the isolated polypeptide or immunogenic combination includes Definition Yo Arrangement No.: 00 Arrangement Identification Number: £6 or a combination thereof. In a preferred embodiment; includes ( yoy

1

Cys deletion dun IY isolate or immunogenic composition Arrangement identification number: terminal polypeptide a! at position Cys where oF) is deleted terminal not at position 1; Definition of Arrangement No.: The isolate or the generating composition, a polypeptide, or a combination thereof. In another representative embodiment, 11 includes the definition of arrangement number: ;; (844); or union thereof. In (ADS) YY immunogenicity, order identification number: one of the embodiments; Includes immunogenic composition to elicit bactericidal antibodies specific for 0 at least one of 0862086 A serogroup 8 subfamily N. meningitides non-lipidic. Look; For example; pyruvylated 62 non-812 (AOS polypeptide .19 - 18 andy) When a representative immunogenic formulation containing at least two ORF2086 non-pyruvylated non-lipidic 0 polypeptides as described above is administered to mammals Amazingly, the inventors discovered that a bactericidal immune response could be induced against serogroup 8 of Neisseria meningitidis in comparison with an immunogenic formulation containing ORF2086, a non-pyruvylated non-lipidic polypeptide. See for example JUG example 19. Accordingly, in one embodiment, the immunogenic composition includes at least one non-pyruvylated non-lipidic ORF2086 polypeptide 0 of the first that acts synergistically with at least a second non-pyruvylated non-lipidic ORF2086 polypeptide to elicit an immune response against the serogroup 8 of meningitides 61556118L.In another aspect, the invention relates to a method for eliciting bactericidal antibodies specific to serogroup C of meningitides in a mammalian organism.The method includes administering to mammalian CA Yo an effective amount of a polypeptide 2086 non pyruvylated non-lipidic isolated from serogroup 8 No meningitides or immunogenic composition cae as described above. Joo lai) for example; Example 22. In an embodiment; A polypeptide includes the amino order 0 mentioned in the order identification number: 71 or the amino acid order selected from the group formed by the order definition number: VY the order identification number: OY the order identification number: VE Yo Arrangement Identification Number: V0 Arrangement Identification Number: OT Arrangement Identification Number: VY Arrangement Identification Number: OA L

s

sequence identification number: 90; sequence definition number: oF and sequence definition number: VY where die cysteine omits position 1. In one embodiment the polypeptide includes the amino acid arrangement mentioned in sequence definition : VY or arrangement identifier acid 807100 picked from the set formed by arrangement identifier number: OY arrangement identifier number: OY arrangement identifier number: VE arrangement identifier number: 10#0 arrangement identifier NT) Configuration ID#: OY Configuration ID#: OA Configuration ID#: 90; Configuration ID#: oo and Configuration ID#: oF) where cysteine at position 1 is substituted with an amino acid which is not retarded 5 no 0. In an embodiment of AT additionally the immunogenic composition includes at least one conjugate selected from 0 encapsulated saccharide conjugate of serogroup A 38:1 (b) conjugate from 0 580008010 encapsulated serogroup ¢Neisseria meningitides C (C) conjugate from serogroup 5800081106 ¢Neisseria meningitides W135 (9) saccharide conjugate encapsulated from serogroup Neisseria meningitides Y containing at least one representative immunogenic composition polypeptide 62 Non-pyruvylated non-lipidic isolated 5) Encapsulated saccharide conjugate of serogroup Neisseria meningitides A Yo (b) Encapsulated saccharide conjugate of serogroup ¢Neisseria 0060109111065 C (c) encapsulated saccharide conjugate from serogroup meningitides W135 61556118l; and (d) saccharide conjugate

Encapsulated Neisseria meningitides serogroup Y on the Ala side. The method involves administering to the mammal an effective amount of non-pyruvylated polypeptide 2086 isolated from serogroup 8 No meningitides or an immunogenic formulation cae as described above. Joo lai) for example; Example 22. In an embodiment; A polypeptide includes the amino arrangement 0 mentioned in arrangement identification number: 71 or the amino acid arrangement selected from group Yo formed by arrangement identification number: OY arrangement identification number: OY arrangement identification number : VE definition

L

Arrangement No. (V0) Arrangement Identification No.: OT Arrangement Identification No.: VY Arrangement Identification No.: OA

Arrangement identification number: 90; Arrangement identification number: oF and Arrangement identification number: VY wherein an amino acid omits the polypeptide arrangement in one embodiment” includes 1. position die cysteine

Mentioned in the definition of the arrangement number: VY or the arrangement of amino acid is selected from the group consisting of

© Arrangement Identification No: OY Arrangement Identification No: OY Arrangement Identification No: VE Arrangement Identification No:

Vo Arrangement Identification Number: OT Arrangement Identification Number: OY Arrangement Identification Number: OA Arrangement Identification No.

ref: 90; order id: oo and order id: oF) where it replaces cysteine at

Position 1 with acid 800100 which is not retarded 5 not 0. in another embodiment; Formula included

Additional immunogenic at least one conjugate material selected from: (1) a saccharide conjugate

0 encapsulated from meningitides serogroup A 38:1 (b) conjugate from

06 encapsulated from serogroup ¢Neisseria meningitides C (c) conjugate

of 5800081106 capsules of serogroup ¢Neisseria meningitides W135 (9)

Encapsulated saccharide conjugate from serogroup Neisseria meningitides Y

in an additional aspect; The invention relates to a method for generating specific bactericidal antibodies

Vo towards the serogroup .meningitides .no in a mammal. The method includes giving

Mammal Organism Effective Amount Of Polypeptide 2086 Non-pyruvylated Non-lipidic Isolated

From serogroup 8 meningitides L. or an immunogenic formulation AIE according to description

above. See for example; Example 22. In an embodiment; Polypeptide includes arrangement

amino acid mentioned in the definition of arrangement number: 71 or acid arrangement 801100 selected from

tad) arrangement definition VY arrangement definition number: VY consisting of arrangement definition number: 0

640; arrangement identification number: V0 arrangement identification number: OT arrangement identification number: OY arrangement identification

No.: OA Arrangement Identification No.: 90; Arrangement Identification No.: 10; Arrangement Identification No.: oF) where

cysteine at position 1 is omitted in one embodiment.”; The polypeptide includes the amino order

0 mentioned in the definition of arrangement number: 71 or the amino acid arrangement selected from the group

Yo Consisting of Arrangement Definition Number: OY Arrangement Definition Number: OY Arrangement Definition Number: VE L Definition

they

Order number: Vo Order number definition: OT Order number definition: OY Order number identification: OA Order number identification: VA Order number definition: oF And order number definition: VY where Replace 6 at position 1 with an amino acid that is not a residual Cys Other embodiment includes __ additionally the immunogenic composition at least one conjugate © selected from: (a) a conjugated sake of a saccharide encapsulated from the group serogroup A meningitides 61556©18; (b) encapsulated saccharide conjugate from serogroup ¢Neisseria 0060109111065 C (c) encapsulated saccharide conjugate from serogroup meningitides W135 61556118L; and (d) a saccharide conjugate

Encapsulated from serogroup Neisseria meningitides Y ya. When a representative immunogenic composition includes four non-pyruvylated non-lipid polypeptides 08572086 and a saccharide conjugate encapsulated from each of the Neisseria meningitides serogroups Y 5 (W135 (W135) C (A) as described above is administered to the sarcastic organism; the inventors surprisingly discovered that a synergistic bactericidal immune response could be elicited at least against serogroup 8 © 5 Y of Neisseria meningitidis - compared with an immunogenic formulation containing polypeptides 01472086 where enzyme saccharide conjugates are not present; and comparison with an immunogenic formulation including Bale conjugate of 606 enzyme for each of the serogroups Neisseria Y 53 W135 (C (A) meningitides where polypeptide is 01472086 Not Found. See for example; Example 22. Accordingly, in one embodiment, the immunogenic composition 0872086 includes at least one non-pyruvylated non-lipidic polypeptide |0 that acts synergistically with ____ conjugate substances Ay PY at least from sac charide encapsulated from serogroup W135 “CA nor Neisseria meningitides to elicit an immune response against Neisseria meningitides. The elicited immune response may be against at least one of the serogroups YC B Yo. From 016010910065 \Neisseria The immunogenic complex may include a protein encoding the sequence

L

-??- polynucleotides (Neisseria 082086 nucleotide) or equivalents thereof as the only active immunogen in the immunogenic combination. Alternatively; the immunogenic composition may additionally include active immunogens; other immunogenic polypeptides from Neisseria csp. or immunoactive proteins from one or more other microbial pathogens (such as a “prion virus”; bacteria or fungus without specification) or a capsular polysaccharide Compositions comprising one or more required proteins; parts or compounds Pharmaceutical on demand for a chosen indication. The present invention foresees any polyantigenic or polyvalent immunogenic combination. For example (JUS) the immunogenic composition includes conjugates of ? or more than <ORF2086 proteins 0 conjugate of ORF2086 protein with Por A proteins one or more; alas) of ORF2086 protein with polysaccharides meningococcal serogroup “CA and W135” and/or polysaccharide conjugates “protein 05472086 conjugates with meningococcus (01601090000005) and pneumococcus (pneumococcus) or a combination of any of the above in a suitable form to give what is required; Sie for delivery to the mucous membrane. 0 skilled in the art will have the ability to formulate such polyantigenic or polyvalent immunogenic formulations. in af sides»; The invention relates to an immunocomplex comprising ORF2086 polypeptides other than 18160/a/0117/a0 and non-lipidic isolated from serogroup 8 “Neisseria meningitidis sala” at least one conjugate selected from: (a) a saccharide conjugate sale Encapsulated from serogroup “Neisseria meningitides A” (b) saccharide conjugate encapsulated from serogroup “Neisseria meningitidis C” encapsulated from serogroup “Neisseria meningitidis C” encapsulated from serogroup Neisseria meningitidis W135 and (d) an encapsulated saccharide conjugate of serogroup Neisseria meningitides Y. in an embodiment; The immunogen construct includes ORF2086 non-cpyruvylated non-lipidic polypeptides isolated from serogroup 8 “Neisseria meningitidis L.”

and at least two conjugates. In another embodiment the composition comprises at least three conjugated substances. (For example Jha combinations may include 58000301065 of: serogroups A and; serogroups / and 135 //; serogroups A and 7; serogroups C and 135 // A; serogroups 135 // A and serogroups M6 and 135//© serogroups M ¢(Y 3C) serogroups W135 (A and 7; serogroup C 5). Compositions including saccharide serogroup © one are preferred. At least (eg YC) in the AT embodiment also the immunocomplex includes a non-0/0/0 non-lipidic polypeptide ORF2086 isolated from serogroup 8 “Neisseria meningitidis 0” and four conjugates; sale Sie encapsulated saccharide conjugate serogroup A ¢Neisseria meningitidis serogroup encapsulated saccharide conjugate ¢Neisseria meningitidis serogroup encapsulated saccharide conjugate ¢Neisseria meningitidis W135 and an encapsulated saccharide conjugate of serogroup meningitides Y 61556118 L. yo In a preferred embodiment the conjugate is a sacchari conjugate de capsule protein carrier. Suitable carrier proteins are known in the art. It is preferable that the carrier protein be 10719 bacteria, “for example, a toxin for diphtheria or tetanus,” or toxoids or mutations thereof. More Sais to be carrier protein s)he than 080/197. For example; in one embodiment; The composition comprises at least one conjugate selected from (a) a saccharide conjugate (1) 00 mcg of serogroup A. meningitides and (1) ¢CRM197 (b) a conjugate from ( V) of encapsulated saccharide of serogroup N.

C (Y) 5 meningitides 0197/40a; (c) saccharide conjugate (1) microcapsulated from total (1) saccharide 000 Wi3say N. meningitides, (7) ¢€CRM197 and (d) saccharide conjugate (1) (encapsulated from Yo of serogroup Y meningitides L. and (1) CRM197 L.

“A and characteristic W135 (CA encapsulated saccharides) Encapsulated A saccharides are known in the art. For example, (a 1—6)-linked N-acetyl The -D- of meningococcal homopolymer is a F04. Can C3 partial at O-acetylation sites 105088m-7180005800116-1; with at position 6-3 in a ratio of 295-75. Conditions used Acetylation to be © a El_sci ezal 886608006 Ai awl at the OAC location (eg under base conditions); but it is useful to keep with 0-0

Av IY (eg at least 50 t Tow this. In some embodiments; it is at least 6-3

A or more) of residues 11801105800106 in 580011811065 original group (740 74. with Acetyl at position 0-3. O-acetylated groups can be substituted as compounds modified 0 are saccharides blocking groups to prevent hydrolysis; these are still within the meaning of the invention. (9<-2 sialic acid (N-acetyl neuraminic acid) sialic at 6-7 and/or 06-8 residues of O-acetyl serogroup © on these 1 groups. O-acetyl but Some clinical isolates lack 0 sialic groups of polymer units that are W135 serogroup saccharide has “C serogroup saccharide with similarity to .acid—galactose disaccharide takes its structure Formula: .518016 acid variant; but at sites "and 5" from O—acetylation .—4)-D-NeupNAc(7/90Ac)-a—-(2—6)-D-Gal-a—-( 1—0 saccharide servogroup 7 is homologous to the saccharide being .galactose instead of glucose. within 5m disaccharide except that the repeat unit —4)~D-NeupNAc(7/90Ac)-a—(2—6)-D- has the formula: Y takes on the serogroup structure variant at O- acetylation has W135c-1)-616-0. Similar to the serogroup sialic acid and 5 of the 17 sites Yo L

-14- As O-acetylated of the invention are the Wh compounds used the saccharides may be saccharides as shown in O-acetylation with the same Sis pattern as described above; partially or wholly when One or more original encapsulated O-acetylated rings; Or the encapsulated saccharides may be removed from them excessively in relation to the 0- acetylated or they may be the “native” saccharide.

in one embodiment; The immunoassay includes “ORF2086 non-0/0/0 non-lipidic polypeptide isolated from serogroup 8” Neisseria meningitidis sala at least one conjugate selected from: (a) a saccharide conjugate encapsulated from serogroup “Neisseria meningitidis A” (b) encapsulated saccharide conjugate from serogroup “Neisseria meningitidis C” (c) encapsulated saccharide conjugate from serogroup Neisseria meningitidis W135 and (d) ) encapsulated saccharide conjugate from serogroup “Neisseria meningitidis 17” where ORF2086 a non-pyruvylated, non-lipidic polypeptide comprises at least one of the following: (Al2 B22 AOS 809 B44 22M 62 824; B15 B16 and 803. In 1 embodiment"; includes the polypeptide containing acid order 8071700 selected from the group formed by the definition of order number: 44 495 00 CTA TT O 15. In another embodiment; includes the polypeptide on an acid order 800100 selected from the group formed by the order identification number: (Yo Te 24 VY where the cysteine is omitted at position 1. In another embodiment the polypeptide includes acid order 800100 selected from the set formed by the order identification number: Ye ge 51:17 0 0 where 01516016 is inferred at the combined location

Cys . which is not an amino acid retarder. The present invention also foresees multiple immunization systems in which any combination useful against a pathogen is combined in or with the formulations of the present invention. For example; without limitation The patient may be given the immunogenic formulation of the present invention and another immunogenic formulation for immunization against, as part of a GARDASIL® regimen called HPV such as human papilloma virus (HPV) YO vaccine

L vy. Multiple immunization. Those skilled in the art will have the ability to select immunogenic formulations for use in combination with immunogenic formulations of the present invention for the purposes of developing and implementing immunization systems. multi 0852086; parts and equivalents as part of the conjugation composition of polypeptides one or more may be used with a carrier to generate an immunogenic polypeptide or protein; So that it is combined with ©.lal meningitidis, a combination that has immunological properties against several serotypes; or serotypes of in particular meningococcal serogroups in particular serogroup 8; and/or against

Ji ORF2086 polypeptides cause various diseases. in an alternative way One of the otters can use a other immunoglobulin. known carrier polypeptides for a protein prepared from these immunogenic compositions. 0 It is preferable that the immunogenic compositions of the invention include an excipient; diluted and/or a medically acceptable carrier. contain warranted material; diluted and/or a carrier of appropriate pharmaceutically acceptable diluents any and all conventional solvents; dispersing medium fillings; solid carriers; aqueous solutions; casings; antibacterial and antifungal agents; surface frequency and adsorption delay factors; etc. Materials include, for example; one or more water; 0 “glycerol” dextrose (phosphate) brine; pH-stable brine etc.; as well as combinations thereof. 6108001 may also include substances as an excipient; a diluent; and/or a carrier Pharmacologically acceptable minor quantities of wetting agents or emulsifying agents; preservatives or pH stabilizers; which Jie adjuvants enhance storage immersion or antibody activity. Well known in the art preparation and use of Substance 0 excipients; diluted and/or a pharmacologically acceptable carrier. So far excluding any conventional non-immunogenic medium or agent of the present invention. sal ge is compatible with the active ingredient; this is expected to be used in formulations. For example, by injection, either subcutaneously or intramuscularly, in addition to the intestine or through the nose. Describe 7 AH

“vy

Wolff et al. Biotechniques;11(4):474-85, (1991). and by Sedegah et al.

PNAS Vol. 91, p. 9866-9870, (1994). Methods of immunization in muscle. Other routes of administration use enteric preparations; rheumatic preparations; downloads; transdermal applications; For example; without limitation. Pharmaceutical grade preparations (JU) include naturally occurring excipients; for example, enteric JUD; for example, magnesium stearate, dactose, mannitol, etc. without limitation. Cmagnesium carbonate (cellulose (sodium) is preferred. saccharine is administered intramuscularly to the immunomodulators. The immunogenic compositions of the present invention also include one or more additional immunomodulators; agents that can modify or perturb the immune system (immunomodulators) 0; as noted in Agents to upregulate or downregulate the cell-involved and/or preferably up-regulate of the cell-involved and/or humoral branches of the humoral pall system. In the embodiment of a booster or JUD. Examples of modulating agents specific immunogens as described in ISCOMATRIX (CSL Limited, Parkville, Australia) or “cytokine US Patent No. 57,547,749 among other agents. 1 Non-exhaustive examples of adjuvants used in the vaccine of the present invention include Jie alum; mineral gels “(Ribi Inc., Hamilton, Mont.) RIBl auxiliary system 0f oil-in-water emulsions; Water-in-oil hydroxide emulsions

Block covalent complete and incomplete copolymer; Freund homologous; for example; Adjuvants (Cambridge Biotech Inc., Cambridge 25-21 ((CytRx, Atlanta Ga.) 0 “AMPHIGEN® Adjuvant” (Chiron, Emeryville Calif.) SAF-M Mass.) and Adjuvant “A monophosphoryl Other Saponin Fat or Quil A Fraction (saponin) Non-Exhaustive Examples of Oil-in-Water Emulsions Useful in the Vaccine of the Invention LAvridine — A Modified Lipid. The 580/62 Modified is a SEAM 1/2 Emulsion Including Preparations 58/0/62 and (vol/vol) detergent 71 squalene (Sigma) (aaa) oil in water containing 725 (vol/ YO L polysorbate © 80 (ana) detergent (vol/ 20.7 ((IC] (surfactants) SPAN® 5 001 (Quil μg/mL 001 m ethanol); 77.2 (vol/v) modified SEAM 1/2 in lecithin (aaa (vol/7 0. sccholesterol) μg/mL (vol/v) 71 squalene (Sigma) oil-in-water emulsion containing 75 (vol/v) polysorbate® 80 detergent ( aaa (vol/70.7 IC) SPAN® 85 Cleanser © ov 5 Quil A 001 µg/mL ethanol (aaa) (vol/77.0 IC) (surfactants)

micrograms/milliliter.cholesterol

Other immunomodulating agents that may be added to the vaccine include one or more cinterferons, interleukins, or other known cytokines. in one embodiment; Ye adjuvant may be a cyclodextrin derivative or a polyanionic polymer (as described in US Patent No. 1118448 and 11103109 respectively. It should be understood that the immunomodulating agent and/or adjuvant used will depend on the person to whom the vaccine is administered or

immune composition; Injection method and number of injections given. In some embodiments the plugin is 58001017. In some embodiments; 0 is a concentration of 5800010 between 1 and YOu μg/milliliter; Between © and 1501 µg/mL or between 0 and 1100 µg/mL. in some embodiments; The concentration of saponin is about 1 µg/mL” about © about µg/gal about 01 µg/Olle about yo µg/mL” about To µg/mL” about 0 © µg/mL” about ov µg/mL” about µg/mL Te” about µg/mL Vo about µg/mL” about Av 0 µg/mL; about 90 micrograms/lill about 001 micrograms/milliliter; Approx. milliliters, about 176 µg/lille about 10 µg/milliliter about 090 µg/milliliter about 7080 µg/lille about 10 µg/milliliter about yY

L

ask µg/l about 771 µg/lle about 0 4 7 µg/mL; Or about YOu μg/milliliter. in selected preferred embodiments; The proteins of this invention are used in an immunogenic formulation for enteral administration that includes a mucosal adjuvant and is used to treat or prevent infection with N. meningitidis in a human host. The mucosal adjuvant may, however, be cholera toxin; It is preferable that mucosal adjuvants other than cholera toxin used according to the present invention include non-toxic derivatives of cholera cholotoxin in which the toxin (A subunit 3, al Las 38) is chemically modified or the resulting related proteins are modified JaY cholera toxin amino acid cholera “10 0 is particularly useful in the preparation of sal ge immunogen formulations of this invention; see the cholera holotoxin E29H mutation as disclosed in the international published application (WO 00/18434) merged here For the entire reference. These compositions may be added to or combined with the polypeptides of this invention. The same techniques may be applied to other molecules having mucosa-assisting sale or heat-transferring Jie toxin delivery properties in Escherichia coli ( LT) 0 Other compounds may be used that have mucosal adjuvant or conducting activity (J fe yellow; DEAE—-dextran J i. polycations and ¢ polyornithine antiseptics; Jie ¢s0dium dodecyl benzene sulphate materials combined with lipid; Jie antibiotics vitamin A streptomycin and may also use other compounds that alter the structural or functional completeness of A mucous membrane surface. Other mucoactive compounds include derivatives of microbial constructions (Ji. na/a; 80110106 IL-5 “MPL (STIMULON™ 05-21 .cimetidine) 2 as described above. The immunogenic compositions of this invention may be delivered in the form of ISCOMS (complexes under ISCOMS (de lid) containing 018; lipid bodies or encapsulated in acrylates Jie L

and 1 or poly (DL -lactide-co-glycoside) to form small globules of suitable size for absorption. Proteins of this invention may also be incorporated into oil emulsions. The quantity (i.e., dose) of the immunogenic combination to be administered to the patient shall be determined according to the standard techniques known to those of ordinary skill in the art; Taking into account such factors as the antigen © of the adjuvant (when present); Age; Type; the weight; factions; Als specific patient and route of administration. For example (JU) may include a dose for an adult human patient of at least 0.1 µg; 1 μg “01 μg” or 0 © μg from Neisseria ORF2086 protein and maximum 80 μg “001 μg” 050 μg 050 μg from Neisseria ORF2086 . protein | 0 Any minimum value and any maximum value may be combined to determine an appropriate range. Adjuvants Immunogenic compositions described herein also include in certain embodiments; one or more adjuvants. An adjuvant is a substance that enhances an immune response at lithe) with an immunoglobulin gene or also an antibody. A number of cytokines or lymphokines have been shown to have Vo-modulating activity of immunity; Thus, they are useful as adjuvants; To name a few (1-0 interleukins 12, 8, 6,7, 5, 4, 2, 1-8 (see eg US Patent No. 13, 16, 17, 18, oVYTIYY). 14,15,14 (and their mutants); ¢interferons—a, B, y granulocyte-macrophage colony-stimulating factor (GM-CSF) (see eg US Patent 50784995 ATCC, entry no. 4980?); ¢ (M-CSF) macrophage colony stimulation Yo-factor stimulation of (G-CSF) granulocyte culture and other factors “© Bs for early tumor programmed death. Other useful adjuvants are included in the immunogenic formulations described here.” Chemokines on Ju mucin-like » as tMadCAM-1 5 GlyCAM-1 (CD34 Lm

17m and 0150.95; member of family <Mac—-1 VLA-1 (LFA-1 (is integrin) and ICAM-2 (CAM-1 family member) of ICAMs (PECAM immunoglobulin asl and CD40 B7-2 BT7-1 co-excitability as lis ¢ (LFA-3 4 CD2 (ICAM-3) growth factors including vascular growth factors; neuronal growth factors; fibroblast growth factors; CD40L 1-A8; vascular endothelial growth factor; receptor molecules including PDGF receptor (PDGF) receptor © “LARD (AIR (Apo-3 (TRAMP) (DR3 WSL-1) p55 (Apo-1 TNF) Fas .Caspase (ICE) 5 tDR6 5 “TRICK2 (TRAIL-R2 (KILLER” NGRF) Other representative adjuvants include, but are not limited to; aluminum hydroxide; aluminum phosphate; STIMULON™ 05-21 (Aquila

Biopharmaceuticals, Inc., Framingham, Mass.); MPL™ (3-O-deacylated 01 monophosphoryl lipid A; Corixa, Hamilton, Mont.), 529 (an amino alkyl glucosamine phosphate compound, Corixa, Hamilton, Mont.), 1-2 (Genetics Institute, Cambridge, Mass.) ; GM-CSF (Immunex Corp.,

Seattle, Wash.); N-acetyl-muramyl-L-theronyl-D-isoglutamine (thr-

MDP); N-acetyl-nor-muramyl-L-alanyl-D-isoglutamine (CGP 11637, 10 referred to as nor-MDP); N-acetylmuramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-(1'-2'-dipalmitoyl-sn—glycero—-3-hydroxyphos—phoryloxy- ethylamin e) (CGP 19835A, referred to as MTP- PE); in selected preferred embodiments; The auxiliary material is . 38 toxin and .5-21 0 “cholera toxin Additional representative adjuvants include non-toxic derivatives of

N. meningitidis and/or engineered couplings or fusions of A subunit including its; 8 subunit (“CTB”) or cholera toxin with a polypeptide “procholeragenoid gyov”

-"171-

fungal polysaccharides; emuramyl dipeptide (schizophyllan) includes derivatives

«E. coli heat variant of the toxin cphorbol esters imuramyl dipeptide ("MDP")

polymers 4 saponins block.

30S aluminum phosphate was used as an aid in a phase 1 analytical experiment to a NYO concentration of 0. + mg/dose; Well below the limit of 0.858 mg/dose specified by

US Code of Federal Regulations [610.15(a)].

Aluminum-containing adjuvants are widely used in humans to enhance the immune response

Antigens when given intramuscularly or subcutaneously. in some embodiments; be concentration

0 in the immunogenic combination between 0091726 5 00 µg/mL; between vet 0.7 0 µg/mL; between 607 and 7.1 µg/mL. in some embodiments; be concentration

0 in the immunogenic formulation is approximately VY Y0 0 μg/mL; about verde

micrograms/milliliter about 175.” micrograms/milliliter about 0.7 µg/milliliter; around

5??. ; micrograms/milliliter about vo YO μg/milliliter; about 0.775 µg/

milliliters about LF is μg/milliliter; about 1.778 micrograms/milliliter; about 0.35 µg/mL Vo; about 375.; micrograms/milliliter about vf μg/milliliter; around

58? vn is micrograms/milliliter; about 0.45 µg/mL; about 0.4975 µg/

milliliters about +0 µg/mL.

in a preferred embodiment; The concentration of aluminum in the immunogenic formulation is about 0.2079

mg/mL and *©.1 mg/mL; between 171 mg/votes Alla mg/milliliter; or between VY Ye mg/mL and 17 mg/mL. in some embodiments; The concentration is 81010101000 in the formula

immunogenic about 0.075 mg/mL; about 1.09 mg/milliliter; about 0.078 mg/

milliliters about 0.71 mg/mL; about 0.7726 mg/mL; about 1.7© mg/mL;

about 0.7175 mg/mL; about 0.71 mg/milliliter; about FY .+ mg/mL; around

0.0 mg/mL; about 0.7978 mg/mL; about 6,400 mg/mL; about 0.4769

L

1 7 _ mg/milliliter; about 6.40 mg/mL; about 0.4978 mg/mL; or about von mg/mL. Contains appropriate adjuvants used to enhance the immune response; To name a few » MPL™ (3-O-deacylated monophosphoryl lipid A, Corixa, Hamilton, (MT)©) described in US Patent 08 HEY Adjuvants suitable for use are also lipid A analogues. Synthetic, aminoalkyl glucosamine phosphate (AGP) compounds, derivatives, or analogues available from Corixa (Hamilton, MT) described in US Patent No. 1,114,917. One of the AGP compounds is 2-[ (R)-3-Tetradecanoyloxytetradecanoylamino] ethyl 2-Deoxy-4-O-phosphono-3-0-[(R)-3-tetradecanoyloxytrade- 0 canoyl]-2-[(R)-3-tetradecanoyloxy — tetradecanoyl-amino]-b-D-glucopyranoside, also known as 529 (previously known as L(RC529) Adjuvant 529 is formulated in aqueous form or as a stable emulsion. Adjuvants include peptides (Sal 0107815; Jie N-acetyl-muramyl-L-threonyl-D-isoglutamine (thr—MDP), 0 N-acetyl-normuramyl-L-alanine-2—(1'-2' dipalmitoyl-sn—glycero -3-hydroxyphosphoryloxy)-ethylamine (MTP-PE); oil-in-water emulsions; such as MESO (US Patent (TYAAAAE) (containing Li 75 polysorbate 80 70.0 “Squalene” and 720.8 85 SPAN (optionally containing various 0 amounts of MTP-PE and formulated into sub-micron particles using a microfluidized device such as the Model 1107¢) (Microfluidics, Newton, MA) and 5381 (containing PLURONIC polymer 75 “polysorbate 80 70.4” Squalene 701 agglomerated with “thr-M DP L121” s) by transformation in a microfluidizer into a sub-micron emulsion or by vortexing to produce larger particle size emulsifier); sale Help Freund is incomplete Support

—VA— aluminum “aluminum hydroxide as ¢(alum) aluminum salts ¢(IFA) ¢tL121/squalene ¢Avridine <AMPHIGEN taluminum sulfate phosphate killed; Stimulon™ QS-21 (Antigenics, Framingham, MA.) 5mths

ISCOMATRIX (CSL Limited, Parkville, US Patent 6V©) and immunostimulating complexes OYO ETT described in US Patent (Australia) Bacterial lipopolysaccharides «Mycobacterium tuberculosis ¢ (ISCOMATRIX) (eg patent CpG motif containing oligonucleotides (synthetic fis; polynucleotides described (IC-31 (Intercell AG, Vienna, Austria) USA No. 47 17)? or a mutation thereof; pertussis toxin (PT) 171357 In European Patents 1,745,117 and 4 0

YYYYIVE or a mutation thereof (e.g., USP.N., aflamagnate cholera toxin, or a mutation thereof; especially (LT) heat-susceptible E.coli or ¢(VFAETE 77110 and L.1) US Patent Numbers 43414 (Lint SE) 1-472 (LT-K63, 171184 and 171184). Methods of Producing Non-Lipidated P2086 Antigens In one aspect; is 011/7/18160/a0 non-lipid The method involves showing a nucleotide arrangement encoding polypeptide 01472086 with a 01472086-deleted L8-terminal cysteine in comparison with the corresponding arrangement from wild-type Yo, and in which the 710160106 arrangement is operationally connected with the system A manifestation that has the potential to appear in a bacterial cell. Representative polypeptides thus produced include any polypeptide described herein. For example, preferably, a polypeptide has the amino acid arrangement mentioned in the definition of arrangement number: 11; arrangement identifier #: 0; arrangement identifier #: 6 1; arrangement identifier #: 630 arrangement identifier #: 71; arrangement identifier #: EVV YO arrangement identifier # : OA arrangement identification number: 91; arrangement identification number: 01; arrangement identification number: supported

-"174- 1 at locus cysteine where (Ve ad) arrangement definition (0A) arrangement identification number: ;71 has an omitted polypeptide; compared to the corresponding wild-type arrangement. In a preferred embodiment Another; is at the cysteine position where the VT is mentioned in the definition of arrangement number: amino acid The amino acid arrangement has an additional representative polypeptide arrangement including polypeptides omitted. Ov configuration definition #: 49; configuration definition #: cE selected from configuration definition #: 0

VE Arrangement Identification Number: VY Arrangement Identification Number: OV Arrangement Identification Number: 00 Arrangement Identification Number: VY Arrangement Identification Number: TA Arrangement Identification Number: TT Arrangement Identification Number: of an amino acid has an additional representative polypeptide arrangement including polypeptide Vo: Vo and (B24) Av, arrangement identification number: 77. Additional examples include arrangement identification number: .polypeptide Lila purification) method includes AY (824) arrangement No.: Vo non-greasy P2086 In some embodiments; the invention provides a method for producing antigens of the heteroform 06472086 in a soluble nucleic acid vector comprising steps Transcribed sequence with BE. coli expression vector without lipid control sequence; Transformation of EL coli virion. In some embodiments; P2086 protein 4 is induced; Expression is induced and isolated 6.

APTG showing with terminal Vo-no in the Cys variation for codon in some embodiments; It is ©16. in some embodiments; codons are omitted. Examples of these include 046 in the variant form 0852086 mutated by N=ill Cys mutagenesis for codon

Gly 5 Ser codons in some embodiments “add Val or Gly (Ala codon ay sil) for a point directly in the stream Gly /Ser stem Alay ORF2086 to the terminal tail-no of divergent form 0 and 5613 in terminal Gly-No. In some embodiments the total number of Cys residues descending for the codon in some embodiments is 11. or 11 1001 9 Ay at least the Gly/Ser stem 11 or 11 00 3 AY terminal - no omitted. In some embodiments, Cys is for

Ser or Gly either N= il retarded (yoy hem

in some embodiments; The no-terminal tail codons of the non-lipid variant ORF2086 are optimized by point mutagenesis. in some embodiments; have terminal tail-l! of variant 05472086 non-fat optimized to match tail N= ahh of variant 809. In some embodiments; The codons of the L1-terminal tail of the ORF2086 variant is optimized by point mutagenesis such that the codon encoding for the fifth acid 0 of the ORF2086 variant is 7000 homologous for the 13-nucleotides 15 Sequence identification number: A and 00001 encoding for the thirteenth amino acid in variant form 0852086 is 7100 identical for 37-39 nucleotides of sequence identification number: 48. In some embodiments; The N-terminal tail shall be of the non-fatty form 0 0452086 dissimilarity in an optimal form such that the nucleic acids 45 57 are identical in proportion to 1-45 NUCeic acids of Order Identification No.: 4. In some embodiments; The no-terminal tail of the non-fatty form 08472086 dissimilarity shall be optimally such that 5 nucleic acids 42 are identical in proportion of 7900 for 4-45 acid 0001616 of arrangement identification No.: 8. in some embodiments”; The N-terminal tail of the ORF2086 variant is non-fatty Yo in an optimal form such that nucleic acids 39 55 are 70000 percent identical to nucleic acids 4-2 of arrangement identification number: 48. In some embodiments ; The tail N= dh of the non-lipid variant P2086 includes one acid 800100 substitution on JN compared with 1-15 acid 800100 of arrangement definition no: AVA some embodiments; The N-terminal tail of the non-lipid variant P2086 includes 1 amino acid substitution Yo-compared with 1-15 acids 80100 of order definition no: VA in some embodiments; The N-terminal tail of the non-lipid variant P2086 includes at least one acid 800100 substitution compared by 2-15 acid 801170 of order definition no: VA in some embodiments; Includes terminal tail-no of variant form P2086 non-fatty ? of amino acid substitutions compared with 2-15 acids 80100 of arrangement definition no.: .18 in some embodiments; be

Yo amino acid substitutions Amino acid substitutions are protective.

(yoy

_— \ - in some embodiments; The non-lipid anisotropic codons are optimal for overexpression. Optimizing 000007 is known in the art. hail, for example; Sastalla et al, Applied and Environmental Microbiology, vol. 75(7): 2099- and Coleman et al, Science, vol. 320: 1784 (2008). © 2110 (2009) In some embodiments; involves making 60007 into a full form matching the use of codon of acid order 807100 with the frequency of codon in all of the selected host with the inclusion of and and/or the exclusion of special DNA rearrangements.In some embodiments, this additionally involves making the codon into a mutated form, minimizing the corresponding secondary mRNA structure in order to reduce translational impediments.4 Some embodiments; Tail codon N= Akl in a is optimized to include any a) of order identification number: YA order definition number: 01 ¢ order definition number: A » and order definition number: Y ¢ In some embodiments ¢ the codon becomes a leg of Gly/Ser optimized to include any a) of the order definition (YA tad) order definition tad) 790 order definition A tad) ; and the definition of order number Yeo for a better understanding of this invention; Remember the following examples. The ARAN is intended to illustrate Vo only and should not be considered as limiting the scope of the invention. sal ge Immunogenic Composition Formulations in special embodiments; The sal gall immunogenic compositions of the invention additionally include at least one adjuvant; pH buffer ¢ (cryoprotectant) salt cation divalent cation inhibitor Lis (non-ionic detergent) ionic 00 for 007 free radical diluent or at least one sale carrier. The immunogenic compositions of the invention may additionally include one or more preservatives as well as a number of meningococcal protein antigens and protein- y.. < polysaccharide conjugates supported

AY — FDA requirements require that vital products in multidose (multidose) bottles contain a preservative; With only a few exceptions. Preservative-containing vaccine products include vaccines containing all) benzethonium chloride (5) virulent “(((anthrax) Polio < Lyme (HepA DTaP) 2-phenoxyethanol (by non-enteric route)); Typhoid (Pneumo) phenol© (other than by the intestinal route); (Vaccinia ro thimerosal -(Rabies Pneumo “(Mening” JE (Influenza (Hib “HepB (Td DT DTaP)) Preservatives approved for use in injectable drugs include; “propylparaben “methylparaben “m—cresol “chlorobutanol” “benzalkonium chloride (benzethonium chloride (2—-phenoxyethanol) thimerosal (phenol < benzyl alcohol < benzoicacid Y. and phenylmercuric nitrate) preparations of the invention may additionally include a stabilizer pH or ch salt; Bll SUS cation non-conic cleaner refrigerant protectant Jie sugar (anti-oxidant) oxidant such as free radical scavenger ) or an agent (SOS 8109a008); or any combination thereof. Selecting any component; sale Mie 1 as parental; may determine whether or not another 1 component is needed ( Hall combinations of these and other components may be empirically determined by a skilled manufacturer for inclusion in the immunogenic compositions of the invention containing preservative depending on a variety of factors such as required storage and administration conditions.

Y. in special embodiments; A preparation of the invention suitable for administration other than by the enteral route includes one or more physiologically acceptable pH stabilizers selected from; without limitation histidine «glycine «citrate (borate «acetate»phosphate (Tris (trimethamine) succinate) in special embodiments; the pH of the preparation is fixed within an acceptable range

pH about 1 to about 9; Preferably from about 104 to about Not

Ed

name

in private embodiments; Adjustment of the pH of an immunogenic composition or preparation of the invention may be required. The pH of a preparation of the invention can be adjusted by techniques standard in the art. Can the product pH be adjusted to be between? and 8. in private embodiments; it may be; or may adjust the pH of the preparation at between TT; and 1 or © and 8. In other 0 incarnations it may be; or he may set the pH of the preparation at about oF about veo about 4; about 4.8» about ©; about 5.©; about 0A about eh about Tio about ov about Veo or about 8. In special embodiments; He or she may adjust the pH die range from glo to Vo from Eo to Helo© to 5.4”; from 5.4 to 5.*; from 9.5 to 0.7 from 9.6 to 2.7 from 5.7 to 5.7"; 5.8 to eed 9.9 to 6” 1 to 0 GY 1.9 to 1.7 1.7 to 1.7 1.7 to ecto 1.5 to » AY 7.5 or

from 7.5 to 4. in special embodiments” the pH of the preparation is about SA in special embodiments; A preparation of the invention suitable for administration other than by the enteral route includes one or more binary cations “SIS” comprising without limitation 1/9012 08012 and 2A00/A at a concentration ranging from about 0.1 mSos to Ae 01 mol;

0 preferably up to about © milligram. in private embodiments; A preparation of the invention suitable for administration other than by the enteral route includes one or more salts; It includes, without limitation, sodium chloride potassium sulfate chloride 0177 for 500” 5 (potassium sulfate) and is present at an ionic intensity that is physiologically acceptable to the organism for administration other than the intestinal route and includes a final concentration to produce an ionic intensity of 0 selected or osmolar ( osmolarity) in the final formulation. The final ionic or osmolarity of the formulation will be determined by various components (eg 10115 of pH-stabilized compound(s) and other non-pH-stabilized salts. A preferred salt, HOI, is present in the range up to ~ to mM; with salt concentrations selected for complementation of other constituents (eg, sugars (509815)) such that the final total osmolarity of the preparation is consistent with YO administration by non-enterial route (eg, intramuscular or subcutaneous injection) It will induce stability (yoy

Longevity of the immunogenic components in the immunogenic formulation across different temperature ranges. Salt-free formulations will tolerate excess ranges from one or more cryoprotectants selected to maintain the desired final osmolarity levels. in private embodiments; A preparation of the invention suitable for administration other than by enteral route includes one or more refrigerant substances selected but not limited to from disaccharides (eg, sucrose «maltose lactose» or polyhydroxy hydrocarbons, trehalose (mannitol «glycerol » dulcitol fi) and sorbitol in special embodiments; the osmolarity of the preparation is in the range from about 0010 to about 0 mOsmol/L and preferably a range from about Yor to about 5000 mOmol/L. l or 0 about 00" to about 506 (Ae osmol/L). A salt-free preparation may contain, for example, from about 75 to about 775 sucrose preferably from about 79 to about 7201 or about to sucrose 717 Ja by way (Aly a preparation free of lal may contain) for example JED from about 77 to about 797 esorbitol preferably from about 4 7 to ZV or about Zo to about 71 sorbitol. When sodium chloride (ic) is added, then 1 the effective range of sucrose or sorbitol is proportionally reduced. These and other considerations of osmolarity are well known to those skilled in the art. In particular embodiments; a preparation of Sis Suitable for administration other than the enteral route One or more free radical oxidation inhibitors and/or LAOS agents A variety of free radical scavengers and chelators are known in the art and used in formulations and methods of use described herein. Examples include, but are not limited to, sodium “glycerol histidine (mannitol) triethanolamine (EDTA/ethanol) ascorbic acid/ascorbate tripolyphosphate “inositol hexaphosphate (citrate) succinic acid/ (DTPA s EDDHA (desferal) malic acid /maleate (succinate) and several conjugates of 1 Yo or more of the above. In special embodiments; a non-reductive free radical scavenger may be added with one hand

—Ao— At least at a concentration that significantly increases the long-term stability of the preparation. One may also be added in different federations; As a scavenger DIS material [a slit OXidation wade or more than] will determine whether or not there is a need to add a bivalent LS sale. The choice of cation is overwhelming. In special embodiments; A preparation of the invention suitable for intravenous administration includes 2 enteric conic comprising one or more non-surfactant (50118018015) surface activators

Polysorbate-80 (polyoxyethylene sorbitan fatty acid esters unspecified) and Polysorbate-40 (Tween 40) “Polysorbate-60 (Tween 60)” (Tween 80) includes without limitation polyoxyethylene alkyl ethers (Polysorbate- 20 (Tween 20) “NP40 (Triton X-114)” (Triton X-100) as well as others such as Brij 35 Brij 58 0 Preferably “(Pluronic 121 Jig) Pluronic ionic and surface doping series other than Span 5 to About 77 (preferably up to 70.0000 at a concentration of about 0.75 Polysorbate—80 constituents (preferably up to 71 at a concentration of about 7,200009 to Polysorbate-40 to about 0.75 7) or 70.5). In other embodiments; a preparation of the invention comprises one or more additional yo-stabilizing agents suitable for administration other than by the enteral route, such as; —SH) includes the group of (reducing agent) “thiosulfate (sodium thioglycolate reductase” glutathione (N-acetyl cysteine or mixtures thereof)” Alternatively or optionally, immunogenic cmonothioglycerol oxygen formulation preparations containing M A preservative of the invention can be additionally fixed by removing 0 from the storage containers; Protect the preparation from light (eg, using amber glass containers). Immunosuppressive formulation preparations containing an acceptable preservative of the invention or excipients include one or more of the diluents; A drug carrier includes any excipient that does not by itself induce an immune response. Includes appropriate citations without YO lam

-A81- Identification of large molecules such as polyglycolic (polylactic acids (saccharides (proteins) polymers “polymeric amino acids” covalent acids “amino acid (copolymers) lactose trehalose”) (Paoletti et al, 2001, Vaccine, 19: 2118) sucrose and fatty acid aggregates (Jie) oil droplets or liposomes © This diluent; the excipient and/or carrier is well known to the skilled manufacturer. The excipient is Pharmacologically acceptable formulations are described, for example, in Gennaro, 2000, Remington: The Science and Practice of Pharmacy, 20th edition, ISBN: 0683306472. The compositions of the invention may be lyophilized or in aqueous form. liquid formulations can usefully be administered directly from their pre-filled form and are thus ideal for injection without the need for sale (forming in aqueous media) as otherwise required with Lyophilized Compositions of the Invention The direct delivery of immunogenic compositions of the invention to an organism may be achieved by administration other than by the intestinal route (intramuscularly; intraperitoneally; intramuscularly). ala subcutaneous dermis; into a vein; or to 0 vesicular space of tissue); or by oral rectal administration; superficial transcutaneous vaginalis; in the nose; in the eye » in the ear; in the lung or other mucous membrane. In the Jamie embodiment the administration is by non-intestinal route by injection into the Ole (Jamal) into the thigh or upper humerus of the subject. Injection may be by needle (eg hypodermic needle); but injection without a needle may be used as an alternative. A typical dose intramuscularly is 1.0 milliliters.The compositions of the invention can be prepared in various forms, eg, for injection, either as solutions or liquid suspensions.In special embodiments, the composition may be prepared as a powder or as a pellet for rheumatic administration.Ole in an inhalation device.In other embodiments; The formulation can be prepared as a suppository or vaginal suppository; or for nasal administration; by ear or eye; by mouth; by drops of gel or powder.

A 7 _ _ The quantities Bd) of the constituents of a specific immunogenic formulation are confirmed by standard studies including monitoring of an appropriate immune response in organisms. after an initial vaccination; Creatures can be given one or more of several immunizations, separated by an appropriate amount of time. Packaging and Dosage Forms 0 The immunogenic composition of the invention may be packaged in unit dose form or in multiple deja (i) two doses”; doses; Or more). For multiple-dose forms; Bottles are typically preferred but not necessarily over prefilled syringes. Various suitable dose formats include but are not limited to m to 11 doses per container at 0.1 to y milliliter per dose. in private embodiments; The dose is v0 milliliter dose. See example International Patent Application WO2007/127668 0 merged here for reference. formulations may be offered in bottles or other suitable storage containers; or may be delivered in pre-packaged deliveries; Sle syringes with one or multiple components; It may or may not have a needle. The syringe typically contains but not necessarily deja one of the immunogenic composition containing a preservative of the invention; Although syringes are multi-dose; Prepackaging is also expected for Vo to use. likewise; It may include a single-dose bottle, but an alternative method includes multiple doses. Binal dose sizes can be routinely established; But a typical dose of the formulation for injection has a volume of 0+ milliliters. in private embodiments; The dose is formulated for administration to humans. In special embodiments the dose Uae SU 3 is formulated for an adult; at the age of YAY a year old; teenager; A crawling child or infant (ie; not more than a year old) is human and in preferred embodiments parenteral administration. Y. The liquid immunogenic compositions of the invention are also suitable for reconstitution for other sal ge immunogenic compositions presented in lyophilized form. when an immunogenic formulation is to be used for such impromptu reconstitution; The invention provides a group with ? or more bottles; SY more than syringes Sl sala or 1 or more of each; With the use of the contents of the syringe to reconstitute the contents of the bottle before injection; or vice versa. uh 7

—AA—

in an alternative way The immunogenic compositions of the present invention may be lyophilized and reconstituted; Ole using one or a number of lyophilization methods well known in the art to form regular shaped dry particles (eg; spherical) such as micropellets or microballs ( Microspheres having distinctive particle features such as average diameter sizes are selected and controlled by varying the exact methods used for their preparation Immunogenic compositions may additionally include an adjuvant that may optionally be prepared with or contained in discrete dry particles of regular (eg, spherical) shape such as microspheres In those embodiments, the present invention further provides a combination immunogenic composition comprising a first component comprising a dry stable immunogenic composition; additionally comprising an optional form 1 or more of the preservatives of the invention; and a second component comprising a sterile aqueous solution For reconstitution of the first component.In special embodiments, the aqueous solution includes 1 or more preservatives; optionally includes at least one adjuvant

(See “WO2009/109550” Jax (merged here for reference)). In the HAT embodiment also a container of multiple potion form is selected from 1 or more of the group consisting of; without limitation general laboratory glassware; decanters » pots » graduated Vo cylinders; bioreactor fermenters; tubes; pipes; bags; intelligence bottles; bottle caps (eg; rubber stopper; screw cap); ampoules; syringes two- or multi-chamber syringes; syringe stoppers; syringe pistons»; rubber stoppers; elastic earplugs; dala stoppers) single-use cartridges and pens; etc. The container of the present invention is not restricted by the material of manufacture; It includes materials such as glass, D4 metals, steel, faa aluminum (stainless steel) 0, etc. and 7/01615A00 SE plastics. thermoplastics elastomers thermoplastic elastomers 8 embodiment of the pala the container is a Schott Type 1 © milliliter glass bottle with butyl stopper the manufacturer will realize The aforementioned formulation is, however, a dense list; but it only serves to guide the manufacturer as to the variety of Yo-facials of the present invention. Additional bodies foreseen for use in the present invention may

L

A q — found in published catalogs from lab equipment vendors and manufacturers such as United States Plastic Corp. (Lima, OH), VWR Examples Example 1: Experimental Procedures © Serum bactericidal assay Syntomological macaques (N = 0 [group]) were immunized intramuscularly with 02020866 or +6 (A + 8 proteins adsorbed to LAIPO4. Synanthropic macaques are an example of a non-human primate. Animals are vaccinated at week 0; 5 (YE) and titers of 6 types ORF2086 and functional antibody are determined at week 0 of 6 and 77 Determines 0 106 titres of serotype 01472086 against A 12086 and 8. Functional antibody titers are screened by a serum bactericidal assay (SBA) against Neisseria meningitidis strains expressing LP2086 with similar rearrangements or Different from those in the vaccine Bacterial killing antibodies in the serum of macaques or rabbits vaccinated with ORF2086 Vo were determined using 58/5 with human complement Immune sera from rabbits or macaques were thermally inactivated to remove complement cross-linked Lalas and subsequently diluted to a series 7:1 dilutions in Dulbecco's PBS with (D-PBS) + Mg2 5 + Ca2 in a 97-well microtitrator plate. The bactericidal activity of serum against WN strains. meningitides bacteria used in the assay were grown in GC media supplemented with Kellogg's complement (GOK) and monitored by optical Yo density at 156 nm. Bacteria for use in the test were harvested at a final 0.0100650 (0.+— 2.00) diluted at 0-5 and CFU 0001-0001 added to the test mixture with complement = AR (yoy

Pa uses human serum that has no detectable bactericidal activity as an exogenously sourced supplement. Sources of the supplement are tested for suitability against each individual test strain. A complement source is used only if the number of live bacteria in the comparison examples without added immunosera is >775. Ten unique complement sources are needed for the 5/58 procedure described in this study.

After incubating for 910 minutes at 717 * deuterium with 70 002; 0-085 is added to the reaction mixture and the complete affinities are transferred to microfiltration dishes filled with 0 GCK 75 media. Microtiter plates are filtered; and incubated overnight at 1?” cloaca with 75 002 microcolonies spotted and quantified. Serum bactericidal titers are determined as a dilution of the elicited serum CFU resulting in a CFU decrease of +75 compared to the CFU in control eyes without immunosera. He specifies

Yo titer of SBA as titer of elicited dilution of test serum causing (alias) 75 in bacterial counts after incubation for 1 min at 717*E. SBA rated; fold or more for immunoglobulin 06 compared to the corresponding primary immunoglobulin sera. Negative sera against the test strain were determined at the initial dilution with a titer half the limit of detection for the test (i.e.; rabbit). 5._Example 2: Copying and displaying variants 05452086 non-fat derived natively derived amino acid arrangement 02086 mature according to the residues YAT-YY from (AOS) 1098250771 N. meningitidis >. by magnification of 04 from .genomic DNA hardens the raw material provided; that have the order TGCCATATGAGCAGCGGAAGCGGAAG (order ID: (YY) to the order of 5 Yo and contain the Ndel site of transcription. 3’ of the gene containing the TAG termination codon followed by the BamHI limiting locus The 799 bp amplified fragment is first cloned into a PCR2.1 (Invitrogen, Carlesbac, CA) intermediate vector that cleaves this 0 with 106l and 1a88© 07; and binds to the expression vector (Novagen, Madison, pET9a WI) Yo that cleaves with (BamHI Ndel) the resulting vector 01100 (which includes identifier 7

Arrangement #94) shows the mature subfamily P2086 5 of lineage 1 without the N-terminal cysteine (see ai Arrangement #: aa VY omitting the N-terminal Cys at position 1 or Tryptic identification number: 55 (which may reside in a lipid protein. The BLR(DE3) E. coli host mop uses hsdSB(rB-mB-) gal dem A(srl-recA)306::Tn10 (TetR ) (DE3)] © A (Novagen) To achieve the expression of fHBP, the same cloning steps are used to prepare the N-terminal omitting heterodimers (B03 2 Cys and Al12 (A04 B44 B24 B22 and 22m). Prepare Je = i is a terminal inequality that does not contain Cys in the same way as the raw materials provided, which 0 also includes a Cys codon (eg the first codon of the definitions of the order numbers: 11-1). Arrangements provided here Those skilled in the art can prepare forward and reverse primers for each of these variations.For example (JE) The following primers are used to enlarge the non-greasy variation 844 followed by cloning in 0198 using Blpl 5 Ndel. Table 1 =a] Wes] Name oa Input Raw Material Introduction | : 5 Y¢ TTTCTTc ccgggAAGGAGatatacatatg TGCAGCAGCGGAGGCGGCGG 3° entry raw sal | Yo 5’ TTTCTTgctcagcaTTATTGC reversed TTGGCGGCAAGACCGAT 3° Aye

q \ — — TTTCTTcccgggAAGGAGatatacatatg AGCAGCGGAGGCGGCGG 3° delete item TTTCTTgctcagcaTTATTGC | iid 5° L inverted TTGGCGGCAAGACCGAT 3' Results Non-lipid plasmid structures are strongly shown but variant forms of lipidic pe protein become 0 forms at the Cys terminal retarder no. See Example 8 and 9 which describe for example a way to show buildings. To overcome this pyruvylation Cys 00000 © omits the terminal WN See eg Example 10. However; Deletion of terminal Cys does not obviate the display of variants A22 and 822. See, for example, Fig. 4. With this the variations AOS 801 are also shown; and 844; Although the terminal Cys retarder WN is omitted see, for example, arrangement identification number: (ADS) VY where the terminal Cys is omitted not at position 1; arrangement identification number: Yo (terminal (BOL N) arrangement identification No.: YY (844), where the terminal Cys is deleted not at position 1. See V, for example, the Lo form. See Example 4. Example 3: Effect of Gly/Ser Stalk on Non-Lipidated Variant Expression to determine the cause of BOL (ADS and 844) in the absence of Cys terminal-no Vo and not showing variants A22 and 822; arrangements of these variants are aligned. The variants (AOS 801; and 844) all possess an extended chain of 01 or 11 retarded Gly Ser immediately after the terminal Cys-N (ie; +58//A6 stalk). Gly and +58. Accordingly, the Gly/Ser leg of the variants A22 and 822 is lengthened by the introduction of additional Ser; Gly residues.

Heteromorphs with a long Gly/[Ser stem are prepared by the methods described in Example 2 using direct starting materials encoding the Gly/Ser stem with either 01 or 11 retarded Ser; Varieties A22 and 822 show long-stemmed 11-01 (Gly/Ser retarded (Gly/Ser) with Cys-deleted terminals-no overexpression than variants A22 and B22 © Short-stemmed Gly/Ser (1 residues) omitting Gly/Ser—terminal Cys-no. These levels of expression, however, are still ALE when compared to levels of the AOS heteromorph B01 and 544. Example 4: Codon Optimization of 600017 The expression of the non-fatty variant 809 is unaffected by the deletion of the Cys-terminal retarded 0 (see definition of order number: OA deleted cysteine aie at position 0; or arrangement identification number: 4. See examples of Fig. 6. Evaluation of the arrangement of variant isoform 809 shows that variant isoform 809 has a [Ser]Gly stem consisting of 1 SerGly residue similar to a stem Gly/Ser in variants 2 822. Indeed, the L-terminal tails of variants 809 and 822 are symmetric at the camino acid level. The L-terminal tails of variants 8509 5 A22 (order definition 0) number: oF and order definition number: oY respectively); on any (Ja) that varies at the nuc level leic 0 with two nucleic acids 15: nucleic acids and 39 of arrangement identification number: A “lal” is an example of form 1. The first amino acids 04 of the L8-terminal tail of the variant form B22 are identical for variants 809 and 822; The terminal tail-l! For isoform 822 it differs only at the fifteenth amino acid. The 1-42 genucleic acids of the variant 822 are identical to the nucleic acids 1-42 0 of the variant of A22. The 1-42 nucleic acids of the variant of the form B22 (see identification of order number: ( OF is identical for 1-42 nucleic acids from 809 (see definition of arrangement no.: oF) but differs at nucleic acids 5 and 39; when aligned optimally. Accordingly » B22 differs from 509 at 15 acids 801100 and 39 from the definition of order number: 8. This last sentence contains an error.

q ¢ —_ _ typographic You must state that B22 differs from 809 at 15 nucleic acids and 39 from the order definition A tad) to determine if the nucleic acid differences affect the expression level of 809 compared to 822 5 (B22) The variants 822 and 822 are mutated by a point mutation to fuse nucleic acids 15© and 39 into the corresponding codons for Gly5 and 713a6. The introduction of these inactive mutations for the nucleic acid significantly increases the expression of the variants A22 and Cys-deleted 822 to levels similar to the Cys-deleted 809 variant, see Se Fig. 7. Accordingly, ld make the codon ie Jal add variant 809 can increase the expression of non-lipid P2086 variants of the Vo-deleted 5-N-terminal Vo. Further analysis of the non-lipid variant rearrangements suggests additional Jaa codons are optimized in the Gly/ stem. Ser to improve display. Gly Ser is optimized for any of the following arrangements: ATGAGCTCTGGAGGTGGAGGAAGCGGGGGCGGTGGA (SEQ ID NO: 28) M SS 5 GG G G G 5 G G G 6 (SEQ ID NO: 29) ATGAGCTCTGGAAGCGGAAGCGGGGGCGGTGGA (SEQ ID NO: 30) 0 M S S 6 5 6 8 G 6 G 68 ID NO: 31) ATGAGCTCTGGAGGTGGAGGA (SEQ ID NO: 32) tyov

q q — M S S G GG G G (SEQ ID NO: 33) ATGAGCAGCGGGGGCGGTGGA (SEQ ID NO: 34) M S S G G G G(SEQID NO: 33) SEQ ID NO) = order identification number) o EXAMPLE 5: Immunogenic Composition Formulation Optimization Al Vaccines prepared with ISCOMATRIX induce a rapid immune response leading to a reduction in the doses required to achieve a response rate greater than ; Fold as measured in SBA groups of 0 Rhesus macaques were incubated with different preparations of rP20 86 z non-divalent fatty acid Ye. The vaccine includes a non-pyruvylated non-lipidized AOS variant (SSID: VV deleted from Cys-terminal at position 1 or SCID: 00 encoded by SCID: 02) and a variant 844 non-pyruvylated non-fatty (configuration definition no: 71 omitted Cys-terminal-no at position 1 or sequence identification number: 4; encoded by sequence definition number: 1©.) The units of matter are The auxiliary sald units for 1004m shown in Tables 5-7 are shown as units in milligrams; therefore, they are shown as YO ( mg) vs. YO μg. pf 0 wk with attrition at 0 wk 11, 77 cf. No increases in SBA titers after the first dose for either group. After the second dose; 0 observed increase in SBA titres and number of respondents as determined by increments;fold in titres 58/8 above baseline for formulations containing adjuvant 5000//8114176A.The tables provide SBA GMTs ¥5 1 observed for AD Family A duc jill and 8 .fH BP The SBA 5 of ISCOMATRIX formulations is 1-7 times greater than that noted in YE 3.

_a q _ of AIPO4 preparation for strains of family A due dll and 8, respectively. Increased titres are also observed after the third dose of ISCOMATRIX preparations at 117-5 and 1-01 for subfamily A strain and 8 fHBP, respectively, compared with 81004. Analysis of response rates; by selection in increments; fold or more in the SBA titer above the sacl line) revealing a similar tendency (Table; and *). Table 7: SBA titers (15/A6) obtained against the immunosera of 102086 008 family de yl strain of rhesus macaques immunized with different preparations of the rP20 86 z bi-ala adjuvant vaccine. Geometric mean For the (GMT) Gl | fatty | ISCOMATRIX® AIPO4 | week | week | week | week zero |; y 71 m 0 I — — — + +++ I Yo — — + ++ 44 m Yoo — — — ++ ++++ 1 WYO — — + +++ Monkeys in each group; Immunization program: 0 + 6 ye weeks; All Zero 6+ Ted and 77 weeks. MnB M98 250771 abs for SBA test ¢A > n_n b"m0(b" except 7 1 7 < "+" «0 1 Y—1 7 9 "4" ¢ 1 0 Table: F (GMTS) SBA titres generated against an immunosera of strain LP2086 008 family H7

de yl) of 8 rhesus macaques immunized with different formulations of rP2086z divalent adjuvant ligand Geometric mean titer (GMT) Gl | fatty | ISCOMATRIX® AIPO4 | week | week | week | Week Zero ¢ 1 YA Tt 01 = = = Bad I A 01 = = Bad I 04 pm Yoo = = = Bad Jab Yoo Yo = = ++ ++ + monkeys in each group; Immunization program: 0 + 6 ye weeks; All Zero 6+ Ted and 77 weeks. Strain 127 CDC1 1008 for SBA Test A Russian Lapis Besiere ie HH Nor meat SVY > b+ table;: the number of rhesus macaques whose SBA titer rises by > ; fold using strain LP2086 1008 subfamily A Gl | ISCOMATRIX® AIPO4 | week | week | week | Week 0 ¢ 1 YA 0 iyov

q A —_ _ ha 01 - zero © o Yo “YO zero zero o Y A05/B44 You - zero zero 2 o You “YO zero zero o Y Table 0 Number of rhesus macaques in which the SBA titer increased by > ; fold using strain 102086 MNB subfamily B Gl | fatty | ISCOMATRIX® AIPO4 | week | week | week | Week 0 ¢ 1 y 88 and “Yo - zero zero 5 5 Yo” Yo zero zero 2 2 A05/B44 Yoo = zero zero; 3 You “YO zero zero o Y Example 6: Immunoprotection conferred by Lipidated and Non-Lipidated Variants Strains presenting different HBP haplotypes (from about 785 to about >2957 in homology) for 02086a rearrangements.

Prepared with LAIPO4. See Table 1 showing SBA response rates for the 1/08 fHBP strain of Family 8 generated by the 7188 bivalent vaccine. The non-lipid vaccine (denoted by a “-” under the “lipidation” column) includes 1 µg of each protein of a non-pyruvylated non-lipid AOS variant (order identification number: VV omitted of it Cys-terminal (no at position 1) and variation 544 non-pyruvylated non-lipid (order identification number: YY omitted from Cys-terminal not at position 1) by an alternative method. The synthetic non-lipid variant of P2086 (B44) induced greater immune responses as measured by SBA titer than the lipid variant (801) against strains bearing similar and different (homogeneity) LP2086 rearrangements (Jil > 797). Higher 0 response rates (as defined by increments; folds or more in SBA titres above baseline) are observed for the 844 rP2086 non-fatty vaccine compared to the lipid rLP2086 BOL vaccine (Table 1) according to Table 6 The non-lipid 844 is a preferred component of the 8 subfamily for THBP in combination to provide broad coverage against (eg producing bactericidal antibodies against) multiple 2086 heteroform strains. The strains of the heteromorph 809 L P2086 is particularly unlikely to have positive SBA response rates for polypeptides (04172086 heterodimer (ue-509)). In particular; The inventors found LP2086 9 to be exceptional for an elective strain described immunogenic formulation 805/5844 in Table 1 as producing bactericidal antibodies against it. So; In a preferred embodiment the immunogenic yo formulation of the invention includes polypeptide 809; particularly in the context of a formulation containing more than one polypeptide ORF2086 of subfamily 8. In a preferred embodiment; The Immunogenic Formula 844 Non Fatty also includes Polypeptide 809 Non Fatty. Table 1: SBA response rates to MnB strains 1180 subfamily 8 produced by immunoserosol 1180 bivalent vaccines from rhesus macaques Lao yam 7 #7 Shallinhdla ol a Help habit Heterogeneously defined with respondents

PD3 to i tll LP2086

YU from subspecies week testing for a non-greasy vaccine component

Va - 4 AH m MY + 0501 AD - 803 AIPO4

As - mg 4 e 8

Jaa ALY + A05/B01 809 4 AH - Safar

Yo ALY + 1 B15

Ar - 4E zero 1/8 + 1 816 8 - 4E zero 1/8 + 1 1 816 . - 4E AoA row + 1 and 1 824 . - 4E 7 Ah

0 \ — \ — 1 A + A05/B01 B44 4E - You You Ya - 4 AOS ISCOMATRIX® V+) You Ya - 4 AOS ISCOMATRIX® (001 μg) ) ISCOMATRIX® 22 m4 - km 01 Ae (μg) ISCOMATRIX® 22 m4 - km 0 (001 μg) monkeys per ic gana ¢ Immunization program: 0 + 6 Ye a week; Zero bleeding program 6 + Ted and 9 weeks. Jt 7: Codon Optimization of the B44 and 809 Variants Although the levels of expression achieved in the previous examples are appropriate for many uses; the codon Jaa in an additional optimal form is required; Constructs showing E. coli codon has additional optimal codons across the full length of order 00018:0. One of those arrangements optimized to show protein 844 without the Cys turned out to be the nucleic acid arrangement mentioned in Arrangement Definition: LEY as shown in Example 9; The display structure that contains the definition of order number: ?; An overexpression is shown compared to that of a non-optimal wild-type arrangement. L

17

Showing protein 809 with no Cys-terminal omitted is improved by making changes to order 0 from structure 844 (order ID: £7) recited above to 809 (order ID: 8;). To generate 809 rearrangements as tie ¢ The DNA sequence present in an optimal form of order 844 (order identification number: £7) first lines up with the BOO allele Jay DNA sequence (order definition © number: 8?). The total non-lipid coding order becomes BOO allele Jal (order definition: EA) is optimized to reflect visible codon changes in optimal allele 844 (order definition: 47) whenever the acids 800100 are between 844 (order definition: ; 4) and 809 (order identification number: £9) is symmetric. The 60000 rearrangements in allele 809 corresponding to amino 5 homologs between the BO9 allele and allele 844 are altered to reflect the codon used in the 0 full order 4 (Order ID: (EY) The codon arrangements for amino acids that vary between 809 (Order ID: £9) and 844 (Order ID: 44) do not change in

DNA arrangement and 809. (Lil) The non-fatty B44 amino acid sequence (order identification number: ; ;) contains 1 consecutive serine—glycine repeat sequence (S-G-G-G-G) (order identification number: 07) (see 1 Also 2 amino acids to 6 of order identification number: 4 4) at the -no end; while the 509 allele contains only one serine-glycine repeat at the -no end (see 2 acids 800100 to 6 and 7 acids 80100 to 11 of the sequence identification number: 49). : ;4) also have a different codon usage (see Y. 00016000654 to 18 and 19 nucleotides to 33 from sequence definition #47) and different conjugates of the optimal serine-glycine 844 repeat i) either 4 nucleotides to 18 of the order identification number: ;; or 19 nucleotides to 33 of sequence identification number: fF or a combination thereof) also used for sequence identification DNA 809 (seq identification number: Ole fA used for nucleotides 4 to 18 of sequence identification number: ($A) in order to test the effect on Show

protein Yo Synthesized. (yoy

-1 0".- ? Various models of the 8509 were constructed in an optimized way: Sequence ID: £0 contains both repeats of (GS1) serine—glycine and (652) 4 nucleic acids (4 to 33) of Sequence Definition #: Arrangement ID: £1 (of 844 optimum” contains Arrangement ID No.: £7 651) 4 nucleic acids to 18 Arrangement Identification No. ¢Y (contains Arrangement Identification No. ai: GS2 ¢v 19) nucleic acids © to 33 of Arrangement Definition No.: 47. The DNA of all sequences sequenced for the above codon is synthesized chemically by standard methods used in the art of chemistry. The resulting DNA is transcribed into standard 01850010 display vectors and has been tested for display in EL coli family cells as described in Example 8 and Example 9. Example 8: Method for Expressing ORF2086, 509 variant 0 WA transforms E. coli K-12 (Derivatives of wild type W3110 (CGSC4474) have removals in (araA fhuA recA with 05063 plasmid includes sequence definition tad) pEBO64 (£0 includes sequence identification number: 47 058065 plasmid includes arrangement identification number: ty or 4sa plasmid includes arrangement identification number: LEA The modification The favored Yo of strain K-12 is useful for fermentation purposes but is not required for the expression of proteins Cells are inoculated into a media defined by —glucose salt. After A hours of incubation at 17 “cloaca, linear glucose feeding is used, and the incubation continues for ? Extra hours. Isopropyl 3-D-1-thiogalactopyranoside (IPTG) was added to the culture to a final concentration (Ake +) gram moss, followed by incubation for 17 hours at 17*Come. Cells were collected by centrifugation at “1” 1,136,888 specific gravity for 10 minutes and decompose with Cell Lysing Kit" from Lienco Technologies (St.

Louis, MO) ™ 56NO-L85 and loading pH stabilizer. Pure lysates for show 809 stain 156 for SDS-PAGE gel analysis and/or Western blotting were quantified with a scanning densitometer. The results from scanning densitometry are shown below in Table 7: L

0 _ \ _ table : Show data in E. coli protein, plasmid family cells, homozygous ratio for total cell protein at VY 1 hour after IPTG induction, as measured by SDS-PAGE. Scanning densitometry E. coli K- 809 | 8063m YAR 12 Arrangement ID No.: £0 809 M E. coli | 8065m JAY 12 0 Arrangement Identification a) A coli K- 809 .8064m 1 12 Arrangement Identification No: £7 809 Ha ARS pLA134 | E. coli 12 0 Configuration definition a) Example 9: Method for Expressing ORF2086 B44 variant (Method for Expressing ORF2086, B44 variant) BLR(DE3) cells with 01014087 (plasmid definition includes arrangement number: SEY) Cells are inoculated into a defined medium with –glucose salt. After A hours of incubation at 17” cloaca a linear glucose inoculation is used and the incubation continues for ? additional hours Isopropyl 3-D-1-thiogalactopyranoside (IPTG) is added to the culture to a final concentration of

mg \_ 8 mM g followed by incubation for VY 1 hour at 17*Choice. Cells were collected by centrifugation at “1,136,888 specific gravity for 10 minutes and lysed by adding Cell Lysing Kit” from Lienco Technologies (St.

Louis, MO) ™ 56NO-L85 and loading pH stabilizer. The supernatants were analyzed for blotting by Coomassie © 809 stain for SDS-PAGE gel analysis and/or Western blotting and quantified with a scanning densimeter. The results from the scanning density measurement are shown below in the tA table. Table 8: Show data in E. coli protein, plasmid family cells, homozygous ratio for total cell protein at VY 1 hour after IPTG induction as measured by SDS-PAGE, scanning densitometry yA pLNO56| E.coliB B44 Arrangement Identification No.: 51 pDKO87 | E. coli K- B44 h12 Arrangement Identification No.: EY Example 10: Pyruvylation The current example shows that the Cys-terminal retarder in ORF2086 proteins Non-fatty can become pyruvylated when exposed; Sie in E. coli Ya monitors heterodimer protein accumulation during production of AOS heteromorphs (order identification number: VF B44 (identification Order #: (Y) using reverse high-performance liquid chromatography (RP-HPLC) Shall this chapter overlap with QTOF-MS to provide a means to monitor the formation of product-specific anisotropies L

0 - \ _ after expression in cells of 8 coli family 5. and/or (K-12) the products derived from these fermentations undergo a purification procedure during which modification of the product is observed. mass +70 daltons; compared to original products of 68 YYTE and 7727/7 daltons for AOS and 844°, respectively.° Published literature indicates that mass transfer of 700+ daltons has been previously observed in proteins and traced back to the pyruvylation of the .amino—_i kl residue The presence and location of the pyruvate clusters is confirmed using mass spectrometry (MS/MS) segmentation data The data indicates that the modification was on the camino— i,k cysteine retard i.e., acid 200100 at position 1 according to the ADS order 844. For 805, the 0-term percentage of pyruvylated polypeptides is about ZY compared to the total number of order 05M polypeptides (order definition no. : LVF of 844 did for pyruvylated polypeptides is about © #7; compared to the total number of B44 polypeptides (order definition number: YY) when purifying the AOS variants (definition Arrangement No.: VY omitted at position 1 0 retarded Cys terminal-no or Arrangement Identification No.: 00) and 844 (Arrangement Identification No.: YY omitted Cys terminal-la at position 1 or Arrangement Definition No.: 44); which do not contain Cys (N= djl there is no pyruvylation definable (Yet daltons). Example 11: Immunogenicity of B09 and B44, individually and in combination Arrangement #: £9 encoded by Arrangement Identification No. EA: ) or a variant B44 (Arrangement Identification No. [m] encoded by Arrangement Identification No.: EY or AOS 809 and 5844 (Arrangement Identification No.: 00) Arrangement ID#: £9 encoded by Arrangement ID#: (EA and Arrangement ID#: 44 encoded by Arrangement ID#: 47; respectively) formulated with YOu 1004 μg per dose.

0 7 _ \ _ monkeys intramuscularly at 4 weeks yolk As with 01 micrograms each of non-fatty THBP alone or in combination as mentioned in Table 5 and Table 01. Serum samples of each of week 0 and 11 on 58/5 against MNB strains with HBP variants either from family A de dl) or from subfamily 8. Respondents are scored as animals with an excess of ; times per caliber. The oo variant 844 tested is the conformational construct (order identification number: ?4) and the widespread response rates observed in previous studies (table above) were retained for the homologous construct (Table 4) of vaccine 844 alone or in combination with 809 Vaccine 809 alone (Table 01) can also generate transient, highly active immune responses (Table 01). the estrous ratio to increase > ; Times against a variant of the ¢PD3 test) Mick Rogerram/ | macaque per group (protein B24 B16 B44 AOS 809 order |rank | order | el | el number: Y 0) number: (Y 1 number: 0 7) number: A tad) (Y 0 0( Ye ta) A 1. B44 + 809 5 + 01 rhesus macaques (n = 01) immunized in muscle at weeks ja ¢ ; Ag With 01 micrograms each of non-fatty THBP alone or in combination as indicated in the vaccine column of the formulation with YOu 1004 micrograms. Serum samples for both weeks 0 and 10 were analyzed on 58/5 against 1 strains 18 / mentioned in the table.The respondents are recorded as animals with a 2 times increase in titer.UH 7

m 0 \ — Table 3 indicates for example; indicated that a formulation containing a combination of AOS 5 809 (B44 without pyruvylated without lipidation) showed greater transient coverage against the tested variants as compared to the transient coverage of formulations containing Unit 844. In light of the results shown in the present application, which are included in particular in Table T and Table 4 together, the comprising compositions (B44 85809 and ©05) singly or in combination are preferred embodiments of the present invention. In particular, the comprising compositions of both 844 and 809 are shown herein. Preferably, that composition additionally includes the polypeptide subfamily Table 01: Resultant response rates to non-fat 509 fHBP vaccine in rhesus macaques (01 micron) vaccine | Ratio of increase > times against a test variant of crawfish/ protein 0 z AD vaccinated in rhesus macaques (n = 0) intramuscularly at yolk weeks; Ast with 01 μg each of non-fatty THBP alone or in combination as indicated In the vaccine column of the preparation with You μg of 1004. Serum samples for both weeks 0 and 10 were lysed on 5/58 against the MNB strains mentioned in the table. BON as animals with an increase of ¢ times in caliber. Vo Example 12: Immunoprotection conferred by Lipidated and Non-Lipidated Variants construct A 01 dal New Zealand white rabbit is pre-examined; Y.o—Y.o kg ; from Charles River Canada by whole-cell ELISA and 01 animals are selected for this study on the basis of their 0 baseline titres against the test strains (isoform fHBP 802 L).

— \ 0 q —

(Arrangement Definition No.: B44 5 (V1) (Arrangement Definition No.: 71) (Table 11.)

* Animals intramuscularly with 100 μg each of protein prepared with 0 © μg

ISCOMATRIX in dose *#.; milliliters at yellow weeks; and 9 (Table 11).

Group 1 with non-fatty B44 (order definition No. ¢£). The study includes a comparison group © inoculated with 801 lipid prepared with YOu 81004 μg). Rabbits drain at weeks

of hua 9 and 10. Separate sera were prepared from week 01 and analyzed with SBA against cocci strains.

meningitis multiple serogroup 8 of subfamily 8 118.

Table 11: Rabbits used in the study

Type: rabbit

Breed: New Zealand white

Charles River Laboratory a: Source

Number of animals in the group: 1

Total number of animals: 9

Age and gender: female

Weight: 0.-09.? kg

11 tables

Aluminum | ISCOMATRIX HBP | number | Adipose form 1 | group

Phosphate | [a as black] | Animals | contrast (µg/ icp microgram/ v0 i ce v0 icon mL) 5 milliliters) (ali char

RYO

I here for 01 93 4 9; Depletion Program: Yellow Weeks of Immunization Program: Yellow Weeks: (BSA) (Serum Bactericidal Assay) Small-colony-based sero-killing test against SBA group meningococcal strains Human subjects are tested Jad) on single serum samples. Multiple VF serotype 8 (Table 8) qualifies as the complement source for the tested strain in the test. The bactericidal titers based on antibody-0 antibody through-complement are elicited and expressed as the inverse of the dilution of the test serum that kills N4. SBA from meningococcal cells in the test. The limit of determination for the test is the titer ; gives a number of 7. Calculates and compares an increase of > 8/58 times in titres in sera of < SBA titer compared to titers in pre-exhaustion samples. 01 week is Alternative SBA Serum bactericidal antibody activity as measured in Ya Immunization with 1185 non-lipid 5508 SBA immunoglobulin for protection against meningococcal disease Determines the level of antibody in a sample SBA measures production of bactericidal antibodies in rabbits Measures serum simulating bacterial degradation due to naturally occurring complement Rabbit serum samples 802 or 844 fHBP against strains with SBA analyzed by 01 collected from week They appear every 01 week after the third immunization (week OF) as shown in Table 018©6. 1

B44.113 The serum samples appeared to have bactericidal activity against each of the test strains. Non-fat table (definition order number: £6) more immunogenic than 801 non-fat in New Zealand rabbits. 801 Jie (N4/N5) against these strains. N 844 is in the same subgroup as N 844 non The liposomal (definition of arrangement No.: ;;) prepared with adjuvant 1500171811 gives against these strains. Aluminum phosphate titers similar to the lipid form 801 with Yo sera before depletion from rabbits generally show no bactericidal activity by antisense. Selected strains. 7 AH

arr

In rabbits THBP 8 table ¥ 0) bactericidal activity in serum against subfamily strains

Bae non-fatty New Zealand white THBP inoculated with tested antibody variant GMT SBA titer tad) variant form of subfamily | 844 (definition of arrangement no: | 802 (definition of arrangement no.) 3 (71 (preparation) 8 non-greasy (definition of | 117758 6 to 4) ( $e arrangement no. { scomatrix) 017 Yo | non-greasy BOI { scomatrix) non-fatty in New Zealand white rabbits H binding factor proteins Example 13: Immunogenesis of six Vaccinated a group of 0 rabbits with heterogeneous morphologies ©1188 Non-fatty as described in Vaccines are given at 0 weeks 4, and 9. Analysis of pooled rabbit serum samples VE Table 0 against strains with 1180 homozygous and heterozygous rearrangements SBA from weeks 0 and 10 are shown by the percentage of respondents after the third immunization One week after the third immunization VE schedule (week 10); all serum samples show bactericidal activity against congenic strains as well as sera previously depleted from FHBP other test strains of the same subfamily rabbits show no preexisting bactericidal activity generally against different breeds. 0 of post-dose responders in New Zealand white rabbits inoculated with 11805 non 14:77 Table 84. Haniyeh a tyov

-17-

fHBP | H.. Hmm AA EERE - 7 53 for Sala -2-11 [A for Thanath MENEN | EEE Wu | | | The CC EERE a [=| 01 | 44 Taos b TT — —T AL2 μg 2 all one/ser | 4

micrograms

Total MNB fHBP Proteins used

Ah

-1- Configuration identification number: 0 where Cys is still at position 1 Definition of arrangement number: (V0 where Cys is still at position 1 809 Configuration identification number YA: ¢ where is Cys at position 3 or sequence identification number: £9 encoded by sequence identification number: m sequence identification number: 109 where Cys at position 1 is still B44 sequence identification number RAVENS BH is still Cys At position 3 or order identification number: 4 encoded by order identification number: 0) dle forms of test in Table 16: AOS B44 B24 B16 809 12m A22 order | to arrange | to arrange | To order the order the order the order (0 o tad) (0 2 number: 0) Y number: (Y 1 tad) (Y 0 number: 7 0 number: (0 A tad) 14: Example 00 is non-lipidic (Arrangement Definition No.: 5

SSGSGSGGGGVAADIGTGLADALTAPLDHKDKGLKSLTLEDSISQNGTLTL

‎SAQGAEKTFKVGDKDNSLNTGKLKNDKISRFDFVQKIEVDGQTITLASGEF ‏ه‎ ‎QIYKQDHSAVVALQIEKINNPDKIDSLINQRSFLVSGLGGEHTAFNQLPSGK ‎AEYHGKAFSSDDAGGKLTYTIDFAAKQGHGKIEHLKTPEQNVELASAELKA ‎DEKSHAVILGDTRYGSEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIA ‎GKQ ‎(10 ‏(تعريف الترتيب رقم:‎ pPEBO42 ٠ troy

-115-

ATGAGCTCTGGAAGCGGAAGCGGGGGCGGTGGAGTTGCAGCAGACAT

TGGAACAGGATTAGCAGATGCACTGACGGCACCGTTGGATCATAAAGA

CAAAGGCTTGAAATCGCTTACCTTAGAAGATTCTATTTCACAAAATGGC

ACCCTTACCTTGTCCGCGCAAGGCGCTGAAAAAACTTTTAAAGTCGGT

GACAAAGATAATAGCTTAAATACAGGTAAACTCAAAAATGATAAAATCT ‏ه‎ ‎CGCGTTTTGATTTCGTGCAAAAAATCGAAGTAGATGGCCAAACCATTAC ‎ATTAGCAAGCGGTGAATTCCAAATATATAAACAAGACCATTCAGCAGTC ‎GTTGCATTGCAAATTGAAAAAATCAACAACCCCGACAAAATCGACAGCC ‎TGATAAACCAACGTTCCTTCCTTGTCAGCGGTTTGGGCGGTGAACATA ‎CAGCCTTCAACCAATTACCAAGCGGCAAAGCGGAGTATCACGGTAAAG ٠

CATTTAGCTCAGATGATTGCAGGCGGTAAATTAACTTATACAATTGACTT

TGCAGCAAAACAAGGACATGGCAAAATTGAACATTTAAAAACACCCGAA

CAGAACGTAGAGCTCGCATCCGCAGAACTCAAAGCAGATGAAAAATCA

CAGGCAGTCATTTTGGGTGACACGCGCTACGGCAGCGAAGAAAAAGG

TACTTACCACTTAGCTCTTTTTGGCGACCGAGCTCAAGAAATCGCAGGT 0

AGCGCAACCGTAAAGATAAGGGAAAAGGTTCACGAAATTGGGATCGCG

GGCAAACAATAA

TT is non-lipidic (Arrangement Definition No.: 2

SSGGGGSGGGGGVAADIGAGLADALTAPLDHKDKSLQSLTLDQSVRKNEK

LKLAAQGAEKTYGNGDSLNTGKLKNDKVSRFDFIRQIEVDGQTITLASGEF 00

QIYKQNHSAWALQIEKINNPDKIDSLINQRSFLVSGLGGEHTAFNQLPDGK

AEYHGKAFSSDDPNGRLHYSIDFTKKQGYGRIEHLKTPEQNVELASAELK

ADEKSHAVILGDTRYGGEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGI

AGKQ

TV (Arrangement ID: 05858043 Yo Insomnia

-11 AH -

ATGAGCTCTGGAGGTGGAGGAAGCGGGGGCGGTGGAGTTGCAGCAG

ACATTGGAGCAGGATTAGCAGATGCACTGACGGCACCGTTGGATCATA

AAACAAAAGTTTGCAGTCGCTTACCTTAGATCAGTCTGTCAGGAAAAA

TGAGAAACTTAAGTTGGCGGCGCAAGGCGCTGAAAAAACTTATGGAAA

© CGGTGACAGCTTAAATACAGGTAAACTCAAAAATGATAAAGTCTCGCGT

TTTGATTTCATTCGTCAAATCGAAGTAGATGGCCAAACCATTACATTAG

CAAGCGGTGAATCCAAATATATAAACAAAACCATTCAGCAGTCGTTGC

ATTGCAAATTGAAAAAATCAACAACCCCGACAAAATCGACAGCCTGATA

AACCAACGTTCCTTCCTTGTCAGCGGTTTGGGCGGTGAACATACAGCC

TTCAACCAATTACCAGACGGCAAAGCGGAGTATCACGGTAAAGCATITT 0

AGCTCAGATGATCCGAACGGTAGGTTACACTATTCCATTGACTTTACCA

AAAAACAAGGATACGGCAGAATTGAACATTTAAAAACGCCCGAACAGA

ACGTAGAGCTCGCATCCGCAGAACTCAAAGCAGATGAAAAATCACACG

CAGTCATTTTGGGTGACACGCGCTACGGCGGCGAAGAAAAAGGTACTT

ACCACTTAGCCCTTTTTGGCGACCGCGCTCAAGAAATCGCAGGTAGCG 00

CAACCGTAAAGATAAGGGAAAAGGTTCACGAAATTGGGATCGCGGGCA

AAACAATAA

2 Non-Lipidic (Arrangement Identification No.: (TA

SSGGGGVAADIGAGLADALTAPLDHKDKSLQSLTLDQSVRKNEKLKLAAQ

GAEKTYGNGDSLNTGKLKNDKVSRFDFIRQIEVDGQLITLESGEFQIYKQD 0

HSAVVALQIEKINNPDKIDSLINQRSFLVSGLGGEHTAFNQLPSGKAEYHG

KAFSSDDAGGKLTYTIDFAAKQGHGKIEHLKTPEQNVELASAELKADEKSH

AVILGDTRYGGEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIAGKQ

8 pm (Definition of arrangement No. 19) Ay

-11-

ATGAGCTCTGGAGGTGGAGGAGTTGCAGCAGACATTGGAGCAGGATT

AGCAGATGCACTGACGGCACCGTTGGATCATAAAGACAAAAGTTTGCA

GTCGCTTACCTTAGATCAGTCTGTCAGGAAAAATGAGAAACTTAAGTTG

GCGGCGCAAGGCGCTGAAAAAACTTATGGAAACGGTGACAGCTTAAAAT

ACAGGTAAACTCAAAAATGATAAAGTCTCGCGTTTTGATTTCATTICGTC©

AAATCGAAGTAGATGGCCAACTTATTACATTAGAAAGCGGTGAATTCCA

AATATATAAACAAGACCATTCAGCAGTCGTTGCATTGCAAATTGAAAAA

ATCAACAACCCCGACAAAATCGACAGCCTGATAAACCAACGTTCCTIC

CTTGTCAGCGGTTTGGGCGGTGAACATACAGCCTTCAACCAATTACCA

AGCGGCAAAGCGGAGTATCACGGTAAAGCATTTAGCTCAGAATGATGCA 0

GGCGGTAAATTAACTTATACAATTGACTTTGCAGCAAAACAAAGGACATG

GCAAAATTGAACATTTAAAAACACCCGAACAGAACGTAGAGCTCGCAT

CCGCAGAACTCAAAGCAGATGAAAAATCACACGCAGTCATTTTGGGTG

ACAGGCGCTACGGCGGCGAAGAAAAAGGTACTTACCACTTAGCTCTTT

TTGGCGACCGAGCTCAAGAAATCGCAGGTAGCGCAACCGTAAAGATAA 0

GGGAAAAGGTTCACGAAATTGGGATCGCGGGCAAACAATAA

ACI46T789.1 Gene Bank: L(V) (Arrangement Identification No.: 2

CSSGGGGVAADIGAGLADALTAPLDHKDKGLQSLTLDQSVRKNEKLKLAA

QGAEKTYGNGDSLNTGKLKNDKVSRFDFIRQIEVDGKLITLESGEFQVYKQ

SHSALTALQTEQVQDSEDSGKMVAKRQFRIGDIAGEHTSFDKLPKGGSAT 0

YRGTAFGSDDAGGKLTYTIDFAAKQGHGKIEHLKTPEQNVELASAELKAD

EKSHAVILGDTRYGGEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIAG

KQ

VY is non-lipidic (Arrangement Identification No.: 2 ay

-119-

SSGGGGVAADIGAGLADALTAPLDHKDKGLQSLTLDQSVRKNEKLKLAAQ

GAEKTYGNGDSLNTGKLKNDKVSRFDFIRQIEVDGKLITLESGEFQVYKQS

HSALTALQTEQVQDSEDSGKMVAKRQFRIGDIAGEHTSFDKLPKGGSATY

RGTAFGSDDAGGKLTYTIDFAAKQGHGKIEHLKTPEQNVELASAELKADE

© KSHAVILGDTRYGGEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIAGK

Q

Important VY (Arrangement Definition No.: 4

ATGAGCAGCGGAGGGGCGGTGTCGCCGCCGAACATCGGTGCGGGGC

TTGCCGATGCACTAACCGCACCGCTCGACCATAAAGACAAAGGTTTTGC

AGTCTTTAACGCTGGATCAGTCCGTCAGGAAAAACGAGAAACTGAAGC 0

TGGCGGCACAAGGTGCGGAAAAAACTTATGGAAACGGCGACAGCCTT

AATACGGGCAAATTGAAGAACGACAAGGTCAGCCGCTTCGACTTTATC

CGTCAAATCGAAGTGGACGGGAAGCTCATTACCTTGGAGAGCGGAGA

GTTCCAAGTGTACAAACAAAGCCATTCCGCCTTAACCGCCCTTCAGAC

CGAGCAAGTACAAGACTCGGAGGATTCCGGGAAGATGGTTGCGAAAC 0

GCCAGTTCAGAATCGGCGACATAGCGGGCGAACATACATCTITTGACA

AGCTTCCCAAAGGCGGCAGTGCGACATATCGCGGGACGGCGTTCGGT

TCAGACGATGCTGGCGGAAAACTGACCTATACTATAGATTTCGCCGCC

AAACAGGGACACGGCAAAATCGAACACTTGAAAACACCCGAGCAAAAT

GTCGAGCTTGCCTCCGCCGAACTCAAAGCAGATGAAAAAATCACACGCC 0

GTCATTTTGGGCAGACGCGCTACGGCGGCGAAGAAAAAGGCACTTA

CCACCTCGCCCTTTTCGGCGACCGCGCCCAAGAAATCGCCGGCTCGG

CAACCGTGAAGATAAGGGAAAAGGTTCACGAAATCGGCATCGCCGGC

AAACAGTAA

VY (Arrangement ID No.: 0016086 Yo Thinner

-11

ATGTCCAGCGGTTCAGGCAGCGGCGGTGGAGGCGTGGCAGCAGATAT

CGGAACAGGTTTAGCAGATGCTCTGACAGCACCCTTAGATCACAAAGA

CAAAGGACTTAAATCACTGACATTGGAAGATTCTATCTCGCAAAATGGT

ACTCTCACTCTTTCAGCCCAAGGCGCAGAAAAAACATTTAAAGTAGGC

GATAAAGATAACTCCTTAAATACAGGTAAATTAAAAAATGACAAAATCTC ‏ه‎ ‎ACGGTTTGATTTCGTTCAGAAAATTGAAGTAGATGGACAAACGATTACA ‎TTAGCAAGCGGCGAATTCCAAATTTATAAACAAGACCATTCAGCAGTAG ‎TAGCATTACAAATCGAAAAAATTAACAACCCGGACAAAATTGATTCTCTT ‎ATTAACCAACGCTCTTTTCTCGTATCAGGACTTGGTGGTGAACATACAG ‎CGTTTAATCAACTGCCGTCAGGAAAAGCAGAATATCATGGTAAAGCATT ٠

TTCATCAGACGACGCAGGTGGCAAACTGACCTATACTATTGACTTTGCA

GCAAAACAGGGACATGGAAAAATTGAACATTTAAAAACACCCGAACAG

AACGTAGAACTGGCCTCAGCAGAATTGAAAGCTGATGAAAAATCCCAT

GCAGTAATTTTAGGCGATACACGTTACGGTAGCGAAGAAAAAGGTACA

TATCACTTAGCTCTTTTTGGCGATCGTGCTCAAGAAATTGCTGGTTCCG 0

CAACAGTTAAAATCCGTGAAAAAGTACATGAAATCGGCATTGCAGGTAA

ACAATAA

VE (Arrangement Definition No.: 9

CSSGGGGSGGGGVAADIGTGLADALTAPLDHKDKGLKSLTLEDSIPQNGT

LTLSAQGAEKTFKAGDKDNSLNTGKLKNDKISRFDFVQKIEVDGQTITLAS 0

GEFQIYKQNHSAVVALQIEKINNPDKIDSLINQRSFLVSGLGGEHTAFNQLP

GDKAEYHGKAFSSDDPNGRLHYTIDFTNKQGYGRIEHLKTPELNVDLASA

ELKADEKSHAVILGDTRYGSEEKGTYHLALFGDRAQEIAGSATVKIGEKVH

EIGIAGKQ

Non-lipidic Vo (arrangement identification number: B22 Yo (voy

-14?-

SSGGGGVAADIGAVLADALTAPLDHKDKSLQSLTLDQSVRKNEKLKLAAQ

GAEKTYGNGDSLNTGKLKNDKVSRFDFIRQIEVDGQLITLESGEFQVYKQS

HSALTALQTEQVQDSEHSGKMVAKRQFRIGDIAGEHTSFDKLPEGGRATY

RGTAFGSDDASGKLTYTIDFAAKQGHGKIEHLKSPELNVDLAASDIKPDKK

© RHAVISGSVLYNQAEKGSYSLGIFGGQAQEVAGSAEVETANGIRHIGLAAK

Q

(PPW1 02) (v1) Non-lipidic acid (Arrangement Identification No.:

CGSSGGGGVAADIGTGLADALTAPLDHKDKGLKSLTLEDSISQNGTLTLSA

QGAEKTFKVGDKDNSLNTGKLKNDKISRFDFVQKIEVDGQTITLASGEFQI

YKQDHSAVVALQIEKINNPDKIDSLINQRSFLVSGLGGEHTAFNQLPSGKA 0

EYHGKAFSSDDAGGKLTYTIDFAAKQGHGKIEHLKTPEQNVELASAELKAD

EKSHAVILGDTRYGSEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIAG

KQ

VY is non-lipidic (Arrangement Identification No.: 5

GSSGGGGVAADIGTGLADALTAPLDHKDKGLKSLTLEDSISQNGTLTLSAQ 0

GAEKTFKVGDKDNSLNTGKLKNDKISRFDFVQKIEVDGQTITLASGEFQIYK

QDHSAVVALQIEKINNPDKIDSLINQRSFLVSGLGGEHTAFNQLPSGKAEY

HGKAFSSDDAGGKLTYTIDFAAKQGHGKIEHLKTPEQNVELASAELKADEK

SHAVILGDTRYGSEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIAGKQ

VA Compatibility Arrangement (Arrangement Definition No.: Ys

CSSGGGGVAADIGAGLADALTAPLDHKDKGLQSLTLDQSVRKNEKLKLAA

QGAEKTYGNGDSLNTGKLKNDKVSRFDFIRQIEVDGQLITLESGEFQIYKQ

SHSALVALQTEQINNSDKSGSLINQRSFRISGIAGEHTAFNQLPKGGKATY

Yes

— \ \ «=

RGTAFSSDDAGGKLTYTIDFAAKQGHGKIEHLKTPEQNVELASAELKADEK

SAVILGDTRYGGEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIAGKQ

ترتيب التوافق (تعريف الترتيب رقم: ‎(V3‏ ‎SSGGGGVAADIGAGLADALTAPLDHKDKGLQSLTLDQSVRKNEKLKLAAQ‏ ‎GAEKTYGNGDSLNTGKLKNDKVSRFDFIRQIEVDGQLITLESGEFQIYKQS ©‏ ‎HSALVALQTEQINNSDKSGSLINQRSFRISGIAGEHTAFNQLPKGGKATYR‏ ‎GTAFSSDDAGGKLTYTIDFAAKQGHGKIEHLKTPEQNVELASAELKADEKS‏ ‎HAVILGDTRYGGEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIAGKQ‏ ‏مثال 15: توليد أشكال متباينة غير ليبيدية من العائلة ‎A rP2086— duc il)‏ ‎٠‏ نسخ Non-lipidic fHBP genes The coding order for the non-lipidic AOS THBP protein (order identification number: 00) lines up to the 844-enhanced expression (order identification number: EY) wherever the amino acids between the two are the same. Use codon of 844 (sequence identification number: EY) to be substituted in gene 5. Recreate the fullest sequence at Genes “!06116; adding nuclease site domain 1 internal BamHI 5 Ndel at the no end “Cy in a row. The resulting gene (sequence identification no: 15) is subtranscribed into 01308 at these sites. The non-synthetic THBP 812 (sequence identification no: V1) of PEBO43 (sequence identification no. Number: (TY) Display of allele 812 by Blue Heron 100001065. This gene is enhanced by the same process used for 805 (PEB042) 0 additionally; the terminal amino codons Blue Heron enhanced with B44 1016 (amino acid residues ? to 11 of order identification number: £8) Jad terminal amino codons SSGGGG Jas .812 The original (1 amino acid residues to 1 from the definition of the arrangement No.: 17). The optimal arrangement is synthesized again at Celtek Genes adding La

-1?1- internal nuclease site domain BamHI; Ndel at the “Cy N” terminus, respectively. The resulting gene (seq identification no: (TV) is subcloned to 01308 at these sites. A synthetic non-lipidic A22 THBP (seq identification no: A) of 8058b is shown (seq identification no: 19). This is improved gene by the same process used for © 055042. Additionally, the terminal amino codons optimized Blue Heron with 544 0 (amino acid residues ? to 1 of sequence identification number: ££) replace the codons 6 22m original (1 amino acid residues to + from sequence identification number: 148) The Jad arrangement is synthesized again at Celtek Genes adding the BamHI 5 Ndel internal nuclease site domain at the no end. “Cy in a row. The resulting gene (0 sequence definition: 19) is subcloned into 01308 at these sites. Synthetic 10180 862 (order identification number: VY) of 018164 (order identification number: () is shown. VY the AG2_002 allele of strain 0167/03 by PCR amplification with an internal nuclease site domain containing primers BamHI;Ndel at the no “Cs” end respectively. The resulting gene ( Arrangement definition number: (VY to 01308 at this Vo sites. Example 16: Show off; fermentation rP2086 proteins dams family de jill of strains showing E. coli using the BLR(DE3) E. coli 8 recA mutant with 018164 (order identification no. tad) (VY) to show 862 (order identification number: 71). Plasmid pEBO42 (order identification number: 71). 15) to 800643 (W3110:DE3 ArecA AfhuA AaraA) E. coli Jile to give Ye strain 800660 to show 805 (order identification no: 00) show A22 (order identification no: (TA) of strain 8100592 consisting of 025058 plasmid (order identification number: 19) located in 80559 host (which repeats -W3110:DE3 ArecA AfhuA AaraA Lay at the end; 8043 plasmid 0 (order identification number: TV) is transformed into 80483 host (W3110:DE3 ArecA) to give strain 810540 to show 812 (arrangement identification number: 17).

-??1- Fermentation The expression strains are fermented in a minimum medium based on ©9110056. Overnight starter culture is inoculated into 01 liter fermenters operating at 1” aeration; 1 v/v/min aeration with dO burst control at 0 77. When batch glucose is depleted from medium sie) approximately = 000600 (Yo © limited linear glucose feeding starts at 7.8 g/L/h. Feeding continues until induction with IPTG dha Avis Ale +0) and during the subsequent protein expression period. To show ADS (order identification no: 00) strain 810660 is induced at Yo = OD600 and fermentation continues for V hours after induction (HPI) check show 822 (order identification no: (TA) and 8012 ( Arrangement identification No.: 17) of strains 810592 and 80540; respectively; by induction at 0 60 - 000600 and fermentation for 101 YE. At the end of fermentation cell pastes are collected by centrifugation. 2 (Arrangement identification No.: (VY) rP2086 proteins are produced as 5 soluble in cytoplasm (E.coli strains). A soluble cytoplasmic extract is typically produced by thawing frozen cells showing a specific heteromorph of subfamily 12086 in a deficient pH buffer. Vo pressure 01(mM) Hepes—NaOH pH Vat containing protease inhibitors and inactivated cells in a Microfluidizer under a pressure of approximately 0,008,691 psi. DNAse RNase was added to digest Nucleic acids and cytoplasmic extract were collected as supernatants after centrifugation at low speed to remove any unbroken cells and then high speed (> JB specification) to remove membranes; cell walls and other larger subcellular components . The cytoplasmic extract was further filtered by serial amendment to 775 and then to saturated 756 ammonium sulfate and removing the precipitate after each amendment by low speed centrifugation. The low molecular weight ionic cell components were then removed by adsorption of 6 + in 0.75 saturated ammonium sulfate in a buffer pH of 01 mM Hepes-NaOH pH”; 1 mM Na2EDTA to YO hydrophobic interfacial column (phenyl sepharose) purchased from GE Healthcare and filtered by Lau

1 pm

6 By linearly reducing the ammonium sulfate concentration to zero with a pH stabilizer of 01 mM Hepes—-NaOH at a pH stabilizer of 1,7.4 mM Na2EDTA. Most of the negatively charged proteins were then removed by modifying Fragments containing 2086© to pH Stabilizer from Pass 01 mM Tris— aha (HCI 0 pH ALT 1 mM Na2EDTA from Fragments Assembled on TMAE Anion Exchange Column) Purchase of EMD balanced with the same pH stabilizer. Then further purified rP2086 by chromatography on ceramic hydroxyapatite (produced from BioRad exchanging stabilizer containing pH 02086 to 01 mM 1165-1120M18 pH 17.4 containing 1 sodium phosphate mM 0 protein adsorbed to ceramic hydroxyapatite then filtered with a linear gradient level of You) sodium phosphate mM at pH 27.4 The unit operations recorded above are often sufficient to produce 202086 members of (ell Alla A) though since the level of manifestation can vary Le is more than 10 times as large; When +6 appears at the lower end of the range or Late there is dala to ultrapure 1© 2086 (at high concentration for AN NMR determinations) the following additional unit operations are added: concentration chromatography followed by hydroxyapatite chromatography 0618116. Replaces pH stabilizer containing protein 202086 from the previous hydroxyapatite step to yo mAs (Tris-acetate) pH AY and adsorbs protein to chromatography concentration column 08894 (yield From GE Healthcare balanced with the same pH 0 stabilizer then filtered with a pH stabilizer of 94-acetate poly-pH stabilizer; pH A7. Proteins 202086 will be filtered at its ~pl The parts containing 000180 were collected. The parts containing 202086 were then replaced with 01 mM Hepes-NaOH pH 7.4 containing sodium (Ak) phosphate molecular weight, adsorbed, and filtered as above. Yo subfamily A prepared by this process is typically homogenous by >795 by

SDS-PAGE analysis, homogenous for the most part, >794.

no

-174- respectively) TA TT 00 (order definitions: A22 12E and AOS) At the end of fermentation, the cell slurry is recovered by continuous centrifugation and resuspended to about

Molecular EDTA eo (Tris mM 01 original fermentation volume in £9 at eg pressure) pH 7. Dissolution of cell suspension achieved by high pressure homogenization to DADMAC concentration psi square). 900-4606060 sale is added to the homogeneous material during that period, 01 sad iome*?5-11 final 0.05 7 is formed. The solution is stirred at (and 22m) A12 AOS proteins precipitate heavy. The solution was purified by continuous centrifugation. Purified using two chromatographic steps followed by exchange of final pH stabilizer. The pH stabilizer of the concentrate (CEX) was adjusted to 5.5, loaded on a sodium column with resin, and filtered in succession using a graded level of protein bound 0.

Vo To a final concentration of sodium citrate CEX is added to the collected material from a column. 06 protein May Filtered. (HIC) Phenyl-650M Gram Molecular; The solution is carried on a column The aggregate is replaced with sodium citrate bound to the resin using a graded level of purifier step in the pH stabilizer of the final drug material by filtration protein containing HIC up to a concentration of 06111056 acetate Regenerates a cassette Ultrafiltration of 5 kD Dual. 1 Filter double use within 161,601,818 mg/mL. Filters target 7-015 to 07

APY = the property at sale prior to bottling for storage. LY Store Micron Filter Example 17: Serological Bactericidal Test vs. (SBA) Functional antibody titers of a Serogroup 8 serogroup 8 germicidal test medium from wild-type or engineered Neisseria meningitidis strains 0 1180 appears either with sequences homologous or heterozygous with those included in the vaccine. Bactericidal antibodies were determined in the serum of rabbits immunized with ©2086 vaccines using a human supplement. The fortified serum of rabbits was inactivated by heat to remove the activity of the essential complement 5

Mg2+ with PBS 00066005+ and 00066005) were diluted sequentially by 2-fold in a 97-well microplate to test the bactericidal activity of serum against (0-085) © Lam

-1?e-s strains WN. meningitidis for union studies with engineered strains; Desired serum is mixed in a ratio of 0:1 prior to the serial dilution described above; Therefore, the effective concentration of each component is half when each of them is tested individually. Bacteria used in the assay were grown in ©66 medium supplemented with (GCK) Kellogg's complement and monitored at optical density at 191 nm. Bacteria were harvested for use in the test at 010650 Aled of 5.0 -0.95; Dilute in D-PBS and 1100-00© CFU to the test mixture. Human serum without detectable bactericidal activity is used as an exogenous supplement source. Complement sources are tested for suitability (sae) against each Adi individually tested. For homozygous strains, a single human serum is homologous and determined for 0 5828 against all homozygous strains. A complement source is used only if the number of surviving bacteria is In control subjects without the addition of Fortified Jame > 775. After incubating for 0 min at 71" with 75 602 and shaking at 0000 rpm on a shaker; 0-8 is added to the reaction mixture and complete affinities are transferred to plates Pre-filled microfiltration with 0 606 medium for wild type strains and GCK 79000 medium for engineered strains Filter Vo microfiltration plates Incubate overnight at 17*cytophore with 602 75 Colonies are stained and quantified Determination of insecticidal titers of bacteria in serum as a cross-reciprocal serum dilution yields a Jo reduction in CFU compared to CFU in eyes compared with no immunized serum Sensitivity to killing by the anti-2086© immunized serum arises if the anti-02086-immune serum rises four-fold or more in the titer SBA for sera immunized against 02086 compared with sera before the counterimmunization 0. Serums that are negative for The ADL Test at Initial Dilution can specify a titer of half the test detection limits. Example 18: Generation of immunogenicity from non-lipidic variants of P2086 proteins from the Farahian family

A

kg) produced from T.0-Y.0 white New Zealand female rabbits were used in two studies. For the first study, a group consisting of? Rabbits (Canada) Charles River +5 Amad

-1?7-

With whether Vo µg or ? μg each of lipidic or non-lipidic AOS FHBP preparation LAOS for study AE A group of © rabbits immunized by intramuscular injection into the right hind leg with a variant of TP2086A at 01 μg/mL adjuvant with 0 µg/mL 81004 (+0 mL/dose/2 sites). Animals inseminated in week © yellow; and 9; You bleed at week 0 (Ty) and become anemic at week 01. The titres are determined

Specific bactericidal antibody 02086 at 02086 yolk 76 and 10 weeks. The aim of these studies is to mimic the reduced responses observed in normally immunocompromised Jie infants. We first compare low-dose and high-dose (? vs. ? micrograms per antigen per dose) of vaccines containing either a lipidic ADS (order definition: OF 0) or a non-lipidic AOS (order definition). No.: 00 (Tables (Vo and 11(b))). Low doses are used so differences in response rate between each vaccination are distinguished. SBA analysis is performed using two groups of strains. The first group consists of wild type strains that cause The second group is a genetically engineered strain with the same ADL background and differing only in the apparent fHBP order as follows: PMB3556 engineered a strain of N. cmenigitidis 10 that shows variant 824 from fHBP so that it replaces the 1700 endogenous Land gene with genes encoding for other THBP variants. Constructs are designed such that the region encoded for the ORF is Ja' (switched) and the surrounding genetic background is left intact. Therefore, SBA analysis Using this strain set allows the evaluation of reactivity against proteins of 1185 different A strains expressed at the same level and with the same genetic background using a single source of 0. Human complement. All strains have expression levels of 1180 that are above the homologous limit by Liang et al (2010) as shown in Tables 11a) and 11(b); both high and low dose levels of the 805 lipid-containing vaccine elicit overall protection towards Variegated ADS variants (li) showed reduced responses at both doses of vaccine containing the non-lipidic ADS variant. Therefore, this side-by-side comparison of Yo shows that, although the ADS variant Non-lipidic is a cross-protective towards

uh 7

-yYV- It is not as immunogenic as the lipid variant of the A (the strains expressing the subfamily A) (B) 011 and (A) 11. It is more likely to form the original form (Tables) micrograms for each variant of the family Farahiya 01 for the subsequent study; the dose level is raised to A. Analysis of non-lipidic WA to evaluate its ability to provide broad coverage against subfamily A strains shows that at this higher dose level all sera from rabbits immunized with SBA 0 Non-lipidic (Arrangement Identification No.: 00) 862 (Arrangement Identification No. AOS HBP) Varieties Inducing Calibers (TA) (Arrangement Identification No.: A225 (TT) (Arrangement Identification No.: 812 (VY)) Homozygous and non-A wild-type strains showing all of the subfamily's heteromorphic forms

homologous Lao dll; It indicates that all of them are cross-protective. At a low dose within the family, it can generate antibodies that can kill (AOS) 181201 strains. Therefore, we note that the vaccine, as well as vaccines from other complexes, these (A12) N2C2 5 (A22) 10162 Variant Form It can kill strains with opposite variant forms. under these conditions; It is noted that the variants accordingly; the higher L(V) across the strains (Table SBA) induce effector rates AG2 5 5, this demonstrates a protective effect through these variants.

SBA Geometric Mean for Calibers | und 805 Table 51(a): Preparation >4xrise | PD3| Previous | 24xrise | PD3 | Previous | ADL (JS name) variant fHBP tyov

—\YA-7| Sleeve YA 76017 RD3040- AOS | Strains AOS i Blas aad) or var] a start ve| RD3044-| Az] +"

Al2

Y| 7 AH 1 YIYYAd| Y¢| RD3042-A22

A22 1941 0 | 8021 AY RD3043- A29

A29

SBA Schedule 11(b): Preparation 805 Non | The geometric mean of the titres of the non-lipidic lipid AQS5 preparation. >4xrise | PD3 | Previous >4xrise| PD3 | Figure | The name of the strain was the previous variant fHBP. | . (yoy

-4?1- Dynasties | 05 PM YoA| 5| RD3040- Yellow 1| 13 | vA Blas 5m al) Az] = albegyuds' ve|pain | 5 11 Al2 YY v| 71 | v¢| RD3042- A22 Adam 11 A22 v1| RD3043- A29 for Love »M Y| viv] ov Table A29 (Ve and 501(b): The geometric mean of the SBA titres against the strains of Meningitidis. For group 8 sera before and after Vo-PD3 (weeks) immunization of rabbits ( Number (T= whether with Te or ?µg vaccines containing lipidic or non-lipidic ADS. The top boxes (tagged “wild-type strains”) from Tables 11(a) and 11(b) © Summaries of activity against those in clinical isolates. The lower frames (parameter 'homozygous strains') from Tables (Vo and 11(b) summarize activity against a group of homozygous treated strains engineered from the L. meningitides source strain (PMB3556) so that it completely replaces the ORF from its own Lindl internal FHBP with either variants ADS (Arrangement Identification No.: A22 (VY) (Arrangement Identification No.: 11 829) Rank number: (VE or 812 (definition of 10 rank 1 tad). The proportion of respondents with an increase of > 7 times

Ad «= \ _ as2| 5 12m A22|Mean 7 Table 16:16 clearly shows the percentage of respondents with at least a four-fold increase in SBA GMT levels over the background of 10 weeks sera taken from rabbits immunized with 10 micrograms of Non-lipidic subfamily A FHBP variants versus strains showing the variant (ADS THBP 62 12m A22 J o) Also note cross protection for all variants using the homozygous strain set described above at an increasing dose of 01 μg; with sera from rabbits vaccinated with variant 862 (order identification number: VY) clearly show the most cross-reactivity; followed by anti-ADS sera (table VY) in addition; sera from rabbits vaccinated with VY Variation Fig. 862 (order identification number: (VY) showing reactivity for both source strain PMB3556 0 and variant strain 809 (Table “(VA” indicating extended cross-reactivity activity for All) sub-8 proteins. The 862 consists of both domains of subfamily A (A22) and subfamily 8 (B09) (Fig. 8). ABA rats versus the KA3011 homozygous progeny group | PMB3556 | RD3044-| RD3043-| RD3042-| RD3040- 05m2m 29m2m (source B24) l= [Fe] Sat Fel Po tyov

-1?”

5 Ratio of Responders (Increase > Fold) Vaccine | KA3011 | PMB3556 | 03044-| RD3043-| RD3042-| RD3040- 05m2m 29m2m Table 17: “A Blas (switched) strains engineered the gene from the source N. meningitidis strain (PMB3556) so that it replaces the full ORF for 1185 intrinsic Lindl (Variation Figure 824) with either Variations AOS (Arrangement Identification No.: A22 (VY ©) (Arrangement Identification No.: Vo 829 (Arrangement Identification No.: VE) or 2 (Arrangement Identification No.: (VE KA3011 is a negative comparator strain (ie source PMB3556 whose M115 gene was deleted). The geometric mean of SBA titres (n = 0) of sera (taken before or after 10) is shown in the upper frame. Weeks of immunization of rabbits with three doses of 01 μg of the non-lipidic subfamily A (FHBP) variants against these strains.

-1“7- The lower frame is the homozygous proportion of respondents that clearly show at least a four-fold increase in response over the background. vs. strains of subfamily 8 homozygous for the SBA gene. Geometric mean of titres.

RD30337-B09 (to source) 6 to 721 603 respondents | Gila the vaccine | Previous 721 003 of respondents (height > fold) (height > fold) 01 weeks of serology (taken before or after SBA Geometric mean of titres: 18 table 01 micrograms not Lepidian A subfamily proteins immunized from rabbits (N-0) with (Location Identification No.: 00) 812 (Location Identification No.: ADS 771; (Location Identification No.: AG2) © vs. two homozygous strains of family TA (seq identification number: A22 7);

The SBA is testing serum consortia to assess impact on the field of inquiry. Test a pair of Ya sera with pre-vaccination versus post-vaccination to ensure that there is no nonspecific killing induced as a result of combination of individual sera and combinations of serotypes across strains GM calculates double elevation 0 serotype double LA is detected Leal homozygous for the four genes representing variation within the growing family of some of the tested consortia to provide evidence that the field of investigation can be increased 7 AH

-?-

By introducing more variants of the A subfamily (Table 19).

It is AOS (order identification number: 00) with 862 (order identification number: VY) or 862 (order identification number:

Arrangement 1 (tad 0) with Al2 (Arrangement Identification No: 1 7) (Table 0")

The Osbua dynasty the week | height | The week is the week of the rise | the week | Weeks high

2 p 01 light 2 p 01 light 2 p 01 light zero f zero f zero f

1 vy 10 Y $9 4A Y | RD3040

-A05

1 AN Y sy q¢ Y oA 5 Y | RD3042 -A22

11 04 a9 | YaA "1 oyyy| via v| RD3043 -A29 yo $0 v YY § AT 7 y4 vv Y | RD3044 -Al2

1 7 7 0 Height GM

uh 9

-?- 0 Calibration 86050 8 2 + 05m2 + 05m2 + 12m AQS509- + 0508-5 -0507m + AQ508-5 -0507m + AQ509-4 4 5 5 Breed week week height week week height week week high zero 10 times zero 10 times times zero 10 times times 4 47 Y 7 yoy A Y¢ VV vy | RD3040- Damham 9) YAY Y Lal 6 Tl eve | TEVA 1 | RD3042-

A22

As $Y A 1 Ya) 11 the Yoo 0.9 Y | RD3043-

A29 oY. If Ah 7 Yi. 17¢y 11A RD3044-

Al2 1 1 7 1 7 1 1 0 High vulnerability GM Table 0) SBA titers of sera from the highest responders of each vaccine group are tested against the homozygous strain group as shown in Table VY The sera are tested in mixtures At a ratio of 0:1 to determine the extent of synergistic activity.

E01 conjugate double height increase of conjugate vaccine against monovalent (order identification no: 00) + [A 0.1 2 (order identification no: 17) 5 (order identification no: 00) + A 0.1 2 Arrangement Identification No.: VY 2 (Array Identification No.: A.4 Vy. + V1 2) Arrangement Identification No.: VY Table 01: Exponential height increase of the tested sera in consortium as It is comparable to each tested by itself (calculated from Table V3). The results presented above in Examples 18-19 show that non-lipid subfamily A proteins are immunogenic and may provide protection against infection with strains. meningitides do not carry either homozygous or heterozygous variants The data presented here demonstrate that select non-lipidic subfamily A variants retain immunogenicity and provide cross protection against heterologous strains; although these responses are lower than the variants We have also shown that the anti-alse 202086 862 (order identification number: VY) has order similarity to family dc yall 8 (see eg JUL figure L) may protect via 0 breadwinner subtypes because the 862 vaccine (order identification number: VY may kill strains showing variants of family de all 8 809 and 824). The data exemplified above demonstrate that not only non-lipidic A de dl family variants warrant the type of synergy observed with fHBP conjugates of gad but they may also provide coverage against subfamily 8 variants. AH 7

-1- A vaccine combined with 1 element binding proteins Example 20: Assessment of immunogenicity from a combination of

New Zealand white calf Meningococcus tetravalent in weight in the range of 7.5-7.5 kg New Zealand white oid The study takes place in before the start of the study; Pre-screen 00 whole-cell rabbits (Charles River Canada (Charles River) against 05M strains) (Table 71) ELISAs to determine the antibodies present using © vaccinated rabbits with relatively low antibody titers (6A titres) andl and 802. after 1 ml per site (0.5 ml per site) intramuscular injection of the hind legs; (Yoo > checked 4; and 9. There are ? rabbits per group. Rabbits bleed hia At week YY (dose; see table) Serum samples are prepared and analyzed 0. At weeks luteal 4; 7 9; and become anemic at weeks 0 and 10 serum samples by /58. The conjugate vaccine with Meningococcal Yo (MENVEO®, meningococcal (Groups A, C, Y and W-135) oligosaccharide diphtheria CRM197 conjugate vaccine, Novartis), non-lipidic bivalent and tetravalent and their conjugates according to rLP2086. Variations of the tables prepare a solution ?: Rabbits used in this study Table 1 Vo Species: Rabbit

New Zeland Breed: White

Charles River Laboratory a Source: 1 Number of animals in each group:

AID Number of animals: lea) Age and gender: Lao males

1 kg Yo-Yo Weight: ?: Rabbits used in this study Table 1 Rabbits are kept according to the guidelines of an accredited Animal Care and Use Institution. za

YY The study design is shown in the experimental design table. YY the preparation table of z group | number | Immune Gene Article Vaccine Auxiliary rabbits Serum A (week) Human dose No jaundice 4; 4 | WEEK 1 1 1; © ARA 1 DOSE [MENVEO 3 . ) milliliter/¥ site sda all 01 week human no jaundice 4; 4 | w deja 001 ?Y © ARA 1 dose [MENVEO 3]: ml/? sda all 01 week Safar 464 week Al PO4 Admia en 1 Y Y 1% yellow . 01 + MENVEO

On 1 5S μg/ | © l. A035) rLP2086-A Interchange: Vie tyov

—\Y¥A- 01 (define order number: | milliliters per week

Ye + (0 7 as

B01) rLP2086-B (Arrangement Identification No.:

Vi en ])) 54 ml/ ¥ zero 6) 4 week Al PO4 water By dose AREA 7 4 yellow ;; 0 + + MENVEO 4 J Early. μg/Al LP -A Exchange: Via, | 0 6 Define alt tab number: p 01 (define the order number of milliliters per week

Ye + (0 7 as

B01) rLP2086-B (Arrangement Identification No.:

Vi en ]))54 mL/¥ position zero 6) 4 week AIPO4 Mick Rogerram Yo 7 ¢¢ yellow vo A05) rLP2086-A 4 : 3 arrangement definition μg/a ) ? microgra + ((VY exchange: 1 fie | PTF ( .B01) rLP2086-B 10 milliliter wk ) (order identification number:

Vi en])) 54 gvoy

1*4 z w 4 ¢ » ver = = [3 4 [3 Baa . : z z

IPO4 | J

A aS ? 1 1 ا Yo. 05) rLP2086-A

H | p i p Arrangement No.: YK Rugram for exchange

NR 7 7 — JR 01 + (\¥

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AIPO4 rLP20 86-A0 9 (1) Definition

Yo. 220) Non-lipidic µg/exchange: Aan 0 wk | : abil. z u ; ; 0.8 SR - } 22 (£9 ta 8” )

Yo. 0 4 Arrangement No.: Hand for microgram per jar: Weekly Yellow 4 4 | Except,

AIPO4 rLP20 86-A0 9 « vo. B09 lipid; H | Z 1 In B vy «B44, 2 LTB fie dose | 7 micrograms per sip

_ \ ¢ "= 0 6 + 44 alpha Al PO4 ademia en 1 7 q fia + MENVEO q (1 rLP2086-A05 µg/8 96 lipid - Exchange: Vien 09 "pad Non +. B44, B22 10 milliliters week 3 micrograms per dose milliliter/? position 1 zero 6 + 44 weeks Al PO4 human subjects from ie 101: 7 A fia + MENVEO q ( 1 rLP2086-A05 µg/B09 eo ean - Exchange: Vien FE “dr + B44, B22

Yo Milliliters per week Micrograms per dose Milliliter/? Site 1 Summary of Preparations Preparations and Immunizations (YY Table of Substance Function Product Appearance/Appearance | Quantity Available

Y JAY

Yo x ¥ | : Lyo A| Novartis zi is active | MENVEO® ine) C A groups (Groups 9 Ah

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GY © and 135-//) Meningococcal vaccine Asyl 1 R-ala | 135-ALA: 17 Na (A mcg) pure phosphorous; solution 135 colorless oligosaccharide diphtheria “CRM197

Novartis » Syringes? 1| Cloudy Suspension ABN rLP2086 Active | AOS) 102086-84 and 8 at homogeneous color |16 (volume A (order identification number: sub vo µg/ | white to | tamper VY rLP2086-B))0 (order definition milliliters per A | white italic | milliliters (801) in yellow protein (COURTS

histidine pH 1; 70...us 0 with .. mg/mmL of AIPO4 144857-50 8 | active 5 (definition of fluff cake | XY vials ely) valence non-order number: (00) | in white; | 0 (17) Ve lipid 4 (definition of lyophilized milliliter volume order number: 4); recon) 2 ( Definition of Arrangement No.: (Vo BO9) (Definition of Arrangement No.: 29) Lad

-1157- at 0 mg/mL formulated in mM 0 gram stabilizer Histidine 1.0 pH 22.1 with LA 116081056

WFI and Malgar Yo | cloudy comment 0 AIPO4 | Auxiliary material, AIPO4, milli-molecular homogeneous in color 0.5 mg/white to milliliters in ?| NaCl Grammy White Italic | Glass vials WF 0 milliliter to yellow mg 0 milliliter in ? Glass vials

XY Flasks | mre solution NA caida milligram 0 a) Yo | net col grams brine

YA Aaa in milliliters).

-16©- liquid conjugate component | Novartis vaccine does not contain MenCYW-135 preservative or adjuvant. (091101) Each dose of vaccine lyophilized conjugate component contains 01 μg MenA o (029011) MenA oligosaccharide (029011) μg of each “MenC and/or MenW135 oligosaccharides and/or.”? to TE ‏

CRM197 Rugram Sue .protein The residue formaldehyde per dose was estimated to be no more than 0 µg (not known a priori). Histidine | CSMD, Pfizer | 962-UPD-(09- rLP2086-A AOS) (Definition | Pearl River, NY 007 v1.0 | pH approx. 1 Arrangement No: PSO Z+.v vo (VY) m rLP2086- B mg/milliliter Al of AIPO4 (BOI 4) Order No.: (OA)

-1654- pH Stabilizer Formulation 1 (Definition 5 MnB As) Histidine Development, | (00 Arrangement No.: L44857-50 1.0 pH | Pearl River, NY | (L35408-140) | Non-Lipidine Tetravalent (L44130-129) Definition 4

Polysorbate 80 (£8) Order No: «(L44130-127) «(L37024-36A)

Trehalose (identification 2 «(L44863-68) (Vo Arrangement No.:

WFI (B|Braun «(L37024-61) (89) Arrangement #: (L43930-80)

AIPO4 i I<| Pfizer Pearl | mg/mL: v0 AIPO4

HO00000606 — River, NY | L44863-86A

D86864M [oe >» Brine Solution WA 9 L44863— H Milliliters (B/Broun #868)

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Pearl River, NY 004 Gram Saline Valence: ely 1/008 Jal Content/ Closure

West Pharmaceuticals 1 Containers: ¥ milliliter bottle of type grey; coated with mm butyl for lyophilization; 11 stoppers: thinner pot stoppers

-E61- Flurotec (WPS V2-F451W 4432/50 Gray 52-104 Westar® RU Verisure Ready-Pack), West Pharmaceuticals Content/Closing for Saline 01 mm: Vessels: dala) capacity? Milliliter Schott (Vendor Part #: 8M002PD-CS) 00 E Plugs: 0777-1 Daikyo Dmi 2-1451 Westar RS West 82-40 (Vendor Part #: 19560180) Content/Seal to order Solutions: AIPO4 Vessels: dala) empty sterile; Volume 0" milliliters-.? milliliters; included Allergy Laboratories stoppers Lot #51/070708 Table YT pH and Appearance of AIPO4 Solutions L44863-86A 9.8 White to off-white suspension; Ale L44863-86B 0.4 White to Off-White Suspension Ale 0 Table Yo Data Analysis Table 5?: Analytical Tests for Lot L44857-50 Non-Lipidic Antigen ol 1 /078 Test 42 “B09 Concentration 822 | Concentration 809 Concentration AOS 544 Concentration AOS 844 | (µg/ | (µg/ | (µg/ | (µg/L))

(lle ans a milliliter).| 1 (Glo milliliter) (μg/milliliter) rh a a a 0 dak 1 15.1 1 HPLC exponent 1.0 1.0 pH Appearance colorless solution Lyo: cake ine) white oll clear: sale) 8: Formation (w/Ale 01 moM):(NaCl) clear colorless solution Reconstitute lyophilized preparation with mobile phase during quantification of AOS 3 809 3 B22 and 844 by IEX-HPLC and with 00 mM NaCl dilute for pH and appearance Use Karl-Fischer (ICH) method for hygrometry (using methanol to reconstitute lyophilized formulations) Monitor non-lipidic tetravalent protein (822) 809 (B44 5 AOS Determination of 7-hour stability at 7-/"C when combined with MENVEO® EXAMPLE 21: Serological Bactericidal Test (SBA) A micron colony-dependent serobacterial test is performed (SBA) vs. 0 meningococcal strains of serogroup 8;© and 7 multiple (Table (YY) on individual serum samples. Human sera from donors can serve as a complementary source for the strains tested in the test. L

AEA

Bactericidal titers based on a complement-mediated antibody appearing as a surrogate for serum dilution of meningococcal cells are intercalated into the test. The detection limit of the test is 75 0 The test kills LY of <; It is a specified number of SBA caliber 58/8 of ;. The titer of sera compared with 01 height is > ; times 8/58 titres per week titres before hemorrhage. oo

SBA strains YY element correlation table 4 "lipid or non-lipid and vaccine proteins Example 22: Generation of immunity to a combination of

New Zealand white rabbit conjugated germicidal antibody in serum is an immunological surrogate for protection against lipidiosis; Non-lipidic; IHBP BOTH immunization with meningococcal SBA. Determined by 0 quadrivalent conjugated vaccines alone or in combination showing germicidal antibodies in rabbits. 7 AH

“VENA

The SBA measures the level of antibodies in a serum sample by simulating naturally occurring complement-mediated bacterial lysis. In humans, SBA le in a ratio of 4:1 is prophylactic; A four-fold increase in titer pre-versus-post immunization is also considered an immunologically relevant immune response. Rabbit serum samples collected from weeks 0 and 10 were analyzed by SBA against © strains from the multimeningococcal serum groups. As shown in Table YA (higher dose) and 79 (lower dose); One week after the third immunization (week 01) only quadrivalent conjugate vaccines show SBA responses against the MnC and 1/097 strains tested. All other serum samples show bactericidal activity against homologous strains as well as other test strains of the same subfamily ©0118 as in vaccine formulations. It is noted that immunization 0 with 805/801 lipid (order definitions: 11 and 8©; respectively) alone at a dose of 30 micrograms each shows germicidal antibody (oY) against similar strains and also Against other tested strains from all HBP subfamilies (YA table with similarity; immunization with non-lipidic A05/B09/B22/BA4 (order definitions Nos.: 4 00 ©2 and 44 respectively) alone also appears Bactericidal antibodies against VO strains from multiple meningococcal serum groups, although SBA titers are ∼ to 2 times lower than for bivalent lipid vaccine (Table 07). by >; in the SBA titer against all strains from the different serum groups

for 1186(andl) high-dose non-lipidic THBP and all conjugates. Table 78: Two-fold increase in SBA titres versus serum group 0 strains of meningococcus Y “CB” using sera from rabbits immunized with the conjugate le dose of 11865

and a conjugate vaccine. MnB strains strains strains - AH 7

—V£4-

BO| Al2| B2| A68| B44 | B1| 80 | 80 AD| The vaccine dose 9 1 4 66 91 a2 5

YY| VeA| Dur Kk 1 1 1 111 dose MENVEO 1 hu

AL 01 | ee No Yes | Av YY AY YE) dose /MENVEO ¢ ALA 1 Y NER EY ERE chu 1 lipid 1 protein

Y.:s microgra ( F<m dose 01| cdo | Ye | 0197| Yao | Ae VE 17| 4 Ye lipid 1 ¢ 1 5 Y 5 ¢ 1 1 µg ( F<m dose Ye ah 1|Alt Yel ya] 01|4 011 A05/B09/B22/

A v a | oe good lipid | microgra 4 ( F< m dose YY YA9 0 1 | for you YA YY 011 erase 4 | 1 dose /MENVEO ° o 1 chu | AQ5/B09/B22 / protein | non-lipidic 4

Y.: s micrograms L

and c-1 M Ca) dose. Pre-bleeding sera in rabbits demonstrates pre-existing bactericidal activity against the tested strains. NZW rabbits (number (Y=) vaccinated at week 0; ; Ag) with v0 vaccine mL intramuscularly data week 01.01 Table 4?: two-fold increase in SBA titres versus serum group strains of meningococcal 8©; nor using sera from rabbits immunized with a low-dose conjugate of 1185 and conjugate vaccine Double increase in titers PD3 SBA MnB strains SNL strains MnY MnC vaccine Dose B2|A6|B4|81|BO|BO|AQ|12m BO|5 A2 9 66 4 8 4 1 9 1 11 Vien MENVEO ark ak vel ga] alm 0 Y 1:1 hu /MENVEO ga VA confirm to count for a ay ed yul ote 1 lipid lo 1:11; Y IO 1 hu protein Ys μg m lca (uh 7

_ \ o \ —_

Ya | ovo free ev] Yes | 6| 11 | ve 0| Lebedy 1 3 1 EY A |; a|. ah |; 0 Y A 0111 | AO5/B09/B22/B 4 non-lipidic chu protein

Ys μg m Ca (dose) Prior hemorrhagic sera in rabbits demonstrate pre-existing bactericidal activity versus the tested strains. NZW rabbits (number (Y=) vaccinated at week 0; Ag with v0 mL vaccine; intramuscularly) Table iY data for week 01.01 Rates of responders to SBA vs. meningococcal serogroup 8© and 7 strains using sera from rabbits vaccinated with a consortium of 111,855 and conjugate vaccines. 7

_ \ o \ —

CNS | MnB strains MnY MnC strains

BO| A12| B2| A6| B4| B1| 80 | BO| AO dose ac 9 1 4 8 4 6 9 2 5

Vol Yoo 01 | Yo ma yo ma yo ma yo ma 1 dose MENVEO : ol hu

Vol Yoo 01 | 01 | Ace ma Ace Aen MENVEO : EY EY EVER EVER EVER EVER 001 hu A Yeo Yo 10 10 1 11 11 Yo dose MENVEO / 0 0 0 0 0 0 0 0 0 hu non A05/B01 protein lipid 5:01 μg m ak_dose Yeo 01 10 10 Yo 11 11 1 icp /MENVEO A A05/B01 lipid 0 0 0 0 0 0 0 0 1 0:1 <hu protein 01: 5 micrograms mk_1 uh

_ \ o — 0 Yen A 0 0 A A A A A 701 Lipidi 1 0 0 0 0 0 0 0 0 micrograms m a | 1 dose 1 lipid 7 10 10 1 aaa aa 0 Yen 0 0 0 0 0 0 0 0 0 micrograms m a | 1 Dose 1 0 1 0*0 1 0 1 0 1 0 1 0 1 0 1 0 1 0 Y 0 A05/B09/B22/B 44 Non-lipidic Microgra 0 0 0 0 0 0 0 0 0 M A | 1 dose A05/B09/B22/B | ¥ No, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no, no. 1 Dose/MENVEO Ve Ve Yo Dose 1 A A A A A 0 Yen 0 0 0 0 0 0 0 0 <h u A05/B09/B22/B 4 Non-lipidic protein

Y.: 8 μg m a | 1 1 Ah

o ¢ —_ \ _ icp /MENVEO Ba A A Ve 1 11 A 11 11 0 0 0 0 0 0 0 1 0 1 A05/B09/B22/B 4 non-lipidic chu protein Y.: 8 μg m lac (dose vaccinated in NZQ rabbits (n = 7) at weeks 0; Ag with v0 mL vaccine; intramuscularly; data wk 01 fHBP Lipidic elicited ef SBA titers of 1180 non-lipidic elicited lipidic FHBP at 0µg per dose elicited SBA titers 1-10-fold higher for all meningococcal strains 8© and 7 tested. Non-lipidic fHBP at 10 μg per dose SBA titers are -77-fold higher for all 8 meningococcal strains tested (Tables 5-748 7). Dose titration is achieved with the 01805 sald vaccine conjugate or conjugates with a high dose of conjugate vaccine (is iad 11805) or their conjugates have greater SBA responses by 0 than with a low dose (Tables Y = YA). A single human dose of conjugate vaccine elicits higher SBA titres by 1-8-fold higher against meningococcal strains ©Ys than the first 10th dose of conjugate vaccine elicits lipid HBP at μg per dose titres SBA higher by -7; fold against all tested chains for ? micrograms per dose. Non-lipidic 0185 at Vo micrograms per dose elicits higher SBA titers than AH 7.

— 00 \ _ by § Yo weakened against all 8 serogroup strains; YC meningococcus for? micrograms per dose. SBA Responses Combined by FHBP Conjugate and Conjugate Vaccines There tends to be higher SBA responses against serogroups 6 and 71© meningococcal strains when a combination of conjugate vaccine and 1118 is used using either component alone”; Especially with the addition of a lower dose of lipidic or non-lipidic THBP (Table 79). In the current study; Functional activity against strains of multiple meningococcal serogroups is evaluated using serum from autoimmune New Zealand white rabbits with non-lipidic or non-lipidic THBP conjugated with preparation with 01004 and sald vaccine) conjugated alone or in combination. Rabbits receiving the conjugate vaccine elicit lost SBA responses against strains of serogroup C Y meningococcus; but not for serogroup 8 strains. Lipidic or non-lipidic THBP in a preparation with L004 elicits serum antibodies that are bactericidal against strains of all meningococcal serogroups tested. New Zealand white rabbits receiving three doses of THBP lipidic or non-0lipidic in a preparation with 004L elicit serum antibodies that are bactericidal against the C ¢ B serogroups and no meningococci strains tested. Yeo 7 validates response rate (> fold increase in SBA titer against all strains tested except for the low Ge jal non-lipidic group) Lipidic FHBP elicits higher bactericidal antibody titers than non-0 forms _ libidin A pronounced dose response is observed with lipidic or non-lipidic THBP and conjugate vaccine alone or in combinations There is a tendency for synergistic SBA responses against serogroup © and meningococcal strains among the 5-HBP vaccine in particular when Add protein, lower dose

_a o \ _ EXAMPLE 23: Evaluation of immunogenic activity of non-lipid factor H binding protein conjugates in New Zealand white rabbits Studies are conducted in New Zealand white rabbits of 3.5-7.5 kg from Charles River, Canada (Table 1 (Y) rabbits vaccinated intramuscularly at the hind legs 0 0 milliliters per site (1 milliliters per dose; see Table YY) at yolks 4 and 9. Report the order identification numbers for each of the tested antigens in Table FY There are 01 rabbits per group.The rabbits are depleted at week 06 and anemic at week 01.Serum samples are prepared and serum samples at weeks 0 and 10 are decomposed in SBA against a frame of isolates N. meningitides. 0 Table ©1: Rabbits used in these studies8 8 Rabbits used in these studies should be kept in accordance with the guidelines of an accredited Animal Care and Use Institution Table 77: Dosage Design Number of Rabbits | For the antigen combination (mg/dose) oo | | or 1 0 uh 7

l o \ — B44 EK + 62M + AOS No 0 mcg Softener per serving

EK + 809 + 62m + 05m only 0 µg finer

B44 per dose Ye + 509 + 62m + 05m except 5 mcg yes

B44 per dose Al2 + 509 + Ye + 05m except 5 mcg Yes

B44 per dose A62 + 809 + 01 + 12m only micrograms finer

B44 per dose EK + 62 + 12m + 05m except © micrograms finer

4 + 509 per serving AOS + 1 EK No 0 µg softer

For each dose 8 rabbits are inoculated intramuscularly (your luteal weeks; 35) and drained (your 6th and 10th yolk weeks) to prepare serum samples for SBA analysis Table vy Alloforms used Protein 1180 Serogroup 8 N. meningitidis rP2086-A05 sequence identification number: VY ¢ where Cys is omitted at position 3 or sequence identification number: 00 for example; encoded by arrangement identification number: of l

m o \ — rP2086-A12 sequence identification number: V¢ ¢ where Cys is omitted at position 3 or arrangement identification number: 1) 0” Sie encodes dail gs sequence identification number: 19 rP2086-A62 configuration identification number: (Ve where Cys is omitted at position 3 or configuration identification number: 1 for example 0” encodes dail gs configuration identification number: YY 2086-9i Arrangement identification number: YA ¢ where Cys is omitted at position 3 or arrangement identification number: 4 2086-4 Arrangement identification number: YY ¢ where Cys is omitted at position 3 or arrangement identification number: 4) for example 0” dail gs encodes arrangement identification number: 7 rLP2086- arrangement identification number: VU A05 rLP2086- arrangement identification number: ON BO1 YE table summarizes the response Le lid) in rabbits for mixtures of non-lipidic THBP proteins compared with the immune response to rLP2086-A05 and 1 rLP2086-B0 pair of lipid antigens. Previously drained serum from rabbits shows no pre-existing bactericidal activity against the tested strains. The immune response is present as a proportion of the animals © in each treatment group that respond to conjugate-specific FHBP antigens after the third immunization with an increase in SBA titer of > ; double. SBA testing is performed using target meningitides strains that display either FHBP variants similar to the vaccine immunogen (A12 (AOS) gene) or strains that display heterozygous HBP variants (A22) 816 824).

— \ o q — Comparative amino acid ranking of the B24 5 B16 HBP variant up to 717 subfamily 8 variants present as antigens. Table 34: Gastrual percentage of New Zealand white rabbits inoculated with 11,805 non-lipidic mice that respond with increased >; fold in the three previous doses of SBA titer 7 responses at 003 with an increase of > ; X SBA titres lipid-generating |dose per B24 816 A22| 12 | A05|adc ill antigen (μg/20+mL) 1 “ou 3 “ou 1 0 no A62 +

B44 at 0 A 0 A 0 4 0 1 0 dom la + 62 + 844

Yen Yen Yen "1 0 hom la + 62 + 509 + 844

Ye 101 4 4 05m no + 62 + 509 + 844 ah 7

Nt.

Te Te Te Te 2 Te 05 pm except +

Al2 + 509 +

B44 For Vou Vou For Veo Only 12 + 62 + 509 +

B44

Te Yoo Yoo 0 Yoo except AOS +

Al2+

A6G2+509+

B44 a Vou a A

Yo.) AIPO4 animals per treatment group; All treatments are formulated with adjuvant (8 0 µg/dose) 0 1 µg of each variant in these groups of rabbits vaccinated with 62m + ADS tested; Serum samples from animals treated with the conjugate of +6 in SBA animals had a high bactericidal response rate. Responsiveness decreases somewhat 844 + 809© treatment with only © μg of each of the same mixture of four non-lipidic 1188 μg isoforms. Dosage at μg HBP is achieved; Other equivalent groups of non-lipid antigens. of the 844 + 809 + /62 + ADS quadruple conjugates in addition to the tested equivalence conjugates; Serum samples from the treatment group containing © 7 μg of AH

-111-

SBA 62m + 809 + 844 non-lipid response rates + Al2 fHBP variants are better for selected test strains. The response rate to the pentavalent non-lipid conjugate is better than the response to any of the tetravalent conjugates 844 + 809 + A62 + 812 + 805 of the tested valences. <110> Rank Lists Pfizer Inc. 11 <Compositions for treating bacterial meningitis and methods for their preparation 120< > 130< 7 <160> 1 Vo <170> PatentIn version 3.5 <210> 1 <211> 780 0 tyov

-١١- <212> DNA <213> Neisseria meningitidis (group B) <400> 1 tgcagcagcg gaggcggagg cggceggtgtc geccgecgaca tcggcacggg gettgecgat ٠ 60 gcactaactg cgccgctcga ccataaagac aaaggtttga aatccctgac attggaagac 120 ٠١ tccattcccc aaaacggaac actgaccctg tcggcacaag gtgcggaaaa aactttcaaa 180 gccggcgaca aagacaacag cctcaacacg ggcaaactga agaacgacaa aatcagccgce 240 10 ttcgacttcg tgcaaaaaat cgaagtggac ggacaaacca tcacactggc aagcggcgaa 300 tttcaaatat acaaacagga ccactccgcc gtcgttgccc tacagattga aaaaatcaac 00 360 gvoy

-١- aaccccgaca aaatcgacag cctgataaac caacgctcct tccttgtcag 9091119096 420 ggagaacata ccgccttcaa ccaactgccc ggcgacaaag ccgagtatca cggcaaagca ٠ 480 ttcagctccg acgatgccgg cggaaaactg acctatacca tagattttgc cgccaaacag 540 ٠١ ggacacggca aaatcgaaca cctgaaaaca cccgagcaaa atgtcgagct tgccgecgec 600 gaactcaaag cagatgaaaa atcacacgcc gtcattttgg gcgacacgcg ctacggcagc 660 eo gaagaaaaag gcacttacca cctcgccctt ttcggcgacc gcgcccaaga aatcgeccggce 720 tcggcaaccg tgaagatagg ggaaaaggtt cacgaaatcg gcatcgccgg caaacagtag 0 780 gvoy

-١١6- <210> 2 <211> 786 <212> DNA ه٠‎ <213> Neisseria meningitidis (group B) <400> 2 tgcagcagcg gaagcggaag cggaggcggce ggtgtcgecg ccgacatcgg cacagggctt 60 0٠ gccgatgcac taactgcgcc gctcgaccat aaagacaaag gtttgaaatc cctgacattg 120 gaagactcca tttcccaaaa cggaacactg accctgtcgg cacaaggtgc ggaaaaaact Vo 180 ttcaaagtcg gcgacaaaga caacagtctc aatacaggca aattgaagaa cgacaaaatc 240 790 gvoy

—-y10- agccgcttcg actttgtgca aaaaatcgaa gtggacggac aaaccatcac gctggcaagc 300 ggcgaatttc aaatatacaa acaggaccac tccgccgtcg ttgccctaca gattgaaaaa 360 ‏هه‎ ‎atcaacaacc ccgacaaaat cgacagcctg ataaaccaac gctccttcct tgtcageggt 420 ttgggcggag aacataccgc cttcaaccaa ctgcccagcg gcaaagccga gtatcacgge ٠٠ 480 aaagcattca gctccgacga tgccggcgga aaactgacct ataccataga ttttgccgcec 540

Vo aaacagggac acggcaaaat cgaacacctg aaaacacccg agcagaatgt cgagcttgcc 600 tccgccgaac tcaaagcaga tgaaaaatca cacgccgtca ttttgggcga cacgcgctac 660 0 (voy

RA ggcagcgaag aaaaaggcac ttaccacctc gctcttttcg gcgaccgagec ccaagaaatc 720 gccggctegg caaccgtgaa gataagggaa aaggttcacg aaatcggcat cgccggcaaa 780 © cagtag 786 <210> 3 0 0٠ <211> 765 <212> DNA <213> Neisseria meningitidis (group B) <400> 3 ١٠ tgcagcagcg 13992099099 tgtcgeccgec gacatcggeg cggggcttge cgatgcacta 60 accgcaccgc tcgaccataa agacaaaagt ttgcagtctt tgacgctgga tcagtccgtc 120 0 gvoy

-17/- aggaaaaacg agaaactgaa gctggcggca caaggtgcgg aaaaaactta tggaaacggc 180 gacagcctca atacgggcaa attgaagaac gacaaggtca gccgcttcga ctttatccgt e

240 caaatcgaag tggacggaca aaccatcacg ctggcaagcg gcgaattica aatatacaaa 300

01 cagaaccact ccgccgtcgt tgccctacag attgaaaaaa tcaacaaccc cgacaaaatc 360 gacagcctga taaaccaacg ctccttcctt gtcageggtt tgggcggaga acataccgcec

420 10 ttcaaccaac tgcctgacgg caaagccgag tatcacggca aagcattcag ctccgacgac 480 ccgaacggca ggctgcacta ctccattgat tttaccaaaa aacagggtta cggcagaatc 0 540 gvoy

-١ ١م‎ gaacacctga aaacgcccga gcagaatgtc gagcttgcct ccgeccgaact 88800881 600 gaaaaatcac acgccgtcat tttgggcgac acgcgctacg gcggcgaaga aaaaggcact © 660 taccacctcg cccttttcgg cgaccgcegec caagaaatcg ccggcetcgge aaccgtgaag 720 ٠١ ataagggaaa aggttcacga aatcggcatc gccggcaaac 09 765 <210> 4 ١٠ <211> 765 <212> DNA <213> Neisseria meningitidis (group B) <400>4 gvoy

-١4- tgcagcagcg gagggggegg tgtcgecgee gacattggtg cggggcttge cgatgecacta 60 accgcaccgc tcgaccataa agacaaaagt ttgcagtctt tgacgctgga tcagtccgtc 120 ‏هه‎ aggaaaaacg agaaactgaa gctggcggca caaggtgcgg aaaaaactta tggaaacggc 180 gacagcctca atacgggcaa attgaagaac gacaaggtca gccgcttcga ctttatcegt ٠ 240 caaatcgaag tggacggaca aaccatcacg ctggcaagcg gcgaattica aatatacaaa 300

Vo cagaaccact ccgccgtegt tgccctacag attgaaaaaa tcaacaaccc cgacaaaatc 360 gacagcctga taaaccaacg ctccttcctt gtcageggtt tgggcggaga acataccgcec 420 0 (voy

_ \ 7 «= ttcaaccaac tgcctgacgg caaagccgag tatcacggca aagcattcag ctccgacgac 480 ccgaacggca ggctgcacta ctccattgat tttaccaaaa aacagggtta cggcagaatc 540 oo gaacacctga aaacgcccga gcagaatgtc gagcttgect ccgccgaact caaagcagat 600 gaaaaatcac acgccgtcat tttgggcgac acgcgctacg gcggcgaaga aaaaggcact ٠ 660 taccacctcg cccttttcgg cgaccgcegec caagaaatcg ccggetcgge aaccgtgaag 720

Vo ataagggaaa aggttcacga aatcggcatc gccggcaaac agtag 765 <210> 5 0 <211> 765 (voy

rar

<212> DNA <213> Neisseria meningitidis (group B) <400> 5 tgcagcagcg gaggcggcegg tgtcgecgee gacatcggeg cggggettge cgatgcacta م٠‎ 60 accgcaccgc tcgaccataa agacaaaagt ttgcagtctt tgacgctgga tcagtcegte 120 ٠١ aggaaaaacg agaaactgaa gctggcggca caaggtgcgg aaaaaactta tggaaacggc 180 gacagcctca atacgggcaa attgaagaac gacaaggtca gccgcttcga ctttatccgt 240 ١٠ caaatcgaag tggacgggca gctcattacc ttggagagcg gagagttcca aatatacaaa 300 caggaccact ccgccgtcgt tgccctacag attgaaaaaa tcaacaaccc cgacaaaatc 0 0 360 gvoy

-١١77- gacagcctga taaaccaacg ctccttectt gtcageggtt tgggtggaga acataccgec 420 ttcaaccaac tgcccagegg caaagcecgag tatcacggca aagcattcag ctccgacgat © 480 gctggcggaa aactgaccta taccatagat ttcgccgcca aacagggaca cggcaaaatc 540 ٠١ gaacacttga aaacacccga gcaaaatgtc gagcttgcct ccgccgaact caaagcagat 600 gaaaaatcac acgccgtcat tttgggcgac acgcgctacg gcggcgaaga aaaaggcact 660 eo taccacctcg cccttttcgg cgaccgcgec caagaaatcg ccggetcgge aaccgtgaag 720 ataagggaaa aggttcacga aatcggcatc gccggcaaac agtag 0 765 gvoy

AR Al <210> 6 <211> 792 <212> DNA ٠ <213> Neisseria meningitidis (group B) <400> 6 tgcagcageg gaggeggegg aageggaggce ggcggtgteg ccgecgacat cggegegggg 60 0٠ cttgccgatg cactaaccgce accgctcgac cataaagaca aaggtttgaa atccctgaca 120 ttggaagact ccatttccca aaacggaaca ctgaccctgt cggcacaaggtgcggaaaga Yo 180 actttcaaag ccggcgacaa agacaacagt ctcaacacag gcaaactgaa gaacgacaaa 240 790 gvoy

-١١/4- atcagccgct tcgactttat ccgtcaaatc gaagtggacg ggcagcetcat taccttggag 300 agcggagagt tccaagtgta caaacaaagc cattccgcct taaccgecct tcagaccgag 360 ‏هه‎ ‎caagtacaag actcggagca ttccgggaag atggttgcga aacgccagtt cagaatcgge 420 gacatagtgg gcgaacatac atcttttgac aagcttccca aagacgtcat ggcgacatat ٠ 480 cgcgggacgg cgttcggttc agacgatgcc ggcggaaaac tgacctacac catagatttc 540

Vo gccgccaagce agggacacgg caaaatcgaa catttgaaat cgcctgaact caatgttgac 600 ctggccgecg ccgatatcaa gccggatgaa aaacaccatg ccgtcatcag cggttecgte 660 0 (voy

— \ 7 c ctttacaacc aagccgagaa aggcagttac tctctaggca tetttggcgg gcaageccag 720 gaagttgccg gcagcgegga agtggaaacc gcaaacggca tacgccatat cggtcttgec 780 © gccaagcaat aa 792 <210> 7 0 <211> 768 <211> 768 Bing Neisseria <212> DNA group Neisser2 <212> 7 10 tgcagcagcg 139190992099 tgtcgecgee gacatcggeg cggggcettge cgatgcacta 60 accgcaccgc tcgaccataa agacaaaagt ttgcagtctt tgacgctgga tcagtcegte 120 0 gvoy

-١75- aggaaaaacg agaaactgaa gctggcggca caaggtgcgg aaaaaactta tggaaacggc 180 gacagcctta atacgggcaa attgaagaac gacaaggtca gccgtttcga ctttatccgt © 240 caaatcgaag tggacgggca gctcattacc ttggagagcg gagagttcca agtgtacaaa 300 ٠١ caaagccatt ccgccttaac cgcccttcag accgagcaag 383-38038106 agagcattce 360 gggaagatgg ttgcgaaacg ccggttcaaa atcggcgaca tagcgggcga acatacatct 420 ١٠ tttgacaagc ttcccaaaga cgtcatggcg acatatcgcg ggacggcegtt cggttcagac 480 gatgccggceg gaaaactgac ctatactata gattitgctg ccaaacaggg acacggcaaa 0 00 540 gvoy

-١ ‏لال‎ ‎atcgaacatt tgaaatcgcc cgaactcaat gtcgagcettg ccaccgcecta tatcaagecg 600 gatgaaaaac accatgccgt catcagcggt tccgtecttt acaatcaaga cgagaaagge ه٠‎ 660 agttactccc tcggtatctt tggcgggcaa gcccaggaag ttgccggecag cgcggaagtg 720 ٠١ gaaaccgcaa acggcataca ccatatcggt cttgccgcca agcaataa 768 <210> 8 ١٠ <211> 768 <212> DNA < 213> Neisseria meningitidis (group B) <400> 8 0 gvoy

-١ ‏حي‎ ‎tgcagcagcg gagggggegg tgtcgecgee gacatcggtg cggggcttge cgatgcacta 60 accgcaccgc tcgaccataa agacaaaggt ttgcagtctt taacgctgga tcagtcegte 120 ‏هه‎ ‎aggaaaaacg agaaactgaa gctggcggca caaggtgcgg aaaaaactta tggaaacggc 180 gacagcctta atacgggcaa attgaagaac gacaaggtca gccgcttcga ctttatccgt ٠ 240 caaatcgaag tggacgggaa gctcattacc ttggagagcg gagagttcca agtgtacaaa 300

Vo caaagccatt ccgccttaac cgcccttcag accgagcaag tacaagactc ggaggattce 360 gggaagatgg ttgcgaaacg ccagttcaga atcggcgaca tagcgggcga acatacatct 420 000 (voy

-١و-‎ tttgacaagc ttcccaaagg cggcagtgeg acatatcgeg ggacggcegtt cggttcagac 480 gatgctggcg gaaaactgac ctatactata gatttcgccg ccaagcaggg acacggcaaa 540 oo atcgaacatt tgaaatcgcc cgaactcaat gtcgagcettg ccaccgcecta tatcaagecg 600 gatgaaaaac gccatgccgt tatcageggt tcegtecttt acaaccaaga cgagaaagge ٠ 660 agttactccc tcggtatctt tggcgggcaa gecccaggaag ttgccggecag cgecggaagtg 720

Vo gaaaccgcaa acggcataca ccatatcggt cttgccgcca agcagtaa 768 <210> 9 0 <211> 768 (voy

_ \ A «= <212> DNA <213> Neisseria meningitidis (group B) <400> 9 tgcagcagcg gaggeggcegg tgtcgecgee gacatcggeg cggtgcttge cgatgecacta © 60 accgcaccgc tcgaccataa agacaaaagt ttgcagtctt tgacgctgga tcagtcegte 120 ٠١ aggaaaaacg agaaactgaa gctggcggca caaggtgcgg aaaaaactta tggaaacggc 180 gacagcctca atacgggcaa attgaagaac gacaaggtca gccgcttcga ctttatccgt 240 10 caaatcgaag tggacgggca gctcattacc ttggagagcg gagagttcca agtgtacaaa 300 caaagccatt ccgccttaac cgcccticag accgagcaag tacaagattc ggagceattca 0 360 gvoy

-١م١-‎ gggaagatgg ttgcgaaacg ccagttcaga atcggcgata tagcgggtga acatacatct 420 tttgacaagc ttcccgaagg cggcagggceg acatatcgeg ggacggcatt cggttcagac ه٠‎ 480 gatgccagtg gaaaactgac ctacaccata gatttcgccg ccaagcaggg acacggcaaa 540 ٠١ atcgaacatt tgaaatcgcc agaactcaat gttgacctgg ccgectecga tatcaagcecg 600 gataaaaaac gccatgccgt catcagcggt tcegtecttt acaaccaage cgagaaaggce 660 eo agttactctc taggcatctt tggcgggcaa gcccaggaag ttgccggecag cgcagaagtg 720 gaaaccgcaa acggcatacg ccatatcggt cttgccgccaagcagtaa 0 768 gvoy

—VAY- <210> 10 <211> 768 <212> DNA ٠ <213> Neisseria meningitidis (group B) <400> 10 tgcagcagcg gagggggtagg tgtcgecgec gacatecggtg cggggcttge cgatgecacta 60 0٠ accgcaccgc tcgaccataa agacaaaggt ttgcagtctt tgacgctgga tcagtcegte 120 aggaaaaacg agaaactgaa gctggcggca caaggtgcgg aaaaaactta tggaaacggt 10 180 gacagcctca atacgggcaa attgaagaac gacaaggtca gccgtttcga ctttatccgce 240 790 gvoy

-١ ‏م‎ ‎caaatcgaag tggacgggca gctcattacc ttggagagtg gagagttcca agtatacaaa 300 caaagccatt ccgccttaac cgcctttcag accgagcaaa tacaagattc ggagcattcc 360 ‏هه‎ ‎gggaagatgg ttgcgaaacg ccagttcaga atcggcgaca tagcgggcga acatacatct 420 tttgacaagc ttcccgaagg cggcagggceg acatatcgeg ggacggcegtt cggttcagac ٠ 480 gatgccggceg gaaaactgac ctacaccata gatttcgcecg ccaagcaggg aaacggcaaa 540

Vo atcgaacatt tgaaatcgcc agaactcaat gtcgacctgg ccgecgecga tatcaagecg 600 gatggaaaac gccatgccgt catcagcggt tcegtecttt acaaccaage cgagaaagge 660 0 (voy

-١م6-‎ agttactccc tcggtatctt tggcggaaaa gcccaggaag ttgccggecag cgcggaagtg 720 aaaaccgtaa acggcatacg ccatatcggc cttgccgcca agcaataa 768 © <210> 11 <211> 792 <212> DNA ٠ <213> Neisseria meningitidis (group B) <400> 11 tgcagcageg gaggeggegg aageggaggce ggcggtgteg cecgecgacat cggegegggg 60 100 cttgccgatg cactaaccgce accgctcgac cataaagaca aaggtttgaa atccctgaca 120 gvoy

— \ A ‏اج‎ ‎ttggaagact ccatttccca aaacggaaca ctgaccctgt cggcacaagg tgcggaaaga 180 actttcaaag ccggcgacaa agacaacagt ctcaacacag gcaaactgaa gaacgacaaa 240 ‏هه‎ ‎atcagccgct tcgactttat ccgtcaaatc gaagtggacg ggcagcetcat taccttggag 300 agcggagagt tccaagtgta caaacaaagc cattccgect taaccgecect tcagaccgag ٠ 360 caagtacaag actcggagca ttccgggaag atggttgcga aacgccagtt cagaatcgge 420

Vo gacatagtgg gcgaacatac atcttttggc aagcttccca aagacgtcat ggcgacatat 480 cgcgggacgg cgttcggttc agacgaatgcc ggcggaaaac tgacctacac catagatttc 540 0 (voy

-١م5-‎ 0-00-0339“ agggacacgg caaaatcgaa catttgaaat cgccagaact caatgttgac 600 ctggccgecg ccgatatcaa gccggatgaa aaacaccatg ccgtcatcag cggtteegte 660 © ctttacaacc aagccgagaa aggcagttac tctctaggca tetttggcgg gcaageccag 720 gaagttgccg gcagcgegga agtggaaacc gcaaacggca tacgccatat cggtettgee ٠ 780 gccaagcaat aa 792

Vo <210> 12 <211> 259 <212> PRT <213> Neisseria meningitidis (group B) 790 gvoy

-1 Lm <400> 12

Cys Ser Ser Gly Gly Gly Gly Gly Gly Val Ala Ala Asp lle Gly Thr 1 5 10 15 lo}

Gly Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly 0

Leu Lys Ser Leu Thr Leu Glu Asp Ser lle Pro Gin Asn Gly Thr Leu

Thr Leu Ser Ala GIn Gly Ala Glu Lys Thr Phe Lys Ala Gly Asp Lys yo 60

Asp Asn Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys lle Ser Arg Ed

1- Ham 65 70 75 80

Phe Asp Phe Val GIn Lys lle Glu Val Asp Gly 60 Thr lle Thr Leu 85 90 95 o

Ala Ser Gly Glu Phe Gin lle Tyr Lys GIn Asp His Ser Ala Val Val 100 105 110 01

Ala Leu GIn lle Glu Lys lle Asn Asn Pro Asp Lys lle Asp Ser Leu 115 120 125

Vo lle Asn Gln Arg Ser Phe Leu Val Ser Gly Leu Gly Gly Glu His Thr 130 135 140 gvoy

1-4m Ala Phe Asn GIn Leu Pro Gly Asp Lys Ala Glu Tyr His Gly Lys Ala 145 150 155 160

Phe Ser Ser Asp Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe ° 165 170 175

Ala Ala Lys Gin Gly His Gly Lys lle Glu His Leu Lys Thr Pro Glu 180 185 190 ye da Asn Val Glu Leu Ala Ala Ala Glu Leu Lys Ala Asp Glu Lys Ser 195 200 205

Vo

His Ala Val lle Leu Gly Asp Thr Arg Tyr Gly Ser Glu Glu Lys Gly 210 215 220 gvoy

Thr Tyr His Leu Ala Leu Phe Gly Asp Arg Ala Gin Glu lle Ala Gly 225 230 235 240 lo}

Ser Ala Thr Val Lys lle Gly Glu Lys Val His Glu lle Gly lle Ala 245 250 255

Gly Lys 60 00 <210> 13 <211> 261 10 <212> PRT <213> Neisseria meningitidis (group B) <400> 13 ed.

-141-

Cys Ser Ser Gly Ser Gly Ser Gly Gly Gly Gly Val Ala Ala Asp lle 1 5 10 15 lo}

Gly Thr Gly Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp

Lys Gly Leu Lys Ser Leu Thr Leu Glu Asp Ser lle Ser Gln Asn Gly 00

Thr Leu Thr Leu Ser Ala Gin Gly Ala Glu Lys Thr Phe Lys Val Gly 60 yo

Asp Lys Asp Asn Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys lle 65 70 75 80 Ed

-147-

Ser Arg Phe Asp Phe Val GIn Lys lle Glu Val Asp Gly 60 Thr lle 85 90 95 lo}

Thr Leu Ala Ser Gly Glu Phe GIn lle Tyr Lys Gln Asp His Ser Ala 100 105 110 0

Val Val Ala Leu GIn lle Glu Lys lle Asn Asn Pro Asp Lys lle Asp 115 120 125

Ser Leu lle Asn GIn Arg Ser Phe Leu Val Ser Gly Leu Gly Gly Glu yo 130 135 140

His Thr Ala Phe Asn GIn Leu Pro Ser Gly Lys Ala Glu Tyr His Gly Evidence

-1- 145 150 155 160

Lys Ala Phe Ser Ser Asp Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle 165 170 175 o

Asp Phe Ala Ala Lys GIn Gly His Gly Lys lle Glu His Leu Lys Thr 180 185 190 01

Pro Glu Gln Asn Val Glu Leu Ala Ser Ala Glu Leu Lys Ala Asp Glu 195 200 205

Vo

Lys Ser His Ala Val lle Leu Gly Asp Thr Arg Tyr Gly Ser Glu Glu 210 215 220 gvoy

Lys Gly Thr Tyr His Leu Ala Leu Phe Gly Asp Arg Ala GIn Glu lle 225 230 235 240

Ala Gly Ser Ala Thr Val Lys lle Arg Glu Lys Val His Glu lle Gly ° 245 250 255 lle Ala Gly Lys GIn 260 AD <210> 14 <211> 254 <212> PRT 10 <213> Neisseria meningitidis (group B) < 400 > 14 end

-1-

Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu 1 5 10 15

Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Ser Leu 60°

Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu ye

Ala Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn 60

Vo

Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 65 70 75 80 gvoy

-141-

GIn lle Glu Val Asp Gly GIn Thr lle Thr Leu Ala Ser Gly Glu Phe 85 90 95 lo}

GIn lle Tyr Lys GIn Asn His Ser Ala Val Val Ala Leu Gin lle Glu 100 105 110

Lys lle Asn Asn Pro Asp Lys lle Asp Ser Leu lle Asn GIn Arg Ser 00 115 120 125

Phe Leu Val Ser Gly Leu Gly Gly Glu His Thr Ala Phe Asn GIn Leu 130 135 140 yo

Pro Asp Gly Lys Ala Glu Tyr His Gly Lys Ala Phe Ser Ser Asp Asp 145 150 155 160 Support

-14-

Pro Asn Gly Arg Leu His Tyr Ser lle Asp Phe Thr Lys Lys GIn Gly 165 170 175 lo}

Tyr Gly Arg lle Glu His Leu Lys Thr Pro Glu Gln Asn Val Glu Leu 180 185 190 0

Ala Ser Ala Glu Leu Lys Ala Asp Glu Lys Ser His Ala Val lle Leu 195 200 205

Gly Asp Thr Arg Tyr Gly Gly Glu Glu Lys Gly Thr Tyr His Leu Ala yo 210 215 220

Leu Phe Gly Asp Arg Ala 60 Glu lle Ala Gly Ser Ala Thr Val Lys

-1 pm- 225 230 235 240 lle Arg Glu Lys Val His Glu lle Gly lle Ala Gly Lys 60 245 250 1) <210> 15 <211> 254 <212> PRT 0 <213> Neisseria meningitidis (group B) <400 >15

Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu Vo 1 5 10 15

Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Ser Leu 60 days

-14-

Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu o

Ala Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn 60 01

Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 65 70 75 80

Vo

GIn lle Glu Val Asp Gly GIn Leu lle Thr Leu Glu Ser Gly Glu Phe 85 90 95 gvoy

-Y a=

GIn lle Tyr Lys Gln Asp His Ser Ala Val Val Ala Leu Gin lle Glu 100 105 110

Lys lle Asn Asn Pro Asp Lys lle Asp Ser Leu lle Asn 0 Arg Ser o 115 120 125

Phe Leu Val Ser Gly Leu Gly Gly Glu His Thr Ala Phe Asn GIn Leu 130 135 140 01

Pro Ser Gly Lys Ala Glu Tyr His Gly Lys Ala Phe Ser Ser Asp Asp 145 150 155 160

Vo

Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys Gln Gly 165 170 175 gvoy

_ \ 0 \ —_

His Gly Lys lle Glu His Leu Lys Thr Pro Glu Gln Asn Val Glu Leu 180 185 190 lo}

Ala Ser Ala Glu Leu Lys Ala Asp Glu Lys Ser His Ala Val lle Leu 195 200 205

Gly Asp Thr Arg Tyr Gly Gly Glu Glu Lys Gly Thr Tyr His Leu Ala 00 210 215 220

Leu Phe Gly Asp Arg Ala 60 Glu lle Ala Gly Ser Ala Thr Val Lys 225 230 235 240 Yo lle Arg Glu Lys Val His Glu lle Gly lle Ala Gly Lys GIn 245 250 Ed

_ \ 0 \ —_ <210> 16 <211> 263 <212> PRT E0 <213> Neisseria meningitidis (group B) <400> 16

Cys Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Val Ala Ala Asp 00 1 5 10 15 lle Gly Ala Gly Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys yo

Asp Lys Gly Leu Lys Ser Leu Thr Leu Glu Asp Ser lle Ser Gln Asn Ed

_ \ 0".-

Gly Thr Leu Thr Leu Ser Ala Gin Gly Ala Glu Arg Thr Phe Lys Ala 60 lo}

Gly Asp Lys Asp Asn Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys 65 70 75 80 "lle Ser Arg Phe Asp Phe lle Arg GIn lle Glu Val Asp Gly 60 Leu 85 90 95 lle Thr Leu Glu Ser Gly Glu Phe Gln Val Tire Lys Gln Ser His Ser yo 100 105 110

Ala Leu Thr Ala Leu GIn Thr Glu GIn Val GIn Asp Ser Glu His Ser Ed

-Y 0 ¢— 115 120 125

Gly Lys Met Val Ala Lys Arg Gin Phe Arg lle Gly Asp lle Val Gly 130 135 140 o

Glu His Thr Ser Phe Asp Lys Leu Pro Lys Asp Val Met Ala Thr Tyr 145 150 155 160 01

Arg Gly Thr Ala Phe Gly Ser Asp Asp Ala Gly Gly Lys Leu Thr Tyr 165 170 175

Vo

Thr lle Asp Phe Ala Ala Lys Gln Gly His Gly Lys lle Glu His Leu 180 185 190 gvoy

_ \ Lys Ser Pro Glu Leu Asn Val Asp Leu Ala Ala Ala Asp lle Lys Pro 195 200 205

Asp Glu Lys His His Ala Val lle Ser Gly Ser Val Leu Tyr Asn 60 ° 210 215 220

Ala Glu Lys Gly Ser Tyr Ser Leu Gly lle Phe Gly Gly Gln Ala GIn 225 230 235 240 Yo

Glu Val Ala Gly Ser Ala Glu Val Glu Thr Ala Asn Gly lle Arg His 245 250 255

All Gly Leu Ala Ala Lys 60 260 gvoy

_ \ 0 - <210> 17 <211> 255 <212> PRT <213> Neisseria meningitidis (group B) © <400> 17

Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu 1 5 10 15 AD

Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Ser Leu 60

Vo

Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu Ed

_ \ 0 l Ala Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn 60 lo}

Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 65 70 75 80

GIn lle Glu Val Asp Gly GIn Leu lle Thr Leu Glu Ser Gly Glu Phe 00 85 90 95

GIn Val Tyr Lys GIn Ser His Ser Ala Leu Thr Ala Leu GIn Thr Glu 100 105 110 yo

Gln Glu GIn Asp Pro Glu His Ser Gly Lys Met Val Ala Lys Arg Arg 115 120 125 Ed

_ \ 0 m Phe Lys lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu 130 135 140 lo}

Pro Lys Asp Val Met Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp 145 150 155 160 0

Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys 60 165 170 175

Gly His Gly Lys lle Glu His Leu Lys Ser Pro Glu Leu Asn Val Glu yo 180 185 190

Leu Ala Thr Ala Tyr lle Lys Pro Asp Glu Lys His His Ala Val lle Ed

_ \ 0 q —_ 195 200 205

Ser Gly Ser Val Leu Tyr Asn GIn Asp Glu Lys Gly Ser Tyr Ser Leu 210 215 220 o

Gly lle Phe Gly Gly Gln Ala Gln Glu Val Ala Gly Ser Ala Glu Val 225 230 235 240 0

Glu Thr Ala Asn Gly lle His His lle Gly Leu Ala Ala Lys 07 245 250 255

Vo <210> 18 <211> 255 <212> PRT <213> Neisseria meningitidis (group B) supported

_ \ \ «= <400> 18

Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu 1 5 10 15 o

Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu 0 01

Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu

Vo

Ala Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn 60 gvoy

11-

Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 65 70 75 80

GIn lle Glu Val Asp Gly Lys Leu lle Thr Leu Glu Ser Gly Glu Phe ° 85 90 95

GIn Val Tyr Lys GIn Ser His Ser Ala Leu Thr Ala Leu 60 Thr Glu 100 105 110 AD Da Val Gln Asp Ser Glu Asp Ser Gly Lys Met Val Ala Lys Arg 60 115 120 125

Vo

Phe Arg lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu 130 135 140 gvoy

11-

Pro Lys Gly Gly Ser Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp 145 150 155 160 lo}

Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys 60 165 170 175

Gly His Gly Lys lle Glu His Leu Lys Ser Pro Glu Leu Asn Val Glu 00 180 185 190

Leu Ala Thr Ala Tyr lle Lys Pro Asp Glu Lys Arg His Ala Val lle 195 200 205 yo

Ser Gly Ser Val Leu Tyr Asn GIn Asp Glu Lys Gly Ser Tyr Ser Leu 210 215 220 Ed

11+

Gly lle Phe Gly Gly Gln Ala Gln Glu Val Ala Gly Ser Ala Glu Val 225 230 235 240 lo}

Glu Thr Ala Asn Gly lle His His lle Gly Leu Ala Ala Lys 60 245 250 255 0 <210> 19 <211> 255 <212> PRT <213> Neisseria meningitidis (group B)

Vo <400> 19

Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Val Leu 1 5 10 15 Ed

-?164-

Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Ser Leu Gin lo}

Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu 0

Ala Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn 60

Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg yo 65 70 75 80

GIn lle Glu Val Asp Gly GIn Leu lle Thr Leu Glu Ser Gly Glu Phe ed

-Y \ c 85 90 95

GIn Val Tyr Lys GIn Ser His Ser Ala Leu Thr Ala Leu GIn Thr Glu 100 105 110 o

Gln Val GIn Asp Ser Glu His Ser Gly Lys Met Val Ala Lys Arg Gin 115 120 125 01

Phe Arg lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu 130 135 140

Vo

Pro Glu Gly Gly Arg Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp 145 150 155 160 gvoy

-? 17-

Asp Ala Ser Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys 60 165 170 175

Gly His Gly Lys lle Glu His Leu Lys Ser Pro Glu Leu Asn Val Asp ° 180 185 190

Leu Ala Ala Ser Asp lle Lys Pro Asp Lys Lys Arg His Ala Val lle 195 200 205 AD

Ser Gly Ser Val Leu Tyr Asn GIn Ala Glu Lys Gly Ser Tyr Ser Leu 210 215 220

Vo

Gly lle Phe Gly Gly Gln Ala Gln Glu Val Ala Gly Ser Ala Glu Val 225 230 235 240 gvoy

-?111/-

Glu Thr Ala Asn Gly lle Arg His lle Gly Leu Ala Ala Lys Gin 245 250 255 lo} <210> 20 <211> 255 <212> PRT <213> Neisseria meningitidis (group B) 0 <400> 20

Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu 1 5 10 15

Vo

Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu 0 support

Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu lo}

Ala Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn 60

Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 00 65 70 75 80

GIn lle Glu Val Asp Gly GIn Leu lle Thr Leu Glu Ser Gly Glu Phe 85 90 95 yo

Gln Val Tyr Lys Gln Ser His Ser Ala Leu Thr Ala Phe GIn Thr Glu 100 105 110 Ed

-?14- Ha lle any Asp Ser Glu His Ser Gly Lys Met Val Ala Lys Arg Gin 115 120 125 lo}

Phe Arg lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu 130 135 140 0

Pro Glu Gly Gly Arg Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp 145 150 155 160

Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys 60 yo 165 170 175

Gly Asn Gly Lys lle Glu His Leu Lys Ser Pro Glu Leu Asn Val Asp

_ \ \ «= 180 185 190

Leu Ala Ala Ala Asp lle Lys Pro Asp Gly Lys Arg His Ala Val lle 195 200 205 o

Ser Gly Ser Val Leu Tyr Asn GIn Ala Glu Lys Gly Ser Tyr Ser Leu 210 215 220 01

Gly lle Phe Gly Gly Lys Ala Gln Glu Val Ala Gly Ser Ala Glu Val 225 230 235 240

Vo

Lys Thr Val Asn Gly lle Arg His lle Gly Leu Ala Ala Lys Gin 245 250 255 gvoy

-?71- <210> 21 <211> 263 <212> PRT <213> Neisseria meningitidis (group B) lo} <400> 21

Cys Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Val Ala Ala Asp 1 5 10 15 0 lle Gly Ala Gly Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys

Vo

Asp Lys Gly Leu Lys Ser Leu Thr Leu Glu Asp Ser lle Ser Gln Asn Ed

-???- Gly Thr Leu Thr Leu Ser Ala Gin Gly Ala Glu Arg Thr Phe Lys Ala 60 55 50 Gly Asp Lys Asp Asn Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys ° 80 75 70 65 lle Ser Arg Phe Asp Phe lle Arg GIn lle Glu Val Asp Gly 60 Leu I 95 90 85 lle Thr Leu Glu Ser Gly Glu Phe Gln Val Tyr Lys Gln Ser His Ser 110 105 100 Vo Ala Leu Thr Ala Leu GIn Thr Glu 60 Val Gln Asp Ser Glu His Ser 125 120 115 gvoy

+1

Gly Lys Met Val Ala Lys Arg Gin Phe Arg lle Gly Asp lle Val Gly 130 135 140 lo}

Glu His Thr Ser Phe Gly Lys Leu Pro Lys Asp Val Met Ala Thr Tyr 145 150 155 160

Arg Gly Thr Ala Phe Gly Ser Asp Asp Ala Gly Gly Lys Leu Thr Tyr 00 165 170 175

Thr lle Asp Phe Ala Ala Lys GIn Gly His Gly Lys lle Glu His Leu 180 185 190 yo

Lys Ser Pro Glu Leu Asn Val Asp Leu Ala Ala Ala Asp lle Lys Pro 195 200 205 hand

Asp Glu Lys His His Ala Val lle Ser Gly Ser Val Leu Tyr Asn 60 220 215 210 lo} Ala Glu Lys Gly Ser Tyr Ser Leu Gly lle Phe Gly Gly Gln Ala GIn 240 235 230 225 0 Glu Val Ala Gly Ser Ala Glu Val Glu Thr Ala Asn Gly lle Arg His 255 250 245 lle Gly Leu Ala Ala Lys 60 yo 260 22 <210> ed

— \ \ Aj <211> 26 <212> DNA <213> Artificial <220> ° <223> Synthetic nucleotide sequence: Forward primer <400> 22 tgccatatga gcagcggaag cggaag 26

Ya <210> 23 <211> 27 <212> DNA <213> Artificial 01 <220> <223> Synthetic nucleotide sequence: Reverse primer (yoy)

<400> 23 cggatcccta ctgtttgccg gegatgce 27 <210> 24 o <211> 49 <212> DNA <213> Artificial <220> 0 0 <223> Synthetic nucleotide sequence: Forward primer <400> 24 tttcttccccg ggaaggagat atacatatgt gcagcagcgg aggeggegg 49

Vo <210> 25 <211> 38 <212> DNA (yoy

—YYV- <213> Artificial <220> <223> Synthetic nucleotide sequence: Reverse primer lo} <400> 25 tttcttgctc agcattattg cttggcggcea agaccgat 38 <210> 26 0 <211> 46 <212> DNA <213> Artificial <220> 1 <223> Synthetic nucleotide sequence: Forward primer <400> 26 tttcttcccg ggaaggagat atacatatga gcagcggagg cggcegg 46 (yoy)

—YYA-

<210> 27

<211> 38 <212> DNA oo

<213> Artificial

<220>

<223> Synthetic nucleotide sequence: Reverse primer 01

<400> 27 tttcttgctc agcattattg cttggcggcea agaccgat 38 <210> 28 10

<211> 36

<212> DNA

<213> Artificial

Lad

<220< Synthetic nucleotide sequence <223< 28 <400< atgagctctg gaggtggagg aagcggggge ggtgga 36 o 29 <210< 12 <211< PRT 0 <212< Artificial <213< <220< Synthetic amino acid sequence <223< Vo 29 <400< Met Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 5 1

—YY.-

<210> 30

<211> 33 <212> DNA H

<213> Artificial

<220>

<223> Synthetic nucleotide sequence 01

<400> 30 atgagctctg gaagcggaag cgggggeggt gga 33 <210> 31 1

<211> 11

<212> PRT

<213> Artificial

AAA

<220> <223> Synthetic amino acid sequence <400> 31 lo}

Met Ser Ser Gly Ser Gly Ser Gly Gly Gly Gly 1 5 10 <210> 32 0 <211> 21 <212> DNA <213> Artificial <220> 10 <223> Synthetic nucleotide sequence <400> 32 atgagctctg gaggtggagg a 21 ( yoy

1 <210> 3 <211> 7 <212> PRT Eh <213> Artificial <220> <223> Synthetic amino acid sequence 01 <400> 3

Met Ser Ser Gly Gly Gly Gly 1 5

Vo <210> 34 <211> 21 <212> DNA gyov

aa Artificial <213< <220< Synthetic nucleotide sequence <223< lo} ‏34 <400< atgagcagcg ggggeggtgg a 21 0 35 <210< 28 <211< PRT <212< Neisseria meningitidis (group B) <213< 0 35 <400< Cys Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Val Thr Ala Asp 10 5 1 ed

—Yye— lle Gly Thr Gly Leu Ala Asp Ala Leu Thr Ala Pro lo} <210> 36 <211> 28 <212> PRT <213> Neisseria meningitidis (group B) 0 <400> 36

Cys Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Val Ala Ala Asp 1 5 10 15

Vo lle Gly Ala Gly Leu Ala Asp Ala Leu Thr Ala Pro 20 25 Ed

AA <210> 7 <211> 27 <212> PRT <213> Neisseria meningitidis (group B) © <400> 37

Cys Ser Ser Gly Ser Gly Ser Gly Gly Gly Gly Val Ala Ala Asp lle 1 5 10 15 AD

Gly Thr Gly Leu Ala Asp Ala Leu Thr Ala Pro

Vo <210> 38 <211> 23 <212> PRT supported

-M? Neisseria meningitidis (group B) <213< 38 <400< Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu ° 10 5 1 Ala Asp Ala Leu Thr Ala Pro 01 20 9 <210> 3 <211> PRT 10 <212> Neisseria meningitidis (group B) <213> 39 <400> gyov

Tafn Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Val Leu 10 5 1 Ala Asp Ala Leu Thr Ala Pro °

<210< 0 23 <211< PRT <212> Neisseria meningitidis (group B) <213< 40 <400< Vo Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu 15 10 5 1 ed

—YYA-

Ala Asp Ala Leu Thr Ala Pro <210> 41 AH <211> 15 <212> PRT <213> Artificial <220> 0 <223> Synthetic amino acid sequence <220> <221> MISC FEATURE 101 <222> (15)..(15) <223> XisGorV <400> 41 gyov

-4+?- 01 <220> Synthetic nucleotide sequence <223< 42 <400> atgagcagcg gaggcggcegg tgtcgecgee gacatcggeg cgggg 01 45 43 <210> gvoy

_ \ ¢ «= <211> 789 <212> DNA <213> Artificial <220> o <223> Synthetic nucleotide sequence <400> 43 agctctggag gtggaggaag cgggggceggt ggagttgcag cagacattgg agcaggatta 60 0٠ gcagatgcac tgacggcacc gttggatcat aaagacaaag gcttgaaatc gcttacctta 120 gaagattcta tttcacaaaa tggcaccctt accttgtccg cgcaaggcge tgaacgtact Yo 180 tttaaagccg gtgacaaaga taatagctta aatacaggta aactcaaaaa tgataaaatc 240 790 gvoy

-?41- tcgcgttttg atttcattcg tcaaatcgaa gtagatggcc aacttattac attagaaagce 300 ggtgaattcc aagtatataa acaatcccat tcagcactta cagcattgca aaccgaacag 360 lo}

gtccaagact cagaacattc cggcaaaatg gtagctaaac gtcaattccg catcggtgac 420 attgtcggtg aacatacaag cttcggaaaa ttaccaaaag atgtgatggc gacctatcge

480 00 ggtacggcat ttggatcaga tgatgcaggc ggtaaattaa cttatacaat tgactttgca 540 gcaaaacaag gacatggcaa aattgaacat ttaaaatctc ccgaacttaa cgtagatctc

600 10 gcagcagcag atattaaacc agatgaaaaa caccacgcag tcatttcagg ttcagtttta 660 tacaatcagg cagaaaaagg ttcgtactct ttaggtattt ttggcgggca agctcaagaa 0 720 gvoy

gttgcaggta gcgcagaagt agaaacggca aatggcattc gtcacattgg gttagcggeg 780 AH 789 aaacaataa 44 <210< 262 <211< PRT 0 <212< Artificial <213< <220< Synthetic amino acid sequence <223< Vo 44 <400< Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Val Ala Ala Asp lle 10 5 1 gvoy

110

Gly Ala Gly Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp lo}

Lys Gly Leu Lys Ser Leu Thr Leu Glu Asp Ser lle Ser Gln Asn Gly 0

Thr Leu Thr Leu Ser Ala Gin Gly Ala Glu Arg Thr Phe Lys Ala Gly 60

Asp Lys Asp Asn Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys lle yo 65 70 75 80

Ser Arg Phe Asp Phe lle Arg Gln lle Glu Val Asp Gly GIn Leu lle Ed

95 90 85 Val Tyr Lys Gln Ser His Ser Ala Ha Thr Leu Glu Ser Gly Glu Phe o 110 105 100 Leu Thr Ala Leu GIn Thr Glu GIn Val Gln Asp Ser Glu His Ser Gly 125 120 115 01 Lys Met Val Ala Lys Arg Gln Phe Arg lle Gly Asp lle Val Gly Glu 140 135 130 Vo His Thr Ser Phe Gly Lys Leu Pro Lys Asp Val Met Ala Thr Tyr Arg 160 155 150 145 gvoy

— \ ¢ Gly Thr Ala Phe Gly Ser Asp Asp Ala Gly Gly Lys Leu Thr Tyr Thr 165 170 175 lle Asp Phe Ala Ala Lys Gln Gly His Gly Lys lle Glu His Leu Lys ° 180 185 190

Ser Pro Glu Leu Asn Val Asp Leu Ala Ala Ala Asp lle Lys Pro Asp 195 200 205 AD

Glu Lys His His Ala Val lle Ser Gly Ser Val Leu Tyr Asn Gin Ala 210 215 220

Vo

Glu Lys Gly Ser Tyr Ser Leu Gly lle Phe Gly Gly Gln Ala Gln Glu 225 230 235 240 gvoy

Val Ala Gly Ser Ala Glu Val Glu Thr Ala Asn Gly lle Arg His lle 255 250 245 lo} Gly Leu Ala Ala Lys GIn 260 © 45 <210< 780 <211< DNA <212< Artificial <213< 10 <220< Synthetic nucleotide sequence <223< <400> hands

—Y¢vV- agctctggag gtggaggaag cgggggceggt ggagttgcag cagacattgg agcaggatta 60 gcagatgcac tgacggcacc gttggatcat aaagacaaag gcttgcagtc gcttacctta 120 ‏هه‎ ‎gatcagtctg tcaggaaaaa tgagaaactt aagttggcgg cgcaaggcgce tgaaaaaact 180 tatggaaacg gtgacagctt aaatacaggt aaactcaaaa atgataaagt ctcgegtttt ٠ 240 gatttcattc gtcaaatcga agtagatggc aagcttatta cattagaaag cggtgaattc 300 caagtatata aacaatccca ttcagcactt acagcattgc aaaccgaaca ggtccaagac 10 360 tcagaagatt ccggcaaaat ggtagctaaa cgtcaattcc gcatcggtga cattgcgggat 420 790 gvoy

-م؛؟- ‎gaacatacaa gcttcgacaa attaccaaaa ggcggcagtg cgacctatcg cggtacggca‏ 480 ‎tttggatcag atgatgcagg cggtaaatta acttatacaa ttgactttgc agcaaaacaa‏ ‎oo‏ 540 ggacatggca aaattgaaca tttaaaatct cccgaactta acgtagagct cgcaaccgca 600 tatattaaac cagatgaaaa acgccacgca gtcatttcag gttcagtttt atacaatcag 660 0٠ gacgaaaaag gttcgtactc tttaggtatt tttggcgggc aagctcaaga agttgcaggt 720 agcgcagaag tagaaacggc aaatggcatt caccacattg ggttagcggc gaaacaataa Vo 780 <210> 46

<211> 765 000 gvoy

DNA <212< Artificial <213< <220< 0e Synthetic nucleotide sequence <223< 46 <400< agctctggag gtggaggagt tgcagcagac attggagcag gattagcaga tgcactgacg 60 01 gcaccgttgg atcataaagaga caaaggcttg cagtcgct1ct2ctgtca g aaaaatgaga aacttaagtt ggcggcgcaa ggcgctgaaa aaacttatgg aaacggtgac 10 180 agcttaaata caggtaaact caaaaatgat aaagtctcgc gttttgattt cattcgtcaa 240 atcgaagtag atggcaagct tattacatta gaaagcggtg avotccaagt atataa3a0 0gay0

_ \ OW — tcccattcag cacttacagc attgcaaacc gaacaggtcc aagactcaga agattccggce 360 aaaatggtag ctaaacgtca attccgcatc ggtgacattg cgggtgaaca tacaagcttc ©

420 gacaaattac caaaaggcgg cagtgcgacc tatcgcggta cggcatttgg atcagatgat 480

01 gcaggcggta aattaactta tacaattgac tttgcagcaa aacaaggaca tggcaaaatt 540 gaacatttaa aatctcccga acttaacgta gagctcgcaa ccgcatatat taaaccagat

600 10 gaaaaacgcc acgcagtcat ttcaggttca gttttataca atcaggacga aaaaggttcg 660 tactctttag gtatttttgg cgggcaagct caagaagttg caggtagcgc agaagtagaa 00 720 gvoy

— \ o \ — acggcaaatg gcattcacca cattgggtta gcggcgaaac aataa 765 <210> 47 o

<211> 765

<212> DNA

<213> Artificial <220> 11

<223> Synthetic nucleotide sequence

<400> 47 agcagcgggg gcggtggagt tgcagcagac attggagcag gattagcaga tgcactgacg 60 100 gcaccgttgg atcataaaga caaaggcttg cagtcgctta ccttagatca gtctgtcagg

120 gvoy

— \ o \ — aaaaatgaga aacttaagtt ggcggcgcaa ggcgctgaaa aaacttatgg aaacggtgac 180 agcttaaata caggtaaact caaaaatgat aaagtctcgc gttttgattt cattcgtcaa 240 lo} atcgaagtag atggcaagct tattacatta gaaagcggtg aattccaagt atataaacaa 300 tcccattcag cacttacagc attgcaaacc gaacaggtcc aagactcaga agattccggce 360 ٠ aaaatggtag ctaaacgtca attccgcatc ggtgacattg cgggtgaaca tacaagcttc 420 gacaaattac caaaaggcgg cagtgcgacc tatcgcggta cggcatitgg atcagatgat ٠ 480 gcaggcggta aattaactta tacaattgac tttgcagcaa aacaaggaca tggcaaaatt 540 790 gvoy

— \ o — gaacatttaa aatctcccga acttaacgta gagctcgcaa ccgcatatat taaaccagat 600 gaaaaacgcc acgcagtcat ttcaggttca gttttataca atcaggacga 388809129 660 © tactctttag gtatttttgg cgggcaagct caagaagttg caggtagcgc agaagtagaa 720 acggcaaatg gcattcacca cattgggtta gcggcgaaac aataa 765 0٠ <210> 48 <211> 765 <212> DNA ٠ <213> Artificial <220> <223> Synthetic nucleotide sequence 790 gvoy

— \ o ¢ — <400> 48 agcagcggag ggggcggtgt cgccgecgac atcggtgegg ggcttgecga tgcactaace 60 gcaccgctcg accataaaga caaaggtttg cagtctttaa cactggatca gtccgtcagg ه٠‎ 120 aaaaacgaga aactgaagct ggcggcacaa ggtgcggaaa aaacttatgg aaacggcgac 180 ٠١ agccttaata cgggcaaatt gaagaacgac aaggtcagcc gcttcgactt tatccgtcaa 240 atcgaagtgg acgggaagct cattaccttg gagagcggag agttccaagt gtacaaacaa 300 ١٠ agccattccg ccttaaccgce ccttcagacc gagcaagtac aagactcgga ggattccggg 360 aagatggttg cgaaacgcca gttcagaatc ggcgacatag cgggcgaaca tacatetttt 0 420 gvoy

_ \ 00 — gacaagcttc ccaaaggcgg cagtgcgaca tatcgcggga cggcegttcgg ttcagacgat 480 gctggcggaa aactgaccta tactatagat ttcgccgecca agcagggaca cggcaaaatc © 540 gaacatttga aatcgcccga actcaatgtc gagcttgcca ccgcectatat caccage 600

I gaaaaacgcc atgccgttat cagcggttcc gtcctttaca accaagacga gaaaggcagt 660 tactcccteg gtatctttgg cgggcaagcec caggaagtty ccggcagegce ggaagtggaa 720 10 accgcaaacg gcatacacca tatcggtctt gccgccaage agtaa 765 <210> 49 0 (voy

— \ o 1- <211> 254 <212> PRT <213> Artificial <220> o <223> Synthetic amino acid sequence <400> 49

Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu Ala 00 1 5 10 15

Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu GIn Ser yo

Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu Ala 40 35 Ed

— \ o 7 —

Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn Thr 60 lo}

Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 60 65 70 75 80 01 lle Glu Val Asp Gly Lys Leu lle Thr Leu Glu Ser Gly Glu Phe 60 85 90 95

Val Tyr Lys GIn Ser His Ser Ala Leu Thr Ala Leu Gln Thr Glu 60 yo 100 105 110

Val GIn Asp Ser Glu Asp Ser Gly Lys Met Val Ala Lys Arg Gin Phe gvoy

— \ o m 115 120 125

Arg lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu Pro 130 135 140 o

Lys Gly Gly Ser Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp Asp 145 150 155 160 01

Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys Gln Gly 165 170 175

Vo

His Gly Lys lle Glu His Leu Lys Ser Pro Glu Leu Asn Val Glu Leu 180 185 190 gvoy

_ \ o q —_

Ala Thr Ala Tyr lle Lys Pro Asp Glu Lys Arg His Ala Val lle Ser 195 200 205

Gly Ser Val Leu Tyr Asn GIn Asp Glu Lys Gly Ser Tyr Ser Leu Gly ° 210 215 220 lle Phe Gly Gly Gln Ala 60 Glu Val Ala Gly Ser Ala Glu Val Glu 225 230 235 240 Yo

Thr Ala Asn Gly lle His His lle Gly Leu Ala Ala Lys 60 245 250

Vo <210> 50 <211> 259 <212> PRT gvoy

Artificial >213< >220< Synthetic amino acid sequence >223< lo}‏ 50 >400< Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Val Ala Ala Asp lle 10 5 1 0 Gly Ala Gly Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp 25 20 Vo Lys Gly Leu GIn Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu 40 35 Ed

-?+1-

Lys Leu Lys Leu Ala Ala Gin Gly Ala Glu Lys Thr Tyr Gly Asn Gly 60

Asp Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe ° 65 70 75 80

Asp Phe lle Arg GIn lle Glu Val Asp Gly Lys Leu lle Thr Leu Glu 85 90 95 I

Ser Gly Glu Phe Ha Val Tyr Lys Gln Ser His Ser Ala Leu Thr Ala 100 105 110

Vo

Leu GIn Thr Glu Mah Val Gln Asp Ser Glu Asp Ser Gly Lys Met Val 115 120 125 gvoy

Ala Lys Arg GIn Phe Arg lle Gly Asp lle Ala Gly Glu His Thr Ser 140 135 130 lo} Phe Asp Lys Leu Pro Lys Gly Gly Ser Ala Thr Tyr Arg Gly Thr Ala 160 155 150 145 Phe Gly Ser Asp Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe 00 175 170 165 Ala Ala Lys GIn Gly His Gly Lys lle Glu His Leu Lys Ser Pro Glu yo 190 185 180 Leu Asn Val Glu Leu Ala Thr Ala Tyr lle Lys Pro Asp Glu Lys Arg 205 200 195 Ed

—Yiyr-

His Ala Val lle Ser Gly Ser Val Leu Tyr Asn GIn Asp Glu Lys Gly 210 215 220 lo}

Ser Tyr Ser Leu Gly lle Phe Gly Gly Gln Ala Gln Glu Val Ala Gly 225 230 235 240 0

Ser Ala Glu Val Glu Thr Ala Asn Gly lle His His lle Gly Leu Ala 245 250 255

Ala Lys 60 yo <210> 1 hand

-4+?- 789 <211< DNA <212< Artificial <213< ht <220< Synthetic nucleotide sequence <223< 1 <400< agcagcggag gcggcggaag cggaggeggce ggtgtcgecg ccgacatcgg cgeggggctt 00 60 gccgatgcac taaccgcacc gctcgaccat aaagacaaag gtttgaaatc cctgacattg 120 gaagactcca tttcccaaaa cggaacactg accctgtcgg cacaaggtgc ggaaagaact Vo 180 ttcaaagccg gcgacaaaga caacagtctc aacacaggca aactgagavoa cgacaaaatc 0 240g 79

-ه)؟- ‎agccgcttcg actttatccg tcaaatcgaa gtggacgggc agctcattac cttggagagce‏ 300 ‎ggagagttcc aagtgtacaa acaaagccat tccgccttaa ccgcccttca gaccgagcaa‏ هه 360 ‎gtacaagact cggagcattc cgggaagatg gttgcgaaac gccagttcag aatcggcgac‏ 420 ‎atagtgggcg aacatacatc ttttggcaag cttcccaaag acgtcatggc gacatatcge ٠‏ 480 ‎gggacggcgt tcggttcaga cgatgccggec ggaaaactga cctacaccat agatttcgec 540 Vo gccaagcagg gacacggcaa aatcgaacat ttgaaatcgc cagaactcaa tgttgacctg 600 gccgecgecg atatcaagcc ggatgaaaaa caccatgccg tcatcagcgg ttccgtectt 0 660 (voy)

-7+?- tacaaccaag ccgagaaagg cagttactct ctaggcatct ttggcgggca agcccaggaa 720 gttgccggca gcgcggaagt ggaaaccgca aacggcatac gccatatcgg tecttgecgec © 780 aagcaataa 789 00 52 >210<2 >211<13 >ArtiDNA212<1 10 <220< Synthetic nucleotide sequence <223< 52 <400< atgagcagcg gaggcggcegg tgtcgecgee gacatcggeg cggtg 45 gvoy

—Yiv- <210> 53 <211> 45 <212> DNA oo <213> Artificial <220> <223> Synthetic nucleotide sequence a <400> 53 atgagcagcg gagggggcegg tgtcgecgece gacatcggtg 09 45 <210> 54 10 <211> 783 < 212> DNA <213> Artificial R

م+؟- >220< ‎Synthetic nucleotide sequence‏ >223< 54 >400< ‎agcagcggaa gcggaagegg aggeggeggt gtcgecgecg acatcggcac agggcettgee ٠‏ 60 ‎gatgcactaa ctgcgccgct cgaccataaa gacaaaggtt tgaaatccct gacattggaa‏ 120 ‎٠١‏ ‎gactccattt cccaaaacgg aacactgacc ctgtcggcac aaggtgcgga aaaaactttc‏ 180 aaagtcggcg acaaagacaa cagtctcaat acaggcaaat tgaagaacga caaaatcagc 10 240 cgcttcgact ttgtgcaaaa aatcgaagtg gacggacaaa ccatcacgct ggcaagcggce 300 gaatttcaaa tatacaaaca ggaccactcc gccgtcgttg0 voctaacagat 300 voctaacagat 3

-4+؟- ‎aacaaccccg acaaaatcga cagcctgata aaccaacgct ccttccttgt cagcggtttg‏ 420 ‎ggcggagaac ataccgcctt caaccaactg cccagcggca aagccgagta tcacggcaaa ٠‏ 480 ‎gcattcagct ccgacgatgc cggcggaaaa ctgacctata ccatagattt tgccgccaaa‏ 540 ‎٠١‏ ‎cagggacacg gcaaaatcga acacctgaaa acacccgagc agaatgtcga gcttgcctcce‏ 600 ‎gccgaactca aagcagatga aaaatcacac gccgtcattt tgggcgacac gcgctacgge eo 660 agcgaagaaa aaggcactta ccacctcgct cttttcggcg accgagccca agaaatcgcec 720 ggctcggcaa ccgtgaagat aagggaaaag gttcacgaaa tcggcatcgc cggcaaacag 0 780 gvoy

_ \ 7 «= tag 783 <210> 55 oo <211> 260 <212> PRT <213> Artificial <220> 0 <223> Synthetic amino acid sequence <400> 5

Ser Ser Gly Ser Gly Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly yo 1 5 10 15

Thr Gly Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Ed

-Except?- 25 20 Gly Leu Lys Ser Leu Thr Leu Glu Asp Ser lle Ser Gln Asn Gly Thr o 45 40 35 Leu Thr Leu Ser Ala GIn Gly Ala Glu Lys Thr Phe Lys Val Gly Asp 60 55 50 01 Lys Asp Asn Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys lle Ser 80 75 70 65 Vo Arg Phe Asp Phe Val GIn Lys lle Glu Val Asp Gly Gin Thr lle Thr 95 90 85 gvoy

—-YVY-

Leu Ala Ser Gly Glu Phe Gin lle Tyr Lys Gln Asp His Ser Ala Val 100 105 110

Val Ala Leu GIn lle Glu Lys lle Asn Asn Pro Asp Lys lle Asp Ser ° 115 120 125

Leu lle Asn Gln Arg Ser Phe Leu Val Ser Gly Leu Gly Gly Glu His 130 135 140 01

Thr Ala Phe Asn GIn Leu Pro Ser Gly Lys Ala Glu Tyr His Gly Lys 145 150 155 160

Vo

Ala Phe Ser Ser Asp Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp 165 170 175 gvoy

—YVY¥-

Phe Ala Ala Lys GIn Gly His Gly Lys lle Glu His Leu Lys Thr Pro 180 185 190 lo}

Glu GIn Asn Val Glu Leu Ala Ser Ala Glu Leu Lys Ala Asp Glu Lys 195 200 205

Ser His Ala Val lle Leu Gly Asp Thr Arg Tyr Gly Ser Glu Glu Lys 00 210 215 220

Gly Thr Tyr His Leu Ala Leu Phe Gly Asp Arg Ala 60 Glu lle Ala 225 230 235 240 Yo

Gly Ser Ala Thr Val Lys lle Arg Glu Lys Val His Glu lle Gly lle 255 250 245 Ed

-?1/4-‎

Ala Gly Lys 60 260 lo} <210> 56 <211> 5 <212> PRT <213> Artificial 0 <220> <223> Synthetic amino acid sequence <400> 56 1

Ser Gly Gly Gly Gly Gly 1 5 gyov

— \ 7 C <210> 7 <211> 9 <212> PRT <213> Artificial © <220> <223> Synthetic amino acid sequence <400> 57 0

Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Val Thr Ala Asp lle 1 5 10 15

Vo

Gly Thr Gly Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Ed

—-YVi-

Lys Gly Leu Lys Ser Leu Thr Leu Glu Asp Ser lle Ser Gln Asn Gly

Thr Leu Thr Leu Ser Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly ° 60

Asp Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe 65 70 75 80 Yo

Asp Phe lle Arg GIn lle Glu Val Asp Gly Gin Leu lle Thr Leu Glu 85 90 95

Vo

Ser Gly Glu Phe Ha Val Tyr Lys Gln Ser His Ser Ala Leu Thr Ala 100 105 110 gvoy

—YVV-

Leu Gln Thr Glu GIn Glu Gln Asp Pro Glu His Ser Glu Lys Met Val 115 120 125 lo}

Ala Lys Arg Arg Phe Arg lle Gly Asp lle Ala Gly Glu His Thr Ser 130 135 140

Phe Asp Lys Leu Pro Lys Asp Val Met Ala Thr Tyr Arg Gly Thr Ala 00 145 150 155 160

Phe Gly Ser Asp Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe 165 170 175 yo

Ala Ala Lys GIn Gly His Gly Lys lle Glu His Leu Lys Ser Pro Glu 180 185 190 support

—YVA-

Leu Asn Val Asp Leu Ala Val Ala Tyr lle Lys Pro Asp Glu Lys His 195 200 205 lo}

His Ala Val lle Ser Gly Ser Val Leu Tyr Asn GIn Asp Glu Lys Gly 210 215 220 0

Ser Tyr Ser Leu Gly lle Phe Gly Glu Lys Ala Gln Glu Val Ala Gly 225 230 235 240

Ser Ala Glu Val Glu Thr Ala Asn Gly lle His His lle Gly Leu Ala yo 245 250 255

Ala Lys 60 hands

—YV4-— <210> 38 <211> 260 E0 <212> PRT <213> Neisseria meningitidis (group B) <400> 38 0

Cys Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Val Thr Ala Asp 1 5 10 15 lle Gly Thr Gly Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys yo

Asp Lys Gly Leu Lys Ser Leu Thr Leu Glu Asp Ser lle Ser 60 Asn Ed

_\A «=

Gly Thr Leu Thr Leu Ser Ala Gln Gly Ala Glu Lys Thr Tyr Gly Asn 60 o

Gly Asp Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg 65 70 75 80 01

Phe Asp Phe lle Arg 60 lle Glu Val Asp Gly GIn Leu lle Thr Leu 85 90 95

Vo

Glu Ser Gly Glu Phe Gln Val Tyr Lys GIn Ser His Ser Ala Leu Thr 100 105 110 gvoy

-?M1-

Ala Leu 60 Thr Glu GIn Glu Gln Asp Pro Glu His Ser Glu Lys Met 115 120 125

Val Ala Lys Arg Arg Phe Arg lle Gly Asp lle Ala Gly Glu His Thr ° 130 135 140

Ser Phe Asp Lys Leu Pro Lys Asp Val Met Ala Thr Tyr Arg Gly Thr 145 150 155 160 01

Ala Phe Gly Ser Asp Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp 165 170 175

Vo

Phe Ala Ala Lys GIn Gly His Gly Lys lle Glu His Leu Lys Ser Pro 180 185 190 gvoy

—YAY-

Glu Leu Asn Val Asp Leu Ala Val Ala Tyr lle Lys Pro Asp Glu Lys 195 200 205 lo}

His His Ala Val lle Ser Gly Ser Val Leu Tyr Asn Gln Asp Glu Lys 210 215 220

Gly Ser Tyr Ser Leu Gly lle Phe Gly Glu Lys Ala Gln Glu Val Ala 00 225 230 235 240

Gly Ser Ala Glu Val Glu Thr Ala Asn Gly lle His His lle Gly Leu 245 250 255 yo

Ala Ala Lys 60 260 hands

—YAY- <210> 59 <211> 255 <212> PRT ee <213> Neisseria meningitidis (group B) <400> 59

Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu 00 1 5 10 15

Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu Gin yo

Ser Leu lle Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu 40 35 Ed

—YAE—

Ala Ala Gln Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn 60 lo}

Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 65 70 75 80"

GIn lle Glu Val Asp Gly GIn Leu lle Thr Leu Glu Ser Gly Glu Phe 85 90 95

Gln Val Tyr Lys GIn Ser His Ser Ala Leu Thr Ala Leu Gln Thr Glu yo 100 105 110

Gln Val GIn Asp Ser Glu His Ser Gly Lys Met Val Ala Lys Arg GIn Ed

— \ A c 115 120 125

Phe Arg lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu 130 135 140 o

Pro Glu Gly Gly Arg Ala Thr Tyr Arg Gly Thr Ala Phe Ser Ser Asp 145 150 155 160 01

Asp Ala Gly Gly Lys Leu lle Tyr Thr lle Asp Phe Ala Ala Lys 60 165 170 175

Vo

Gly His Gly Lys lle Glu His Leu Lys Ser Pro Glu Leu Asn Val Asp 180 185 190 gvoy

-1 m?- Phe Gly Gly Lys Ala Gln Glu Val Ala Gly Ser Ala Glu Val Yo 240 235 230 225 Lys Thr Val Asn Gly lle Arg His lle Gly Leu Ala Ala Lys GIn 255 250 245 Vo 60 <210< 255 <211> PRT <212> R

—YAV-<213>Neisseria meningitidis (group B) <400>60

Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu ° 1 5 10 15

Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Ser Leu 60 01

Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu

Vo

Ala Ala Gln Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn 60 Ed

—YAA-

Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 65 70 75 80 lo}

GIn lle Glu Val Asp Gly GIn Leu lle Thr Leu Glu Ser Gly Glu Phe 85 90 95

Gln Val Tyr Lys GIn Ser His Ser Ala Leu Thr Ala Leu Gln Thr Glu 00 100 105 110 What Val GIn Asp Ser Glu His Ser Gly Lys Met Val Ala Lys Arg 60 115 120 125 yo

Phe Arg lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu 130 135 140 support

—YA4—

Pro Glu Gly Gly Arg Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp 145 150 155 160 lo}

Asp Ala Ser Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys Gin 165 170 175 0

Gly His Gly Lys lle Glu His Leu Lys Ser Pro Glu Leu Asn Val Asp 180 185 190

Leu Ala Ala Ser Asp lle Lys Pro Asp Lys Lys Arg His Ala Val lle yo 195 200 205

Ser Gly Ser Val Leu Tyr Asn Gln Ala Glu Lys Gly Ser Tyr Ser Leu Ed

—Y4.— 210 215 220

Gly lle Phe Gly Gly GIn Ala Gln Glu Val Ala Gly Ser Ala Glu Val 225 230 235 240 o

Glu Thr Ala Asn Gly lle Arg His lle Gly Leu Ala Ala Lys 60 245 250 255 0 <210> 1 <211> 768 <212> DNA <213> Artificial 0 <220> <223> Synthetic nucleic acid sequence Ed.

-??1-

<400< 1 ggcagcagcg gaggcggcgg 01-0-0026 gacatcggceg cggtgcettge cgatgcacta 60 accgcaccgc tcgaccataa agacaaaagt ttgcagtctt tgacgctgga tcagtccgtc © 120 aggactaaaaacg agaaactgaa gacggaacggca 1 agggtg cggtg

01 gacagcctca atacgggcaa attgaagaac gacaaggtca gccgcttcga 111816001 240 caaatcgaag tggacgggca gctcattacc ttggagagcg gagagttcca agtgtacaaa

300 10 caaagccatt ccgccttaac cgcccttcag accgagcaag tacaagattc ggagcattca 360 gggaagatgg ttgcgaaacg ccagttcaga atcggcgata tagcgggtga acatacatct 0 420 gvoy

‎tttgacaagc ttcccgaagg cggcagggcg acatatcgcg ggacggcatt cggttcagac‏ 480 ‎gatgccagtg gaaaactgac ctacaccata gatttcgccg ccaagcaggg acacggcaaa ©‏ 540 ‎atcgaacatt tgaaatcgcc agaactcaat gttgacctgg ccgcctccga tatcaagcecg‏ 600 ‎٠١‏ ‎gataaaaaac gccatgccgt catcagcggt tccgtecttt 32-64880083802 93038838096‏ 660 ‎agttactctc taggcatctt tggcgggcaa gcccaggaag ttgccggcag cgcagaagtg‏ 10 720 acggcatacg ccatatcggt cttgccgceca accagtaa 028838 768 790 62 <210< gvoy

6+ <211> 255 <212> PRT <213> Artificial <220> o <223> Synthetic amino acid sequence <400> 62

Gly Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Val Leu 00 1 5 10 15

Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Ser Leu 60 yo

Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu Ed

Ala Ala Gln Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn 60 55 50 lo} Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 80 75 70 65 " GIn lle Glu Val Asp Gly GIn Leu lle Thr Leu Glu Ser Gly Glu Phe 95 90 85 Gln Val Tyr Lys GIn Ser His Ser Ala Leu Thr Ala Leu Gln Thr Glu yo 110 105 100 Val GIn Asp Ser Glu His Ser Gly Lys Met Val Ala Lys Arg 0 What is supported

—-Y4o- 115 120 125

Phe Arg lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu 130 135 140 o

Pro Glu Gly Gly Arg Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp 145 150 155 160 01

Asp Ala Ser Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys Gin 165 170 175

Vo

Gly His Gly Lys lle Glu His Leu Lys Ser Pro Glu Leu Asn Val Asp 180 185 190 gvoy

Leu Ala Ala Ser Asp lle Lys Pro Asp Lys Lys Arg His Ala Val lle 205 200 195 Ser Gly Ser Val Leu Tyr Asn Gln Ala Glu Lys Gly Ser Tyr Ser Leu ° 220 215 210 Gly lle Phe Gly Gly GIn Ala Gln Glu Val Ala Gly Ser Ala Glu Val 01 240 235 230 225 Glu Thr Ala Asn Gly lle Arg His lle Gly Leu Ala Ala Lys Gin 255 250 245 Vo 63 <210< <211< DNA <212> ard

—Yav- <213> Artificial <220> <223> Synthetic nucleic acid sequence lo} <400> 63 ggcagcagcg gaggcggcegg tgtcgccgec gacatcggeg cggggcettge cgatgcacta 60 accgcaccgc tcgaccataa agacaaaagt ttgcagtctt tgacgctgga tcagtccgtc ٠ 120 aggaaaaacg agaaactgaa gctggcggca caaggtgcgg aaaaaactta tggaaacggc 180

Vo gacagcctca atacgggcaa attgaagaac gacaaggtca gccgcttcga ctttatccgt 240 caaatcgaag tggacgggca gctcattacc ttggagagcg gagagttcca aatatacaaa 300 0 gvoy

—Y4A- caggaccact ccgccgtcgt tgccctacag attgaaaaaa tcaacaaccc cgacaaaatc 360 gacagcctga taaaccaacg ctccttcctt gtcagcggtt tgggtggaga acataccgec ه٠‎ 420 ttcaaccaac tgcccagcgg caaagccgag tatcacggca aagcattcag ctccgacgat 480 ٠١ gctggcggaa aactgaccta taccatagat ttcgccgcca aacagggaca cggcaaaatc 540 gaacacttga aaacacccga gcaaaatgtc gagcttgcct ccgeccgaact caaagcagat 600 ١٠ gaaaaatcac acgccgtcat tttgggcgac acgcgctacg gcggcgaaga aaaaggcact 660 taccacctcg cccttttcgg cgaccgcgecc caagaaatcg ccggcetcgge aaccgtgaag 0 720 gvoy

aggttcacga aatcggcatc gccggcaaac agtaa 28838 765 lo} 64 <210< 254 <211< PRT <212< Artificial <213< 01 <220< Synthetic amino acid sequence <223< 64 <400< Vo Gly Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu 10 5 1 gvoy

As Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Ser Leu 60 25 20 Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu ° 40 35 Ala Ala Gln Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn 01 60 55 50 Thr Gly Lys Leu Lys Asp Lys Val Ser Arg Phe Asp Phe lle Arg 80 75 70 65 Vo GIn lle Glu Val Asp Gly GIn Leu lle Thr Leu Glu Ser Gly Glu Phe 95 90 85 gvoy

0 \ —_ as GIn lle Tyr Lys GIn Asp His Ser Ala Val Val Ala Leu 60 lle Glu 110 105 100 lo} Lys lle Asn Asn Pro Asp Lys lle Asp Ser Leu lle Asn 60 Arg Ser 125 120 115 Phe Leu Val Ser Gly Leu Gly Gly Glu His Thr Ala Phe Asn 0 Leu 00 140 135 130

Pro Ser Gly Lys Ala Glu Tyr His Gly Lys Ala Phe Ser Ser Asp Asp 145 150 155 160 Yo

Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys GIn Gly

175 170 165 support

0 \ —_ As His Gly Lys lle Glu His Leu Lys Thr Pro Glu 60 Asn Val Glu Leu 190 185 180 lo} Ala Ser Ala Glu Leu Lys Ala Asp Glu Lys Ser His Ala Val lle Leu 205 200 195 0 Gly Asp Thr Arg Tyr Gly Gly Glu Glu Lys Gly Thr Tyr His Leu Ala 220 215 210 Leu Phe Gly Asp Arg Ala 60 Glu lle Ala Gly Ser Ala Thr Val Lys yo 240 235 230 225 lle Arg Glu Lys Val His Glu lle Gly lle Ala Gly Lys GIn Ed

0".- since 250 245

<210> 65 <211> 786 A.H

<212> DNA

‎<213> Artificial Sequence

<220> <223> Synthetic nucleic acid sequence 00

<400> 65

atgagctctg gaagcggaag cggggggceggt ggagttgcag cagacattgg aacaggatta

60 Vo

gcagatgcac tgacggcacc gttggatcat aaagacaaag gcttgaaatc gcttacctta

120

gaagattcta tttcacaaaa tggcaccctt accttgtccg cgcaaggcgc tgaaaaaact 180 0

gvoy

‎٠ _‏ اذ ‎tttaaagtcg gtgacaaaga taatagctta aatacaggta aactcaaaaa tgataaaatc‏ 240 ٠ه ‎tcgcgttttg atttcgtgca aaaaatcgaa gtagatggcc aaaccattac attagcaagc‏ 300 ‎ggtgaattcc aaatatataa acaagaccat tcagcagtcg ttgcattgca aattgaaaaa‏ 360 ‎٠١‏ ‎atcaacaacc ccgacaaaat cgacagcctg ataaaccaac gttccttcct tgtcageggt‏ 420 ‎ttgggcggtg aacatacagc cttcaaccaa ttaccaagcg gcaaagcgga gtatcacggt 10 480 aaaagcattta gctcagatga tgcaggcggt aaattaactt atacaattga ctttgcagca 540 aaaacaaggac atggcaaaat tgaacattta aaaacacccg aacagaacgt agagctcgca 0 600 gvoy

مج اذ ‎tccgcagaac tcaaagcaga tgaaaaatca cacgcagtca ttttgggtga cacgcgctac‏ 660 ‎ggcagcgaag aaaaaggtac ttaccactta gctctttttg gcgaccgagc tcaagaaatc‏ © 720 ‎gcaggtagcg caaccgtaaa gataagggaa aaggttcacg aaattgggat cgcgggcaaa‏ 780 ‎caataa 786 ٠‏ 66 >210< 8 >211< ‎PRT ٠‏ >212< Artificial Sequence <213< <220< Synthetic amino acid sequence <223< 790 gvoy

©, <400> 66

Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Val Ala Ala Asp lle 1 5 10 15 lo}

Gly Ala Gly Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp 0

Lys Ser Leu 60 Ser Leu Thr Leu Asp 60 Ser Val Arg Lys Asn Glu

Lys Leu Lys Leu Ala Ala Gin Gly Ala Glu Lys Thr Tyr Gly Asn Gly yo 60

Asp Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Ed

0 7 _ As 80 75 70 65 Thr lle Thr Leu Ala (6 Asp Phe lle Arg GIn lle Glu Val Asp Gly o 95 90 85 lle Tyr Lys Gln Asn His Ser Ala Val Val Ala Ser Gly Glu Phe 110 105 100 01 Leu Gln lle Glu Lys lle Asn Asn Pro Asp Lys lle Asp Ser Leu lle 125 120 115 Vo Asn Gln Arg Ser Phe Leu Val Ser Gly Leu Gly Gly Glu His Thr Ala 140 135 130 gvoy

M 0 As Phe Asn GIn Leu Pro Asp Gly Lys Ala Glu Tyr His Gly Lys Ala Phe 160 155 150 145 Ser Ser Asp Asp Pro Asn Gly Arg Leu His Tyr Ser lle Asp Phe Thr ° 175 170 165 Lys Lys GIn Gly Tyr Gly Arg lle Glu His Leu Lys Thr Pro Glu 60 ye 190 185 180 Asn Val Glu Leu Ala Ser Ala Glu Leu Lys Ala Asp Glu Lys Ser His 205 200 195 Vo Ala Val lle Leu Gly Asp Thr Arg Tyr Gly Gly Glu Glu Lys Gly Thr 220 215 210 gvoy

0 q —_ as Tyr His Leu Ala Leu Phe Gly Asp Arg Ala GIn Glu lle Ala Gly Ser 240 235 230 225 lo} Ala Thr Val Lys lle Arg Glu Lys Val His Glu lle Gly lle Ala Gly 255 250 245

Lys GIn 00 <210> 67

‎<211> 780 10 <212> DNA <213> Artificial Sequence <220>

Ed

=« \ اذ ‎Synthetic nucleic acid sequence‏ >223< 67 >400< ‎atgagctctg gaggtggagg aagcgggggce ggtggagttg cagcagacat tggagcagga‏ ‎o‏ 60 ‎ttagcagatg cactgacggc accgttggat cataaagaca aaagtttgca gtcgcttacc‏ 120 ‎ttagatcagt ctgtcaggaa aaatgagaaa cttaagttgg cggcgcaagg cgctgaaaaa ٠‏ 180 ‎acttatggaa acggtgacag cttaaataca ggtaaactca aaaatgataa agtctcgcgt 240 Vo tttgatttca ttcgtcaaat cgaagtagat ggccaaacca ttacattagc aagcggtgaa 300 ttccaaatat ataaacaaaa ccattcagca gtcgttgcat tgcaaattga aaaatcaac 360 gvoy9

-+١١- aaccccgaca aaatcgacag cctgataaac caacgttcct tccttgtcag cggtttggge 420 ggtgaacata cagccttcaa ccaattacca gacggcaaag cggagtatca cggtaaagca 480 © tttagctcag atgatccgaa cggtaggtta cactattcca ttgactttac caaaaaacaa 540 ggatacggca gaattgaaca tttaaaaacg cccgaacaga acgtagagct cgcatccgca 600 ٠ gaactcaaag cagatgaaaa atcacacgca gtcattttgg gtgacacgcg ctacggcggc 660 gaagaaaaag gtacttacca cttagccctt tttggcgacc gcgctcaaga aatcgcaggt Vo 720 agcgcaaccqg taaagataag ggaaaaggtt cacgaaattg ggatcgcggg caaacaataa 780 790 gvoy

©17- <210> 68 <211> 253 <212> PRT <213> Artificial Sequence lo} <220> <223> Synthetic amino acid sequence <400> 68 0

Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu Ala 1 5 10 15

Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Ser Leu GIn Ser yo

Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu Ala Ed

Add 45 40 35 Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn Thr o 60 55 50 Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 60 80 75 70 65 01 lle Glu Val Asp Gly 60 Leu lle Thr Leu Glu Ser Gly Glu Phe 60 95 90 85 Vo lle Tyr Lys GIn Asp His Ser Ala Val Val Ala Leu 60 lle Glu Lys 110 105 100 gvoy ‏

+16 lle Asn Asn Pro Asp Lys lle Asp Ser Leu lle Asn Gln Arg Ser Phe 115 120 125

Leu Val Ser Gly Leu Gly Gly Glu His Thr Ala Phe Asn Gin Leu Pro ° 130 135 140

Ser Gly Lys Ala Glu Tyr His Gly Lys Ala Phe Ser Ser Asp Asp Ala 145 150 155 160 01

Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys GIn Gly His 165 170 175

Vo

Gly Lys lle Glu His Leu Lys Thr Pro Glu Gln Asn Val Glu Leu Ala 180 185 190 gvoy

—Y¥yo-—

Ser Ala Glu Leu Lys Ala Asp Glu Lys Ser His Ala Val lle Leu Gly 195 200 205 lo}

Asp Thr Arg Tyr Gly Gly Glu Glu Lys Gly Thr Tyr His Leu Ala Leu 210 215 220

Phe Gly Asp Arg Ala GIn Glu lle Ala Gly Ser Ala Thr Val Lys lle 00 225 230 235 240

Arg Glu Lys Val His Glu lle Gly lle Ala Gly Lys 60 245 250 yo <210> 69 <211> 765 ed

-+١- <212> DNA <213> Artificial Sequence <220> <223> Synthetic nucleic acid sequence © <400> 69 atgagctctg gaggtggagg agttgcagca gacattggag caggattagc agatgcactg 60 ٠١ acggcaccgt tggatcataa agacaaaagt ttgcagtcgc ttaccttaga tcagtctgtc 120 aggaaaaatg agaaacttaa gttggcggcg caaggcgctg aaaaaactta tggaaacggt 180 10 gacagcttaa atacaggtaa actcaaaaat gataaagtct cgcgttttga tttcattcgt 240 caaatcgaag tagatggcca acttattaca ttagaaagcg gtgaattcca aatatataaa 300 0 gvoy

+١ /- caagaccatt cagcagtcgt tgcattgcaa attgaaaaaa tcaacaaccc cgacaaaatc 360 gacagcctga taaaccaacg ttccttcctt gtcagcggtt tgggcggtga acatacagcc ه٠‎ 420 ttcaaccaat taccaagcgg caaagcggag tatcacggta aagcatttag ctcagatgat 480 ٠١ gcaggcggta aattaactta tacaattgac tttgcagcaa aacaaggaca tggcaaaatt 540 gaacatttaa aaacacccga acagaacgta gagctcgcat ccgcagaact caaagcagat 600 ١٠ gaaaaatcac acgcagtcat tttgggtgac acgcgctacg gcggcgaaga aaaaggtact 660 taccacttag ctctttttgg cgaccgagct caagaaatcg caggtagcgc aaccgtaaag 0 720 gvoy

—¥\A- ataagggaaa aggttcacga aattgggatc gcgggcaaac aataa 765 <210> 70 ee <211> 255 <212> PRT <213> Neisseria meningitidis (group B) <400> 70 0

Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu 1 5 10 15

Vo

Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu 60 gvoy

-1 and -

Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu

Ala Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn ° 60

Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 65 70 75 80 Yo

GIn lle Glu Val Asp Gly Lys Leu lle Thr Leu Glu Ser Gly Glu Phe 85 90 95

Vo

Gln Val Tyr Lys GIn Ser His Ser Ala Leu Thr Ala Leu Gln Thr Glu 100 105 110 gvoy

©. Da Val GIn Asp Ser Glu Asp Ser Gly Lys Met Val Ala Lys Arg Gin 115 120 125 lo}

Phe Arg lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu 130 135 140

Pro Lys Gly Gly Ser Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp 00 145 150 155 160

Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys 0 165 170 175 yo

Gly His Gly Lys lle Glu His Leu Lys Thr Pro Glu 60 Asn Val Glu 180 185 190 Ed

©1-

Leu Ala Ser Ala Glu Leu Lys Ala Asp Glu Lys Ser His Ala Val lle 195 200 205 lo}

Leu Gly Asp Thr Arg Tyr Gly Gly Glu Glu Lys Gly Thr Tyr His Leu 210 215 220 0

Ala Leu Phe Gly Asp Arg Ala GIn Glu lle Ala Gly Ser Ala Thr Val 225 230 235 240

Lys lle Arg Glu Lys Val His Glu lle Gly lle Ala Gly Lys Gin yo 245 250 255 <210> 71 Ed

-?© 4 <211< PRT <212> Artificial Sequence <213< ht <220< Synthetic amino acid sequence <223< 1 <400< Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu Ala 00 10 5 1 Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu GIn Ser yo 30 25 20 Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu Ala 40 35 ed

A Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn Thr 60 55 50 lo} Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 60 80 75 70 65 " lle Glu Val Asp Gly Lys Leu lle Thr Leu Glu Ser Gly Glu Phe 60 95 90 85 Val Tyr Lys GIn Ser His Ser Ala Leu Thr Ala Leu 60 Thr Glu 60 yo 110 105 100 Val GIn Asp Ser Glu Asp Ser Gly Lys Met Val Ala Lys Arg Gin Phe Ed

+76 115 120 125

Arg lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu Pro 130 135 140 o

Lys Gly Gly Ser Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp Asp 145 150 155 160 01

Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys GIn Gly 165 170 175

Vo

His Gly Lys lle Glu His Leu Lys Thr Pro Glu 60 Asn Val Glu Leu 180 185 190 gvoy

- E?+ Gly Asp Arg Ala GIn Glu lle Ala Gly Ser Ala Thr Val Lys Yo 240 235 230 225 lle Arg Glu Lys Val His Glu lle Gly lle Ala Gly Lys GIn 250 245 Vo 72 <210 < 768 > 211 < DNA < 212 < gvoy

+- 7? 0 120 agggaaaaacg agaaactgaa gctggcggca caaggtgcgg aaaaaactta tggaaacggc 180 Vo gacagcctta atacgggcaa attgaagaac gacaaggtca gccgcttcga ctttatccgt 240 caaatcgaag tggacgggaa gctcattacc ttggagagcg 0ggagataa3 0ggagagcg gagagttcay0

مصففة ‎caaagccatt ccgccttaac cgcccttcag accgagcaag tacaagactc ggaggattcc‏ 360 ‎gggaagatgg ttgcgaaacg ccagttcaga atcggcgaca tagcgggcga acatacatct ©‏ 420 ‎tttgacaagc ttcccaaagg cggcagtgcg acatatcgcg ggacggcegtt cggttcagac‏ 480 ‎٠١‏ ‎gatgctggcg gaaaactgac ctatactata gatttcgccg ccaaacaggg acacggcaaa‏ 540 ‎atcgaacact tgaaaacacc cgagcaaaat gtcgagcttg cctccgccga actcaaagca‏ ‎ 10 600 gatgaaaaat cacacgccgt cattttgggc gacacgcgct acggcggcga agaaaaaggc 660 acttaccacc tcgccctttt cggcgaccge gcccaagaaa tcgccggcetc ggcaaccgtg 0 720 gvoy

tambour aaaaggttca cgaaatcggc atcgccggca aacagtaa 2889 768 lo} 73 <210< 786 <211< DNA >212< Artificial Sequence <213< 01 <220< Synthetic nucleic acid sequence >223 < 73 <400< atgtccagcg gttcaggcag cggcggtgga ggcgtggcag cagatatcgg aacaggttta Vo 60 gcagatgctc tgacagcacc cttagatcac aaagacaaag gacttaaatc actgacattg 120 790 gvoy

-74؟+- ‎gaagattcta tctcgcaaaa tggtactctc actctttcag cccaaggcgc agaaaaaaca‏ 180 ‎tttaaagtag gcgataaaga taactcctta aatacaggta aattaaaaaa tgacaaaatc‏ هه 240 ‎tcacggtttg atttcgttca gaaaattgaa gtagatggac aaacgattac attagcaagc‏ 300 ‎ggcgaattcc aaatttataa acaagaccat tcagcagtag tagcattaca aatcgaaaaa ٠‏ 360 ‎attaacaacc cggacaaaat tgattctctt attaaccaac gctcttttct cgtatcagga 420 cttggtggtg aacatacagc gtttaatcaa ctgccgtcag gaaaagcaga atatcatggt Vo 480 aaagcatttt catcagacga cgcaggtggc aaactgacct atactattga ctttgcagca 540 790 gvoy

gif aaacagggac atggaaaaat tgaacattta aaaacacccg aacagaacgt agaactggcc

600 tcagcagaat tgaaagctga tgaaaaatcc catgcagtaa ttttaggcga tacacgttac 660 © ggtagcgaag aaaaaggtac atatcactta gctctttttg gcgatcgtgc tcaagaaatt 720 gctggttccg caacagttaa aatccgtgaa aaagtacatg aaatcggcat tgcaggtaa7 4006a200 780 <4006a 780

<211> 262

<212> PRT

<213> Neisseria meningitidis (group B) <400> 74 | 0 gvoy

D harmful Cys Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Val Ala Ala Asp 10 5 1 lo} lle Gly Thr Gly Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys 25 20 Asp Lys Gly Leu Lys Ser Leu Thr Leu Glu Asp Ser lle Pro GIn Asn 00 40 35

Gly Thr Leu Thr Leu Ser Ala Gln Gly Ala Glu Lys Thr Phe Lys Ala 60 yo

Gly Asp Lys Asp Asn Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys 65 70 75 80 Ed

Add lle Ser Arg Phe Asp Phe Val GIn Lys lle Glu Val Asp Gly 60 Thr 95 90 85 lo} lle Thr Leu Ala Ser Gly Glu Phe GIn lle Tyr Lys Gln Asn His Ser 110 105 100 0 Ala Val Val Ala Leu GIn lle Glu Lys lle Asn Asn Pro Asp Lys lle 125 120 115 Asp Ser Leu lle Asn GIn Arg Ser Phe Leu Val Ser Gly Leu Gly Gly yo 140 135 130 Glu His Thr Ala Phe Asn Gln Leu Pro Gly Asp Lys Ala Glu Tyr His supported

Lav 160 155 150 145 Gly Lys Ala Phe Ser Ser Asp Asp Pro Asn Gly Arg Leu His Tyr Thr o 175 170 165 lle Asp Phe Thr Asn Lys GIn Gly Tyr Gly Arg lle Glu His Leu Lys 190 185 180 01

Thr Pro Glu Leu Asn Val Asp Leu Ala Ser Ala Glu Leu Lys Ala Asp 195 200 205

Vo

Glu Lys Ser His Ala Val lle Leu Gly Asp Thr Arg Tyr Gly Ser Glu 210 215 220 gvoy

A A Glu Lys Gly Thr Tyr His Leu Ala Leu Phe Gly Asp Arg Ala Gln Glu 240 235 230 225 lle Ala Gly Ser Ala Thr Val Lys lle Gly Glu Lys Val His Glu lle ° 255 250 245 Gly lle Ala Gly Lys GIn I 260 <210< 254 <211< PRT 0 <212< Artificial Sequence <213< <220< Synthetic amino acid sequence <223> supported

Add <400< Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Val Leu Ala o 15 10 5 1 Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Ser Leu GIn Ser 25 20 01 Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu Ala 40 35 Vo Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn Thr 60 55 50 gvoy

Add Gly Lys Leu Lys Asp Lys Val Ser Arg Phe Asp Phe lle Arg 60 80 75 70 65 lle Glu Val Asp Gly 60 Leu lle Thr Leu Glu Ser Gly Glu Phe Gin ° 95 90 85 Val Tyr Lys GIn Ser His Ser Ala Leu Thr Ala Leu 60 Thr Glu 60 ye 110 105 100 Val Gln Asp Ser Glu His Ser Gly Lys Met Val Ala Lys Arg Gln Phe 125 120 115 Vo Arg lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu Pro 140 135 130 gvoy

Fa Glu Gly Gly Arg Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp Asp 160 155 150 145 lo} Ala Ser Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys Gln Gly 175 170 165 His Gly Lys lle Glu His Leu Lys Ser Pro Glu Leu Asn Val Asp Leu 00 190 185 180 Ala Ala Ser Asp lle Lys Pro Asp Lys Lys Arg His Ala Val lle Ser yo 205 200 195 Gly Ser Val Leu Tyr Asn GIn Ala Glu Lys Gly Ser Tyr Ser Leu Gly 220 215 210 Ed

R lle Phe Gly Gly Gln Ala Gln Glu Val Ala Gly Ser Ala Glu Val Glu 240 235 230 225 lo} Thr Ala Asn Gly lle Arg His lle Gly Leu Ala Ala Lys 60 250 245 0 76 <210< 8 <211< PRT <212< Artificial Sequence <213> Vo <220> Synthetic amino acid sequence <223< 6 <400> support

4m Cys Gly Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Thr Gly 10 5 1 lo} Leu Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu 25 20 Lys Ser Leu Thr Leu Glu Asp Ser lle Ser Gln Asn Gly Thr Leu Thr 00 40 35

Leu Ser Ala Gln Gly Ala Glu Lys Thr Phe Lys Val Gly Asp Lys Asp 60 yo

Asn Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys lle Ser Arg Phe 65 70 75 80 Ed

Asp Phe Val GIn Lys lle Glu Val Asp Gly Gln Thr lle Thr Leu Ala 85 90 95 lo}

Ser Gly Glu Phe Sa lle Tyr Lys Gln Asp His Ser Ala Val Val Ala 100 105 110 0

Leu Gin lle Glu Lys lle Asn Asn Pro Asp Lys lle Asp Ser Leu lle 115 120 125

Asn Gln Arg Ser Phe Leu Val Ser Gly Leu Gly Gly Glu His Thr Ala yo 130 135 140

Phe Asn GIn Leu Pro Ser Gly Lys Ala Glu Tyr His Gly Lys Ala Phe Support

EA

145 150 155 160

Ser Ser Asp Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala 165 170 175 o

Ala Lys GIn Gly His Gly Lys lle Glu His Leu Lys Thr Pro Glu 60 180 185 190 01

Asn Val Glu Leu Ala Ser Ala Glu Leu Lys Ala Asp Glu Lys Ser His 195 200 205

Vo

Ala Val lle Leu Gly Asp Thr Arg Tyr Gly Ser Glu Glu Lys Gly Thr 210 215 220 gvoy

EAL

Tyr His Leu Ala Leu Phe Gly Asp Arg Ala GIn Glu lle Ala Gly Ser 225 230 235 240

Ala Thr Val Lys lle Arg Glu Lys Val His Glu lle Gly lle Ala Gly ° 245 250 255

Lys GIn 0 <210> 7 <211> 257 <212> PRT 0 <213> Artificial Sequence <220> <223> Synthetic amino acid sequence supported

Da 7 <400< Gly Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Thr Gly Leu o 15 10 5 1 Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu Lys 25 20 01 Ser Leu Thr Leu Glu Asp Ser lle Ser 60 Asn Gly Thr Leu Thr Leu 40 35 Vo Ser Ala GIn Gly Ala Glu Lys Thr Phe Lys Val Gly Asp Lys Asp Asn 60 55 50 gvoy

+46

Ser Leu Asn Thr Gly Lys Leu Lys Asn Asp Lys lle Ser Arg Phe Asp 65 70 75 80

Phe Val 60 Lys lle Glu Val Asp Gly GIn Thr lle Thr Leu Ala Ser ° 85 90 95

Gly Glu Phe GIn lle Tyr Lys Gln Asp His Ser Ala Val Val Ala Leu 100 105 110 ye

Gln lle Glu Lys lle Asn Asn Pro Asp Lys lle Asp Ser Leu lle Asn 115 120 125

Vo

Gln Arg Ser Phe Leu Val Ser Gly Leu Gly Gly Glu His Thr Ala Phe 130 135 140 gvoy

—Yto—

Asn GIn Leu Pro Ser Gly Lys Ala Glu Tyr His Gly Lys Ala Phe Ser 145 150 155 160 lo}

Ser Asp Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala 165 170 175

Lys GIn Gly His Gly Lys lle Glu His Leu Lys Thr Pro Glu Gln Asn 00 180 185 190

Val Glu Leu Ala Ser Ala Glu Leu Lys Ala Asp Glu Lys Ser His Ala 195 200 205 yo

Val lle Leu Gly Asp Thr Arg Tyr Gly Ser Glu Glu Lys Gly Thr Tyr 210 215 220 Ed

EA

His Leu Ala Leu Phe Gly Asp Arg Ala 60 Glu lle Ala Gly Ser Ala 225 230 235 240 lo}

Thr Val Lys lle Arg Glu Lys Val His Glu lle Gly lle Ala Gly Lys 245 250 255 0

Gln <210> 78 10 <211> 255 <212> PRT <213> Artificial Sequence supported

EA

<220> <223> Synthetic amino acid sequence <400> 78 lo}

Cys Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu 1 5 10 15

Ala Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu 60 00

Ser Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu yo

Ala Ala Gln Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn 60 55 50 Ed

Ya

Thr Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 65 70 75 80 lo}

GIn lle Glu Val Asp Gly GIn Leu lle Thr Leu Glu Ser Gly Glu Phe 85 90 95 0

Gln lle Tyr Lys L6 Ser His Ser Ala Leu Val Ala Leu 60 Thr Glu 100 105 110 Mau lle Asn Asn Ser Asp Lys Ser Gly Ser Leu lle Asn Gln Arg Ser yo 115 120 125

Phe Arg lle Ser Gly lle Ala Gly Glu His Thr Ala Phe Asn Gin Leu Ed

+44 130 135 140

Pro Lys Gly Gly Lys Ala Thr Tyr Arg Gly Thr Ala Phe Ser Ser Asp 145 150 155 160 o

Asp Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys 0 165 170 175 01

Gly His Gly Lys lle Glu His Leu Lys Thr Pro Glu L 60 Asn Val Glu 180 185 190

Vo

Leu Ala Ser Ala Glu Leu Lys Ala Asp Glu Lys Ser His Ala Val lle 195 200 205 gvoy

Leu Gly Asp Thr Arg Tyr Gly Gly Glu Glu Lys Gly Thr Tyr His Leu 220 215 210 Ala Leu Phe Gly Asp Arg Ala GIn Glu lle Ala Gly Ser Ala Thr Val ° 240 235 230 225 Lys lle Arg Glu Lys Val His Glu lle Gly lle Ala Gly Lys GIn I 255 250 245 79 <210< 254 <211< PRT 010 <212< Artificial Sequence <213< <220< Synthetic amino acid sequence <223> endorsed

o \ — as 9 >400< Ser Ser Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu Ala o 15 10 5 1 Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu GIn Ser 25 20 01 Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu Ala 40 35 Vo Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn Thr 60 55 50 gvoy

o \ —_ As Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 60 80 75 70 65 lle Glu Val Asp Gly 60 Leu lle Thr Leu Glu Ser Gly Glu Phe GIn o 95 90 85 lle Tyr Lys GIn Ser His Ser Ala Leu Val Ala Leu 60 Thr Glu GIn ye 110 105 100 lle Asn Asn Ser Asp Lys Ser Gly Ser Leu lle Asn Gln Arg Ser Phe 125 120 115 Vo Arg lle Ser Gly lle Ala Gly Glu His Thr Ala Phe Asn 0 Leu Pro 140 135 130 gvoy

o — As Lys Gly Gly Lys Ala Thr Tyr Arg Gly Thr Ala Phe Ser Ser Asp Asp 160 155 150 145 lo} Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys GIn Gly 175 170 165 His Gly Lys lle Glu His Leu Lys Thr Pro Glu 60 Asn Val Glu Leu 00 190 185 180 Ala Ser Ala Glu Leu Lys Ala Asp Glu Lys Ser His Ala Val lle Leu yo 205 200 195 Gly Asp Thr Arg Tyr Gly Gly Glu Glu Lys Gly Thr Tyr His Leu Ala 220 215 210 gvoy

o ¢ — as Leu Phe Gly Asp Arg Ala Gln Glu lle Ala Gly Ser Ala Thr Val Lys 240 235 230 225 lo} lle Arg Glu Lys Val His Glu lle Gly lle Ala Gly Lys GIn 250 245 0 80 <210< 254 <211< PRT <212< Artificial Sequence <213< Vo <220< Synthetic amino acid sequence <223< 80 <400> support

Ser 10 5 1 lo} Asp Ala Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu GIn Ser 25 20 Leu Thr Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu Ala 00 40 35

Ala GIn Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn Thr 60 yo

Gly Lys Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg 60 65 70 75 80 gvoy

_ A o As lle Glu Val Asp Gly 60 Leu lle Thr Leu Glu Ser Gly Glu Phe 60 95 90 85 lo} Val Tyr Lys GIn Ser His Ser Ala Leu Thr Ala Phe GIn Thr Glu 60 110 105 100 01 lle Gln Asp Ser Glu His Ser Gly Lys Met Val Ala Lys Arg Gln Phe 125 120 115 Arg lle Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu Pro yo 140 135 130 Glu Gly Gly Arg Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp Asp gvoy

o 7 — As 160 155 150 145 Ala Gly Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys GIn Gly o 175 170 165 Asn Gly Lys lle Glu His Leu Lys Ser Pro Glu Leu Asn Val Asp Leo 190 185 180

01

Ala Ala Ala Asp lle Lys Pro Asp Gly Lys Arg His Ala Val lle Ser

195 200 205

Vo

‎Gly Ser Val Leu Tyr Asn GIn Ala Glu Lys Gly Ser Tyr Ser Leu Gly

210 215 220 goy

M o As lle Phe Gly Gly Lys Ala GIn Glu Val Ala Gly Ser Ala Glu Val Lys 240 235 230 225 Thr Val Asn Gly lle Arg His lle Gly Leu Ala Ala Lys GIn o 250 245 1 <210< 0 252 <211< PRT <212< Artificial Sequence <213< <220< Synthetic amino acid sequence 0 <223< 81 <400< Gly Gly Gly Gly Val Ala Ala Asp lle Gly Ala Gly Leu Ala Asp Ala support

o q — Since 10 5 1 Leu Thr Ala Pro Leu Asp His Lys Asp Lys Gly Leu GIn Ser Leu Thr o 30 25 20 Leu Asp GIn Ser Val Arg Lys Asn Glu Lys Leu Lys Leu Ala Ala 60 40 35

01

Gly Ala Glu Lys Thr Tyr Gly Asn Gly Asp Ser Leu Asn Thr Gly Lys

50 55 60

Vo

Leu Lys Asn Asp Lys Val Ser Arg Phe Asp Phe lle Arg Gin lle Glu

65 70 75 80

gvoy a=

Val Asp Gly GIn Leu lle Thr Leu Glu Ser Gly Glu Phe GIn Val Tyr 85 90 95

Lys GIn Ser His Ser Ala Leu Thr Ala Phe GIn Thr Glu Gin lle Gln ° 100 105 110

Asp Ser Glu His Ser Gly Lys Met Val Ala Lys Arg Gin Phe Arg lle 115 120 125 ye

Gly Asp lle Ala Gly Glu His Thr Ser Phe Asp Lys Leu Pro Glu Gly 130 135 140

Vo

Gly Arg Ala Thr Tyr Arg Gly Thr Ala Phe Gly Ser Asp Asp Ala Gly 145 150 155 160 gvoy

Fy -

Gly Lys Leu Thr Tyr Thr lle Asp Phe Ala Ala Lys GIn Gly Asn Gly 165 170 175 lo}

Lys lle Glu His Leu Lys Ser Pro Glu Leu Asn Val Asp Leu Ala Ala 180 185 190

Ala Asp lle Lys Pro Asp Gly Lys Arg His Ala Val lle Ser Gly Ser 00 195 200 205

Val Leu Tyr Asn GIn Ala Glu Lys Gly Ser Tyr Ser Leu Gly lle Phe 210 215 220 yo

Gly Gly Lys Ala GIn Glu Val Ala Gly Ser Ala Glu Val Lys Thr Val 225 230 235 240 Ed

+7-

Asn Gly lle Arg His lle Gly Leu Ala Ala Lys 60 245 250 lo} Ed

Claims (1)

© Claims polypeptide -1 other than 18160/a/7117/a0 and an isolated non-lipidic one that includes an amino acid sequence that has a homology of at least 795 with sequence identification number: VY where the terminal 6 is omitted but not at the position 1 in comparison with arrangement definition No.: .710 polypeptide -" pursuant to claim 1 where arrangement includes arrangement definition No.: VY 0 where the N-terminal cysteine at position 1 is omitted Compared to Arrangement Definition #: .700 polypeptide — according to claim 0 where the polypeptide is an immunogen .(immunogenic);- polypeptide according to the claim)¢ where the polypeptide does not show an ABS transition of about + Dalton compared to the non-lipidic polypeptide as measured by mass spectrometry. polypeptide -#« 0 isolated under claim 0 where the polypeptide includes an amino acid arrangement (@Mino acid) at position 1854-1 of arrangement identification number: YY polypeptide —1 Isolated according to claim 1 wherein the amino acid arrangement (@MINo acid) is of the arrangement stated in the arrangement definition: YY = sal ge immunogenic composition comprising a polypeptide As in Ve any of Claims 1 9-7 f. A — the immunoogenic composition according to claim 1; also includes a polypeptide other than 18160/a/7111/a0 and an isolated non-lipid having the amino acid arrangement mentioned in the definition of arrangement no. : £9 where the terminal cysteine is omitted at position 1 compared to the definition of arrangement number: AA 01 ed F form (1) Nucleic acid arrangements non-fat contrast form P2086 )١ ‏(تعريف الترتيب رقم:‎ PAO ‏شكل متباين‎ Nucleic Acid ‏ترتيب‎ ‎1 ‏تخكتؤذااى “اهتاذ اطغناتا إن إ واتاات تاضقانت ناذا انافاا مات‎ GGGCTTOGCCGATGLCACTAACTGCGCCGUTCGACCATAAAGACAAAGGTT TGAAATCCCTGACATTGGAAGACTCCATTCCCCAAAACGGAACACTGAC CCTGTCGGCACAAGGTGCGUAAAAAACTTITCAAAGUCCGGUGACAAAGA CAACAGCCTCAACACGGGCAAACTGAAGAACGACAAAATCAGCCGCTT COGACTTCGTGCAAAAAATCGAAGTGOGACGGATCAAACCATCACACTGGC AAGCGGCGAATTTICAAATATACAAACAGGACCACTCCGCCGTCGTTGEC CTACAGATTGAAAAAATCAACAACCCCGACAAAATCGACAGOCTGATA AACCAACGUTCCTTCCTTGTCAGCGGTTIGGGCGGAGAACATACCGUCT TCAACCAACTGCCOCGOCGACAAAGCCGAGTATCACGGCAAAGCATTICA GCTCCGACGATGUCCGGUCGGAAAACTGACCTATACCATAGATTITGCCGC CAAACAGGGACACGGCAAAATCOAACACCTGAAAACACCCGAGCAAAA TGTCGAGCTTGCCGUCGUCGAACTCAAAGCAGATGAAAAATCACACGCC GTCATTTTGGGCGACACGCGCTACGGCAGCGAAGAAAAAGGCACTTACC ACCTCQCCCTTITCGGCGACCGUGCCCAAGAAATCGCCGGCTCGGCAAC CGTGAAGATAGGGGAAAAGGTTCACGAAATCGGCATCGCCGGCAAACA GTAG ترتيب ‎Nucleic Acid‏ شكل متباين 205< (تعريق الترتيب رقم: ؟) ‎TGCAGCAGCGGAAGCGGAAGCGGAGGCGGUGHTETCGCCGCCGACATC‏ ‎GGCACAGGGCTTGCCCGATGCACTAACTGCGCCGCTCGACCATAAAGACA‏ ‎AAGGTTTGAAATCCCTGACATTGGAAGACTCCATTTCCCAAAACGGAAC‏ ‎ACTGACCCTOTCGGCACAAGGTGCGGAAAAAACTTITCAAAGTCGGCGAC‏ ‎AAAGACAACAGTCTCAATACAGGCAAATTGAAGAACGACAAAATCAGC‏ ‎CGQUTTCGACTTTGTGCAAAAAATCGAAGTGGACGGACAAACCATCACGC‏ ‎TGGCAAGCGGCOAATTTCAAATATACAAACAGGACCACTCCGCCOTCAT‏ ‎TGCCCTACAGATTGAAAAAATCAACAACCCCGACAAAATCGACAGCCTG‏ ‎ATAAACCAACGCTCCTTCCTTIGTCAGCOGGTTTGGGCGGAGAACATACCG‏ ‎CCTTCAACCAACTGUCCAGCGGUCAAAGUUGAGTATCACGGCAAAGUATT‏ ‎CAGCTCCGACGATGCCGGUGOAAAACTGACCTATACCATAGATTITGCC‏ ‎ GCCAAACAGGGACACGGCAAAATCOGAACACCTGAAAACACCCGAGCAG AATGTCGAGCTTGCCTCCGUCGAACTCAAAGCAGATGAAAATCACACG CCGTCATTTTGGGCGACGCGCTACGGLAGCGAAGAAAAAGGCACTTA CCACCTCGCTCTTITCGGUGACCGAGCCCAAGAAATCGCCGGCTCGGCA ACCGTGAAGATAAGGCAAACAGGCAACGATCGATCG uh 7 + 1- )ب(١ ‏شكل‎ ‏رقم: ؟)‎ md all ‏(تعريف‎ ALR ‏شكل متباين‎ Nucleic Acid ‏ترتيب‎ ‎TGCAGCAGUGGAGGCGGCOGETETCGCCGCOGACATCGGCGECGGOGGOTT ‎GCCGATGCACTAACCGCACCGUTCGACCATAAAGACAAAAGTTTGCAGT ‎CTTTGACGUTGGATCAGTCCGTCAGGAAAAACGAGAAACTGAAGCTOGGC ‎GGCACAAGGTGUCGGAAAAAACTTATGGAAACGGCGACAGCUTCAATAC ‎GGGCAAATTGAAGAACGACAAGGTCAGCCGUTTCGACTTTATCCGTCAA ‎ATCGAAGTGGACGGACAAACCATCACGCTGGCAAGCGGUGAATTTICAA ‎ATATACAAACAGAACCACTCCGCCOTCGTTGUCCTACAGATTGAAAAAA ‎TCAACAACCCCGACAAAATCGACAGCCTGATAAACCAACGCTCUTTCCE ‎TGTCAGCGGTTTGGGCGGAGAACATACCGCCTTCAACCAACTGCCTGAC ‎GOCAAAGCCOGAGTATCACGGCAAAGCATTCAGCTCCGACGACCCGAAC ‎GOGCAGGUCTGCACTACTCCATTGATTTITACCAAAAAACAGGGTTACGGCA ‎GAATCGAACACCTGAAAACGCCCGAGCAGAATGOTCGAGCTTGCCTCCGLC ‎CGAACTCAAAGCAGATGAAAAATCACACGCCGTCATTTTGGGCGACACG ‎CGCTACGGCGOGCOGAAGAAAAAGGCACTTACCACCTCGCCCTTTTICGGCG ‎ACCGCGCCCAAGAAATCOGCCGGCTCGGCAACCOGTGAAGATAAGGGAAA ‎AGGTTCACGAAATCGGCATCGCCGGCAAACAGTAG ‎(§ ‏(تعريف الترتيب رقم:‎ >AL12-2 ‏شكل متباين‎ Nucleic Acid ‏ترثيب‎ ‎TGULAGCAGCGGAGGGOGGCOGGTOTCGCUGCCGACA TTGOTGCGGGGUTT ‎GUCGATGCACTAACCGCACCGUTCGACCATAAAGACAAAAGTTTGCAGT ‎CTTTGACGCTGGATCAGTCCGETCAGGAAAAACGAGAAACTGAAGCTGGC ‎GOGCACAAGGTGCGOGAAAAAACTTATGGAAACGGUGACAGCCTCAATAC ‎GOGCAAATTOAAGAACGACAAGOTCAGCUGCTTICGACTTTATCCOGTCAA ‎ATCGAAGTGGACGGACAAACCATCACGUTGGCAAGCGGCGAATTICAA ‎ATATACAAACAGAACCACTCCGCCGTCGTTGCCCTACAGATTGAAAAAA ‎TCAACAACCCCGACAAAATCGACAGCCTGATAAACCAACGCTCCTTCCY ‎TOTCAGCGOTTTGGGCGGAGAACATACCGCCTTCAACCAACTGCCTGALC ‎GGCAAAGCCGAGTATCACGGCAAAGCATTCAGCTCCGACGACTCGAAL ‎GGCAGGCTGCACTACTCCATTGATTITACCAAAAAACAGGGTTACGGCA ‎GAATCOGAACACCTGAAAACGCCCGAGUCAGAATOGTCGAGCTTGCCTCCGC ‎CGAACTCAAAGCAGATGAAAAATCACACGUCCGTCATTTITGGGCGACALCG ‎CGCTACGGCGGCGAAGAAAAAGGCACTTACCACCTCGCCCTTTTCGGCG ‎ACCGCOGCCCAAGAAATCGCCOGGCTCGGCAACCGTGAAGATAAGGGAAA ‎AGGTTCACGAAATCOGCATCGCCGGTCAAACAGTAG ‎tyov -??- appearance )-( )# ‏شكل متباين 22< (تعريف الترتيب رقم:‎ Nucleic Acid ‏ترتيب‎ ‎TGCAGCAGCGGAGGUGGCOGTGTCGCCGCCGACATCGGCGCGGEGCTT ‎GCCGATGCACTAACCGCACCGCTCGACCATAAAGACAAAAGTTTGCAGT ‎CTTTGACGCTGGATCAGTCOCGTCAGGAAAAACGAGAAACTGAAGCTGGC ‎GGCACAAGGTGOGGAAAAAACTTATGGAAACGGCGACAGCCTCAATAC ‎GGGCAAATTGAAGAACGACAAGGTCAGUCGCTTCGACTTTATCOGTCAA ‎ATCGAAGTGGACGGGCAGCTCATTACCTTGGAGAGCGGAGAGTTCCAA ‎ATATACAAACAGGACCACTCCGUCCOGTCGTTGCCCTACAGATTGAAAAAA ‎TCAACAACCCUGACAAAATCGACAGCCTGATAAACCAACGUTCCTTCCT ‎TGTCAGCGGTTTGGGTGGAGAACATACCGCCTTCAACCAACTGCCCAGT ‎GGUAAAGCCGAGTATCACGGCAAAGCATTCAGUTCCGACGATGCTGGO ‎GQGAAAACTGACCTATACCATAGATTITCGCCGCCAAACAGGGACALCGGC ‎AAAATCGAACACTTGAAAACACCCGAGCAAAATOGTCGAGCTTGCCTCCG ‎COGAACTCAAAGCAGATGAAAAATCACACGCCGTCATTTTGGGCGACAC ‎GCGLTACGGUGGUCGAAGAAAAAGGCACTTACCACCTCGCCCTTTTCGGC ‎GACCGCGUCCAAGAAATCOGCCGOGUTCGOCAACCGTGAAGATAAGGGAA ‎AAGOTTCACGAAATCGGCATCGCCGGCAAACAGTAG (7 ‏(تعريف الترتيب رقم:‎ PBO2 ‏شكل متباين‎ Nucleic Acid ‏ترتيب‎ ‎TGCAGCAGUCGGAGGUGGCGEHGAAGCGGAGGCGGCGOTGTCGCCGCTGAC ‎ATCGGCGCGOOGUTTGCUOGATGCACTAACCGUCACCGUTCGACCATAAAG ‎ACAAAGGTTTGAAATCCCTGACATTGGAAGACTCCATTTCCCAAAACGG ‎AACACTGACCCTGTOGGCACAAGGTGUGGAAAGAACTTTICAAAGCCGG ‎CGACAAAGACAACAQGTCTCAACACAGGCAAACTGAAGAACGACAAAAT ‎CAGUOGCTTCGACTTTATCCGTCAAATCGAAGTGGACGGGCAGCTCATT ‎ACCTTGGAGAGCGGAGAGTTCCAAGTGTACAAACAAAGCCATTCCGCCT ‎TAACCGCCUCTTCAGACCGAGCAAGTACAAGACTCGGAGCATTCCOGGGAA ‎GATGGTTGCGAAACGCCAGTICAGAATCGGCGACATAGTGGGCGAACA ‎TACATCTTTTGACAAGUTTCCCAAAGACGTCATGGCGACATATCGUGGG ‎ACGGUGTTCOGTTCAGACGATGUCGGCGOGAAAACTGACCTACACCATAG ‎ATTTCGCCGUCAAGCAGGGACACGGCAAAATCGAACATTTGAAATCGCC ‎TGAACTCAATGTTGACCTGGCCGCCGCCGATATCAAGCUCGGATGAAAAA ‎CACCATGCCGTCATCAGCOGGTTCCOGTCUTTTACAACCAAGCCGAGAAAG ‎GCAGTTACTCTCTAGGUCATCTTTGGCGGGCAAGCCCAGGAAGTTGCCGH ‎ CAGCGCGGAAGTGGAAACCGCAAACGGCATACGCCATATCGGTCTITGCC GCCAAGCAATAA tyov -yiv-{oft-form {Vp ‏(تعريف الترتيب‎ BOS ‏شكل مثباين‎ Nucleic Acid ‏ترتيب‎ ‏ا لفان اا الى ل تأجاا ات وااف تا اتات فدات‎ GUUGATOGCACTAACCGUACUGUTCGACTATAAAGACAAAAGTTTLGOAGLT CTTTCACGUTGUATUAGTUUGTCAGGAAAAACGAGAAACTODAAGUTGOU CET ACAAGHTGUOGARAAAACTTATGGAAATGGUGACAGUCTTAATAC GOGUAAATTIGAAGAACGACAAGGTICAGUOGTTITOGACTITATCOGTUAA ATOGAAGTGUHACGLOGUAGU NAT TACUTTOGAGAGUOGAGAGTTUUAA GTOGTACAAACAAAGOCATTOCOOOTTAACCGCOCTTCAGALCGAGUAAD AACAAGATOCAGAGUATTOOOGGAAGATGOTTOUGAAACGUOGGTICA AMATOGGUGACATAGUGGEUGAAUATACATOTTITIGACAAGOTTOOU AA AGACGTUATGOGUGACATATOGUGGHALGLGOGTTOGG TT CAG ADGA THO GOUGGAAAACTGACCTATACTATAGATTT TOUT GCC AAACAGGGALC ACG GUAAAATUGAACATITGAAATOGUCCGAAUTUAATOTOGAGUTTLUCAL COUUTATATCAADCCGOATGAAAAAL ALL ATOUOGTCATUAGUGHL THC GTCOTTTACAATCAAGACOAGAAAGGCAGTTACTOCCTOGGTATCTTTG GUGHGGUAAGUOU AGEAAGTTGUOGHOAGOGUGEAAGTGEAAALCGUAA ACGUULTACATATOGCAGUATACOATAGUAA {8 ‏شكل متباين 109< (لعريف الترتيب رقم:‎ Nucleic Acid ‏ترتيب‎ ‎1 ‏ا تاكن فا ااانا انا 61 1 )اذا انض تلات‎ GULOATOOACTAACCGUACCHUTCOACCA TAAAGACAAAGLT I TOCALT CTTITAACGUTGUATCAGTOUGTCAGGAAAAADGAGAAACUTGAAGUTGGT GUOACAAGGTOGOGGAARAAAUT TATGGAAACGGUGACAGOCTTAATAL GUOCAAATTGAAGAACGAVAALGICAGUOGUTTUGATT TATOO CAA ATCGAAGTGOACGGGAAGUTCATTACCOTTGGAGAGUGGAGAGTTOCAA GTGTACAAACAAAGUCATTCOGLUTTAACCGUOCTTUAGACTGAGUAAG TACAAGAUTOLGAGUATTOCHGLAAGATGHTTHUGAAACGUOALTTCA GAATOGGUGACATAGUGGOUGAATATACATOTTTIGACAAGUTTOOOAA AGGUGGUAGTGUGACATATOGEGOGGACGHUGT TOUGHT 1 GOGUGGAAAACTOADCTATACTATAGATTTCGUOGOCAAGCAGGGACADG GUAAAATCGAACATTTGAAATOGOCCGAAUTCAATGTUGAGOTTGOC AC COCOTATATCAAGUCOGATOAAAAACGOUCATGUCGTTATOCAGOGG TTC GTOOTTTACAACCAAGACGAGAAAGLOAGTTAUTOOUTUGHTATCITIG GUOGGGUAABCCCAGGA AGT TGUOGLOUAGUGOGOAAGTOGAAACCGUAA ACGOUATACACCATATOGGIUTTQUUGUUAAGCAGTAA uh 7 حداف شكل ا(ها ترتيب ‎Nucleic Acid‏ شكل متباين 822< (تعريف الترثيب رقم: 4) ‎TGCAGCAGCGGAGGUGGCGGETGTCGCOGCUGACATCGGCGCGGTGCTTG‏ ‎CCGATGUACTAACCGCACCGCTCGACCATAAAGACAAAAGTTTGCAGTC‏ ‎TTTGACGCTGGATCAGTCOGTUCAGGAAAAACGAGAAACTGAAGCTGGO‏ ‎GGCACAAGGTGUGGAAAAAACTTATGGAAACGGCGACAGCCTCAATALC‏ ‎GOGUAAATTGAAGAACGACAAGGTCAGCCGUTTCGACTTTATCCGTICAA‏ ‎ATCGAAGTGGACGGGUCAGUTCATTACCTTIGGAGAGCGULAGAGTTCCAA‏ ‎GTOGTACAAACAAAGCCATTCCGCCTTAACCGCCCTTCAGACCGAGCAAG‏ ‎TACAAGATTCGGAGCATTCAGGGAAGATGGTITGCGAAACGCCAGTTCAG‏ ‎AATCOGGCGATATAGCGGGETGAACATAUATCTTITGACAAGUTTCCCGAA‏ ‎GOCGUCAGGOGUGACATATCGCLHGLACGGUATTCGHTTCAGACGATGOC ‏ ‎AGTGGAAAACTGACCTACACCATAGATTICGCCGCCAAGCAGGGACALC‏ ‎GGUAAAATCGAACATTTGAAATCGCCAGAACTCAATGEITGACCTGGCOG‏ ‎CCTCCGATATCAAGUCUGGATAAAAAACGUCATGQUCGTCATCAGUGGTTC‏ ‎COTCCTTTACAACCAAGUCCGAGAAAGGCAGTTACTCICTAGGCATCTTT‏ ‎GGCGOGGCAAGCCCAGGAAGTTGCCGGCAGCGCAGAAGTGGAAACCGCA‏ ‎AACGGCATACGUCATATCGGEICTTGCUCGCCAAGUAGTAA‏ ‏ترتيب ‎Nucleic Acid‏ شكل مثباين ‎>B24‏ (تعريف الترتيب رقم: ‎)٠١‏ ‎TGCAGCAGUCGGAGGGGGTGGE TGTCGUCGUCGACATCGGTGLGGGGCTT‏ ‎GCCGATGCACTAACCGCACCGUTCGACCATAAAGACAAAGGTTTGCAGT‏ ‎CTTTGACGCTGGATCAGTCOGTCAGGAAAAACGAGAAACTGAAGOTGGC‏ ‎GGCACAAGGTGUGGAAAAAACTTATGGAAACGGTGACAGCUTCAATAC‏ ‎GGGCAAATTGAAGAACGACAAGGTCAGUCGTTTOGACTTTATCCGCCAA‏ ‎ATCGAAGTOGACGGGUAGCTCATTACCTTGGAGAGTGGAGAGTTCCAAG‏ ‎TATACAAACAAAGUCATTCCGLCTTAACUGUCOTTTCAGACTUGAGCAAAT‏ ‎ACAAGATTCGGAGCATTCCGGGAAGATGGTTGCGAAACGCCAGTTCAG‏ ‎AATCGGCGACATAGCGGGUGAACATACATCTTITTIGACAAGUTTCUCGAA‏ ‎GGCGGCAGGGCGACATATCGOGGGACGGCGTTCGGTITCAGACGATGCT‏ ‎GGUOGGAAAACTGACCTACACCATAGATTICGCCGUCAAGUAGGGAAALC‏ ‎GGUAAAATCGAACATTITGAAATOGCCAGAACTCAATGTOGACCTGGLCG‏ ‎CCGCCGATATCAAGCCGGATGGAAAACGCCATGCCGTCATCAGCGOGTTC‏ ‎CGTCCTTTACAACCAAGCCGAGAAAGGCAGTTACTCCCTCGGTATCTTT‏ ‎GGCGGAAAAGCCCAGGAAGTTGCCGGCAGCGUGGAAGTGAAAACCGETA‏ ‎AACGGCATACGCCATATCGGCCTTGUCGUCAAGCAATAA‏ ‏ااي -and1*+ Fig. 1(f). (VY ‏شكل متباين 1344< (تعريف الترتيب رقم:‎ Nucleic Acid ‏ترتيب‎ ‎TOCAGCAGUOGGAGLHOGGUGGAAGUGHAGHUGGOGH IGTOGCUGUCGAC ATCGOGUGLCGGHOUTTOCUGATGUCACTAAUCGCACCGUTUGACCATAAAG ACAAAGGTTTGAAATCCCTGACATTGUGAAGACTCCATTTCCCAAAACGG AACACTGACCUCTGTCGOCACAAGUTGCOGAAAGAACTTTCAAAGCCGOG CUACAAAGACAACAGTCTCAACACAGUGUCAAACTGAAGAACGACAAAAT CAGUCGCTTOOGACTTTATCCGTCAAATOHAAGTOGGACGGGUAGUTCATT ACCTTGGAGAGUGGAGAGTTCCAAGTGTACAAACAAAGCCATTCCGCCT TAACCGCCCTTCAGACCGAGCAAGTACAAGACTCGGAGCATTCCGGHGAA GATGOTTGCOAAACGCCAGTTCAGAATCGGLGACATAGTOUGOUGAACA TACATCTTTTOGCAAGCTTCCCAAAGACGTUCATOGCGACATATCGOGGH ACGOOGTICGGTTCAGACGATOUCGGUGGAAAACTGACCTACACCATAG ATTTCGCOGCCAAGCAGHGACACGGCAAAATCHAACATTTGAAATCGCC AGAACTCAATGTTCACCTGGUCGUCGCCGATATCAAGCCGGATGAAAAA CACCATGCCGTCATCAGCHGGTTCCOTCCTTTACAACCAAGCCGAGAAAG GUAGTTACTCTCTAGGUATCTTTGGOGGGUAAGCCCAGOAAGTTGUCGG CAGCGCGOGAAGTGOAAACCGLCAAACGGUATACGUCATATCOGTCTTIGCC GCCAAGCAATAA Lad No form? () Amino acid arrangements P2086 fatty gas anisotropy (VY ‏شكل مثباين 04< (تعريف الترتيب رقم:‎ Amine Acid ‏ترتيب‎ ‎CSSGGGGGGVAADIGTGLADALTAPLDHKDKGLKSLTLEDSIPONGTLTLS ‎AQGAEKTFEAGDKDNSLNTGKLENDKISRFDFVORKIEVDGQTITLASGEFQI ‎YRKQDHSAVVALQIEKINNPDRIDSLINQRSFLVSGLGGEHTAFNQLPGDKAE ‎YHGKAFSSDDAGGKLTYTIDFAAKQGHGKIEHLKTPEQNVELAAAELKAD ‎EKSHAVILGDTRYGSEEKGTYHLALFGDRAQEIAGSATVKIGEKVHEIGIAG ‎KQ (VY ‏(تعريف الترتيب رقم:‎ ADS ‏شكل متباين‎ Amino Acid ‏ترتيب‎ ‎CSSGSGSGGOGOGVAADIGTGLADALTAPLDHKDKGLESLTLEDSISONGTLT ‎LSAQGAEKTFKVGDEDNSENTGKLKNDKISRFDFVQKIEVDGQTITLASGEF ‎QIYKQDHSAVVALQIEKINNPDKIDSLINQRSFLVSGLGGEHTAFNQLPSGK ‎AEYHGKAFSSDDAGGKLTYTIDFAAKQGHGKIEHLKTPEQONVELASAELKA ‎DEKSHAVILGDTRYGSEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIA ‎GKQ (VE ‏شكل متباين 12< (تلعريف الترتيب رقم:‎ Amino Acid ‏ترتيب‎ ‎CSSGOGGYVAADIGAGLADALTAPLDHKDKSLOSLTLDQSVRKNEKLK LAA QGAEKTYGNGDSLNTGKLKNDKVSRFDFIRQIEVDGQTITLASGEFQIYKON HSAVVALQIEKINNPDKIDSLINQRSFLVSGLGGEHTAFNQLPDGKAEYHGK AFSSDDPNGRLHYSIDFIKKQGYGRIEHLKTPEQNVELASAELKADEKSHA VILGDTRYGGEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIAGKQ (Ve ‏شكل متباين 22< (تعريف الترتيب رقم:‎ Amino Acid ‏ترتيب‎ ‎CSSGGGGVAADIGAGLADAL TAPLDHKDKSLOSLTLDOQSVRENEKLKLAA QGAEKTYGNGDSLNTGKLENDKVSREDFIRQIEVDGQLITLESGEFQIVKQD HSAVVALQIEKINNPDKIDSLINQRSFLVSGLGGEHTAFNQLPSGKAEYHGK AFSSDDAGGKLTYTIDFAAKQGHGKIEHLKTPEQNVELASARL KADEKSHA VILGDTRYGGEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIAGKQ tyov - Mother form? (B) )17 ‏شكل متباين 502< (تغعريف الثرتيب رقم:‎ Amino Acid af CSSGHGGESGGEGVAADIGAGLADAL TAPLDHKDKGLKSETLEDSISQNGTL TLSAQGAERTFKAGDKDNSINTGKLKNDKISRFDFIRQIEVDGQLITLESGEF QVYKQSHSALTALQTEQVOQDSEHSGKMVAKRQFRIGDIVGEHTSFDKLPKD VMATYRGTAFGSDDAGGKLTYTIDFAAKQGHGKIEHLKSPELNVDLAAAD IKPDEKHHAVISGSVLEYNQAEKGSYSLGIFGGQAQEVAGSAEVETANGIRHI GLAAKQ (VY ‏(تعريف الترتيب رقم:‎ >B03 ‏شكل متباين‎ Amine Acid ‏ترتيب‎ ‎CSSGGGGVAADIGAGLADALTAPLDHKDKSLOSLTLDQSVRKNEKLKLAA ‎QGAEKTYGNGDSLNTGKLKNDKVSREDFIRQIEVDGQUITLESGEFQVYKQ ‎SHSALTALQTEQEQDPEHSGKMVAKRRFKIGDIAGEHTSFDKLPKDVMATY ‎RGTAPGSDDAGGKLTYTIDFAAKQGHGKIEHLKSPEINVELATAYIKPDEK ‎HHAVISGSVLYNQDEKGSYSLGIFGGQAQEVAGSAEVETANGIHHIGLAAK ‎Q (YA ‏شكل متباين 309< (تعريف الترتيب رقم:‎ Amino Acid «ad 5 CSSGGGGVAADIGAGLADALTAPLDHKDKGLOQSLTLDOSVREKNEKLKLAA QGAEKTYGNGDSINTGKLKNDKVSREDFIRQIEVDGKLITLESGEFQVYKQ SHSALTALQTREQVOQDSEDSGKMVARKRQFRIGDIAGEHTSFDKLPKGGSATY RGTAFGSDDAGGKLTYTIDFAAKQGHGKIEHLKSPELNVELATAYIKPDEK RHAVISGSVLYNQDEKGSYSLGIFGGQAQEVAGSAEVETANGIHHIGLAAK Q (V3 ‏(تعريف الترتيب رقم:‎ >B22 ‏شكل متاين‎ Amino Acid ‏ترتيب:‎ ‎CSSGGLGGVAADIGAVLADALTAPLDHKDESLOSLTLDOQSVRENEKLKLAA ‎QGAEKTYGNGDSENTGKLKNDKVSREDFIRQIEVDGQLITLESGEFQVYKQ ‎SHSALTALQTEQVQDSEHSGKMVAKRQFRIGDIAGEHTSFDRKLPEGGRATY ‎RGTAFGSDDASGKLTYTIDFAAKQGHGKIEHLKSPELNVDLAASDIKPDKK ‎RHAVISGSVLYNQAEKGSYSLGIFGGQAQEVAGSAEVETANGIRHIGLAAK ‎Q tyov -Yvy- (—)Y shape (V+ ‏شكل متباين 1324< (تعريف الترثيب رقم:‎ Amino Acid ‏ترتيب‎ ‎CSSGGGGVAADIGAGLADALTAPLDHKDKGLQSLTLDQSVRKNEKLKLAA ‎QGAEKTYGNGDSLNTGKLKNDKVSRFDFIRQIEVDGQLITLESGEFQVYKQ ‎SHSALTAFQTEQIQDSEHSGKMVAKRQFRIGDIAGEHTSFDKLPEGGRATYR ‎GTAFGSDDAGGKLTYTIDFAAKQGNGKIEHLKSPELNVDLAAADIKPDGKR ‎HAVISGSVLYNQAEKGSYSLGIFGGKAQEVAGSAEVKTVNGIRHIGLAAKQ (YY) ‏رقم:‎ cui i ‏(تعريف‎ >B44 ‏شكل متثباين‎ Amino Acid ‏ترتيب‎ ‎CSSGGGGSGGGGVAADIGAGLADALTAPLDHKDKGLKSLTLEDSISQNGTL ‎TLSAQGAERTFKAGDKDNSLNTGKLKNDKISRFDFIRQIEVDGQLITLESGEF ‎QVYKQSHSALTALQTEQVQDSEHSGKMVAKRQFRIGDIVGEHTSFGKLPKD ‎VMATYRGTAFGSDDAGGKLTYTIDFAAKQGHGKIEHLKSPELNVDLAAAD ‎IKPDEKHHAVISGSVLYNQAEKGSYSLGIFGGQAQEVAGSAEVETANGIRHI ‎GLAAKQ any Al apical view doubt lateral view vy > 0 OF ING ae A ES Ll fy a Vane ) 8 ag RE 0 WN Na \ 1 NY \ a prematurity NY RRR AN 3 WEI © LAH | Q "1 d . A > NN + SEES TFT scarcity so a ur + no code RR va! NR X aN l l wR ; a) Na ti Ar NN TY Ry Ry Aah WS FF | ra = Na LF {CaO 153-2610 Line Recorder 8# 0 (141-154) AN Arrived no form; Cys Ad is terminal N resulting in loss of visibility in E. coli wie an EEE EEE NY ae | | BEBEEEEEE | Ed So Sis: AA BR a. = d0 BR produces similar results for a Cys-minus A22 Ju iyov build —Yyo-a Non-lipophilic form ORF2086 on Gly/Ser isoform expression Gly/Ser stalk length effect without Coomassie Protein on excess heteroform? Cy 1 a terminal Cys 1 CSSGGGGRGGGRVTADIGTGLADAL TAP (ot) NO NO YES YES TELEPHONE GENERATOR [ENT TRANSMISSION TAN NAKA A A (¢ +) Yes Yes ADS ; COSSGSGSGUGUGVAADIGTGLADALTAP bY 0 AZZ COSSGGHLTGVAADIGAGLADALTAP n B22 CUSSGDAGVAAD 19GDAGHVAAD IGAVVAAD LO SBGGGOGVAADIGAGLADALTAP Terminal 88 again Cys Yes when adding * UL I< = 7 levels high to show BOY is non-gony J g Gly/Ser stalk short 1 7 3 4 º + uh 0 *a to induce PYG a to d TE + 0 for oo = p + _- 1 wo = = aha NoCys Bog | 0 0 0 - | | ¥ OA * # bit ATE AGC AGU OR GI oF ST STC S00 IN GAC ATC B60 S08 oR A 31 801 828 a alg “a + 8” > 1 his “the dX * MN NR + lga A + 8 NNR NR God > “NWR No” BOY Me and EO EO 7 - Lala Vo KK 5 Improvement Co don Increases contrast 2 3 A22 non-fat o “YA ¢ ? 10 1 Surah inductor om * e. + . + a on 1 a Ta = __ _ _ .... a TEE a aE a a Sn EN i a Ce on . Te AN |a Co = a Naa a a LL ; a a a a a gan here a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a a ENE ESE aE ARENA i : a =. = a ' Le “0 ! : changes of B09 Gly codon terminal B22 AN and A22 voy —YYA- (ha form EV ‏تعريف الترتيب رقم:‎ > AGCTCTGGAGGTGGAGGAAGCGGGGGCGGTGGAGTTGCAGCAGACATT GGAGCAGGATTAGCAGATGCACTGACGGCACCGTTGGATCATAAAGAC AAAGGCTTGAAATCGCTTACCTTAGAAGATTCTATTTCACAAAATGGCA CCCTTACCTTGTCCGCGCAAGGCGCTGAACGTACTTITAAAGCCGGTGA CAAAGATAATAGCTTAAATACAGGTAAACTCAAAAATGATAAAATCTCG CGTTITGATTTCATTCGTCAAATCGAAGTAGATGGCCAACTTATTACATT AGAAAGCGGTGAATTCCAAGTATATAAACAATCCCATTCAGCACTTACA GCATTGCAAACCGAACAGGTCCAAGACTCAGAACATTCCGGCAAAATG GTAGCTAAACGTCAATTCCGCATCGGTGACATTGTCGGTGAACATACAA GCTTCGGAAAATTACCAAAAGATGTGATGGCGACCTATCGCGGTACGGC ATTTGGATCAGATGATGCAGGCGGTAAATTAACTTATACAATTGACTTT GCAGCAAAACAAGGACATGGCAAAATTGAACATTTAAAATCTCCCGAA CTTAACGTAGATCTCGCAGCAGCAGATATTAAACCAGATGAAAAACACC ACGCAGTCATTTCAGGTTCAGTTTTATACAATCAGGCAGAAAAAGGTTC GTACTCTTTAGGTATTTTTGGCGGGCAAGCTCAAGAAGTTGCAGGTAGC GCAGAAGTAGAAACGGCAAATGGCATTCGTCACATTGGGTTAGCGGCG AAACAATAA i ad) ‏الترتيب‎ hy ml > SSGGGGSGGGGVAADIGAGLADALTAPLDHKDKGLKSLTLEDSISQNGTLT LSAQGAERTFKAGDKDNSLNTGKLKNDKISRFDFIRQIEVDGQLITLESGEF QVYKQSHSALTALQTEQVQDSEHSGKMVAKRQFRIGDIV GEH TSFGKLPKD VMATYRGTAFGSDDAGGKLTYTIDFAAKQGHGKIEHLKSPELNVDLAAAD IKPDEKHHAVISGSVLYNQAEKGS YSLGIFGGQAQEVAGSAEVETANGIRHI GLAAKQ, Yes No form (A 5١ ‏تعريف الترثيب رقم:‎ > AGCAGCGGAGGLGGCGGAAGCGGAGGCGGCGGTGTCGCCGCCGACATC GGCGCGGGGCTTGCCGATGCACTAACCGCACCGUTCGACCATAAAGACA AAGGTTTGAAATCCCTGACATTGGAAGACTCCATTITCCCAAAACGGAAC ACTGACCCTGTCGGCACAAGGTGUGGAAAGAACTTTCAAAGCCGGCGA CAAAGACAACAGTCTCAACACAGGCAAACTGAAGAACGACAAAATCAG CCGCTTCGACTTTATCCGTUAAATCGAAGTGGACGGGCAGCTCATTACC TTGGAGAGCGGAGAGTTCCAAGTGTACAAACAAAGCCATTCCGCCTTAA CCGCCCTTCAGACCGAGCAAGTACAAGACTCGGAGCATTCOGGGAAGAT GGTTGCGAAACGCCAGTTCAGAATCGGCGACATAGTGGGUGAACATAC ATCTTTTGGCAAGCTTCCCAAAGACGTCATGGUGACATATOGCGGGACG GCGTTCGOGTTCAGACGATGUCOGGUGOAAAACTGACCTACACCATAGATT TCGCCGCCAAGCAGGGACAUGGCAAAATCGAACATTTGAAATCGCCAG AACTCAATGTTGACCTGGCCGCCGCCGATATCAAGCCGGATGAAAAACA CCATGCCGTCATCAGCGGTTCCGTCCTTTACAACCAAGCCGAGAAAGGC AGTTACTCTCTAGGCATCTTTGGCGGGCAAGCCCAGGAAGTTGOCGGCA GCGCGGAAGTGGAAACCGCAAACGGCATACGUCCATATCGGTCTTGCCGC CAAGCAATAA £0 ‏تعريف الترثيب رقم:‎ > AGCTCTGGAGGTGOAGGAAGCGGGGOCGUTGCAGT TGC AGCAGACATT GGAGCAGGATTAGCAGATGCACTGACGGCACCGTTGGATCATAAAGAC AAAGGCTTGCAGTCGCTTACCTTAGATCAGTCTGTCAGGAAAAATGAGA AACTTAAGTTGGCGGCGCAAGGCGCTGAAAAAACTTATGGAAACGGTG ACAGCTTAAATACAGGTAAACTCAAAAATGATAAAGTCTCGCGTTTTGA TTTCATTCGTCAAATCGAAGTAGATGGCAAGCTTATTACATTAGAAAGC GOTOGAATTCCAAGTATATAAACAATCCCATTCAGCACTTACAGCATTGC AAACCGAACAGGTCCAAGACTCAGAAGATTCCGLGCAAAATGGTAGCTA AACGTCAATTCCGUATCGGTGACATIGCGGGTGAACATACAAGCTTCGA CAAATTACCAAAAGGCGGCAGTGCGALCTATCGUGGTACGGCATTTGGA TCAGATGATGCAGGUGGTAAATTAACTTATACAATTGACTTTGCAGCAA AACAAGGACATGGCAAAATTGAACATTTAAAATCTCCCGAACTTAACGT AGAGUTCGCAACCGUCATATATTAAACCAGATGAAAAACGCCACGCAGT CATTTCAGGTTCAGTTTTATACAATCAGGACGAAAAAGGTTCGTACTCTT TAGGTATTTTTGGCGGGCAAGCTCAAGAAGTTGCAGGTAGCGCAGAAGT AGAAACGGCAAATGGCATTCACCACATTGGGTTAGCGGCGAAACAATA A club _ Ad A «= (—=) A form 56 ‏تعريف الترتيب رقم:‎ > SSGLGOGGSGGGOVAADIGAGLADALTAPLDHKDKGLOQSLTLDOQSVRKNEKL KLAAQGAEKTYGNGDSLNTGKLKNDKVSRFDFIRQIEVDGKLITLESGEFQ VYKQSHSALTALQTEQVODSEDSGKMVAKRQFRIGDIAGEHTSFDKLPKGG SATYRGTAFGSDDAGGKLTYTIDFAAKQGHGKIEHLKSPELNVELATAYIK PDEKRHAVISGSVLYNQDEKGSYSLGIFGGQAQEVAGSAEVETANGIHHIGL AAKQ £7 ‏تعريف الترتيب رقم:‎ > AGCTCTGGAGGTGGAGGAGTTGCAGCAGACATTGGAGCAGGATTAGCA GATGCACTGACGGCACCGTTGGATCATAAAGACAAAGGCTTGCAGTCGC TTACCTTAGATCAGTCTGTCAGGAAAAATOAGAAACTTAAGTTGGCGGC GCAAGGUCGUTGAAAAAACTTATGGAAACGGTGACAGUTTAAATACAGG TAAACTCAAAAATGATAAAGTICICGUGTTTTCATTTCATTCGICAAATCO AAGTAGATGGCAAGCTTATTACATTAGAAAGCGGTGAATTCCAAGTATA TAAACAATCCCATTCAGCACTTACAGCATTGCAAACCGAACAGGTCCAA GACTCAGAAGATTCCOGGCAAAATGOTAGCTAAACGTCAATTCCGCATCG GTGACATTGUCGOGGTGAACATACAAGCTTCGACAAATTACCAAAAGHGCG GCAGTGCGACCTATCGCGGTACGGCATTTGGATCAGATGATGCAGGCGG TAAATTAACTTATACAATTOGACTTTGCAGCAAAACAAGCGACATGOCAAA ATTGAACATTTAAAATCTCCCGAACTTAACGTAGAGCTCGCAACCGCAT ATATTAAACCAGATGAAAAACGCCACGCAGTCATTTCAGGTICAGTTTT ATACAATCAGGACGAAAAAGGTTCGTACTCTTTAGGTATTTITGGCGGG CAAGCTCAAGAAGTTGCAGGTAGCGCAGAAGTAGAAACGGCAAATGGC ATTCACCACATTGGGTTAGCGGUGAAACAATAA (voy م شكل ‎{mh‏ ‏> تريب ‎Arad yaad ih‏ ‎GLCAGCAGUGCAGELGEREETETOCCOGLCRACARTCHGCLLEGEEETTECOCATCLAL TAALCECAICENTUCALCATARAG‏ ‎ACARBAGTTIGCAGT CTT TEACGUTGGAT CAG TOC GT CAGEARMARCEAGBAATT CRAGCTGGUGEUACRAGGI CUGEAARAA‏ ‎ARDTTRTGEAAACGGUGREAGLCTCARTACCGECAAAT TCARGAATGACAR GAT CAGCCECTTCGACT TTATOCGTCARATC‏ ‎GEARS TT CORARTATALBAACAGGACCACTCCEUCHTCGTPEOCUTAL‏ ممم سق ا لمم ‎CAT‏ موت ياج ‎CABG TG EAR‏ ‎AGAT TGARBARATCAACAACTCUGACAMRRTCRACAGCCT GRTARACTARCGLTCETTCCT IGT CAGCLUTT TERE TGRAGA‏ ‎TOAGCTCCGACGATRETEGLAGAARA.‏ لتقا ممق موق ‎ACATACCGLCTTCARCCAACTGCUCAGCEROAAAGUCEAG TAT‏ ‎CIGACTTATACCATAGATTTCECCGCCARACAGRGEACACARCARMAT COAAIACT THARRACACTCGAGCARARATETCHART‏ ‎TTECOTCCEOCERARUTCABAGCAGA TCAABARTCACAC GOI TCATT TTRGGUAALACECECTACGRCCECEARSRARBAGS‏ ‎CACTTACCACCTCGUCCT TTICGECGALC EUG CARGRANTCRUCEGC TC RGCARCCETGRAGR TAAGG GARBAGE TAC‏ ‎GRANTOGECATCECCECCAANCAGTRA > Definition of Arrangement 24 YEN EDS LNT EGR LRNDKVERFOFIR]Y 1 YN 90 3F 3 YN 8 EVDGOLITLESCERQ IY KL DH ERVVALGI ER INN PLE I DS LING RS FLVEAGLGCEH TAF ROL PGRARYHEEA FS SLDAGGEK ‎LEY TID FARKQGHGR IEH LNT FEONVE LAS AE LEADERS HAVI LGDTRYGEEERGT Y HLALFGDRAQE TAGEATVK IT RERVH EIGIAGKD Aye —YAY- ( 3) + fig 2١ ‏تعريف الترثيب رقم:‎ > AGCAGCGGGOGUGEGTGOAGTTGCAGUAGACATIGGAGCAGUATTAGCA GATGCACTGACGGCACCGTTIGGATCATAAAGACAAAGGCTTGCAGTCGC TTACCTTAGATCAGTCTGTCAGGAAAAATGAGAAACTTAAGTTGGCGGC GCAAGGCGCTGAAAAAACTTATGGAAACGGTGACAGCTTAAATACAGG TAAACTCAAAAATGATAAAGICTCGCGTTTITGATTICATTCGTCAAATOG AAGTAGATGGCAAGCTTATTACATTAGAAAGCGGTGAATTCCAAGTATA TAAACAATCCCATTCAGUACTTACAGCATTGCAAACCGAACAGGTCCAA GACTCAGAAGATTCCGGUAAAATOGTAGUTAAACGTCAATTCCGCATCG GTGACATTGOGGGTGAACATACAAGUTTCGACAAATTACCAAAAGGCG GCAGTGCGACCTATCGUGGTACGGCATTTGGATCAGATGATGCAGGOGG TAAATTAACTTATACAATTGACTTITGCAGUAAAACAAGGACATGGCAAA ATTGAACATTTAAAATCTCCCOGAACTTAACGTAGAGCTCGCAACCGCAT ATATTAAACCAGATGAAAAACGCCACGCAGTCATTTCAGGTTCAGTTTT ATACAATCAGGACGAAAAAGGTTCGTACTCTITTAGGTATTITIGGCGLG CAAGCTCAAGAAGTTGCAGGTAGCGCAGAAGTAGAAACGGCAAATGGC ATTCACCACATTGGGTTAGCGGOGAAACAATAA CA ‏تعريف الترثيب رقم:‎ > AGCAGCGGAGGGGOCGETGTCGCCGCCGACATCGOTGCGGOOCTTGCC GATGCACTAACCGCACCGUTCGACCATAAAGACAAAGGTTTIGCAGTICTT TAACACTGGATCAGTCOGTUAGGAAAAACGAGAAACTGAAGCTGGEGG CACAAGGTGUCGGAAAAAALTTATGGAAACGGUGACAGUCTTAATALCGG GCAAATTGAAGAACGACAAGOGTCAGCCGUTICGAUTTTATCCGTCAAAT COAAGTGOACGGGAAGCTCATTACCTTGOAGAGCGGAGAGTTCCAAGT GTACAAACAAAGCCATTCCGOCCTTAACCGCCCTTCAGACCGAGCAAGTA CAAGACTCGGAGGATTCCGGGAAGATGGTTGUGAAACGCCAGTTCAGA ATCGGCGACATAGUGGGUGAACATACATCTTTTIGACAAGUTTCCCAAAG GCGGUAGTGCGACATATCGUGGGACGGUGTTCGUTTCAGACGATGCTOG COGAAAACTGACCTATACTATAGATTTICGCCGLCAAGCAGGGACACGGU AAAATCGAACATTTIGAAATCGCCCGAACTCAATGTOGAGCTTGCCACCG CCTATATCAAGCCGGATGAAAAACGCCATGCOGTTATCAGCGGTTCCGT CCTTTACAACCAAGACGAGAAAGGCAGTTACTCCCTCGGTATCTTTGGC GGGCAAGCCCAGGAAGTTGCCGGLAGCGLOGAAGTOGGAAACCGCAAAC GGUATACACCATATCGGTCTTGUCOGCCAAGCAGTAA tyov (Mm Shaq 38) > Definition of Trib No.: 8+ A R A A A A A A A A 0 A A B A A A B A R A 6 A A A A A 5 A B R 6 A A A A A A A A A A A A A A A A A A B A A A A A A B A A R A 6 A A 5 A 1 Teqina Mama Bethan Nataha 0 Rathant Dlakh LTTA Mutammam Mahmarj 006 TE TACASACBARGCIRT TACO PTAA GAB PEACE SAR TORT TACO PTAA GAG AEACCGAGUARSTALARAGATTCCGAGCAT TCAGERRAGR TEST THCAARACRCCAGT 6 TC EUCEATGCEGGT GR CCEMATRCATARG TECRACAATARG TESTRACATRACA TTRACA TTOLGT TCAGRI GRTGUCAGT SGA‏ ان المع عد ‎ARRCTARCOTACACCATAGR PT TOHCCGOCABGIARGEACACHECARRATC ERAT TT TEARAT‏ ‎CRAGCCHEATFARBARCCLCAT GUCCI CATCAGUEGT TCQET CCT TTACAALCARGICGAGAR‏ اا ااانا ان ناوا لاا ‎AGGOASTIACTOTCTAGGUATCT TT RECHEGHCARGUCCAGEAART TECUGLCASCLCAGAAGT BRARATCHCARALGEIATA‏ ‎CEECATRICAGTOTTEOLGUCABGRBE TRA‏ ‎RRIF SERRE URES‏ ‎GRSGRGEVAADIGAVIADALTATLDRK DRS LUST LOQSVRENEK LR LAR GREET YONG IS IN TRE LENDEVERFDFIROT ‎EVIL IT LES BR FQVY KG SB SALTALGTE VO IS EHS GHMVARR PR IGRI AGER TS FOE L PEGGRATY RGTAFGEIIDAS G SLOT FURORORVAGEREVRIANGY KP IER AAV KLTV FARR QORCKT RH URS PEL REIGLAAKG AH 7 -YAt- Figure 4 (e). £0 Set order definition: > SSGGOGVAADIGAGLADALTAPLDHKDKGLOSLTLDOSVRENEKLKLAAQ GAEKTYOGNGDSLNTGKLEKNDKVSREDFIRQIEVDGKLITL. ESGEFQVYK(S HSALTALQTEQVODSEDSGKMVAKROQFRIGDIAGEHTSFDKLPKGGSATYR GTAFGSD DAGGKLTYTIDFAAKQGHGKIEHLKSPELNVELATAYIKPDEKRHAVISGSY LYNQDEKGSYSLGIFGGQAQEVAGSAEVETANGIHHIGLAAKQ 5: ‏تعريف الترتيب رقم:‎ > AGCAGUGGAAGCGGAAGCGOGAGGCGGCOGTGTCGUCGCCOACATIGGC ACAGGGUTTGUCCGATGCACTAACTGUGCCGUTCGACCATAAAGACAAAG GTTTGAAATCCCTGACATTGOAAGACTCCATTTCCCAAAACGGAACACT GACCCTGTCGGCACAAGGTGOGGAAAAAACTTTCAAAGTCGGUOGACAA AGACAACAGTCTCAATACAGGCAAATTGAAGAACGACAAAATCAGCCG CTICGACTTTGTGCAAAAAATOGAAGTOGADGGATCAAACCATCACGCTG GCAAGCGGCGAATTTCAAATATACAAACAGGACCACTCCGCCGTOGTTIG COCTACAGATTGAAAAAATCAACAACCCCGACAAAATCGACAGUUTGAT AAACCAACGCTCCTTCCTTGICAGCGGTTTGGOUGGAGAACATACCECC TTCAACCAACTGCCCAGUGGCAAAGCUGAGTATCACGGCAAAGCATTCA GCTCCGACGATGUCGGUGGAAAACTGACCTATACCATAGATTTTIGCCGE CAAACAGGGACACGGCAAAATCGAACACCTGAAAACACCCGAGUAGAA TGTCGAGCTTGCCTCCGCCGAACTCAAAGCAGATGAAAAATCACACGCC GTCATITIGGGUGACACGUGUTACGGLCAGUGAAGAAAAAGGCACTTACC ACCTCOCTCTTTTCGOCGACCGAGUCCAAGAAATCGCCGGCTCGGCAAC COTOAAGATAAGGGAAAAGGTTICACGAAATCGOCATCGCCOGGLAAACA GTAG > Definition of arrangement No.: 20 ‎DHKDKGLKSLTLEDSISONGTLTLS‏ The god of my devotion! و)ابنخم بأ واوا ادناه اك 016111000111150 57101717 لاي لاه 1 از اتن زرات تتا الااطذتايلمخ ‎YRKOQDHSAVVALQIEKINNPDKIDSLINQRSFLVSGLGGEHTAFNQLPSGKAE‏ ‎YHORKAFSSDDAGGKL TY TIDFAAKQGHGKIEHL KTPEQNVELASAFLKADE‏ ‎KSHAVILGDTRYGSEEKGTYHLALFGDRAQEIAGSATVKIREKVHEIGIAGK‏ ‏0 uh 7 + { BR ‏شكل‎ ‎SY a8 5 adi J ‏تعريف‎ > SRGLGGESGOGOVTADIGTEL ADALTAPLDHEDRGLESLTLEDSISUNGTLY LSAQGARKTYGNGDSINTGKLINDKVSRFDFIRGIEVDGOLITLESGERQVY ROSIBALTALOTEQLOQDPEHAERKMVAKRRFRIGIIAGEHTSFDRIPEDVMA TYRGIAPGSDDAGLK 7 TIDFAAKQUHGKIEHEKSPEERVDLAVAYIR PD ERHHAVISGEVLYNOQDEKGRY SLGIFGERADEVAGRAEVETANGIHHIGL A ARQ {2% ad ‏(تعزيت الترتيب‎ GenBank Ay330406 > CSRGGGH SOGGGVTADIGTGLADIAL TAPLDHKDKGLESLTT FNRISONGTL TISAQGARKRTYGNOGDSLNTOR LENDER VSREDFRQIEVDGOLITLESGLIQY YEOQSHSALTALQTEQEQDPEHSEKMVAKRRFRIGDIAGEHTSFDKLPKDVM ATYRGTAFGSNDAGHGRL TY TIDFAARQGHGKTEHLESFEINVDLAVAYIKE DERHHAVISGEVEYROQDERGEY SLGIFGERAQEVAGS ALEVE TANGHIIGL AARG (9 tad; ‏(تعريف الترتيب‎ GenBank FI184191 > CSSGHOLOGVAAIGAGLADALTAPLDHEDRGLOSLILDOSVRENEKLELAA QUAEKTY GNGISENTGRLENDRVEREDFIRCHEVDGULITLESGEFUNVY RQ SHSAT TALOTEQVODRFHSGEMVARKRQFRIGINAGEHTSFDRIPEGGRATY RGTAFSSRDAGUGRLIY TIF AAKQGHGRIEHLESPELKVDLAAADKPDERH ‏ا لح ا ا‎ ) ١ tad ‏(لتعريف: الترتيب‎ GenBank ‏كم‎ 330335 > CSSGHUOVAADIGAGLADALTAPLDHEKOESLOSLTLDOSVERNE RLKEL AA QGAEKTYGNGUSINTOR LENDR VERFDFIRGHEVDGOLITLESGEFQVYRG SHEALTALOTEQVODSEHRGEMVARROFRIGINAGETTSFDKILPEGGRATY RGTAFGSDDASGEKLUTY TIDFAAKQGHGEIEHLES PELNVELAASDIKPDRE REAVISGEVLYNQAERKGSYSLGIFGVOGOAQEVAGSAEVET 3 g -0/5- )( 4 Fig 1 b - ~ hd b orev kK Lo oe ahd ob lod ago Rs ADS (4) SeediECVEANT ST ATRLTAPLINKD pein Pi 1 1... 5 110810 Sp LE PAVE rier QUEAE aan 4) 4 ~~ BOGVARDI HGLADALTAPLDHKD Srp DORVREN HANA QUAL; 62 £4 2 {3GGVARDI GAS LADALTAPLDHK DRE DS LP LORY 1 TH GAEKT, A i pl A D HEDR LBL DORE RE HL OGRE 809 1 8-93 HE Toh Nf ip 824 1 ... s PELANALTAPLIHKD 5 s ... s 3 0 1 s q q q {vi C88 CEGVAADTGAGTADALTAPLDHRDKGLOS LIL IOS VRENER LKLARQGAE 98 a i “3 ts : L A A B 0 31 i 0 LA AOS 1) BRVEORONEINTORLKNDRER FORUpK] EVD RIO § Ee hi ML 0 ei i mm yash lamaa fl Except HONG ~-BLNTGKLKNDHVEREDE LASHIN º i SERCH) it Tet) LF 62 {oY} 3 4 1 1_1) REDE b;3 EVDGE LH x oS . i 1 1 0 809 OVA Reply to Spe wa gs. ke Hh 2 B24 (ov; jronpo---ELsrcrLKNDRVERFDETRGEVDEDLT TLE ce rd kos ARO compatibility f1}) YGNGD 070 tyov + L form 4 (b) ny YA BOS A 2 TET ROR N SCL ORE BTA GY 5 2 3063 pop dL nay 212 ARES hob A 3 Cenk x 0 m oo mille! 222 Khad Roa Messam Kadt hid i RPE 5 1 8 262 SERINE A 5 Gp 63 Hef COE AGERTERD HH FIRE Sh di is i #09 SEES 2 58 ES 8 DEA SEHD Ki 0 L ; DDR LE 3 8334 SAEED HER AA AMOD 8 COMPATIBILITY RUN INNSDKSGSLINGRSFRISCIAGEHTAFNQLE 0 LE YAY Yio 205 LE 812 TA RxoaERIEAL 5 9 ane EHH av Hr TR) i 11][ 222 between RRO HH EHLHTE 0 mn EEK 8 dri AN 3 0 1 positioned for a ha ha a ani BOO 807: Fa S54 een i FEDER) RV ISEEVHYNGEE iis 1 raha B24 No. No. 0 NAN SEBVIERQARK GRY:} 3 gee 05 071010577006 ) Yi 17 AS AED) FRSA TRERVEES PH (VF) FRSA TRERVEES PH (VF) i] ERLE Arrangement wy yas} and 22AD 157 3 19 (Arrangement Definition No.: 262 Arrangement Definition is GIG) 01 Arrangement Definition No.: 809 (i PPOSARVETAN 8) Arrangement Definition No.: 14 (and 324 iY 0 3 IR GORE VNGIRED) Arrangement: ¥ (0 m i wi) Compatibility rang: 0570717613550 (VA) (voy) The validity period of this patent is twenty years from the date of filing the application, provided that the annual financial fee is paid for the patent and that it is not invalid or forfeited for violating any of the provisions of the patent system, layout designs of integrated circuits, plant varieties, and industrial designs, or its executive regulations issued by King Abdulaziz City for Science and Technology; Saudi Patent Office P.O. Box TAT Riyadh 57??11 ¢ Kingdom of Saudi Arabia Email: Patents @kacst.edu.sa