Hybrid expression of neisserial proteins

Abstract

Two or more Neisserial proteins (e.g. A and B) are expressed as a single hybrid protein which can be represented simply by the formula NH2-A-B—COOH.

Description

[0001] All documents cited herein are incorporated by reference in their entirety.

TECHNICAL FIELD

[0002] This invention is in the field of protein expression. In particular, it relates to the heterologous expression of proteins from Neisseria (e.g. N. gonorrhoeae or, preferably, N. meningitidis).

BACKGROUND ART

[0003] International patent applications WO99/24578, WO99/36544, WO99/57280 and WO00/22430 disclose proteins from Neisseria meningitidis and Neisseria gonorrhoeae. These proteins are typically described as being expressed in E. coli (i.e. heterologous expression) as either N-terminal GST-fusions or C-terminal His-tag fusions, although other expression systems, including expression in native Neisseria, are also disclosed.

[0004] It is an object of the present invention to provide alternative and improved approaches for the heterologous expression of these proteins. These approaches will typically affect the level of expression, the ease of purification, the cellular localisation of expression, and/or the immunological properties of the expressed protein.

DISCLOSURE OF THE INVENTION

[0005] In accordance with the invention, two or more (e.g. 3, 4, 5, 6 or more) proteins of the invention are expressed as a single hybrid protein. It is preferred that no non-Neisserial fusion partner (e.g. GST or poly-His) is used.

[0006] This offers two advantages. Firstly, a protein that may be unstable or poorly expressed on its own can be assisted by adding a suitable hybrid partner that overcomes the problem. Secondly, commercial manufacture is simplified—only one expression and purification need be employed in order to produce two separately-useful proteins.

[0007] Thus the invention provides a method for the simultaneous heterologous expression of two or more proteins of the invention, in which said two or more proteins of the invention are fused (i.e. they are translated as a single polypeptide chain).

[0008] The method will typically involve the steps of: obtaining a first nucleic acid encoding a first protein of the invention; obtaining a second nucleic acid encoding a second protein of the invention; ligating the first and second nucleic acids. The resulting nucleic acid may be inserted into an expression vector, or may already be part of an expression vector.

[0009] Where just two proteins are joined, the hybrid protein can be represented simply by the formula NH2-A-B—COOH. A and B can each be selected from any Neisserial proteins, and in particular those represented by SEQ#s 1-4326. The method is well suited to the expression of proteins orf1, orf4, orf25, orf40, Orf46146.1, orf83, 233, 287, 292L, 564, 687, 741, 907, 919, 953, 961 and 983.

[0010] The 42 hybrids indicated by ‘X’ in the following table of form NH2-A-B—COOH are preferred: 1 B→ ↓A ORF46.1 287 741 919 953 961 983 ORF46.1 X X X X X X 287 X X X X X X 741 X X X X X X 919 X X X X X X 953 X X X X X X 961 X X X X X X 983 X X X X X X

[0011] Preferred proteins to be expressed as hybrids are thus ORF46.1, 287, 741, 919, 953, 961 and 983. These may be used in their essentially full-length form, or poly-glycine deletions (&Dgr;G) forms may be used (e.g. &Dgr;G-287, &Dgr;GTbp2, &Dgr;G741, &Dgr;G983 etc.), or truncated forms may be used (e.g. &Dgr;1-287, &Dgr;2-287 etc.), or domain-deleted versions may be used (e.g. 287B, 287C, 287BC, ORF461-433, ORF46433-608, ORF46, 961c etc.) and so on.

[0012] Particularly preferred are: (a) a hybrid protein comprising 919 and 287; (b) a hybrid protein comprising 953 and 287; (c) a hybrid protein comprising 287 and ORF46.1; (d) a hybrid protein comprising ORF1 and ORF46.1; (e) a hybrid protein comprising 919 and ORF46.1; (f) a hybrid protein comprising ORF46.1 and 919; (g) a hybrid protein comprising ORF46.1, 287 and 919; (h) a hybrid protein comprising 919 and 519; and (i) a hybrid protein comprising ORF97 and 225.

[0013] Further embodiments are shown in the drawings and include &Dgr;G287-919, &Dgr;G287-953, &Dgr;G287-961, &Dgr;G983-ORF46.1, &Dgr;G983-741, &Dgr;G983-961, &Dgr;G983-961C, &Dgr;G741-961, &Dgr;G741-961C, &Dgr;G741-983, &Dgr;G741-ORF46.1, ORF46.1-741, ORF46.1-961, ORF46.1-961C, 961-ORF46.1, 961-741, 961-983, 961C-ORF46.1, 961C-741, 961C-983, 961CL-ORF46.1, 961CL-741, and 961CL-983.

[0014] Where 287 is used, it is preferably at the C-terminal end of a hybrid; if it is to be used at the N-terminus, if is preferred to use a &Dgr;G form of 287 is used (e.g. as the N-terminus of a hybrid with ORF46.1, 919, 953 or 961).

[0015] Where 287 is used, this is preferably from strain 2996 or from strain 394/98.

[0016] Where 961 is used, this is preferably at the N-terminus. Domain forms of 961 may be used.

[0017] Alignments of polymorphic forms of ORF46, 287, 919 and 953 are disclosed in WO00/66741. Any of these polymorphs can be used according to the present invention.

[0018] Preferably, the constituent proteins (A and B) in a hybrid protein according to the invention will be from the same strain.

[0019] The fused proteins in the hybrid may be joined directly, or may be joined via a linker peptide e.g. via a poly-glycine linker (i.e. Gn where n=3, 4, 5, 6, 7, 8, 9, 10 or more) or via a short peptide sequence which facilitates cloning. It is evidently preferred not to join a &Dgr;G protein to the C-terminus of a poly-glycine linker.

[0020] The fused proteins may lack native leader peptides or may include the leader peptide sequence of the N-terminal fusion partner.

[0021] Host

[0022] It is preferred to utilise a heterologous host. The heterologous host may be prokaryotic or eukaryotic. It is preferably E. coli, but other suitable hosts include Bacillus subtilis, Vibrio cholerae, Salmtonella typhi, Salmonenna typhimiurium, Neisseria meningitidis, Neisseria gonorrhoeae, Neisseria lactamica, Neisseria cinerea, Mycobateria (e.g. Mtuberculosis), yeast etc.

[0023] Vectors, Hosts etc.

[0024] As well as the methods described above, the invention provides (a) nucleic acid and vectors useful in these methods (b) host cells containing said vectors (c) proteins expressed or expressable by the methods (d) compositions comprising these proteins, which may be suitable as vaccines, for instance, or as diagnostic reagents, or as immunogenic compositions (e) these compositions for use as medicaments (e.g. as vaccines) or as diagnostic reagents (f) the use of these compositions in the manufacture of (1) a medicament for treating or preventing infection due to Neisserial bacteria (2) a diagnostic reagent for detecting the presence of Neisserial bacteria or of antibodies raised against Neisserial bacteria, and/or (3) a reagent which can raise antibodies against Neisserial bacteria and (g) a method of treating a patient, comprising administering to the patient a therapeutically effective amount of these compositions.

[0025] Sequences

[0026] The invention also provides a protein or a nucleic acid having any of the sequences set out in the following examples. It also provides proteins and nucleic acid having sequence identity to these. As described above, the degree of ‘sequence identity’ is preferably greater than 50% (eg. 60%, 70%, 80%, 90%, 95%, 99% or more).

[0027] Nomenclature Herein

[0028] The 2166 protein sequences disclosed in WO99/24578, WO99/36544 and WO99/57280 are referred to herein by the following SEQ# numbers: 2 Application Protein sequences SEQ# herein WO99/24578 Even SEQ IDs 2-892 SEQ#s 1-446 WO99/36544 Even SEQ IDs 2-90 SEQ#s 447-491 WO99/57280 Even SEQ IDs 2-3020 SEQ#s 492-2001 Even SEQ IDs 3040-3114 SEQ#s 2002-2039 SEQ IDs 3115-3241 SEQ#s 2040-2166

[0029] In addition to this SEQ# numbering, the naming conventions used in WO99/24578, WO99/36544 and WO99/57280 are also used (e.g. ‘ORF4’, ‘ORF40’, ‘ORF40-1’ etc. as used in WO99/24578 and WO99/36544; ‘m919’, ‘g919’ and ‘a919’ etc. as used in WO99/57280).

[0030] The 2160 proteins NMB0001 to NMB2160 from Tettelin et al. [Science (2000) 287:1809-1815] are referred to herein as SEQ#s 2167-4326 [see also WO00/66791].

[0031] The term ‘protein of the invention’ as used herein refers to a protein comprising:

[0032] (a) one of sequences SEQ#s 1-4326; or

[0033] (b) a sequence having sequence identity to one of SEQ#s 1-4326; or

[0034] (c) a fragment of one of SEQ#s 1-4326.

[0035] The degree of ‘sequence identity’ referred to in (b) is preferably greater than 50% (eg. 60%, 70%, 80%, 90%, 95%, 99% or more). This includes mutants and allelic variants [e.g. see WO00/667,41]. Identity is preferably determined by the Smith-Waterman homology search algorithm as implemented in the MPSRCH program (Oxford Molecular), using an affine gap search with parameters gap open penalty=12 and gap extension penalty=1. Typically, 50% identity or more between two proteins is considered to be an indication of functional equivalence.

[0036] The ‘fragment’ referred to in (c) should comprise at least n consecutive amino acids from one of SEQ#s 1-4326 and, depending on the particular sequence, n is 7 or more (eg. 8, 10, 12, 14, 16, 18, 20, 25, 30, 35, 40, 50, 60, 70, 80, 90, 100 or more). Preferably the fragment comprises an epitope from one of SEQ#s 1-4326. Preferred fragments are those disclosed in WO00/71574 and WO01/04316.

[0037] Preferred proteins of the invention are found in N. meningitidis serogroup B.

[0038] Preferred proteins for use according to the invention are those of serogroup B N. meningitidis strain 2996 or strain 394/98 (a New Zealand strain). Unless otherwise stated, proteins mentioned herein are from N. meningitidis strain 2996. It will be appreciated, however, that the invention is not in general limited by strain. References to a particular protein (e.g. ‘287’, ‘919’ etc.) may be taken to include that protein from any strain.

[0039] It will be appreciated that references to “nucleic acid” includes DNA and RNA, and also their analogues, such as those containing modified backbones, and also peptide nucleic acids (PNA) etc.

BRIEF DESCRIPEON OF DRAWINGS

[0040] FIGS. 1 to 26 show hybrid proteins according to the invention.

MODES FOR CARRYING OUT THE INVENTION

EXAMPLE 1

Hybrids of ORF46

[0041] The complete ORF46 protein from N. meningitidis (serogroup B, strain 2996) has the following sequence: 3 1 LGISRKISLI LSILAVCLPM HAHASDLAND SFIRQVLDRQ HFEPDGKYHL 51 FGSRGELAER SGHIGLGKIQ SHQLGNIMIQ QAAIKGNIGY IVRFSDHGHE 101 VHSPFDNHAS HSDSDEAGSP VDGFSLYRIH WDGYEHHPAD GYDGPQGGGY 151 PAPKGARDIY SYDIKGVAQN IRLNLTDNRS TGQRLADRFH NAGSMLTQGV 201 GDGFKRATRY SPELDRSGNA AEAFNGTADI VKNIIGAAGE IVGAGDAVQG 251 ISEGSNIAVM HGLGLLSTEN KMARINDLAD MAQLKDYAAA AIRDWAVQNP 301 NAAQGIEAVS NIFMAAIPIK GIGAVRGKYG LGGITAHPIK RSQMGAIALP 351 KGKSAVSDNF ADAAYAKYPS PYHSPNIRSN LEQRYGKENI TSSTVPPSNG 401 KNVKLADQRH PKTGVPFDGK GFPNFEKHVK YDTKLDIQEL SGGGIPKAXP 451 VSDAKPRWEV DRKLNKLTTR EQVEENVQEI RNGNKNSNFS QHAQLEREIN 501 KLKSADEINF ADGMGKFTDS MNDKAFSELV KSVKENGFTN PVVEYVEING 551 KAYIVRGNNR VFAAEYLGRI HELKFKKVDF PVPNTSWKNP TDVLNESGNV 601 KRPRYRSK*

[0042] The leader peptide is underlined.

[0043] The sequences of ORF46 from other strains can be found in WO00/66741.

[0044] ORF46 has been fused at its C-terminus and N-terminus with 287, 919, and ORF1. The hybrid proteins were generally insoluble, but gave some good ELISA and bactericidal results (against the homologous 2996 strain): 4 Protein ELISA Bactericidal Ab Orf1-Orf46.1-His 850 256 919-Orf46.1-His 12900 512 919-287-Orf46-His n.d. n.d. Orf46.1-287His 150 8192 Orf46.1-919His 2800 2048 Orf46.1-287-919His 3200 16384

[0045] For comparison, ‘triple’ hybrids of ORF46.1, 287 (either as a GST fusion, or in &Dgr;G287 form) and 919 were constructed and tested against various strains (including the homologous 2996 strain) versus a simple mixture of the three antigens. FCA was used as adjuvant: 5 2996 BZ232 MC58 NGH38 F6124 BZ133 Mixture 8192 256  512 1024 >2048  >2048 ORF46.1-287- 16384  256 4096 8192 8192  8192 919his &Dgr;G287-919- 8192  64 4096 8192 8192 16384 ORF46.1his &Dgr;G287- 4096 128  256 8192  512  1024 ORF46.1- 919his

[0046] Again, the hybrids show equivalent or superior immunological activity.

[0047] Hybrids of two proteins (strain 2996) were compared to the individual proteins against various heterologous strains: 6 1000 MC58 F6124 (MenA) ORF46.1-His <4 4096 <4 ORF1-His 8 256 128 ORF1 - ORF46.1-His 1024 512 1024

[0048] Again, the hybrid shows equivalent or superior immunological activity.

EXAMPLE 2

Hybrids of &Dgr;G287

[0049] The deletion of the (Gly)6 sequence in 287 was found to have a dramatic effect on protein expression. The protein lacking the N-terminal amino acids up to GGGGGG is called ‘&Dgr;G287’. In strain MC58, its basic sequence (leader peptide underlined) is: 7     SPDVKS ADTLSKPAAP VVSEKETEAK EDAPQAGSQG QGAPSAQGSQ DMAAVSEENT GNGGAVTADN PKNEDEVAQN DMPQNAAGTD SSTPNHTPDP NMLAGNMENQ ATDAGESSQP ANQPDMANAA DGMQGDDPSA GGQNAGNTAA QGANQAGNNQ AAGSSDPIPA SNPAPANGGS NFGRVDLANG VLIDGPSQNI TLTHCKGDSC SGNNFLDEEV QLKSEFEKLS DADKISNYKK DGKNDKFVGL VADSVQMKGI NQYIIFYKPK PTSPARFRRS ARSRRSLPAE MPLIPVNQAD TLIVDGEAVS LTGHSGNIFA PEGNYRYLTY GAEKLPGGSY ALRVQGEPAK GEMLAGAAVY NGEVLHFHTE NGRPYPTRGR FAAKVDFGSK SVDGIIDSGD DLHMGTQKFK AAIDGNGFKG TWTENGSGDV SGKFYGPAGE EVAGKYSYRP TDAEKGGFGV FAGKKEQD*

[0050] &Dgr;G287, with or without His-tag (‘&Dgr;G287-His’ and ‘&Dgr;G287K’, respectively), are expressed at very good levels in comparison with the ‘287-His’ or ‘287untagged’.

[0051] On the basis of gene variability data, variants of &Dgr;G287-His were expressed in E. coli from a number of MenB strains, in particular from strains 2996, MC58, 1000, and BZ232. The results were also good—each of these gave high ELISA titres and also serum bactericidal titres of >8192. &Dgr;G287K, expressed from pET-24b, gave excellent titres in ELISA and the serum bactericidal assay.

[0052] Deletion of poly-Gly sequences is also applicable to Thp2 (NMB0460), 741 (NMB 1870) and 983 (NMB1969). When cloned in pET vector and expressed in E. coli without the sequence coding for their leader peptides and without poly-Gly (i.e. as “&Dgr;G forms”), the same effect was seen—expression was good in the clones carrying the deletion of the poly-glycine stretch, and poor or absent if the glycines were present in the expressed protein.

[0053] &Dgr;G287 was fused directly in-frame upstream of 919, 953, 961 (sequences shown below) and ORF46.1: 8 &Dgr;G287-919 ATGGCTAGCCCCGATGTTAAATCGGCGGACACGCTGTCAAAACCGGCCGCTCCTGTTGTTGCTGAAAAAGAGACAGAG GTAAAAGAAGATGCGCCACAGGCAGGTTCTCAAGGACAGGGCGCGCCATCCACACAAGGCAGCCAAGATATGGCGGCA GTTTCGGCAGAAAATACAGGCAATGGCGGTGCGGCAACAACGGACAAACCCAAAAATGAAGACGAGGGACCGCAAAAT GATATGCCGCAAAATTCCGCCGAATCCGCAAATCAAACAGGGAACAACCAACCCGCCGATTCTTCAGATTCCGCCCCC GCGTCAAACCCTGCACCTGCGAATGGCGGTAGCAATTTTGGAAGGGTTGATTTGGCTAATGGCGTTTTGATTGATGGG CCGTCGCAAAATATAACGTTGACCCACTGTAAAGGCGATTCTTGTAATGGTGATAATTTATTGGATGAAGAAGCACCG TCAAAATCAGAATTTGAAAATTTAAATGAGTCTGAACGAATTGAGAAATATAAGAAAGATGGGAAAAGCGATAAATTT ACTAATTTGGTTGCGACAGCAGTTCAAGCTAATGGAACTAACAAATATGTCATCATTTATAAAGACAAGTCCGCTTCA TCTTCATCTGCGCGATTCAGGCGTTCTGCACGGTCGAGGAGGTCGCTTCCTGCCGAGATGCCGCTAATCCCCGTCAAT CAGGCGGATACGCTGATTGTCGATGGGGAAGCGGTCAGCCTGACGGGGCATTCCGGCAATATCTTCGCGCCCGAAGGG AATTACCGGTATCTGACTTACGGGGCGGAAAAATTGCCCGGCGGATCGTATGCCCTCCGTGTGCAAGGCGAACCGGCA AAAGGCGAAATGCTTGCTGGCACGGCCGTGTACAACGGCGAAGTGCTGCATTTTCATACGGAAAACGGCCGTCCGTAC CCGACTAGAGGCAGGTTTGCCGCAAAAGTCGATTTCGGCAGCAAATCTGTGGACGGCATTATCGACAGCGGCGATGAT TTGCATATGGGTACGCAAAAATTCAAAGCCGCCATCGATGGAAACGGCTTTAAGGGGACTTGGACGGAAAATGGCGGC GGGGATGTTTCCGGAAGGTTTTACGGCCCGGCCGGCGAGGAAGTGGCGGGAAAATACAGCTATCGCCCGACAGATGCG GAAAAGGGCGGATTCGGCGTGTTTGCCGGCAAAAAAGAGCAGGATGGATCCGGAGGAGGAGGATGCCAAAGCAAGAGC ATCCAAACCTTTCCGCAACCCGACACATCCGTCATCAACGGCCCGGACCGGCCGGTCGGCATCCCCGACCCCGCCGGA ACGACGGTCGGCGGCGGCGGGGCCGTCTATACCGTTGTACCGCACCTGTCCCTGCCCCACTGGGCGGCGCAGGATTTC GCCAAAAGCCTGCAATCCTTCCGCCTCGGCTGCGCCAATTTGAAAAACCGCCAAGGCTGGCAGGATGTGTGCGCCCAA GCCTTTCAAACCCCCGTCCATTCCTTTCAGGCAAAACAGTTTTTTGAACGCTATTTCACGCCGTGGCAGGTTGCAGGC AACGGAAGCCTTGCCGGTACGGTTACCGGCTATTACGAGCCGGTGCTGAAGGGCGACGACAGGCGGACGGCACAAGCC CGCTTCCCGATTTACGGTATTCCCGACGATTTTATCTCCGTCCCCCTGCCTGCCGGTTTGCGGAGCGGAAAAQCCCTT GTCCGCATCAGGCAGACGGGAAAAAACAGCGGCACAATCGACAATACCGGCGGCACACATACCGCCGACCTCTCCCGA TTCCCCATCACCGCGCGCACAACGGCAATCAAAGGCAGGTTTGAAGGAAGCCGCTTCCTCCCCTACCACACGCGCAAC CAAATCAACGGCGGCGCGCTTGACGGCAAAGCCCCGATACTCGGTTACGCCGAAGACCCCGTCGAACTTTTTTTTATG CACATCCAAGGCTCGGGCCGTCTGAAAACCCCGTCCGGCAAATACATCCGCATCGGCTATGCCGACAAAAACGAACAT CCCTACGTTTCCATCGGACGCTATATGGCGGACAAAGGCTACCTCAAGCTCGGGCAGACCTCGATGCAGGGCATCAAA GCCTATATGCGGCAAAATCCGCAACGCCTCGCCGAAGTTTTGGGTCAAAACCCCAGCTATATCTTTTTCCGCGAGCTT GCCGGAAGCAGCAATGACGGTCCCGTCGGCGCACTGGGCACGCCGTTGATGGGGGAATATGCCGGCGCAGTCGACCGG CACTACATTACCTTGGGCGCGCCCTTATTTGTCGCCACCGCCCATCCGGTTACCCGCAAAGCCCTCAACCGCCTGATT ATGGCGCAGGATACCGGCAGCGCGATTAAAGGCGCGGTGCGCGTGGATTATTTTTGGGGATACGGCGACGAAGCCGGC GAACTTGCCGGCAAACAGAAAACCACGGGTTACGTCTGGCAGCTCCTACCCAACGGTATGAAGCCCGAATACCGCCCG TAACTCGAG 1 MASPDVKSAD TLSKPAAPVV AEKETEVKBD APQAGSQGQG APSTQGSQDM 51 AAVSAENTGN GGAATTDKPK NEDEGPQNDM PQNSAESANQ TGNNQPADSS 101 DSAPASNPAP ANGGSNFGRV DLANGVLIDG PSQNITLTHC KGDSCNGDNL 151 LDEEAPSKSE FENLNESERI EKYKKDGKSD KFTNLVATAV QANGTNKYVI 201 IYKDKSASSS SARFRRSAPS RRSLPAEMPL IPVNQADTLI VDGEAVSLTG 251 HSGNIFAPEG NYRYLTYGAE KLPGGSYALR VQGEPAKGEM LAGTAVYNGE 301 VLHFRTENGR PYPTRGFPAA KVDFGSKSVD GIIDSGDDLH MGTQKFKAAI 351 DGNGFKGTWT ENGGGDVSGR FYGPAGEEVA GKYSYRPTDA EKGGFGVFAG 401 KKEQDGSGGG GCQSKSIQTF PQPDTSVING PDRPVGIPDP AGTTVGGGGA 451 VYTVVPHLSL PHWAAQDRAK SLQSFRLGCA NLKNRQGWQD VCAQAFQTPV 501 HSFQAKQFFE RYFTPWQVAG NGSLAGTVTG YYEPVLKGDD RRTAQARPPI 551 YGIPDDFISV PLPAGLRSGK ALVRIRQTGK NSGTIDNTGG THTADLSRFP 601 ITARTTAIKG RFEGSRFLPY HTRNQINGGA LDGKAPILGY AEDPVELFFM 651 HIQGSGRLKT PSGKYIRIGY ADKNEHPYVS IGRYHADKGY LKLGQTSMQG 701 IKAYMRQNPQ RLAEVLGQNP SYIFFRELAG SSNDGPVGAL GTPLMGEYAG 751 AVDRHYITLG APLFVATMIP VTEXALNRLI MAQDTGSAIK GAVRVDYFWG 801 YGDEAGELAG KQKTTGYVWQ LLPNGMKPEY RP* AG287-953 ATGGCTAGCCCCGATGTTAAATCGGCGGACACGCTGTCAAAACCGGCCGCTCCTGTTGTTGCTGAAAAAGAGACAGAG GTAAAAGAAGATGCGCCACAGGCAGGTTCTCAAGGACAGGGCGCGCCATCCACACAAGGCAGCCAAGATATGGCGGCA GTTTCGGCAGAAAATACAGGCAATGGCGGTGCGGCAACAACGGACAAACCCAAAAATGAAGACGAGGGACCGCAAAAT GATATGCCGCAAAATTCCGCCGAATCCGCAAATCAAACAGGGAACAACCAACCCGCCGATTCTTCAGATTCCGCCCCC GCGTCAAACCCTGCACCTGCGAATGGCGGTAGCAATTTTGGAAGGGTTGATTTGGCTAATGGCGTTTTGATTGATGGG CCGTCGCAAAATATAACGTTGACCCACTGTAAAGGCGATTCTTGTAATGGTGATAATTTATTGGATGAAGAAGCACCG TCAAAATCAGAATTTGAAAATTTAAATGAGTCTGAACGAATTGAGAAATATAAGAAAGATGGGAAAAGCGATAAATTT ACTAATTTGGTTGCGACAGCAGTTCAAGCTAATGGAACTAACAAATATGTCATCATTTATAAAGACAAGTCCGCTTCA TCTTCATCTGCGCGATTCAGGCGTTCTGCACGGTCGAGGAGGTCGCTTCCTGCCGAGATGCCGCTAATCCCCGTCAAT CAGGCGGATACGCTGATTGTCGATGGGGAAGCGGTCAGCCTGACGGGGCATTCCGGCAATATCTTCGCGCCCGAAGGG AATTACCGGTATCTGACTTACGGGGCGGAAAAATTGCCCGGCGGATCGTATGCCCTCCGTGTGCAAGGCGAACCGGCA AAAGGCGAAATGCTTGCTGGCACGGCCGTGTACAACGGCGAAGTGCTGCATTTTCATACGGAAAACGGCCGTCCGTAC CCGACTAGAGGCAGGTTTGCCGCAAAAGTCGATTTCGGCAGCAAATCTGTGGACGGCATGATCGACAGCGGCGATGAT TTGCATATGGGTACGCAAAAATTCAAAGCCGCCATCGATGGAAACGGCTTTAAGGGGACTTGGACGGAAAATGGCGGC GGGGATGTTTCCGGAAGGTTTTTACGGCCCGGCCGGCGAGGAAGTGGCGGAAAATACAGCTATCGCCCGACAGATGCG GAAAAGGGCGGATTCGGCGTGTTTGCCGGCAAAAAAGAGCAGGATGGATCCGGAGGAGGAGGAGCCACCTACAAAGTG GACGAATATCACGCCAACGCCCGTTTCGCCATCGACCATTTCAACACCAGCACCAACGTCGGCGGTTTTTACGGTCTG ACCGGTTCCGTCGAGTTCGACCAAGCAAAACGCGACGGTAAAATCGACATCACCATCCCCGTTGCCAACCTGCAAAGC GGTTCGCAACACTTTACCGACCACCTGAAATCAGCCGACATCTTCGATGCCGCCCAATATCCGGACATCCGCTTTGTT TCCACCAAATTCAACTTCAACGGCAAAAAACTGGTTTCCGTTGACGGCAACCTGACCATGCACGGCAAAACCGCCCCC GTCAAACTCAAAGCCGAAAAATTCAACTGCTACCAAAGCCCGATGGCGAAAACCGAAGTTTGCGGCGGCGACTTCAGC ACCACCATCGACCGCACCAAATGGGGCGTGGACTACCTCGTTAACGTTGGTATGACCAAAAGCGTCCGCATCGACATC CAAATCGAGGCAGCCAAACAATAACTCGAG 1 MASPDVKSAD TLSKPAAPVV AEKETBVKED APQAGSQGQG APSTQGSQDM 51 AAVSAENTGN GGAATTDKPK NEDEGPQNDM PWNSAESANQ TGNNQPADSS 101 DSAPASNPAP ANGGSNFGRV DLANGVLIDG PSQNITLTHC KGDSCNGDNL 151 LDEEAPSKSE FENLNESERI KKYKKDGKSD KFTNLVATAV QANGTNKYVI 201 IYKDKSASSS SARFRRSARS RRSLPAEMPL IPVNQADTLI VDGEAVSLTG 251 HSGNIFAPEG NYRYLTYGAB KLPGGSYALR VQGEPAKGEM LAGTAVYNGE 301 VLHFHTENGR PYPTRGRFAA KVDFGSKSVD GIIDSGDDLH MGTQKFKAAI 351 DGNGFXGTWT ENGGGDVSGR FYGPAGEBVA GKYSYRPTDA EKGGFGVFAG 401 KKEQDGSGGG GATYKVDEYH ANARFAIDHF NTSTNVGGFY GLTGSVEFDQ 451 AXRDGKIDIT IPVANLQSGS QHFTDHLKSA DIFDAAQYPD IRFVSTKFNF 501 NGKXLVSVDG NLTHHGKTAP VKLKAEKFNC YQSPMAKTEV CGGDFSTTID 551 RTKWGVDYLV NVGMTKSVRI DIQIEAAKQ* AG287-961 ATGGCTAGCCCCGATGTTAAATCGGCGGACACGCTGTCAAAACCGGCCGCTCCTGTTGTTGCTGAAAAAGAGACAGAG GTAAAAGAAGATGCGCCACAGGCAGGTTCTCAAGGACAGGGCGCGCCATCCACACAAGGCAGCCAAGATATGGCGGCA GTTTCGGCAGAAAATACAGGCAATGGCGGTGCGGCAACAACGGACAAACCCAAAAATGAAGACGAGGGACCGCAAAAT GATATGCCGCAAAATTCCGCCGAATCCGCAAATCAAACAGGGAACAACCAACCCGCCGATTCTTCAGATTCCGCCCCC GCGTCAAACCCTGCACCTGCGAATGGCGGTAGCAATTTTGGAAGGGTTGATTTGGCTAATGGCGTTTTGATTGATGGG CCGTCGCAAAATATAACGTTGACCCACTGTAAAGGCGATTCTTGTAATGGTGATAATTTATTGGATGAAGAAGCACCG TCAAAATCAGAATTTGAAAATTTAAATGAGTCTGAACGAATTGAGAAATATAAGAAAGATGGGAAAAGCGATAAATTT ACTAATTTGGTTGCGACAGCAGTTCAAGCTAATGGAACTAACAAATATGTCATCATTTATAAAGACAAGTCCGCTTCA TCTTCATCTGCGCGATTCAGGCGTTCTGCACGGTCGAGGAGGTCGCTTCCTGCCGAGATGCCGCTAATCCCCGTCAAT CAGGCGGATACGCTGATTGTCGATGGGGAAGCGGTCAGCCTGACGGGGCATTCCGGCAATATCTTCGCGCCCGAAGGG AATTACCGGTATCTGACTTACGGGGCGGAAAAATTGCCCGGCGGATCGTATGCCCTCCGTGTGCAAGGCGAACCGGCA AAAGGCGAAATGCTTGCTGGCACGGCCGTGTACAACGGCGAAGTGCTGCATTTTCATACGGAAAACGGCCGTCCGTAC CCGACTAGAGGCAGGTTTGCCGCAAAAGTCGATTTCGGCAGCAAATCTGTGGACGGCATTATCGACAGCGGCGATGAT TTGCATATGGGTACGCAAAAATTCAAAGCCGCCATCGATGGAAACGGCTTTAAGGGGACTTGGACGGAAAATGGCGGC GGGGATGTTTCCGGAAGGTTTTACGGCCCGGCCGGCGAGGAAGTGGCGGGAAAATACAGCTATCGCCCGACAGATGCG GAAAAGGGCGGATTCGGCGTGTTTGCCGGCAAAAAAGAGCAGGATGGATCCGGAGGAGGAGGAGCCACAAACGACGAC GATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATCAACGGTTTCAAAGCTGGA GAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCCGATGTTGAAGCCGACGAC TTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAACAAACAAAACGTCGATGCC AAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGATGCCGCTTTAGCAGATACT GATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACATTTGCTGAAGAGACTAAG ACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCATGCCGAAGCATTCAACGAT ATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAATGAAGCCAAACAGACGGCC GAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAAGCCGAAGCTGCCGCTGGC ACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAAGCTGATATCGCTACGAAC AAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGACAGCAAATTTGTCAGAATT GATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCCATTGCCGATCACGATACT CGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTTGCAGAACAAGCCGCGCTC TCCGGTCTGTTCCAACCTTACAACGTGGGTCGGTTCAATGTAACGGCTGCAGTCGGCGGCTACAAATCCGAATCGGCA GTCGCCATCGGTACCGGCTTCCGCTTTACCGAAAACTTTGCCGCCAAAGCAGGCGTGGCAGTCGGCACTTCGTCCGGT TCTTCCGCAGCCTACCATGTCGGCGTCAATTACGAGTGGTAACTCGAG 1 HASPDVKSAD TLSKPAAPVV AEXETEVKED APQAGSQGQG APSTQGSQDM 51 AAVSAENTGN GGAATTDKPK NEDEGPQNDM PQNSABSANQ TGNNQPADSS 101 DSAPASNPAP ANGGSNFGRV DLANGVLIDG PSQNITLTHC KGDSCNGDNL 151 LDEEAPSKSE FENLNESERI EKYKKDGKSD KFTNLVATAV QANGTNKYVI 201 IYKDKSASSS SARFRRSARS RRSLPAENPL IPVNQADTLI VDGEAVSLTG 251 HSGNIFAPEG NYRYLTYGAB KLPGGSYALR VQGEPAKGEM LAGTAVYNGE 301 VLHFHTENGR PYPTRGRFAA KVDFGSKSVD GIIDSGDDLH MGTQKFKAAI 351 DGNGFKGTWT ENGGGDVSGR PYGPAGEEVA GKYSYRPTDA EKGGFGVFAG 401 KKEQDGSGGG GATNDDDVKK AATVAIAAAY NNGQEINGFK AGETIYDIDE 451 DGTITKKDAT AADVEADDFK GLGLKKVVTN LTKTVNENKQ NVDAKVKAAE 501 SEIEKLTTKL ADTDAALADT DAALDATTNA LNKLGENITT FAEETKTNIV 551 KIDEKLEAVA DTVDKHAEAF NDIADSLDET NTKADEAVKT ANEAKQTAEE 601 TKQNVDAKVK AAETAAGKAE AAAGTANTAA DKABAVAAKV TDIKADIATN 651 KDNIAKKANS ADVYTREESD SKFVRIDGLN ATTEKLDTRL ASAEKSIADH 701 DTRLNGLDKT VSDLRXETRQ GLAEQAALSG LFQPYNVGRF NVTAAVGGYK 751 SESAVAIGTG FRFTENFAAK AGVAVGTSSG SSAAYHVGVN YEW*

[0054] 9 ELISA Bactericidal &Dgr;G287-953-His 3834 65536 &Dgr;G287-961-His 108627 65536

[0055] The bactericidal efficacy (homologous strain) of antibodies raised against the hybrid proteins was compared with antibodies raised against simple mixtures of the component antigens (using 287-GST) for 919 and ORF46.1: 10 Mixture with 287 Hybrid with &Dgr;G287 919 32000 128000 ORF46.1 128 16000

[0056] Data for bactericidal activity against heterologous MenB strains and against serotypes A and C were also obtained: 11 919 ORF46.1 Strain Mixture Hybrid Mixture Hybrid NGH38 1024 32000 — 16384 MC58 512 8192 — 512 BZ232 512 512 — — MenA (F6124) 512 32000 — 8192 MenC (C11) >2048 >2048 — — MenC (BZ133) >4096 64000 — 8192

[0057] The hybrid proteins with &Dgr;G287 at the N-terminus are therefore immunologically superior to simple mixtures, with &Dgr;G287-ORF46.1 being particularly effective, even against heterologous strains. &Dgr;G287-ORF46.1K may be expressed in pET-24b.

[0058] The same hybrid proteins were made using New Zealand strain 394/98 rather than 2996: 12 AG2B7NZ-919 ATGGCTAGCCCCGATGTCAAGTCGGCGGACACGCTGTCAAAACCTGCCGCCCCTGTTGTTTCTGAAAAAGAGACAGAG GCAAAGGAAGATGCGCCACAGGCAGGTTCTCAAGGACAGGGCGCGCCATCCGCACAAGGCGGTCAAGATATGGCGGCG GTTTCGGAAGAAAATACAGGCAATGGCGGTGCGGCAGCAACGGACAAACCCAAAAATGAAGACGAGGGGGCGCAAAAT GATATGCCGCAAAATGCCGCCGATACAGATAGTTTGACACCGAATCACACCCCGGCTTCGAATATGCCGGCCGGAAAT ATGGAAAACCAAGCACCGGATGCCGGGGAATCGGAGCAGCCGGCAAACCAACCGGATATGGCAAATACGGCGGACGGA ATGCAGGGTGACGATCCGTCGGCAGGCGGGGAAAATGCCGGCAATACGGCTGCCCAAGGTACAAATCAAGCCGAAAAC AATCAAACCGCCGGTTCTCAAAATCCTGCCTCTTCAACCAATCCTAGCGCCACGAATAGCGGTGGTGATTTTGGAAGG ACGAACGTGGGCAATTCTGTTGTGATTGACGGGCCGTCGCAAAATATAACGTTGACCCACTGTAAAGGCGATTCTTGT AGTGGCAATAATTTCTTGGATGAAGAAGTACAGCTAAAATCAGAATTTGAAAAATTAAGTGATGCAGACAAAATAAGT AATTACAAGAAAGATGGGAAGAATGACGGGAAGAATGATAAATTTGTCGGTTTGGTTGCCGATAGTGTGCAGATGAAG GGAATCAATCAATATATTATCTTTTATAAACCTAAACCCACTTCATTTGCGCGATTTAGGCGTTCTGCACGGTCGAGG CGGTCGCTTCCGGCCGAGkTGCCGCTGATTCCCGTCAATCAGGCGGATACGCTGATTGTCGATGGGGAAGCGGTCAGC CTGACGGGGCATTCCGGCAATATCTTCGCGCCCGAkGGGAATTACCGGTATCTGACTTACGGGGCGGAAAAATTGCCC GGCGGATCGTATGCCCTCCGTGTTCAAGGCGAACCTTCAAAAGGCGAAATGCTCGCGGGCACGGCAGTGTACAACGGC GAAGTGCTGCATTTTCATACGGAAAACGGCCGTCCGTCCCCGTCCAGAGGCAGGTTTGCCGCAAAAGTCGATTTCGGC AGCAAATCTGTGGACGGCATTATCGACAGCGGCGATGGTTTGCATATGGGTACGCAAAAATTCAAAGCCGCCATCGAT GGAAACGGCTTTAAGGGGACTTGGACGGAAAATGGCGGCGGGGATGTTTCCGGAAAGTTTTACGGCCCGGCCGGCGAG GAAGTGGCGGGAAAATACAGCTATCGCCCAACAGATGCGGAAAAGGGCGGATTCGGCGTGTTTGCCGGCAAAAAGGAG CAGGATGGATCCGGAGGAGGAGGATGCCAAAGCAAGAGCATGCAAACCTTTCCGCAACCCGACACATCCGTCATCAAC GGCCCGGACCGGCCGGTCGGCATCCCCGACCCCGCCGGAACGACGGTCGGCGGCGGCGGGGCCGTCTATACCGTTGTA CCGCACCTGTCCCTGCCCCACTGGGCGGCGCAGGATTTCGCCAAAAGCCTGCAATCCTTCCGCCTCGGCTGCGCCAAT TTGAAAAACCGCCAAGGCTGGCAGGATGTGTGCGCCCAAGCCTTTCAAACCCCCGTCCATTCCTTTCAGGCAAAACAG TTTTTTGAACGCTATTTCACGCCGTGGCAGGTTGCAGGCAACGGAAGCCTTGCCGGTACGGTTACCGGCTATTACGAG CCGGTGCTGAAGGGCGACGACAGGCGGACGGCACAAGCCCGCTTCCCGATTTACGGTATTCCCGACGATTTTATCTCC GTCCCCCTGCCTGCCGGTTTGCGGAGCGGAAAAGCCCTTGTCCGCATCAGGCAGACGGGAAAAAACAGCGGCACAATC GACAATACCGGCGGCACACATACCGCCGACCTCTCCCGATTCCCCATCACCGCGCGCACAACGGCAATCAAAGGGAGG TTTGAAGGAAGCCGCTTCCTCCCCTACCACACGCGCAACCAAATCAACGGCGGCGCGCTTGACGGCAAAGCCCCGATA CTCGGTTACGCCGAAGACCCCGTCGAACTTTTTTTTATGCACATCCAAGGCTCGGGCCGTCTGAAAACCCCGTCCGGC AAATACATCCGCATCGGCTATGCCGACAAAAACGAACATCCCTACGTTTCCATCGGACGCTATATGGCGGACAAAGGC TACCTCAAGCTCGGGCAGACCTCGATGCAGGGCATCAAAGCCTATATGCGGCAAAATCCGCAACGCCTCGCCGAAGTT TTGGGTCAAAACCCCAGCTATATCTTTTTCCGCGAGCTTGCCGGAAGCAGCAATGACGGTCCCGTCGGCGCACTGGGC ACGCCGTTGATGGGGGAATATGCCGGCGCAGTCGACCGGCACTACATTACCTTGGGCGCGCCCTTATTTGTCGCCACC GCCCATCCGGTTACCCGCAAAGCCCTCAACCGCCTGATTATGGCGCAGGATACCGGCAGCGCGATTAAAGGCGCGGTG CGCGTGGATTATTTTTGGGGATACGGCGACGAAGCCGGCGAACTTGCCGGCAAACAGAAAACCACGGGTTACGTCTGG CAGCTCCTACCCAACGGTATGAAGCCCGAATACCGCCCGTAAAAGCTT 1 HASPDVKSAD TLSKPAAPVV SEKETEAXED APQAGSQGQG APSAQGGQDM 51 AAVSEENTGN GGAAATDKPK NEDEGAQNDM PQNAADTDSL TPNHTPASNM 101 PAGNMENQAP DAGESEQPAN QPDMANTADG MQGDDPSAGG ENAGNTAAQG 151 TNQAENNQTA GSQNPASSTN PSATNSGGDF GRTNVGNSVV IDGPSQNITL 201 THCKGDSCSG NNFLDEEVQL KSEFEKLSDA DKISNYKKDG KNDGKNDKFV 251 GLVADSVQMK GINQYIIFYK PKPTSFARFR RSARSRRSLP AENPLIPVNQ 301 ADTLIVDGEA VSLTGHSGNI FAPEGNYRYL TYGAEKLPGG SYALRVQGEP 351 SKGEMLAGTA VYNGEVLHFH TENGRPSPSR GRFAAKVDFG SKSVDGIIDS 401 GDGLHHGTQK FKAAIDGNGF KGTWTENGGG DVSGKFYGPA GEEVAGKYSY 451 RPTDAEKGGF GVFAGKKEQD GSGGGGCQSK SIQTFPQPDT SVINGPDRPV 501 GIPDPAGTTV GGGGAVYTVV PHLSLPHWAA QDFAKSLQSF RLGCANLKNR 553 QGWQDVCAQA FQTPVHSFQA KQFFERYFTP WQVAGNGSLA GTVTGYYEPV 601 LKGDDRRTAQ ARFPIYGIPD DFISVPLPAG LRSGKALVRI RQTGKNGSTI 651 DNTGGTHTAD LSRFPITART TAIKGRFEGS RPLPYHTRNQ INGGALDGKA 701 PILGYAEDPV ELFFMHIQGS GRLKTPSGKY IRIGYADKNB HPYVSIGRYM 751 ADKGYLKLGQ TSMQGIKAYM RQNPQRLAEV LGQNPSYIFF RELAGSSNDG 801 PVGALGTPLM GEYAGAVDRH YITLGAPLFV ATAHPVTEKA LNRLIMAQDT 851 GSAIKGAVRV DYFWGYGDEA GELAGKQKTT GYVWQLLPNG MKPEYRP* &Dgr;G287NZ-953 ATGGCTAGCCCCGATGTCAAGTCGGCGGACACGCTGTCAAAACCTGCCGCCCCTGTTGTTTCTGAAAAAGAGACAGAG GCAAAGGAAGATGCGCCACAGGCAGGTTCTCAAGGACAGGGCGCGCCATCCGCACAAGGCGGTCAAGATATGGCGGCG GTTTCGGAAGAAAATACAGGCAATGGCGGTGCGGCAGCAACGGACAAACCCAAAAATGAAGACGAGGGGGCGCAAAAT GATATGCCGCAAAATGCCGCCGATACAGATAGTTTGACACCGAATCACACCCCGGCTTCGAATATGCCGGCCGGAAAT ATGGAAAACCAAGCACCGGATGCCGGGGAATCGGAGCAGCCGGCAAACCAACCGGATATGGCAAATACGGCGGACGGA ATGCAGGGTGACGATCCGTCGGCAGGCGGGGAAAATGCCGGCAATACGGCTGCCCAAGGTACAAATCAAGCCGAAAAC AATTACAAGAAAGATGGGAAGAATGACGGGAAGAATGATAAATTTGTCGGTTTGGTTGCCGATAGTGTGCAGATGAAG ACGAACGTGGGCAATTCTGTTGTGATTGACGGGCCGTCGCAAAATATAACGTTGACCCACTGTAAAGGCGATTCTTGT AGTGGCAATAATTTCTTGGATGAAGAAGTACAGCTAAAATCAGAATTTGAAAAATTAAGTGATGCAGACAAAATAAGT AATTACAAGAAAGATGGGAAGAATGACGGGAAGAATGATAAATTTGTCGGTTTGGTTGCCGATAGTGTGCAGATGAAG GGAATCAATCAATATATTATCTTTTATAAACCTAAACCCACTTCATTTGCGCGATTTAGGCGTTCTGCACGGTCGAGG CGGTCGCTTCCGGCCGAGATGCCGCTGATTCCCGTCAATCAGGCGGATACGCTGATTGTCGATGGGGAAGCGGTCAGC CTGACGGGGCATTCCGGCAATATCTTCGCGCCCGAAGGGAATTACCGGTATCTGACTTACGGGGCGGAAAAATTGCCC GGCGGATCGTATGCCCTCCGTGTTCAAGGCGAACCTTCAAAAGGCGAAATGCTCGCGGGCACGGCAGTGTACAACGGC GAAGTGCTGCATTTTCATACGGAAAACGGCCGTCCGTCCCCGTCCAGAGGCAGGTTTGCCGCAAAAGTCGATTTCGGC AGCAAATCTGTGGACGGCATTATCGACAGCGGCGATGGTTTGCATATGGGTACGCAAAAATTCAAAGCCGCCATCGAT GGAAACGGCTTTAAGGGGACTTGGACGGAAAATGGCGGCGGGGATGTTTCCGGAAAGTTTTACGGCCCGGCCGGCGAG GAAGTGGCGGGAAAATACAGCTATCGCCCAACAGATGCGGAAAAGGGCGGATTCGGCGTGTTTGCCGGCAAAAAAGAG CAGGATGGATCCGGAGGAGGAGGAGCCACCTACAAAGTGGACGAATATCACGCCAACGCCCGTTTCGCCATCGACCAT TTCAACACCAGCACCAACGTCGGCGGTTTTTACGGTCTGACCGGTTCCGTCGAGTTCGACCAAGCAAAACGCGACGGT AAAATCGACATCACCATCCCCGTTGCCAACCTGCAAAGCGGTTCGCAACACTTTACCGACCACCTGAAATCAGCCGAC ATCTTCGATGCCGCCCAATATCCGGACATCCGCTTTGTTTCCACCAAATTCAACTTCAACGGCAAAAAACTGGTTTCC GTTGACGGCAACCTGACCATGCACGGCAAAACCGCCCCCGTCAAACTCAAAGCCGAAAAATTCAACTGCTACCAAAGC CCGATGGCGAAAACCGAAGTTTGCGGCGGCGACTTCAGCACCACCATCGACCGCACCAAATGGGGCGTGGACTACCTC GTTAACGTTGGTATGACCAAAAGCGTCCGCATCGACATCCAAATCGAGGCAGCCAAACAATAAAAGCTT 1 NASPDVKSAD TLSKPAAPVV SEKETEAKED APQAGSQGQG APSAQGGQDM 51 AAVSEBNTGN GGAAATDKPK NEDEGAQNDM PQNAADTDSL TPNHTPASNM 101 PAGNMENQAP DAGESEQPAN QPDMANTADG MQGDDPSAGG ENAGNTAAQG 151 TNQAENNQTA GSQNPASSTN PSATNSGGDF GRTNVGNSVV IDGPSQNITL 201 THCKGDSCSG NNFLDEEVQL KSEPEKLSDA DKISNYKKDG KNDGKNDKFV 251 GLVADSVQMK GINQYIIFYK PKPTSFAPFR RSARSRRSLP AEMPLIPVNQ 301 ADTLIVDGEA VSLTGHSGNI FAPEGNYRYL TYGAEKLPGG SYALRVQGEP 351 SKGEMLAGTA VYNGEVUIFH TENGRPSPSR GRFAAKVDFG SKSVDGIIDS 401 GDGLHMGTQK FKAAIDGNGF KGTWTENGGG DVSGKFYGPA GEEVAGKYSY 451 RPTDAEKGGF GVFAGKKEQD GSGGGGATYK VDEYHANARF AIDHFNTSTN 501 VGGFYGLTGS VEFDQAKRDG KIDITIPVAN LQSGSQHFTD HLKSADIFDA 551 AQYPDIRFVS TKFNFNGKKL VSVDGNLTMH GKTAPVKLKA EKFNCYQSPM 601 AKTEVCGGDF STTIDRTKWG VDYLVNVGMT KSVRIDIQIE AAKQ* &Dgr;G287NZ-961 ATGGCTAGCCCCGATGTCAAGTCGGCGGACACGCTGTCAAAACCTGCCGCCCCTGTTGTTTCTGAAAAAGAGACAGAG GCAAAGGAAGATGCGCCACAGGCAGGTTCTCAAGGACAGGGCGCGCCATCCGCACAAGGCGGTCAAGATATGGCGGCG GTTTCGGAAGAAAATACAGGCAATGGCGGTGCGGCAGCAACGGACAAACCCAAAAATGAAGACGAGGGGGCGCAAAAT GATATGCCGCAAAATGCCGCCGATACAGATAGTTTGACACCGAATCACACCCCGGCTTCGAATATGCCGGCCGGAAAT ATGGAAAACCAAGCACCGGATGCCGGGGAATCGGAGCAGCCGGCAAACCAACCGGATATGGCAAATACGGCGGACGGA ATGCAGGGTGACGATCCGTCGGCAGGCGGGGAAAATGCCGGCAATACGGCTGCCCAAGGTACAAATCAAGCCGAAAAC AATCAAACCGCCGGTTCTCAAAATCCTGCCTCTTCAACCAATCCTAGCGCCACGAATAGCGGTGGTGATTTTGGAAGG ACGAACGTGGGCAATTCTGTTGTGATTGACGGGCCGTCGCAAAATATAACGTTGACCCACTGTAAAGGCGATTCTTGT AGTGGCAATAATTTCTTGGATGAAGAAGTACAGCTAAAATCAGAATTTGAAAAATTAAGTGATGCAGACAAAATAAGT AATTACAAGAAAGATGGGAAGAATGACGGGAAGAATGATAAATTTGTCGGTTTGGTTGCCGATAGTGTGCAGATGAAG GGAATCAATCAATATATTATCTTTTATAAACCTAAACCCACTTCATTTGCGCGATTTAGGCGTTCTGCACGGTCGAGG CGGTCGCTTCCGGCCGAGATGCCGCTGATTCCCGTCAATCAGGCGGATACGCTGATTGTCGATGGGGAAGCGGTCAGC CTGACGGGGCATTCCGGCAATATCTTCGCGCCCGAAGGGAATTACCGGTATCTGACTTACGGGGCGGAAAAATTGCCC GGCGGATCGTATGCCCTCCGTGTTCAAGGCGAACCTTCAAAAGGCGAAATGCTCGCGGGCACGGCAGTGTACAACGGC GAAGTGCTGCATTTTCATACGGAAAACGGCCGTCCGTCCCCGTCCAGAGGCAGGTTTGCCGCAAAAGTCGATTTCGGC AGCAAATCTGTGGACGGCATTATCGACAGCGGCGATGGTTTGCATATGGGTACGCAAAAATTCAAAGCCGCCATCGAT GGAAACGGCTTTAAGGGGACTTGGACGGAAAATGGCGGCGGGGATGTTTCCGGAAAGTTTTACGGCCCGGCCGGCGAG GAAGTGGCGGGAAAATACAGCTATCGCCCAACAGATGCGGAAAAGGGCGGATTCGGCGTGTTTGCCGGCAAAAAAGAG CAGGATGGATCCGGAGGAGGAGGAGCCACAAACGACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCC TACAACAATGGCCAAGAAATCAACGGTTTCAAAGCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACC AAAAAAGACGCAACTGCAGCCGATGTTGAAGCCGACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTG ACCAAAACCGTCAATGAAAACAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACA ACCAAGTTAGCAGACACTGATGCCGCTTTAGCAGATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAA TTGGGAGAAAATATAACGACATTTGCTGAAGAGACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTG GCTGATACCGTCGACAAGCATGCCGAAGCATTCAACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGAC GAAGCCGTCAAAACCGCCAATGAAGCCAAACAGACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCT GCAGAAACTGCAGCAGGCAAAGCCGAAGCTGCCGCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCT GCAAAAGTTACCGACATCAAAGCTGATATCGCTACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTG TACACCAGAGAAGAGTCTGACAGCAAATTTGTCAGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGC TTGGCTTCTGCTGAAAAATCCATTGCCGATCACGATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGC AAAGAAACCCGCCAAGGCCTTGCAGAACAAGCCGCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTCGGTTCAAT GTAACGGCTGCAGTCGGCGGCTACAAATCCGAATCGGCAGTCGCCATCGGTACCGGCTTCCGCTTTACCGAAAACTTT GCCGCCAAAGCAGGCGTGGCAGTCGGCACTTCGTCCGGTTCTTCCGCAGCCTACCATGTCGGCGTCAATTACGAGTGG TAAAAGCTT 1 MASPDVKSAD TLSKPAAPVV SEKETEAKED APQAGSQGQG APSAQGGQDM 51 AAVSEENTGN GGAAATDKPK NEDEGAQNDM PQNAADTDSL TPNHTPASNM 101 PAGNMENQAP DAGESEQPAN QPDMANTADG MQGDDPSAGG ENAGNTAAQG 151 TNQAENNQTA GSQNPASSTN PSATNSGGDF GRTNVGNSVV IDGPSQNITL 201 THCKGDSCSG NNFLDEEVQL KSEFEKLSDA DKISNYKKDG KNDGKNDKFV 251 GLVADSVQMK GINQYIIFYK PKPTSFARFR RSARSRRSLP AEMPLIPVNQ 301 ADTLIVDGEA VSLTGHSGNI FAPEGNYRYL TYGAEKLPGG SYALRVQGEP 351 SKGEMLAGTA VYNGEVLHFH TENGRPSPSR GRFAAKVDFG SKSVDGIIDS 401 GDGLHMGTQK FKAAIDGNGF KGTWTENGGG DVSGKFYGPA GEEVAGKYSY 451 RPTDAEKGGF GVFAGKKEQD GSGGGGATND DDVKKAATVA IAAAYNNGQE 501 INGFKAGETI YDIDEDGTIT KKDATAADVE ADDFKGLGLK KVVTNLTKTV 551 NENKQNVDAK VKAAESEIEK LTTKLADTDA ALADTDAALD ATTNALNKLG 601 ENITTFAEET KTNIVKIDEK LEAVADTVDK HAEAFNDIAD SLDETNTKAD 651 EAVKTANEAK QTAEETKQNV DAKVKAAETA AGKAEAAAGT ANTAADKAEA 701 VAAKVTDIKA DIATNKDNIA KKANSADVYT REESDSKFVR IDGLNATTEK 751 LDTRLASAEK SIADHDTRLN GLDKTVSDLR KETRQGLAEQ AALSGLFQPY 801 NVGRFNVTAA VGGYKSESAV AIGTGFRFTE NFAAKAGVAV GTSSGSSAAY 851 HVGVNYEW*

EXAMPLE 3

Hybrids of &Dgr;G983

[0059] Protein 983 has the following sequence: 13 983                                         &Dgr;G983 1 MRTTPTFPTK TFKPTAMALA VATTLSACLG GGGGGTSAPD FNAGGTGIGS 51 NSRATTAKSA AVSYAGIKNE HCKDRSNLCA GRDDVAVTDR DAKINAPPPN 101 LHTGDFPNPN DAYKNLINLK PAIEAGYTGR GVEVGIVDTG ESVGSISFPE 151 LYGRKERGYN ENYKNYTAYN RKEAPEDGGG KDIEASFDDE AVIETEAKPT 201 DIRHVKEIGH IDLVSHIIGG RSVDGRPAGG IAPDATLHIM NTNDETKNEM 251 MVAAIRNAWV KLGERGVRIV NNSFGTTSRA GTADLFQIAN SEEQYRQALL 301 DYSGGDKTDE GIRIMQQSDY GNLSYHIRNK NNLFIFSTGN DAQAQPNTYA 351 LLPFYEKDAQ KGIITVAGVD RSGEKFKREM YGEPGTEPLE YGSNHCGITA 401 MWCLSAPYEA SVRFTRTNPI QIAGTSFSAP IVTGTAALLL QKYPWMSNDN 451 LRTTLLTTAQ DIGAVGVDSK FGWGLLDAGK ANNGPASFPF GDFTADTKGT 501 SDIAYSFRND ISGTGGLIKK GGSQLQLHGN NTYTGKTIIE GGSLVLYGNN 551 KSDMRVETKG ALIYNGAASG GSLNSDGIVY LADTDQSGAN ETVHIKGSLQ 601 LDGKGTLYTR LGKLLKVDGT AIIGGKLYMS ARGKGAGYLN STGRRVPFLS 651 AAKIGQDYSF FTNIETDGGL LASLDSVEKT AGSEGDTLSY YVRRGNAART 701 ASAAAHSAPA GLKHAVEQGG SNLENLMVEL DASESSATPE TVETAAADRT 751 DMPGIRPYGA TFRAAAAVQR ANAADGVRIF NSLAATVYAD STAAHADHQG 801 RRLKAVSDGL DHNGTGLRVI AQTQQDGGTW EQGGVEGKNR GSTQTVGIAA 851 KTGENTTAAA TLGMGRSTWS ENSANAKTDS ISLFAGIRHD AGDIGYLKGL 901 FSYGRYKNSI SRSTGADEHA EGSVNGTLMQ LGALGGVNVP FAATGDLTVE 951 GGLRYDLLKQ DAPAEKGSAL GWSGNSLTBG TLVGLAGLKL SQPLSDKAVL 1001 FATAGVERDL NGRDYTVTGG FTGATAATGK TGARNMPHTR LVAGLGADVE 1051 FGNGWNGLAR YSYAGSKQYG NHSGRVGVGY RF*

[0060] &Dgr;G983 thus has the following basic sequence: 14                                       TSAPD FNAGGTGIGS NSRATTAKSA AVSYAGIKNE MCKDRSMLCA GRDDVAVTDR DAKINAPPPN LHTGDFPNPN DAYKNLINLK PAIEAGYTGR GVEVGIVDTG ESVGSISFPE LYGRKEHGYN ENYKNYTAYM RKEAPEDGGG KDIEASFDDE AVIETEAKPT DIRHVKEIGH IDLVSHIIGG RSVDGRPAGG IAPDATLHIM NTNDETKNEM MVAAIRNAWV KLGERGVRIV NNSFGTTSRA GTADLFQIAN SEEQYRQALL DYSGGDKTDE GIRLMQQSDY GNLSYHIRNK NMLFIFSTGN DAQAQPNTYA LLPFYEKDAQ KGIITVAGVD RSGEKFKREM YGEPGTEPLE YGSNHCGITA MWCLSAPYEA SVRFTRTNPI QIAGTSFSAP IVTGTAALLL QKYPWMSNDN LRTTLLTTAQ DIGAVGVDSK FGWGLLDAGK AMNGPASFPF GDFTADTKGT SDIAYSFRND ISGTGGLIKK GGSQLQLHGN NTYTGKTIIE GGSLVLYGNN KSDMRVETKG ALIYNGAASG GSLNSDGIVY LADTDQSGAN ETVHIKGSLQ LDGKGTLYTR IGKLLKVDGT AIIGGKLYHS ARGKGAGYLN STGRRVPFLS AAKIGQDYSF FTNIETDGGL LASLDSVEKT AGSEGDTLSY YVRRGNAART ASAAAHSAPA GLKHAVEQGG SNLENLMVEL DASESSATPE TVETAAADRT DMPGIRPYGA TFRAAAAVQH ANAADGVRIF NSLAATVYAD STAAHADMQG RRLKAVSDGL DHNGTGLRVI AQTQQDGGTW EQGGVEGKMR GSTQTVGIAA KTGENTTAAA TLGMGRSTWS ENSANAKTDS ISLFAGIRHD AGDIGYLKGL FSYGRYKNSI SRSTGADEHA EGSVNGTLMQ LGALGGVNVP FAATGDLTVE GGLRYDLLKQ AFABKGSAL GWSGNSLTEG TLVGLAGLKL SQPLSDKAVL FATAGVERDL NGRDYTVTGG FTGATAATGK TGARNMPHTR LVAGLGADVE FGNGWNGLAR YSYAGSKQYG NHSGRVGVGY RF*

[0061] &Dgr;G983 was expressed as a hybrid, with ORF46.1, 741, 961 or 961c at its C-terminus: 15 AG983-ORF46.1 ATGACTTCTGCGCCCGACTTCAATGCAGGCGGTACCGGTATCGGCAGCAACAGCAGAGCAACAACAGCGAAATCAGCA GCAGTATCTTACGCCGGTATCAAGAACGAAATGTGCAAAGACAGAAGCATGCTCTGTGCCGGTCGGGATGACGTTGCG GTTACAGACAGGGATGCCAAAATCAATGCCCCCCCCCCGAATCTGCATACCGGAGACTTTCCAAACCCAAATGACGCA TACAAGAATTTGATCAACCTCAAACCTGCAATTGAAGCAGGCTATACAGGACGCGGGGTAGAGGTAGGTATCGTCGAC ACAGGCGAATCCGTCGGCAGCATATCCTTTCCCGAACTGTATGGCAGAAAAGAACACGGCTATAACGAAAATTACAAA AACTATACGGCGTATATGCGGAAGGAAGCGCCTGAAGAQGGAGGCGGTAAAGAATTGAAGCTTCTTTCGACGATGAAG GCCGTTATAGAGACTGAAGCAAAGCCGACGGATATCCGCCACGTAAAAGAAATCGGACACATCGATTTGGTCTCCCAT ATTATTGGCGGGOGTTCCGTGGACGGCAGACCTGCAGGCGGTATTGCGCCCGATGCGACGCTACACATAATGAATACG AATGATGAAACCAAGAACGAAATGATGGTTGCAGCCATCCGCAATGCATGGGTCAAGCTGGGCGAACGTGGCGTGCGC ATCGTCAATAACAGTTTTGGAACAACATCGAGGGCAGGCACTGCCGACCTTTTCCAAATAGCCAATTCGGAGGAGCAG TACCGCCAAGCGTTGCTCGACTATTCCGGCGGTGATAAAACAGACGAGGGTATCCGCCTGATGCAACAGAGCGATTAC GGCAACCTGTCCTACCACATCCGTAATAAAAACATGCTTTTCATCTTTTCGACAGGCAATGACGCACAAGCTCAGCCC AACACATATGCCCTATTGCCATTTTATGAAAAAGACGCTCAAAAAGGCATTATCACAGTCGCAGGCGTAGACCGCAGT GGAGAAAAGTTCAAACGGGAAATGTATGGAGAACCGGGTACAGAACCGCTTGAGTATGGCTCCAACCATTGCGGAATT ACTGCCATGTGGTGCCTGTCGGCACCCTATGAAGCAAGCGTCCGTTTCACCCGTACAAACCCGATTCAAATTGCCGGA ACATCCTTTTCCGCACCCATCGTAACCGGCACGGCGGCTCTGCTGCTGCAGAAATACCCGTGGATGAGCAACGACAAC CTGCGTACCACGTTGCTGACGACGGCTCAGGACATCGGTGCAGTCGGCGTGGACAGCAAGTTCGGCTGGGGACTGCTG GATGCGGGTAAGGCCATGAACGGACCCGCGTCCTTTCCGTTCGGCGACTTTACCGCCGATACGAAAGGTACATCCGAT ATTGCCTACTCCTTCCGTAACGACATTTCAGGCACGGGCGGCCTGATCAAAAAAGGCGGCAGCCAACTGCAACTGCAC GGCAACAACACCTATACGGGCAAAACCATTATCGAkGGCGGTTCGCTGGTGTTGTACGGCAACAACAAATCGGATATG CGCGTCGAAACCAAAGGTGCGCTGATTTATAACGGGGCGGCATCCGGCGGCAGCCTGAACAGCGACGGCATTGTCTAT CTGGCAGATACCGACCAATCCGGCGCAAACGAAACCGTACACATCAAAGGCAGTCTGCAGCTGGACGGCAAAGGTACG CTGTACACACGTTTGGGCAAACTGCTGAAAGTGGACGGTACGGCGATTATCGGCGGCAAGCTGTACATGTCGGCACGC GGCAAGGGGGCAGGCTATCTCAACAGTACCGGACGACGTGTTCCCTTCCTGAGTGCCGCCAAAATCGGGCAGGATTAT TCTTTCTTCACAAACATCGAAACCGACGGCGGCCTGCTGGCTTCCCTCGACAGCGTCGAAAAAACAGCGGGCAGTGAA GGCGACACGCTGTCCTATTATGTCCGTCGCGGCAATGCGGCACGGACTGCTTCGGCAGCGGCACATTCCGCGCCCGCC GGTCTGAAACACGCCGTAGAACAGGGCGGCAGCAATCTGGAAAACCTGATGGTCGAACTGGATGCCTCCGAATCATCC GCAACACCCGAGACGGTTGAAACTGCGGCAGCCGACCGCACAGATATGCCGGGCATCCGCCCCTACGGCGCAACTTTC CGCGCAGCGGCAGCCGTACAGCATGCGAATGCCGCCGACGGTGTACGCATCTTCAACAGTCTCGCCGCTACCGTCTAT GCCGACAGTACCGCCGCCCATGCCGATATGCAGGGACGCCGCCTGAAAGCCGTATCGGACGGGTTGGACCACAACGGC ACGGGTCTGCGCGTCATCGCGCAAACCCAACAGGACGGTGGAACGTGGGAACAGGGCGGTGTTGAAGGCAAAATGCGC GGCAGTACCCAAACCGTCGGCATTGCCGCGAAAACCGGCGAAAATACGACAGCAGCCGCCACACTGGGCATGGGACGC AGCACATGGAGCGAAAACAGTGCAAATGCAAAAACCGACAGCATTAGTCTGTTTGCAGGCATACGGCACGATGCGGGC GATATCGGCTATCTCAAAGGCCTGTTCTCCTACGGACGCTACAAAAACAGCATCAGCCGCAGCACCGGTGCGGACGAA CATGCGGAAGGCAGCGTCAACGGCACGCTGATGCAGCTGGGCGCACTGGGCGGTGTCAACGTTCCGTTTGCCGCAACG GGAGATTTGACGGTCGAAGGCGGTCTGCGCTACGACCTGCTCAAACAGGATGCATTCGCCGAAAAAGGCAGTGCTTTG GGCTGGAGCGGCAACAGCCTCACTGAAGGCACGCTGGTCGGACTCGCGGGTCTGAAGCTGTCGCAACCCTTGAGCGAT AAAGCCGTCCTGTTTGCAACGGCGGGCGTGGAACGCGACCTGAACGGACGCGACTACACGGTAACGGGCGGCTTTACC GGCGCGACTGCAGCAACCGGCAAGACGGGGGCACGCAATATGCCGCACACCCGTCTGGTTGCCGGCCTGGGCGCGGAT GTCGAATTCGGCAACGGCTGGAACGGCTTGGCACGTTACAGCTACGCCGGTTCCAAACAGTACGGCAACCACAGCGGA CGAGTCGGCGTAGGCTACCGGTTCCTCGACGGTGGCGGAGGCACTGGATCCTCAGATTTGGCAAACGATTCTTTTATC CGGCAGGTTCTCGACCGTCAGCATTTCGAACCCGACGGGAAATACCACCTATTCGGCAGCAGGGGGGAACTTGCCGAG CGCAGCGGCCATATCGGATTGGGAAAAATACAAAGCCATCAGTTGGGCAACCTGATGATTCAACAGGCGGCCATTAAA GGAAATATCGGCTACATTGTCCGCTTTTCCGATCACGGGCACGAAGTCCATTCCCCCTTCGACAACCATGCCTCACAT TCCGATTCTGATGAAGCCGGTAGTCCCGTTGACGGATTTAGCCTTTACCGCATCCATTGGGACGGATACGAACACCAT CCCGCCGACGGCTATGACGGGCCACAGGGCGGCGGCTATCCCGCTCCCAAAGGCGCGAGGGATATATACAGCTACGAC ATAAAAGGCGTTGCCCAAAATATCCGCCTCAACCTGACCGACAACCGCAGCACCGGACAACGGCTTGCCGACCGTTTC CACAATGCCGGTAGTATGCTGACGCAAGGAGTAGGCGACGGATTCAAACGCGCCACCCGATACAGCCCCGAGCTGGAC AGATCGGGCAATGCCGCCGAAGCCTTCAACGGCACTGCAGATATCGTTAAAAACATCATCGGCGCGGCAGGAGAAATT GTCGGCGCAGGCGATGCCGTGCAGGGCATAAGCGAAGGCTCAAACATTGCTGTCATGCACGGCTTGGGTCTGCTTTCC ACCGAAAACAAGATGGCGCGCATCAACGATTTGGCAGATATGGCGCAACTCAAAGACTATGCCGCAGCAGCCATCCGC GATTGGGCAGTCCAAAACCCCAATGCCGCACAAGGCATAGAAGCCGTCAGCAATATCTTTATGGCAGCCATCCCCATC AAAGGGATTGGAGCTGTTCGGGGAAAATACGGCPTGGGCGGCATCACGGCACATCCTATCAAGCGGTCGCAGATGGGC GCGATCGCATTGCCGAAAGGGAAATCCGCCGTCAGCGACAATTTTGCCGATGCGGCATACGCCAAATACCCGTCCCCT TACCATTCCCGAAATATCCGTTCAAACTTGGAGCAGCGTTACGGCAAAGAAAACATCACCTCCTCAACCGTGCCGCCG TCAAACGGCAAAAATGTCAAACTGGCAGACCAACGCCACCCGAAGACAGGCGTACCGTTTGACGGTAAAGGGTTTCCG AATTTTGAGAAGCACGTGAAATATGATACGCTCGAGCACCACCACCACCACCACTGA 1 MTSAPDFNAG GTGIGSNSRA TTAKSAAVSY AGIKNEMCKD RSMLCAGRDD 51 VAVTDRDAKI NAPPPNLHTG DFPNPNDAYK NLINLKPAIE AGYTGRGVEV 101 GIVDTGESVG SISFPELYGR KEHGYNENYK NYTAYMRKEA PEDGGGKDIE 151 ASFDDEAVIE TEAKPTDIRH VKEIGHIDLV SHIIGGRSVD GRPAGGIAPD 201 ATLHIMNTND ETKNEMMVAA IRNAWVKLGE RGVRIVNNSF GTTSRAGTAD 251 LFQIANSEEQ YRQALLDYSG GDKTDEGIRL MQQSDYGNLS YHIRNKNMLF 301 IFSTGNDAQA QPNTYALLPF YEKDAQKGII TVAGVDRSGE KFKREMYGEP 351 GTEPLEYGSN HCGITAMWCL SAPYEASVRF TRTNPIQIAG TSFSAPIVTG 401 TAALLLQKYP WMSNDNLRTT LLTTAQDIGA VGVDSKFGWG LLDAGKAMNG 451 PASFPFGDFT WMTKGTSDIA YSFRNDISGT GGLIKKGGSQ LQLHGNNTYT 501 GKTIIEGGSL VLYGNNKSDM RVETKGALIY NGAASGGSLN SDGIVYLADT 551 DQSGANETVH IKGSLQLDGR GTLYTRLGKL LKVDGTAIIG GKLYMSARGK 601 GAGYLNSTGR RVPFLSAAKI GQDYSFFTNI ETDGGLLASL DSVEKTAGSE 651 GDTLSYYVRR GNAARTASAA AHSAPAGLKH AVEQGGSNLE NLMVELDASE 701 SSATPETVET AAADRTDNPG IRPYGATFRA AAAVQHANAA DGVRIFNSLA 751 ATVYADSTAA HADMQGRBLK AVSDGLDHNG TGLRVIAQTQ QDGGTWEQGG 801 VEGKMRGSTQ TVGIAAKTGE NTTAAATLGM GRSTWSENSA NAKTDSISLF 851 AGIRHDAGDI GYLKGLFSYG RYKNSISRST GADEHAEGSV NGTLMQLGAL 901 GGVNVPFAAT GDLTVEGGLR YDLLKQDAFA EKGSALGWSG NSLTEGTLVG 951 LAGLKLSQPL SDKAVLFATA GVERDLNGRD YTVTGGFTGA TAATGKTGAR 1001 NMPHTRLVAG LGADVEFGNG WNGLARYSYA GSKQYGNHSG RVGVGYRFLD 1051 GGGGTGSSDL ANDSFIRQVL DRQHFEPDGK YHLFGSRGEL AERSGHIGLG 1101 KIQSHQLGNL MIQQAAIKGN IGYIVRFSDH GHBVHSPFDN HASHSDSDEA 1151 GSPVDGFSLY RIHWDGYEHH PADGYDGPQG GGYPAPKGAR DIYSYDIKGV 1201 AQNIRLNLTD NRSTGQRLAD RFHNAGSMLT QGVGDGFKRA TRYSPELDRS 1251 GNAAEAFNGT ADIVKNIIGA AGEIVGAGDA VQGISEGSNI AVMHGLGLLS 1301 TENKMARIND LADMAQLKDY AAAAIRDWAV QNPNAAQGIE AVSNIFMAAI 1351 PIKGIGAVRG KYGLGGITAH PIKRSQMGAI ALPKGKSAVS DNFADAAYAK 1401 YPSPYHSRNI RSNLEQRYGK ENITSSTVPP SNGKNVKLAD QRHPKTGVPF 1451 DGKGFPNFBK HVKYDTLEHH HHHH* &Dgr;G983-741 ATGACTTCTGCGCCCGACTTCAATGCAGGCGGTACCGGTATCGGCAGCAACAGCAGAGCAACAACAGCGAAATCAGCA GCAGTATCTTACGCCGGTATCAAGAACGAAATGTGCAAAGACAGAAGCATGCTCTGTGCCGGTCGGGATGACGTTGCG GTTACAGACAGGGATGCCAAAATCAATGCCCCCCCCCCGAATCTGCATACCGGAGACTTTCCAAACCCAAATGACGCA TACAAGAATTTGATCAACCTCAAACCTGCAATTGAAGCAGGCTATACAGGACGCGGGGTAGAGGTAGGTATCGTCGAC ACAGGCGAATCCGTCGGCAGCATATCCTTTCCCGAACTGTATGGCAGAAAAGAACACGGCTATAACGAAAATTACAAA AACTATACGGCGTATATGCGGAAGGAAGCGCCTGAAGACGGAGGCGGTAAAGACATTGAAGCTTCTTTCGACGATGAG GCCGTTATAGAGACTGAAGCAAAGCCGACGGATATCCGCCACGTAAAAGAAATCGGACACATCGATTTGGTCTCCCAT ATTATTGGCGGGCGTTCCGTGGACGGCAGACCTGCAGGCGGTATTGCGCCCGATGCGACGCTACACATAATGAATACG AATGATGAAACCAAGAACGAAATGATGGTTGCAGCCATCCGCAATGCATGGGTCAAGCTGGGCGAACGTGGCGTGCGC ATCGTCAATAACAGTTTTGGAACAACATCGAGGGCAGGCACTGCCGACCTTTTCCAAATAGCCAATTCGGAGGAGCAG TACCGCCAAGCGTTGCTCGACTATTCCGGCGGTGATAAAACAGACGAGGGTATCCGCCTGATGCAACAGAGCGATTAC GGCAACCTGTCCTACCACATCCGTAATAAAAACATGCTTTTCATCTTTTCGACAGGCAATGACGCACAAGCTCAGCCC AACACATATGCCCTATTGCCATTTTATGAAAAAGACGCTCAAAAAGGCATTATCACAGTCGCAGGCGTAGACCGCAGT GGAGAAAAGTTCAAACGGGAAATGTATGGAGAACCGGGTACAGAACCGCTTGAGTATGGCTCCAACCATTGCGGAATT ACTGCCATGTGGTGCCTGTCGGCACCCTATGAAGCAAGCGTCCGTTTCACCCGTACAAACCCGATTCAAATTGCCGGA ACATCCTTTTCCGCACCCATCGTAACCGGCACGGCGGCTCTGCTGCTGCAGAAATACCCGTGGATGAGCAACGACAAC CTGCGTACCACGTTGCTGACGACGGCTCAGGACATCGGTGCAGTCGGCGTGGACAGCAAGTTCGGCTGGGGACTGCTG GATGCGGGTAAGGCCATGAACGGACCCGCGTCCTTTCCGTTCGGCGACTTTACCGCCGATACGAAAGGTACATCCGAT ATTGCCTACTCCTTCCGTAACGACATTTCAGGCACGGGCGGCCTGATCAAAAAAGGCGGCAGCCAACTGCAACTGCAC GGCAACAACACCTATACGGGCAAAACCATTATCGAAGGCGGTTCGCTGGTGTTGTACGGCAACAACAAATCGGATATG CGCGTCGAAACCAAAGGTGCGCTGATTTATAACGGGGCGGCATCCGGCGGCAGCCTGAACAGCGACGGCATTGTCTAT CTGGCAGATACCGACCAATCCGGCGCAAACGAAACCGTACACATCAAAGGCAGTCTGCAGCTGGACGGCAAAGGTACG CTGTACACACGTTTGGGCAAACTGCTGAAAGTGGACGGTACGGCGATTATCGGCGGCAAGCTGTACATGTCGGCACGC GGCAAGGGGGCAGGCTATCTCAACAGTACCGGACGACGTGTTCCCTTCCTGAGTGCCGCCAAAATCGGGCAGGATTAT TCTTTCTTCACAAACATCGAAACCGACGGCGGCCTGCTGGCTTCCCTCGACAGCGTCGAAAAAACAGCGGGCAGTGAA GGCGACACGCTGTCCTATTATGTCCGTCGCGGCAATGCGGCACGGACTGCTTCGGCAGCGGCACATTCCGCGCCCGCC GGTCTGAAACACGCCGTAGAACAGGGCGGCAGCAATCTGGAAAACCTGATGGTCGAACTGGATGCCTCCGAATCATCC GCAACACCCGAGACGGTTGAAACTGCGGCAGCCGACCGCACAGATATGCCGGGCATCCGCCCCTACGGCGCAACTTTC CGCGCAGCGGCAGCCGTACAGCATGCGAATGCCGCCGACGGTGTACGCATCTTCAACAGTCTCGCCGCTACCGTCTAT GCCGACAGTACCGCCGCCCATGCCGATATGCAGGGACGCCGCCTGAAAGCCGTATCGGACGGGTTGGACCACAACGGC ACGGGTCTGCGCGTCATCGCGCAAACCCAACAGGACGGTGGAACGTGGGAACAGGGCGGTGTTGAAGGCAAAATGCGC GGCAGTACCCAAACCGTCGGCATTGCCGCGAAAACCGGCGAAAATACGACAGCAGCCGCCACACTGGGCATGGGACGC AGCACATGGAGCGAAAACAGTGCAAATGCAAAAACCGACAGCATTAGTCTGTTTGCAGGCATACGGCACGATGCGGGC GATATCGGCTATCTCAAAGGCCTGTTCTCCTACGGACGCTACAAAAACAGCATCAGCCGCAGCACCGGTGCGGACGAA CATGCGGAAGGCAGCGTCAACGGCACGCTGATGCAGCTGGGCGCACTGGGCGGTGTCAACGTTCCGTTTGCCGCAACG GGAGATTTGACGGTCGAAGGCGGTCTGCGCTACGACCTGCTCAAACAGGATGCATTCGCCGAAAAAGGCAGTGCTTTG GGCTGGAGCGGCAACAGCCTCACTGAAGGCACGCTGGTCGGACTCGCGGGTCTGAAGCTGTCGCAACCCTTGAGCGAT AAAGCCGTCCTGTTTGCAACGGCGGGCGTGGAACGCGACCTGAACGGACGCGACTACACGGTAACGGGCGGCTTTACC GGCGCGACTGCAGCAACCGGCAAGACGGGGGCACGCAATATGCCGCACACCCGTCTGGTTGCCGGCCTGGGCGCGGAT GTCGAATTCGGCAACGGCTGGAACGGCTTGGCACGTTACAGCTACGCCGGTTCCAAACAGTACGGCAACCACAGCGGA CGAGTCGGCGTAGGCTACCGGTTCCTCGAGGGATCCGGAGGGGGTGGTGTCGCCGCCGACATCGGTGCGGGGCTTGCC GATGCACTAACCGCACCGCTCGACCATAAAGACAAAGGTTTGCAGTCTTTGACGCTGGATCAGTCCGTCAGGAAAAAC GAGAAACTGAAGCTGGCGGCACAAGGTGCGGAAAAAACTTATGGAAACGGTGACAGCCTCAATACGGGCAAATTGAAG AACGACAAGGTCAGCCGTTTCGACTTTATCCGCCAAATCGAAGTGGACGGGCAGCTCATTACCTTGGAGAGTGGAGAG TTCCAAGTATACAAACAAAGCCATTCCGCCTTAACCGCCTTTCAGACCGAGCAAATACAAGATTCGGAGCATTCCGGG AAGATGGTTGCGAAACGCCAGTTCAGAATCGGCGACATAGCGGGCGAACATACATCTTTTGACAAGCTTCCCGAAGGC GGCAGGGCGACATATCGCGGGACGGCGTTCGGTTCAGACGATGCCGGCGGAAAACTGACCTACACCATAGATTTCGCC GCCAAGCAGGGAAACGGCAAAATCGAACATTTGAAATCGCCAGAACTCAATGTCGACCTGGCCGCCGCCGATATCAAG CCGGATGGAAAACGCCATGCCGTCATCAGCGGTTCCGTCCTTTACAACCAAGCCGAGAAAGGCAGTTACTCCCTCGGT ATCTTTGGCGGAAAAGCCCAGGAAGTTGCCGGCAGCGCGGAAGTGAAAACCGTAAACGGCATACGCCATATCGGCCTT GCCGCCAAGCAACTCGAGCACCACCACCACCACCACTGA 1 MTSAPDFNAG GTGIGSNSRA TTAKSAAVSY AGIKNEMCKD RSMLCAGRDD 51 VAVTDRDAKI NAPPPNLHTG DFPNPNDAYK NLINLKPAIE AGYTGRGVEV 101 GIVDTGESVG SISFPELYGR KEHGYNENYK NYTAYMRKEA PEDGGGKDIE 151 ASFDDEAVIE TEAKPTDIRH VKBIGHIDLV SHIIGGRSVD GRPAGGIAPD 201 ATLHIMNTND ETKNEMMVAA IRNAWVKLGE RGVRIVNNSF GTTSRAGTAD 251 LFQIANSEEQ YRQALLDYSG GDKTDEGIRL MQQSDYGNLS YHIRNKNMLF 301 IFSTGNDAQA QPNTYALLPF YEKDAQKGII TVAGVDRSGE KFKRBMYGEP 351 GTEPLEYGSN HCGITAMWCL SAPYEASVRF TRTNPIQIAG TSFSAPIVTG 401 TAALLLQKYP WMSNDNLRTT LLTTAQDIGA VGVDSKFGWG LLDAGKAMNG 451 PASFPFGDFT ADTKGTSDIA YSFRNDISGT GGLIKKGGSQ LQLHGNNTYT 501 GKTIIEGGSL VLYGNNKSDM RVETKGALIY NGAASGGSLN SDGIVYLADT 551 DQSGANETVH IKGSLQLDGK GTLYTRLGKL LKVDGTAIIG GKLYMSARGK 601 GAGYLNSTGR RVPFLSAAKI GQDYSFFTNI ETDGGLLASL DSVEKTAGSE 651 GDTLSYYVRR GNAARTASAA AHSAPAGLKH AVBQGGSNLE NLMVELDASE 701 SSATPETVET AAADRTDHPG IRPYGATFRA AAAVQHANAA DGVRIFNSLA 751 ATVYADSTAA HADMQGRBLK AVSDGLDHNG TGLRVIAQTQ QDGGTWEQGG 801 VEGKMRGSTQ TVGIAAKTGE NTTAAATLGM GRSTWSENSA NAKTDSISLF 851 AGIPHDAGDI GYLKGLFSYG RYKNSISRST GADEHAEGSV NGTLNQLGAL 901 GGVNVPFAAT GDLTVEGGLR YDLLKQDAFA EKGSALGWSG NSLTEGTLVG 951 LAGLKLSQPL SDKAVLFATA GVERDLNGRD YTVTGGFTGA TAATGKTGAR 1001 NMPHTRLVAG LGADVEFGNG WNGLARYSYA GSKQYGNHSG RVGVGYRFLE 1051 GSGGGGVAAD IGAGLADALT APLDHKDKGL QSLTLDQSVR KNEKLKLAAQ 1101 GAEKTYGNGD SLNTGKLKND KVSRFDFIRQ IEVDGQLITL ESGEFQVYKQ 1151 SHSALTAPQT EQIQDSEHSG KMVAKRQFRI GDIAGEHTSF DKLPEGGRAT 1201 YRGTAFGSDD AGGKLTYTID FAAKQGNGKI EHLKSPELNV DLAAADIKPD 1251 GK~HAVISGS VLYNQAEKGS YSLGIFGGKA QEVAGSAEVK TVNGIRHIGL 1301 AAKQLEHHHH HH* &Dgr;G983 -961 ATGACTTCTGCGCCCGACTTCAATGCAGGCGGTACCGGTATCGGCAGCAACAGCAGAGCAACAACAGCGAAATCAGCA GCAGTATCTTACGCCGGTATCAAGAACGAAATGTGCAAAGACAGAAGCATGCTCTGTGCCGGTCGGGATGACGTTGCG GTTACAGACAGGGATGCCAAAATCAATGCCCCCCCCCCGAATCTGCATACCGGAGACTTTCCAAACCCAAATGACGCA TACAAGAATTTGATCAACCTCAAACCTGCAATTGAAGCAGGCTATACAGGACGCGGGGTAGAGGTAGGTATCGTCGAC ACAGGCGAATCCGTCGGCAGCATATCCTTTCCCGAACTGTATGGCAGAAAAGAACACGGCTATAACGAAAATTACAAA AACTATACGGCGTATATGCGGAAGGAAGCGCCTGAAGACGGAGGCGGTAAAGACATTGAAGCTTCTTTCGACGATGAG GCCGTTATAGAGACTGAAGCAAAGCCGACGGATATCCGCCACGTAAAAGAAATCGGACACATCGATTTGGTCTCCCAT ATTATTGGCGGGCGTTCCGTGGACGGCAGACCTGCAGGCGGTATTGCGCCCGATGCGACGCTACACATAATGAATACG AATGATGAAACCAAGAACGAAATGATGGTTGCAGCCATCCGCAATGCATGGGTCAAGCTGGGCGAACGTGGCGTGCGC ATCGTCAATAACAGTTTTGGAACAACATCGAGGGCAGGCACTGCCGACCTTTTCCAAATAGCCAATTCGGAGGAGCAG TACCGCCAAGCGTTGCTCGACTATTCCGGCGGTGATAAAACAGACGAGGGTATCCGCCTGATGCAACAGAGCGATTAC GGCAACCTGTCCTACCACATCCGTAATAAAAACATGCTTTTCATCTTTTCGACAGGCAATGACGCACAAGCTCAGCCC AACACATATGCCCTATTGCCATTTTATGAAAAAGACGCTCAAAAAGGCATTATCACAGTCGCAGGCGTAGACCGCAGT GGAGAAAAGTTCAAACGGGAAATGTATGGAGAACCGGGTACAGAACCGCTTGAGTATGGCTCCAACCATTGCGGAATT ACTGCCATGTGGTGCCTGTCGGCACCCTATGAAGCAAGCGTCCGTTTCACCCGTACAAACCCGATTCAAATTGCCGGA ACATCCTTTTCCGCACCCATCGTAACCGGCACGGCGGCTCTGCTGCTGCAGAAATACCCGTGGATGAGCAACGACAAC CTGCGTACCACGTTGCTGACGACGGCTCAGGACATCGGTGCAGTCGGCGTGGACAGCAAGTTCGGCTGGGGACTGCTG GATGCGGGTAAGGCCATGAACGGACCCGCGTCCTTTCCGTTCGGCGACTTTACCGCCGATACGAAAGGTACATCCGAT ATTGCCTACTCCTTCCGTAACGACATTTCAGGCACGGGCGGCCTGATCAAAAAAGGCGGCAGCCAACTGCAACTGCAC GGCAACAACACCTATACGGGCAAAACCATTATCGAAGGCGGTTCGCTGGTGTTGTACGGCAACAACAAATCGGATATG CGCGTCGAAACCAAAGGTGCGCTGATTTATAACGGGGCGGCATCCGGCGGCAGCCTGAACAGCGACGGCATTGTCTAT CTGGCAGATACCGACCAATCCGGCGCAAACGAAACCGTACACATCAAAGGCAGTCTGCAGCTGGACGGCAAAGGTACG CTGTACACACGTTTGGGCAAACTGCTGAAAGTGGACGGTACGGCGATTATCGGCGGCAAGCTGTACATGTCGGCACGC GGCAAGGGGGCAGGCTATCTCAACAGTACCGGACGACGTGTTCCCTTCCTGAGTGCCGCCAAAATCGGGCAGGATTAT TCTTTCTTCACAAACATCGAAACCGACGGCGGCCTGCTGGCTTCCCTCGACAGCGTCGAAAAAACAGCGGGCAGTGAA GGCGACACGCTGTCCTATTATGTCCGTCGCGGCAATGCGGCACGGACTGCTTCGGCAGCGGCACATTCCGCGCCCGCC GGTCTGAAACACGCCGTAGAACAGGGCGGCAGCAATCTGGAAAACCTGATGGTCGAACTGGATGCCTCCGAATCATCC CATGCGGAAGGCAGCGTCAACGGCACGCTGATGCAGCTGGGCGCACTGGGCGGTGTCAACGTTCCGTTTGCCGCAACG GGAGATTTGACGGTCGAAGGCGGTCTGCGCTACGACCTGCTCAAACAGGATGCATTCGCCGAAAAAGGCAGTGCTTTG GGCTGGAGCGGCAACAGCCTCACTGAAGGCACGCTGGTCGGACTCGCGGGTCTGAAGCTGTCGCAACCCTTGAGCGAT AAAGCCGTCCTGTTTGCAACGGGGGGCGTGGAACGCGACCTGAACGGACGCGACTACACGGTAACGGGCGGCTTTACC GGCGCGACTGCAGCAACCGGCAAGACGGGGGCACGCAATATGCCGCACACCCGTCTGGTTGCCGGCCTGGGCGCGGAT GTCGAATTCGGCAACGGCTGGAACGGCTTGGCACGTTACAGCTACGCCGGTTCCAAACAGTACGGCAACCACAGCGGA CGAGTCGGCGTAGGCTACCGGTTCCTCGAGGGTGGCGGAGGCACTGGATCCGCCACAAACGACGACGATGTTAAAAAA GCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATCAACGGTTTCAAAGCTGGAGAGACCATCTAC GACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCCGATGTTGAAGCCGACGACTTTAAAGGTCTG GGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAACAAACAAAACGTCGATGCCAAAGTAAAAGCT GCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGATGCCGCTTTAGCAGATACTGATGCCGCTCTG GATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACATTTGCTGAAGAGACTAAGACAAATATCGTA AAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCATGCCGAAGCATTCAACGATATCGCCGATTCA TTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAATGAAGCCAAACAGACGGCCGAAGAAACCAAA CAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAAGCCGAAGCTGCCGCTGGCACAGCTAATACT GCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAAGCTGATATCGCTACGAACAAAGATAATATT GCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGACAGCAAATTTGTCAGAATTGATGGTCTGAAC GCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCCATTGCCGATCACGATACTCGCCTGAACGGT TTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTTGCAGAACAAGCCGCGCTCTCCGGTCTGTTC CAACCTTACAACGTGGGTCGGTTCAATGTAACGGCTGCAGTCGGCGGCTACAAATCCGAATCGGCAGTCGCCATCGGT ACCGGCTTCCGCTTTACCGAAAACTTTGCCGCCAAAGCAGGCGTGGCAGTCGGCACTTCGTCCGGTTCTTCCGCAGCC TACCATGTCGGCGTCAATTACGAGTGGCTCGAGCACCACCACCACCACCACTGA 1 MTSAPDFNAG GTGIGSNSRA TTAKSAAVSY AGIKNEMCKD RSMLCAGRDD 51 VAVTDRDAKI NAPPPNLHTG DFPNPNDAYK NLINLKPAIE AGYTGRGVBV 101 GIVDTGESVG SISFPELYGR KEHGYNENYK NYTAYMBKEA PEDGGGKDIE 151 ASFDDEAVIE TEAKPTDIRH VKEIGHIDDV SHIIGGRSVD GRPAGGIAPD 201 ATLHIMNTND ETKNBNMVAA IRNAWVKLGE RGVRIVNNSF GTTSRSGTAD 251 LFQIANSEEQ YRQALLDYSG GDKTDEGIHL MQQSDYGNLS YHIRNKIMLP 301 IFSTGNDAQA QPNTYALLPF YEKDAQKGII TVAGVDRSGE KFKREMYGEP 351 GTEPLEYGSN HCGITAMWCL SAPYEASVRF TRTNPIQIAG TSFSAPIVTG 401 TAALLLQKYP WHSNDNLRTT LLTTAQDIGA VGVDSKFGWG LLDAGKAMNG 451 PASFPFGDFT ADTKGTSDIA YSFRNDISGT GGLIKKGGSQ LQLHGNNTYT 501 GKTIIEGGSL VLYGNNKSDH RVETKGALIY NGAASGGSLN SDGIVYLADT 551 DQSGANETVH IKGSLQLDGK GTLYTRLGKL LKVDGTAIIG GKLYMSARGK 601 GAGYLNSTGR RVPFLSAAKI GQDYSFFTNI ETDGGLLASL DSVEKTAGSE 651 GDTLSYYVRR GNAARTASAA AHSAPAGLKE AVBQGGSNLE NLMVELDASE 701 SSATPETVET AAADRTDHPG IRPYGATFRA AAAVQHANAA DGVRIFNSLA 751 ATVYADSTAA HADMQGRRLK AVSDGLDHNG TGLRVIAQTQ QDGGTWBQGG 801 VEGKMRGSTQ TVGIAAKTGE NTTAAATLGM GRSTWSENSA NAKTDSISLF 851 AGIRHDAGDI GYLKGLFSYG RYKNSISRST GADEHAEGSV NGTLMQLGAL 901 GGVNVPFAAT GDLTVEGGLR YDLLKQDAFA EKGSALGWSG NSLTEGTLVG 951 LAGLKLSQPL SDKAVLFATA GVERDLNGRD YTVTGGFTGA TAkTGKTGAR 1001 NMPHTRLVAG LGADVEFGNG WNGLARYSYA GSKQYGNHSG RVGVGYRFLE 1051 GGGGTGSATN DDDVKKAATV AIAAAYNNGQ EINGFKAGET IYDIDEDGTI 1101 TKKDATAADV EADDFKGLGL KKVVTNLTKT VNENKQNVDA KVKAAESEIE 1151 KLTTKLADTD AALADTDAAL DATTNALNKL GENITTFABE TKTNIVKIDE 1201 KLEAVADTVD KHAEAFNDIA DSLDETNTKA DEAVKTANEA KQTAEETKQN 1251 VDAKVKAAET AAGKAEAAAG TANTAADKAE AVAMCVTDIK ADIATNKDNI 1301 AKKANSADVY TREBSDSKFV RIDGLNATTE KLDTRLASAE KSIADHDTEL 1351 NGLDKTVSDL RKETRQGLAE QAALSGLPQP YNVGRFNVTA AVGGYKSESA 1401 VAIGTGFRPT ENFAAKAGVA VGTSSGSSAA YHVGVNYEWL EHHHHHH* AG983 -961c ATGACTTCTGCGCCCGACTTCAATGCAGGCGGTACCGGTATCGGCAGCAACAGCAGAGCAACAACAGCGAAATCAGCA GCAGTATCTTACGCCGGTATCAAGAACGAAATGTGCAAAGACAGAAGCATGCTCTGTGCCGGTCGGGATGACGTTGCG GTTACAGACAGGGATGCCAAAATCAATGCCCCCCCCCCGAATCTGCATACCGGAGACTTTCCAAACCCAAkTGACGCA TACIAGAATTTGATCAACCTCAAACCTGCAATTGAAGCAGGCTATACAGGACGCGGGGTAGAGGTAGGTATCGTCGAC ACAGGCGAATCCGTCGGCAGCATATCCTTTCCCGAACTGTATGGCAGAAAAGAACACGGCTATAACGAAAATTACAAA AACTATACGGCGTATATGCGGAAGGAAGCGCCTGAAGACGGAGGCGGTAAAGACATTGAAGCTTCTTTCGACGATGAG GCCGTTATAGAGACTGAAGCAAAGCCGACGGATATCCGCCACGTAAAAGAAATCGGACACATCGATTTGGTCTCCCAT ATTATTGGCGGGCGTTCCGTGGACGGCAGACCTGCAGGCGGTATTGCGCCCGATGCGACGCTACACATAATGAATACG AATGATGAAACCAAGAACGAAATGATGGTTGCAGCCATCCGCAATGCATGGGTCAAGCTGGGCGAACGTGGCGTGCGC ATCGTCAATAACAGTTTTGGAACAACATCGAGGGCAGGCACTGCCGACCTTTTCCAAATAGCCAATTCGGAGGAGCAG TACCGCCAAGCGTTGCTCGACTATTCCGGCGGTGATAAAACAGACGAGGGTATCCGCCTGATGCAACAGAGCGATTAC GGCAACCTGTCCTACCACATCCGTAATAAAAACATGCTTTTCATCTTTTCGACAGGCAATGACGCACAAGCTCAGCCC AACACATATGCCCTATTGCCATTTTATGAAAAAGACGGTCAAAAAGGCATTATCACAGTCGCAGGCGTAGACCGCAGT GGAGAAAAGTTCAAACGGGAAATGTATGGAGAACCGGGTACAGAACCGCTTGAGTATGGCTCCAACCATTGCGGAATT ACTGCCATGTGGTGCCTGTCGGCACCCTATGAAGCAAGCGTCCGTTTCACCCGTACAAACCCGATTCAAATTGCCGGA ACATCCTTTTCCGCACCCATCGTAACCGGCACGGCGGCTCTGCTGCTGCAGAAATACCCGTGGATGAGCAACGACAAC CTGCGTACCACGTTGCTGACGACGGCTCAGGACATCGGTGCAGTCGGCGTGGACAGCAAGTTCGGCTGGGGACTGCTG GATGCGGGTAAGGCCATGAACGGACCCGCGTCCTTTCCGTTCGGCGACTTTACCGCCGATACGAAAGGTACATCCGAT ATTGCCTACTCCTTCCGTAACGACATTTCAGGCACGGGCGGCCTGATCAAAAAAGGCGGCAGCCAACTGCAACTGCAC GGCAACAACACCTATACGGGCAAAACCATTATCGAAGGCGGTTCGCTGGTGTTGTACGGCAACAACAAATCGGATATG CGCGTCGAAACCAAAGGTGCGCTGATTTATAACGGGGCGGCATCCGGCGGCAGCCTGAACAGCGACGGCATTGTCTAT CTGGCAGATACCGACCAATCCGGCGCAAACGAAACCGTACACATCAAAGGCAGTCTGCAGCTGGACGGCAAAGGTACG CTGTACACACGTTTGGGCAAACTGCTGAAAGTGGACGGTACGGCGATTATCGGCGGCAAGCTGTACATGTCGGCACGC GGCAAGGGGGCAGGCTATCTCAACAGTACCGGACGACGTGTTCCCTTCCTGAGTGCCGCCAAAATCGGGCAGGATTAT TCTTTCTTCACAAACATCGAAACCGACGGCGGCCTGCTGGCTTCCCTCGACAGCGTCGAAAAAACAGCGGGCAGTGAA GGCGACACGCTGTCCTATTATGTCCGTCGCGGCAATGCGGCACGGACTGCTTCGGCAGCGGCACATTCCGCGCCCGCC GGTCTGAAACACGCCGTAGAACAGGGCGGCAGCAATCTGGAAAACCTGATGGTCGAACTGGATGCCTCCGAATCATCC GCAACACCCGAGACGGTTGAAACTGCGGCAGCCGACCGCACAGATATGCCGGGCATCCGCCCCTACGGCGCAACTTTC CGCGCAGCGGCAGCCGTACAGCATGCGAATGCCGCCGACGGTGTACGCATCTTCAACAGTCTCGCCGCTACCGTCTAT GCCGACAGTACCGCCGCCCATGCCGATATGCAGGGACGCCGCCTGAAAGCCGTATCGGACGGGTTGGACCACAACGGC ACGGGTCTGCGCGTCATCGCGCAAACCCAACAGGACGGTGGAACGTGGGAACAGGGCGGTGTTGAAGGCAAAATGCGC GGCAGTACCCAAACCGTCGGCATTGCCGCGAAAACCGGCGAAAATACGACAGCAGCCGCCACACTGGGCATGGGACGC AGCACATGGAGCGAAAACAGTGCAAATGCAAAAACCGACAGCATTAGTCTGTTTGCAGGCATACGGCACGATGCGGGC GATATCGGCTATCTCAAAGGCCTGTTCTCCTACGGACGCTACAAAAACAGCATCAGCCGCAGCACCGGTGCGGACGAA CATGCGGAAGGCAGCGTCAACGGCACGCTGATGCAGCTGGGCGCACTGGGCGGTGTCAACGTTCCGTTTGCCGCAACG GGAGATTTGACGGTCGAAGGCGGTCTGCGCTACGACCTGCTCAAACAGGATGCATTCGCCGAAAAAGGCAGTGCTTTG GGCTGGAGCGGCAACAGCCTCACTGAAGGCACGCTGGTCGGACTCGCGGGTCTGAAGCTGTCGCAACCCTTGAGCGAT AAAGCCGTCCTGTTTGCAACGGCGGGCGTGGAACGCGACCTQAACGGACGCGACTACACGGTAACGGGCGGCTTTACC GGCGCGACTGCAGCAACCGGCAAGACGGGGGCACGCAATATGCCGCACACCCGTCTGGTTGCCGGCCTGGGCGCGGAT GTCGAATTCGGCAACGGCTGGAACGGCTTGGCACGTTACAGCTACGCCGGTTCCAAACAGTACGGCAACCACAGCGGA CGAGTCGGCGTAGGCTACCGGTTCCTCGAGGGTGGCGGAGGCACTGGATCCGCCACAAACGACGkCGATGTTAAAAAA GCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATCAACGGTTTCAAAGCTGGAGAGACCATCTAC GACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCCGATGTTGAAGCCGACGACTTTAAAGGTCTG GGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAACAAACAAAACGTCGATGCCAAAGTAAAAGCT GCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGATGCCGCTTTAGCAGATACTGATGCCGCTCTG GATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACATTTGCTGAAGAGACTAAGACAAATATCGTA AAAkTTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCATGCCGAAGCATTCAACGATATCGCCGATTCA TTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAATGAAGCCAAACAGACGGCCGAAGAAACCAAA CAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAAGCCGAAGCTGCCGCTGGCACAGCTAATACT GCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAAGCTGATATCGCTACGAACAAAGATAATATT GCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGACAGCAAATTTGTCAGAATTGATGGTCTGAAC GCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCCATTGCCGATCACGATACTCGCCTGAACGGT TTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTTGCAGAACAAGCCGCGCTCTCCGGTCTGTTC CAACCTTACAACGTGGGTCTCGAGCACCACCACCACCACCACTGA 1 MTSAPDFNAG GTGIGSNSRA TTAKSAAVSY AGIKNEMCKD RSMLCAGRDD 51 VAVTDRDAKI NAPPPNLHTG DFPNPNDAYK NLINLKPAIB AGYTGRGVEV 101 GIVDTGESVG SISFPELYGR KEEGYNENYK NYTAYMRKEA PEDGGGKDIE 151 ASFDDEAVIE TEAKPTDIBH VKEIGHIDLV SHIIGGRSVD GRPAGGIAPD 201 ATLHINNTND ETKNEMMVAA IRNAWVKLGE RGVRIVNNSF GTTSRAGTAD 251 LFQIANSEEQ YRQALLDYSG GDKTDEGIRL MQQSDYGNLS YHIRNKNMLF 301 IFSTGNDAQA QPNTYALLPF YEKDAQKGII TVAGVDRSGE KFKREMYGEP 351 GTEPLEYGSN HCGITAMWCL SAPYEASVRF TRTNPIQIAG TSFSAPIVTG 401 TAALLLQKYP WMSNDNLRTT LLTTAQDIGA VGVDSKFGWG LLDAGKAMNG 451 PASFPFGDFT ADTKGTSDIA YSFRNDISGT GGLIKKGGSQ LQLHGNNTYT 501 GKTIIEGGSL VLYGNNKSDM RVETKGALIY NGAASGGSLN SDGIVYLADT 551 DQSGANETVH IKGSLQLDGK GTLYTRLGKL LKVDGTAIIG GKLYHSARGK 601 GAGYLNSTGR RVPFLSAAKI GQDYSFFTNI ETPGGLLASL DSVEKTAGSE 651 GDTLSYYVRR GNAARTASAA AHSAPAGLKH AVEQGGSNLE NLMVELDASE 701 SSATPETVET AAADRTDMPG IRPYGATFRA AAAVQHANAP DGVRIFNSLA 751 ATVYADSTAA HADMQGRRLK AVSDGLDHNG TGLRVIAQTQ QDGGTWEQGG 801 VEGKMRGSTQ TVGIAAKTGE NTTAAATLGN GRSTWSENSA NAKTDSISLF 851 AGIEHDAGDI GYLKGLFSYG RYKNSISRST GADEHAEGSV NGTLNQLGAL 901 GGVNVPFAkT GDLTVEGGLR YDLLKQDAFA EKGSALGWSG NSLTEGTLVG 951 LAGLKLSQPL SDKAVLPATA GVERDLNGRD YTVTGGFTGA TAATGKTGAP 1001 NMPHTRLVAG LGADVEFGNG WNGLARYSYA GSKQYGNHSG RVGVGYRFLE 1051 GGGGTGSATN DDDVKKAATV AIAAAYNNGQ EINGFKAGET IYDIDEDGTI 1101 TKKDATAADV EADDFKGLGL KKVVTNLTKT VNENKQNVDA KVKAAESEIE 1151 KLTTKLADTD AALADTDAAL DATTNALNKL GENITTFAEE TKTNIVKIDE 1201 KLEAVADTVD KHAEAFNDIA DSLDETNTKA DEAVKTANEA KQTAEETKQN 1251 VDAKVKAAET AAGKABAAAG TANTAADKAE AVAAKVTDIK ADIATNKDNI 1301 AKKANSADVY TREESDSKFV RIDGLNATTE KLDTRLASAE KSIADHDTRL 1351 NGLDKTVSDL RKETRQGLAE QAALSGLFQP YNVGLEHHHH HH*

EXAMPLE 4

Hybrids of &Dgr;G741

[0062] Protein 741 has the following sequence: 16 1 VNRTAFCCLS LTTALILTAC SSGGGGVAAD IGAGLADALT APLDHKDKGL 51 QSLTLDQSVR KNEKLKLAAQ GAEKTYGNGD SLNTGKLKND KVSRPDFIRQ 101 IEVDGQLITL ESGBFQVYKQ SHSALTAFQT EQIQDSBHSG KMVAKRQFRI 151 GDIAGEHTSF DKLPEGGRAT YRGTAFGSDD AGGKLTYTID FAAKQGNGKI 201 EHLKSPELNV DLAAADIKPD GKRHAVISGS VLYNQAEKGS YSLGIFGGKA 251 QEVAGSAEVK TVNGIRHIGL AAKQ*

[0063] &Dgr;G741 thus has the following basic sequence: 17                           VAAD IGAGLADALT APLDHKDKGL QSLTLDQSVR KNEKLKLAAQ GAEKTYGNGD SLNTGKLKND KVSRPDFIRQ IEVDGQLITL ESGEFQVYKQ SHSMJTAPQT EQIQDSEHSG KMVAKRQFRI GDIAGEHTSF DKLPEGGRAT YRGTAFGSDD AGGKLTYTID FAAKQGNGKI EHLKSPBLNV DLAAADIKPD GKRHAVISGS VLYNQAEKGS YSLGIFGGKA QEVAGSAEVK TVNGIRHIGL AAKQ*

[0064] &Dgr;G741 was fused directly in-frame upstream of proteins 961, 961c, 983 and ORF46.1: 18 &Dgr;G741-961 ATGGTCGCCGCCGACATCGGTGCGGGGCTTGCCGATGCACTAACCGCACCGCTCGACCATAAAGACAAAGGTTTGCAG TCTTTGACGCTGGATCAGTCCGTCAGGAAAAACGAGAAACTGAAGCTGGCGGCACAAGGTGCGGAAAAAACTTATGGA AACGGTGACAGCCTCAATACGGGCAAATTGAAGAACGACAAGGTCAGGCGTTTCGACTTTATCCGCCAAATCGAAGTG GACGGGCAGCTCATTACCTTGGAGAGTGGAGAGTTCCAAGTATACAAACAAAGCCATTCCGCCTTAACCGCCTTTCAG ACCGAGCAAATACAAGATTCGGAGCATTCCGGGAAGATGGTTGCGAAACGCCAGTTCAGAATCGGCGACATAGCGGGC GAACATACATCTTTTGACAAGCTTCCCGAAGGCGGCAGGGCGACATATCGCGGGACGGCGTTCGGTTCAGACGATGCC GGCGGAAAACTGACCTACACCATAGATTTCGCCGCCAAGCAGGGAAACGGCAAAATCGAACATTTGAAATCGCCAGAA CTCAATGTCGACCTGGCCGCCGCCGATATCAAGCCGGATGGAAAACGCCATGCCGTCATCAGCGGTTCCGTCCTTTAC AACCAAGCCGAGAAAGGCAGTTACTCCCTCGGTATCTTTGGCGGAAAAGCCCAGGAAGTTGCCGGCAGCGCGGAAGTG AAAACCGTAAACGGCATACGCCATATCGGCCTTGCCGCCAAGCAACTCGAGGGTGGCGGAGGCACTGGATCCGCCACA AACGACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATCAACGGTTTC AAAGCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCCGATGTTGAA GCCGACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAACAAACAAAAC GTCGATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGATGCCGCTTTA GCAGATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACATTTGCTGAA GAGACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCATGCCGAAGCA TTCAACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAATGAAGCCAAA CAGACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAAGCCGAAGCT GCCGCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAAGCTGATATC GCTACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGACAGCAAATTT GTCAGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCCATTGCCGAT CACGATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTTGCAGAACAA GCCGCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTCGGTTCAATGTAACGGCTGCAGTCGGCGGCTACAAATCC GAATCGGCAGTCGCCATCGGTACCGGCTTCCGCTTTACCGAAAACTTTGCCGCCAAAGCAGGCGTGGCAGTCGGCACT TCGTCCGGTTCTTCCGCAGCCTACCATGTCGGCGTCAATTACGAGTGGCTCGkGCACCACCACCACCACCACTGA 1 MVAJWIGAGL ADALTAPLDH KDKGLQSLTL DQSVRRNBKL KLAAQGAEKT 51 YGNGDSLNTG KLKNDKVSRF DFIRQIEVDG QLITLESGEF QVYKQSHSAL 101 TAFQTEQIQD SEHSGKMVAK RQFRIGDIAG EHTSFDKLPE GGRATYRGTA 151 PGSDDAGGKL TYTIDFAAKQ GNGKIEHLKS PELNVDLAAA DIKPDGKRHA 201 VISGSVLYNQ AEKGSYSLGI FGGKAQEVAG SAEVKTVNGI RHIGLAAKQL 251 EGGGGTGSAT NDDDVKKAAT VAIAAAYNNG QEINGFKAGE TIYDIDEDGT 301 ITKKDATAAD VEADDFKGLG LKKVVTNLTK TVNENKQNVD AKVKAAESEI 351 EKLTTKLADT DAALADTDAA LDATTNALNK LGENITTFAE ETKTNIVKID 401 EKLEAVADTV DKHAEAFNDI ADSLDETNTK ADEAVKTANE AKQTAEETKQ 451 NVDAKVKAAE TAAGKAEAAA GTANTAADKA EAVAAKVTDI KADIATNKDN 501 IAKKANSADV YTREESDSKF VRIDGLNATT EKLDTRLASA EKSIADHDTR 551 LNGLDKTVSD LRKETRQGLA EQAALSGLFQ PYNVGRFDVT AAVGGYKSES 601 AVAIGTGFRF TENFAAKAGV AVGTSSGSSA AYHVGVNYEW LEHHHHHH* &Dgr;G741-961c ATGGTCGCCGCCGACATCGGTGCGGGGCTTGCCGATGCACTAACCGCACCGCTCGACCATAAAGACAAAGGTTTGCAG TCTTTGACGCTGGATCAGTCCGTCAGGAAAAACGAGAAACTGAAGCTGGCGGCACAAGGTGCGGAAAAAACTTATGGA AkCGGTGACAGCCTCAATACGGGCAAATTGAAGAACGACAAGGTCAGCCGTTTCGACTTTATCCGCCAAATCGAAGTG GACGGGCAGCTCATTACCTTGGAGAGTGGAGAGTTCCAAGTATACAAACAAAGCCATTCCGCCTTAACCGCCTTTCAG ACCGAGCAAATACAAGATTCGGAGCATTCCGGGAAGATGGTTGCGAAACGCCAGTTCAGAATCGGCGACATAGCGGGC GAACATACATCTTTTGACAAGCTTCCCGAAGGCGGCAGGGCGACATATCGCGGGACGGCGTTCGGTTCAGACGATGCC GGCGGAAAACTGACCTACACCATAGATTTCGCCGCCAAGCAGGGAAACGGCAAAATCGAACATTTGAAkTCGCCAGAA CTCAATGTCGACCTGGCCGCCGCCGATATCAAGCCGGATGGAAAACGCCATGCCGTCATCAGCGGTTCCGTCCTTTAC AACCAAGCCGAGAAAGGCAGTTACTCCCTCGGTATCTTTGGCGGAAAAGCCCAGGAAGTTGCCGGCAGCGCGGAAGTG AAAACCGTAAACGGCATACGCCATATCGGCCTTGCCGCCAAGCAACTCGAGGGTGGCGGAGGCACTGGATCCGCCACA AACGACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATCAACGGTTTC AAAGCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCCGATGTTGAA GCCGACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAACAAACAAAAC GTCGATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGATGCCGCTTTA GCAGATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACATTTGCTGAA GAGACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCATGCCGAAGCA TTCAACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAATGAAGCCAAA CAGACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAAGCCGAAGCT GCCGCTGGCACAGCTAATACTGCAGCCGAGAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAAGCTGATATC GCTACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGACAGCAAATTT GTCAGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCCATTGCCGAT CACGATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTTGCAGAACAA GCCGCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTCTCGAGCACCACCACCACCACCACTGA 1 MVAADIGAGL ADALTAPLDH KDKGLGSLTL DQSVRKNEKL KLAAGGAEKT 51 YGNGDSLNTG KLKNDKVSRF DFIRGIEVDG GLITLESGEF GVYKGSHSAL 101 TAFGTEGIGD SEHSGKMVAK RGFRIGDIAG EETSFDKLPE GGRATYRGTA 151 FGSDDAGGKL TYTIDFAAKQ GNGKIEHLKS PELNVDLAAA DIKPDGKRHA 201 VISGSVLYNG AEKGSYSLGI FGGKAGEVAG SAEVKTVNGI RHIGLAAKGL 251 EGGGGTGSAT NDDDVKKAAT VAIAAAYNNG GEINGFKAGE TIYDIDEDGT 301 ITKKDATAAD VEADDFKGLG LKKVVTNLTK TVNENKGNVD AKVKAAESEI 351 EKLTTKLADT DAALADTDAA LDATTNALNK LGENITTFAE ETKTNIVKID 401 EKLEAVADTV DKHAEAFNDI ADSLDETNTK ADEAVKTANE AKGTAEETKG 451 NVDAKVKAAE TAAGKAEAAA GTANTAADKA EAVAAKVTDI KADIATNKDN 501 IAKKANSADV YTREESDSKF VRIDGLNATT EKLDTRLASA EKSIADHDTR 551 LNGLDKTVSD LRKETRGGLA EGAALSGLFG PYNVGLEHHH HHH* &Dgr;G741-983 ATGGTCGCCGCCGACATCGGTGCGGGGCTTGCCGATGCACTAACCGCACCGCTCGACCATAAAGACAAAGGTTTGGAG TCTTTGACGCTGGATCAGTCCGTCAGGAAAAACGAGAAACTGAAGCTGGCGGCACAAGGTGCGGAAAAAACTTATGGA AACGGTGACAGCCTCAATACGGGCAAATTGAAGAACGACAAGGTCAGCCGTTTCGACTTTATCCGCCAAATCGAAGTG GACGGGCAGCTCATTACCTTGGAGAGTGGAGAGTTCCAAGTATACAAACAAAGCCATTCCGCCTTAACCGCCTTTCAG ACCGAGCAAATACAAGATTCGGAGCATTCCGGGAAGATGGTTGCGAAACGCCAGTTCAGAATCGGCGACATAGCGGGC GAACATACATCTTTTGACAAGCTTCCCGAAGGCGGCAGGGCGACATATCGCGGGACGGCGTTCGGTTCAGACGATGCC GGCGGAAAACTGACCTACACCATAGATTTCGCCGCCAAGCAGGGAAACGGCAAAATCGAACATTTGAAATCGCCAGAA CTCAATGTCGACCTGGCCGCCGCCGATATCAAGCCGGATGGAAAACGCCATGCCGTCATCAGCGGTTCCGTCCTTTAC AACCAAGCCGAGAAAGGCAGTTACTCCCTCGGTATCTTTGGCGGAAAAGCCCAGGAAGTTGCCGGCAGCGCGGAAGTG AAAACCGTAAACGGCATACGCCATATCGGCCTTGCCGCCAAGCAACTCGAGGGATCCGGCGGAGGCGGCACTTCTGCG CCCGACTTCAATGCAGGCGGTACCGGTATCGGCAGCAACAGCAGAGCAACAACAGCGAAATCAGCAGCAGTATCTTAC GCCGGTATCAAGAACGAAATGTGCAAAGACAGAAGCATGCTCTGTGCCGGTCGGGATGACGTTGCGGTTACAGACAGG GATGCCAAAATCAATGCCCCCCCCCCGAATCTGCATACCGGAGACTTTCCAAACCCAAATGACGCATACAAGAATTTG ATCAACCTCAAACCTGCAATTGAkGCAGGCTATACAGGACGCGGGGTAGAGGTAGGTATCGTCGACACAGGCGAATCC GTCGGCAGCATATCCTTTCCCGAACTGTATGGCAGAAAAGAACACGGCTATAACGAAAATTACAAAAACTATACGGCG TATATGCGGAAGGAAGCGCCTGAAGACGGAGGCGGTAAAGACATTGAAGCTTCTTTCGACGATGAGGCCGTTATAGAG ACTGAAGCAAAGCCGACGGATATCCGCCACGTAAAAGAAATCGGACACATCGATTTGGTCTCCCATATTATTGGCGGG CGTTCCGTGGACGGCAGACCTGCAGGCGGTATTGCGCCCGATGCGACGCTACACATAATGAATACGAATGATGAAACC AAGAACGAAATGATGGTTGCAGCCATCCGCAATGCATGGGTCAAGCTGGGCGAACGTGGCGTGCGCATCGTCAATAAC AGTTTTGGAACAACATCGAGGGCAGGCACTGCCGACCTTTTCCAAATAGCCAATTCGGAGGAGCAGTACCGCCAAGCG TTGCTCGACTATTCCGGCGGTGATAAAACAGACGAGGGTATCCGCCTGATGCAACAGAGCGATTACGGCAACCTGTCC TACCACATCCGTAATAAAAACATGCTTTTCATCTTTTCGACAGGCAATGACGCACAAGCTCAGCCCAACACATATGCC CTATTGCCATTTTATGAAAAAGACGCTCAAAAAGGCATTATCACAGTCGCAGGCGTAGACCGCAGTGGAGAAAAGTTC AAACGGGAAATGTATGGAGAACCGGGTACAGAACCGCTTGAGTATGGCTCCAACCATTGCGGAATTACTGCCATGTGG TGCCTGTCGGCACCCTATGAAGCAAGCGTCCGTTTCACCCGTACAAACCCGATTCAAATTGCCGGAACATCCTTTTCC GCACCCATCGTAACCGGCACGGCGGCTCTGCTGCTGCAGAAATACCCGTGGATGAGCAACGACAACCTGCGTACCACG TTGCTGACGACGGCTCAGGACATCGGTGCAGTCGGCGTGGACAGCAAGTTCGGCTGGGGACTGCTGGATGCGGGTAAG GCCATGAACGGACCCGCGTCCTTTCCGTTCGGCGACTTTACCGCCGATACGAAAGGTACATCCGATATTGCCTACTCC TTCCGTAACGACATTTCAGGCACGGGCGGCCTGATCAAAAAAGGCGGCAGCCAACTGCAACTGCACGGCAACAACACC TATACGGGCAAAACCATTATCGAAGGCGGTTCGCTGGTGTTGTACGGCAACAACAAATCGGATATGCGCGTCGAAACC AAAGGTGCGCTGATTTATAACGGGGCGGCATCCGGCGGCAGCCTGAACAGCGACGGCATTGTCTATCTGGCAGATACC GACCAATCCGGCGCAAACGAAACCGTACACATCAAAGGCAGTCTGCAGCTGGACGGCAAAGGTACGCTGTACACACGT TTGGGCAAACTGCTGAAAGTGGACGGTACGGCGATTATCGGCGGCAAGCTGTACATGTCGGCACGCGGCAAGGGGGCA GGCTATCTCAACAGTACCGGACGACGTGTTCCCTTCCTGAGTGCCGCCAAAATCGGGCAGGATTATTCTTTCTTCACA AACATCGAAACCGACGGCGGCCTGCTGGCTTCCCTCGACAGCGTCGAAAAAACAGCGGGCAGTGAAGGCGACACGCTG TCCTATTATGTCCGTCGCGGCAATGCGGCACGGACTGCTTCGGCAGCGGCACATTCCGCGCCCGCCGGTCTGAAACAC GCCGTAGAACAGGGCGGCAGCAATCTGGAAAACCTGATGGTCGAACTGGATGCCTCCGAATCATCCGCAACACCCGAG ACGGTTGAAACTGCGGCAGCCGACCGCACAGATATGCCGGGCATCCGCCCCTACGGCGCAACTTTCCGCGCAGCGGCA GCCGTAGAACAGGGCGGCAGCAATCTGGAAAACCTGATGGTCGAACTGGATGCCTCCGAATCATCCGCAACACCCGAG GCCGCCCATGCCGATATGCAGGGACGCCGCCTGAAAGCCGTATCGGACGGGTTGGACCACAACGGCACGGGTCTGCGC GTCATCGCGCAAACCCAACAGGACGGTGGAACGTGGGAACAGGGCGGTGTTGAAGGCAAAATGCGCGGCAGTACCCAA ACCGTCGGCATTGCCGCGAAAACCGGCGAAAATACGACAGCAGCCGCCACACTGGGCATGGGACGCAGCACATGGAGC GAAAACAGTGCAAATGCAAAAACCGACAGCATTAGTCTGTTTGCAGGCATACGGCACGATGCGGGCGATATCGGCTAT CTCAAAGGCCTGTTCTCCTACGGACGCTACAAAAACAGCATCAGCCGCAGCACCGGTGCGGACGAACATGCGGAAGGC AGCGTCAACGGCACGCTGATGCAGCTGGGCGCACTGGGCGGTGTCAACGTTCCGTTTGCCGCAACGGGAGATTTGACG GTCGAAGGCGGTCTGCGCTACGACCTGCTCAAACAGGATGCATTCGCCGAAAAAGGCAGTGCTTTGGGCTGGAGCGGC AACAGCCTCACTGAAGGCACGCTGGTCGGACTCGCGGGTCTGAAGCTGTCGCAACCCTTGAGCGATAAAGCCGTCCTG TTTGCAACGGCGGGCGTGGAACGCGACCTGAACGGACGCGACTACACGGTAACGGGCGGCTTTACCGGCGCGACTGCA GCAACCGGCAAGACGGGGGCACGCAATATGCCGCACACCCGTCTGGTTGCCGGCCTGGGCGCGGkTGTCGAATTCGGC AACGGCTGGAACGGCTTGGCACGTTACAGCTACGCCGGTTCCAAACAGTACGGCAACCACAGCGGACGAGTCGGCGTA GGCTACCGGTTCCTCGAGCACCACCACCACCACCACTGA 1 MVAADIGAGL ADALTAPLDH KDKGLGSLTL DGSVRKNEKL KLAAGGAEKT 51 YGNGDSLNTG KLKNDKVSRF DFIRGIEVDG GLITLESGEF GVYKGSHSAL 101 TAFGTEGIGD SBHSGKNVAK RGFRIGDIAG EHTSFDKLPE GGRATYRGTA 151 FGSDDAGGKL TYTIDFAAKG GNGKIEHLKS PELNVDLAAA DIKPDGKRHA 201 VISGSVLYNG AEKGSYSLGI FGGKAGEVAG SABVKTVNGI RHIGLAAKGL 251 EGSGGGGTSA PDFNAGGTGI GSNSRATTAR SAAVSYAGIK NEHCKDRSML 301 CAGRDDVAVT DRDAKINAPP PNLHTGDFPN PNDAYKNLIN LKPAIEAGYT 351 GRGVEVGIVD TGESVGSVGF PELYGRKEHG YNENYKNYTA YMRKEAPEDG 401 GGKDIEASFD DEAVIETEAK PTDIRHVKEI GHIDLVSHII GGRSVDGRPA 451 GGIAPDATLH IMNTNDETKN EMMVAAIRNA WVKLGERGVR IVNNSFGTTS 501 RAGTADLFGI LNSEEGYRGA LLDYSGGDKT DEGIRLMQQS DYGNLSYHIR 551 NKNMLFIFST GNDAGAGPNT YALLPFYEKD AGKGIITVAG VDRSGEKFKR 601 EMYGEPGTEP LEYGSNHCGI TAMWCLSAPY EASVRFTRTN PIGIAGTSFS 651 APIVTGTAAL LLGKYPWMSN DNLRTTLLTT AGDIGAVGVD SKFGWGLLDA 701 GKAHNGPASF PFGDFTADTK GTSDIAYSFR NDISGTGGLI KKGGSGLGLH 751 GNNTYTGKTI IEGGSLVLYG NNKSDMRVET KGALIYNGAA SGGSLNSDGI 801 VYLADTDGSG ANETVHIKGS LGLDGKGTLY TELGKLLKVD GTAIIGGKLY 851 MSARGKGAGY LNSTGRRVPF LSAAKIGGDY SFFTNIETDG GLLASLDSVE 901 KTAGSEGDTL SYYVRRGNAA RTASAAAHSA PAGLKHAVEG GGSNLENLMV 951 ELDASESSAT PETVETAAAD RTDMPGIRPY GATFRAAAAV GHANAADGVR 1001 IFNSLAATVY ADSTAAHADN GGRPLKAVSD GLDHNGTGLR VIAGTGGDGG 1051 TWEGGGVBGK MRGSTGTVGI AAKTGENTTA AATLGMGRST WSENSANAKT 1101 DSISLFAGIR HDAGDIGYLK GLFSYGRYKN SISRSTGADE HAEGSVNGTL 1151 MGLGALGGVN VPFAATGDLT VEGGLRYDLL KGDAFAEKGS ALGWSGNSLT 1201 EGTLVGLAGL KLSGPLSDKA VLFATAGVER DLNGRDYTVT GGFTGATAAT 1251 GKTGARNMPH TRLVAGLGAD VEFGNGWNGL ARYSYAGSKG YGNHSGRVGV 1301 GYRFLEHHHH HH* &Dgr;G741-ORF46.1 ATGGTCGCCGCCGACATCGGTGCGGGGCTTGCCGATGCACTAACCGCACCGCTCGACCATAAAGACAAAGGTTTGCAG TCTTTGACGCTGGATCAGTCCGTCAGGAAAAACGAGAAACTGAAGCTGGCGGCACAAGGTGCGGAAAAAACTTATGGA AACGGTGACAGCCTCAATACGGGCAAATTGAAGAACGACAAGGTCAGCCGTTTCGACTTTATCCGCCAAATCGAAGTG GACGGGCAGCTCATTACCTTGGAGAGTGGAGAGTTCCAAGTATACAAACAAAGCCATTCCGCCTTAACCGCCTTTCAG ACCGAGCAAATACAAGATTCGGAGCATTCCGGGAAGATGGTTGCGAAACGCCAGTTCAGAATCGGCGACATAGCGGGC GAACATACATCTTTTGACAAGCTTCCCGAAGGCGGCAGGGCGACATATCGCGGGACGGCGTTCGGTTCAGACGATGCC GGCGGAAAACTGACCTACACCATAGATTTCGCCGCCAAGCAGGGAAACGGCAAAATCGAACATTTGAAATCGCCAGAA CTCAATGTCGACCTGGCCGCCGCCGATATCAAGCCGGATGGAAAACGCCATGCCGTCATCAGCGGTTCCGTCCTTTAC AACCAAGCCGAGAAAGGCAGTTACTCCCTCGGTATCTTTGGCGGAAAAGCCCAGGAAGTTGCCGGCAGCGCGGAAGTG AAAACCGTAAACGGCATACGCCATATCGGCCTTGCCGCCAAGCAACTCGACGGTGGCGGAGGCACTGGATCCTCAGAT TTGGCAAACGATTCTTTTATCCGGCAGGTTCTCGACCGTCAGCATTTCGAACCCGACGGGAAATAGCACCTATTCGGC AGCAGGGGGGAACTTGCCGAGCGCAGCGGCCATATCGGATTGGGAAAAATACAAAGCCATCAGTTGGGCAACCTGATG ATTCAACAGGCGGCCATTAAAGGAAATATCGGCTACATTGTCGGCTTTTCCGATCACGGGCACGAAGTCCATTCCCCC TTCGACAACCATGCCTCACATTCCGATTCTGATGAAGCCGGTAGTCCCGTTGACGGATTTAGCCTTTACCGCATCCAT TGGGACGGATACGAACACCATCCCGCCGACGGCTATGACGGGCCACAGGGCGGCGGCTATCCCGCTCCCAAAGGCGCG AGGGATATATACAGCTACGACATAAAAGGCGTTGCCCAAAATATCCGCCTCAACCTGACCGACAACCGCAGCACCGGA CAACGGCTTGCCGACCGTTTCCACAATGCCGGTAGTATGCTGACGCAAGGAGTAGGCGACGGATTCAAACGCGCCACC CGATACAGCCCCGAGCTGGACAGATCGGGCAATGCCGCCGAAGCCTTCAACGGCACTGCAGATATCGTTAAAAACATC ATCGGCGCGGCAGGAGAAATTGTCGGCGCAGGCGATGCCGTGCAGGGCATAAGCGAAGGCTGAAACATTGCTGTGATG CACGGCTTGGGTCTGCTTTCCACCGAAAACAAGATGGCGCGCATCAACGATTTGGCAGATATGGCGCAACTCAAAGAC TATGCCGCAGCAGCCATCCGCGATTGGGCAGTCCAAAACCCCAATGCCGCACAAGGCATAGAAGCCGTCAGCAATATC TTTATGGCAGCCATCCCCATCAAAGGGATTGGAGCTGTTCGGGGAAAATACGGCTTGGGCGGCATCACGGGACATCCT ATCAAGCGGTCGCAGATGGGCGCGATCGCATTGCCGAAAGGGAAATCCGCCGTCAGGGACAATTTTGCCGATGCGGCA TACGCCAAATACCCGTCCCCTTACCATTCCCGAAATATCCGTTCAAACTTGGAGCAGCGTTACGGCAAAGAAAACATC ACCTCCTCAACCGTGCCGCCGTCAAACGGCAAAAATGTCAAACTGGCAGACCAACGCCACCCGAAGACAGGCGTACCG TTTGACGGTAAAGGGTTTCCGAATTTTGAGAAGCACGTGAAATATGATACGCTCGAGCACCACCACCACCACCACTGA 1 MVAADIGAGL ADALTAPLDH KDKGLGSLTL DGSVRKNEKL KLAAGGAEKT 51 YGNGDSLNTG KLKNDKVSRF DFIRGIEVDG GLITLESGEF GVYKGSHSAL 101 DAFGTEGIGD SEHSGKMVAK RGFRIGDIAG EHTSFDKLPE GGRATYRGTA 151 FGSDDAGGKL TYTIDFAAKG GNGRIEHLKS PELNVDLAAA DIKPDGKRHA 201 VISGSVLYNG AEKGSYSLGI FGGKAGEVAG SAEVKTVNGI RHIGLAAKGL 251 DGGGGTGSSD LANDSFIRGV LDRGHFBPDG KYHLFGSRGE LAERSGHIGL 301 GKIGSHGLGN LMIGGAAIKG NIGYIVRFSD HGHEVHSPFD NHASHSDSDE 351 AGSPVDGPSL YRIHWDGYEH HPADGYDGPG GGGYPAPKGA RPIYSYDIKG 401 VAGNIRLNLT DNRSTGQRLA DRFHNAGSML TGGVGDGFKR ATRYSPELDR 451 SGNAAEAFNG TADIVKNIIG AAGEIVGAGD AVQGISEGSN IAVHHGLGLL 501 STENKMARIN DLADMAGLKD YAAAAIRDWA VGNPNAAGGI EAVSNIFNAA 551 IPIKGIGAVR GKYGLGGITA HPIKRSGMGA IALPKGKSAV SDNFADAAYA 601 KYPSPYHSRN IRSNLEGRYG KENITSSTVP PSNGKNVKLA DGRHPKTGVP 651 FDGKGFPNFE KHVKYDTLRH HHHHH*

EXAMPLE 5

Hybrids of 287

[0065] Expression of 287 as full-length with a C-terminal His-tag, or without its leader peptide but with a C-terminal His-tag, gives fairly low expression levels. Better expression is achieved using a N-terminal GST-fusion. As an alternative to using GST as an N-terminal fusion partner, 287 was placed at the C-terminus of protein 919 ('919-287′), of protein 953 (‘953-287’), and of proteins ORF46.1 (‘ORF46.1-287’). In both cases, the leader peptides were deleted, and the hybrids were direct in-frame fusions.

[0066] To generate the 953-287 hybrid, the leader peptides of the two proteins were omitted by designing the forward primer downstream from the leader of each sequence; the stop codon sequence was omitted in the 953 reverse primer but included in the 287 reverse primer. For the 953 gene, the 5′ and the 3′ primers used for amplification included a NdeI and a BamHI restriction sites respectively, whereas for the amplification of the 287 gene the 5′ and the 3′ primers included a BamHI and a XhoI restriction sites respectively. In this way a sequential directional cloning of the two genes in pET21b+, using NdeI-BamHI (to clone the first gene) and subsequently BamHI-XhoI (to clone the second gene) could be achieved.

[0067] The 919-287 hybrid was obtained by cloning the sequence coding for the mature portion of 287 into the XhoI site at the 3′-end of the 919-His clone in pET21b+. The primers used for amplification of the 287 gene were designed for introducing a SalI restriction site at the 5′- and a XhoI site at the 3′- of the PCR fragment. Since the cohesive ends produced by the SalI and XhoI restriction enzymes are compatible, the 287 PCR product digested with SalI-XhoI could be inserted in the pET21b-919 clone cleaved with XhoI.

[0068] The ORF46.1-287 hybrid was obtained similarly.

[0069] The bactericidal efficacy (homologous strain) of antibodies raised against the hybrid proteins was compared with antibodies raised against simple mixtures of the component antigens: 19 Mixture with 287 Hybrid with 287 919 32000 16000 953 8192 8192 ORF46.1 128 8192

[0070] Data for bactericidal activity against heterologous MenB strains and against serotypes A and C were also obtained for 919-287 and 953-287: 20 919 953 ORF46.1 Strain Mixture Hybrid Mixture Hybrid Mixture Hybrid MC58  512 1024  512 1024  — 1024 NGH38 1024 2048 2048 4096  — 4096 BZ232  512  128 1024  16 — — MenA (F6124)  512 2048 2048  32 — 1024 MenC (C11) >2048  n.d. >2048  n.d. — n.d. MenC (BZ133) >4096  >8192  >4096  <16 — 2048

[0071] Hybrids of ORF46.1 and 919 were also constructed. Best results (four-fold higher titre) were achieved with 919 at the N-terminus.

[0072] Hybrids 919-519His, ORF97-225His and 225-ORF97His were also tested. These gave moderate ELISA fitres and bactericidal antibody responses.

[0073] As hybrids of two proteins A & B may be either NH2-A-B—COOH or NH2-B-A—COOH, the “reverse” hybrids with 287 at the N-terminus were also made, but using &Dgr;G287. A panel of strains was used, including homologous strain 2996. FCA was used as adjuvant: 21 287 & 919 287 & 953 287 & ORF46.1 Strain &Dgr;G287-919 919-287 &Dgr;G287-953 953-287 &Dgr;G287-46.1 46.1-287 2996 128000 16000 65536 8192 16384 8192 BZ232 256 128 128 <4 <4 <4 1000 2048 <4 <4 <4 <4 <4 MC58 8192 1024 16384 1024 512 128 NGH38 32000 2048 >2048 4096 16384 4096 394/98 4096 32 256 128 128 16 MenA (F6124) 32000 2048 >2048 32 8192 1024 MenC (BZ133) 64000 >8192 >8192 <16 8192 2048

[0074] Better bactericidal titres are generally seen with 287 at the N-terminus.

[0075] When fused to protein 961 [NH2-&Dgr;G287-961-COOH—sequence shown above], the resulting protein is insoluble and must be denatured and renatured for purification. Following renaturation, around 50% of the protein was found to remain insoluble. The soluble and insoluble proteins were compared, and much better bactericidal titres were obtained with the soluble protein (FCA as adjuvant): 22 2996 BZ232 MC58 NGH38 F6124 BZ133 Soluble 65536 128 4096 >2048 >2048 4096 Insoluble  8192  <4  <4   16 n.d. n.d.

[0076] Titres with the insoluble form were, however, improved by using alum adjuvant instead: 23 Insoluble 32768 128 4096 >2048 >2048 2048

[0077] 961c was also used in hybrid proteins (see above). As 961 and its domain variants direct efficient expression, they are ideally suited as the N-terminal portion of a hybrid protein.

EXAMPLE 23

Further Hybrids

[0078] Further hybrid proteins of the invention are shown in the drawings and have the sequences set out below. These are advantageous when compared to the individual proteins: 24 ORF46.1-741. ATGTCAGATTTGGCAAACGATTCTTTTATCCGGCAGGTTCTCGACCGTCAGCATTTCGAACCCGACGGGAAATACCAC CTATTCGGCAGCAGGGGGGAACTTGCCGAGCGCAGCGGCCATATCGGATTGGGAAAAATACAAAGCCATCAGTTGGGC AACCTGATGATTCAACAGGCGGCCATTAAAGGAAATATCGGCTACATTGTCCGCTTTTCCGATCACGGGCACGAAGTC CATTCCCCCTTCGACAACCATGCCTCACATTCCGATTCTGATGAAGCCGGTAGTCCCGTTGACGGATTTAGCCTTTAC CGCATCCATTGGGACGGATACGAACACCATCCCGCCGACGGCTATGACGGGCCACAGGGCGGCGGCTATCCCGCTCCC AAAGGCGCGAGGGATATATACAGCTACGACATAAAAGGCGTTGCCCAAAATATCCGCCTCAACCTGACCGACAACCGC AGCACCGGACAACGGCTTGCCGACCGTTTCCACAATGCCGGTAGTATGCTGACGCAAGGAGTAGGCGACGGATTCAAA CGCGCCACCCGATACAGCCCCGAGCTGGACAGATCGGGCAATGCCGCCGAAGCCTTCAACGGCACTGCAGATATCGTT AAAAACATCATCGGCGCGGCAGGAGAAATTGTCGGCGCAGGCGATGCCGTGCAGGGCATAAGCGAAGGCTCAAACATT GCTGTCATGCACGGCTTGGGTCTGCTTTCCACCGAAAACAAGATGGCGCGCATCAACGATTTGGCAGATATGGCGCAA CTCAAAGACTATGCCGCAGCAGCCATCCGCGATTGGGCAGTCCAAAACCCCAATGCCGCACAAGGCATAGAAGCCGTC AGCAATATCTTTATGGCAGCCATCCCCATCAAAGGGATTGGAGCTGTTCGGGGAAAATACGGCTTGGGCGGCATCACG GCACATCCTATCAAGCGGTCGCAGATGGGCGCGATCGCATTGCCGAAAGGGAAATCCGCCGTCAGCGACAATTTTGCC GATGCGGCATACGCCAAATACCCGTCCCCTTACCATTCCCGAAATATCCGTTCAAACTTGGAGCAGCGTTACGGCAAA GAAAACATCACCTCCTCAACCGTGCCGCCGTCAAACGGCAAAAATGTCAAACTGGCAGACCAACGCCACCCGAAGACA GGCGTACCGTTTGACGGTAAAGGGTTTCCGAATTTTGAGAAGCACGTGAAATATGATACGGGATCCGGAGGGGGTGGT GTCGCCGCCGACATCGGTGCGGGGCTTGCCGATGCACTAACCGCACCGCTCGACCATAAAGACAAAGGTTTGCAGTCT TTGACGCTGGATCAGTCCGTCAGGAAAAACGAGAAACTGAAGCTGGCGGCACAAGGTGCGGAAAAAACTTATGGAAAC GGTGACAGCCTCAATACGGGCAAATTGAAGAACGACAAGGTCAGCCGTTTCGACTTTATCCGCCAAATCGAAGTGGAC GGGCAGCTCATTACCTTGGAGAGTGGAGAGTTCCAAGTATACAAACAAAGCCATTCCGCCTTAACCGCCTTTCAGACC GAGCAAATACAAGATTCGGAGCATTCCGGGAAGATGGTTGCGAAACGCCAGTTCAGAATCGGCGACATAGCGGGCGAA CATACATCTTTTGACAAGCTTCCCGAAGGCGGCAGGGCGACATATCGCGGGACGGCGTTCGGTTCAGACGATGCCGGC GGAAAACTGACCTACACCATAGATTTCGCCGCCAAGCAGGGAAACGGCAAAATCGAACATTTGAAATCGCCAGAACTC GGAAAACTGACCTACACCATAGATTTCGCCGCCAAGCAGGGAAACGGCAAAATCGAACATTTGAAATCGCCAGAACTC AATGTCGACCTGGCCGCCGCCGATATCAAGCCGGATGGAAAACGCCATGCCGTCATCAGCGGTTCCGTCCTTTACAAC CAAGCCGAGAAAGGCAGTTACTCCCTCGGTATCTTTGGCGGAAAAGCCCAGGAAGTTGCCGGCAGCGCGGAAGTGAAA ACCGTAAACGGCATACGCCATATCGGCCTTGCCGCCAAGCAACTCGAGCACCACCACCACCACCACTGA 1 HSDLANDSFI RGVLDRWHFE PDGKYHLFGS RGELAERSGH IGLGKIGSHG 51 LGNLMIGGAA IKGNIGYIVR FSDHGHBVHS PFDNHASHSD SDEAGSPVDG 101 FSLYRIHWDG YEHHPADGYD GPGGGGYPAP KGARDIYSYD IKGVAGNIRL 151 NLTDNRSTGG RLADRFHNAG SMLTGGVGDG FKRATRYSPE LDRSGNAAFA 201 FNGTADIVKN IIGAAGEIVG AGDAVGGISE GSNIAVNHGL GLLSTENKMA 251 RINDLADMAG LKDYAAAAIR DWAVGNPNAA GGIRAVSNIF MAAIPIKGIG 301 AVRGKYGLGG ITAHPIKRSG MGAIALPKGK SAVSDNFADA AYAKYPSPYH 351 SBNIRSNLEG RYGKENITSS TVPPSNGKNV KLADGRHPKT GVPFDGKGFP 401 NFBKHVKYDT GSGGGGVAAD IGAGLADALT APLDHKDKGL GSLTLDGSVR 451 KNEKLKLAAG GAEKTYGNGD SLNTGKLKND KVSRPDFIRG IEVDGGLITL 501 ESGEFGVYKG SHSALTAPGT EGIGDSEHSG KMVAKRGFRI GDIAGEHTSF 551 DKLPEGGRAT YRGTAPGSDD AGGKLTYTID FAAKGGNGKI EHLKSPELNV 601 DLAAADIKPD GKRHAVISGS VLYNGAEKGS YSLGIFGGKA GEVAGSAEVE 651 TVNGIRHIGL AAKGLBHHHH HH* IRF46.1-961 ATGTCAGATTTGGCAAACGATTCTTTTATCCGGCAGGTTCTCGACCGTCAGCATTTCGAACCCGACGGGAAATACCAC CTATTCGGCAGCAGGGGGGAACTTGCCGAGCGCAGCGGCCATATCGGATTGGGAAAAATACAAAGCCATCAGTTGGGC AACCTGATGATTCAACAGGCGGCCATTAAAGGAAATATCGGCTACATTGTCCGCTTTTCCGATCACGGGCACGAAGTC CATTCCCCCTTCGACAACCATGCCTCACATTCCGATTCTGATGAAGCCGGTAGTCCCGTTGACGGATTTAGCCTTTAC CGCATCCATTGGGACGGATACGAACACCATCCCGCCGACGGCTATGACGGGCCACAGGGCGGCGGCTATCCCGCTCCC AAAGGCGCGAGGGATATATACAGCTACGACATAAAAGGCGTTGCCCAAAATATCCGCCTCAACCTGACCGACAACCGC AGCACCGGACAACGGCTTGCCGACCGTTTCCACAATGCCGGTAGTATGCTGACGCAAGGAGTAGGCGACGGATTCAAA CGCGCCACCCGATACAGCCCCGAGCTGGACAGATCGGGCAATGCCGCCGAAGCCTTCAACGGCAGTGCAGATATCGTT AAAAACATCATCGGCGCGGCAGGAGAAATTGTCGGCGCAGGCGATGCCGTGCAGGGCATAAGCGAAGGCTCAAACATT GCTGTCATGCACGGCTTGGGTCTGCTTTCCACCGAAAACAAGATGGCGCGCATCAACGATTTGGCAGATATGGCGCAA CTCAAAGACTATGCCGCAGCAGCCATCCGCGATTGGGCAGTCCAAAACCCCAATGCCGCACAAGGCATAGAAGCCGTC AGCAATATCTTTATGGCAGCCATCCCCATCAAAGGGATTGGAGCTGTTCGGGGAAAATACGGCTTGGGCGGCATCACG GCACATCCTATCAAGCGGTCGCAGATGGGCGCGATCGCATTGCCGAAAGGGAAATCCGCCGTCAGCGACAATTTTGCC GATGCGGCATACGCCAAATACCCGTCCCCTTACCATTCCCGAAATATCCGTTCAAACTTGGAGCAGCGTTACGGCAAA GAAAACATCACCTCCTCAACCGTGCCGCCGTCAAACGGCAAAAATGTCAAACTGGCAGACCAACGCCACCCGAAGACA GGCGTACCGTTTGACGGTAAAGGGTTTCCGAATTTTGAGAAGCACGTGAAATATGATACGGGATCCGGAGGAGGAGGA GCCACAAACGACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATCAAC GGTTTCAAAGCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCCGAT GTTGAAGCCGACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAACAAA CAAAACGTCGATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGATGCC GCTTTAGCAGATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACATTT GCTGAAGAGACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCATGCC GAAGCATTCAACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAATGAA GCCAAACAGACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAAGCC GAAGCTGCCGCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAAGCT AAATTTGTCAGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCCATT GCCGATCACGATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTTGCA GAAGCTGCCGCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAAGCT GATATCGCTACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGACAGC AAATCCGAATCGGCAGTCGCCATCGGTACCGGCTTCCGCTTTACCGAAAACTTTGCCGCCAAAGCAGGCGTGGCAGTC GGCACTTCGTCCGGTTCTTCCGCAGCCTACCATGTCGGCGTCAATTACGAGTGGCTCGAGCACCACCACCACCACCAC TGA 1 MSDLANDSFI RGVLDRGHFE PDGKYHLFGS RGELABRSGH IGLGKIGSHG 51 LGNLHIGGAA IKGNIGYIVR FSDHGHEVHS PFDNHASHSD SDEAGSPVDG 101 FSLYRIHWDG YEHHPADGYD GPGGGGYPAP KGARDIYSYD IKGVAGNIRL 151 NLTDNRSTGG RLADRFHNAG SMLTGGVGDG FKRATRYSPB LDRSGNAAEA 201 FNGTADIVKN IIGAAGEIVG AGDAVGGISE GSNIAVMHGL GLLSTENKMA 251 RINDLADNAG LKDYAAAAIR DWAVGNPNAA GGIEAVSNIF MAAIPIKGIG 301 AVRGKYGLGG ITAHPIKRSG MGAIMaPKGK SAVSDNFADA AYAKYPSPYH 351 SRNIRSNLBG RYGKENITSS TVPPSNGKNV KLADQRHPKT GVPFDGKGFP 401 NFEKHVKYDT GSGGGGATND DDVKKAATVA IAAAYNNGGE INGFKAGETI 451 YDIDEDGTIT KKDATAADVE ADDFKGLGLK KVVTNLTKTV NENKGNVDAK 501 VKAABSEIEK LTTKLADTDA ALADTDAALD ATTNALNKLG ENITTFAEET 551 KTNIVKIDEK LEAVADTVDK HAEAFNDIAD SLDETNTKAD EAVKTANEAK 601 GTAEETKGNV DAKVKAAETA AGKAEAAAGT ANTAADKAEA VAAKVTDIKA 651 DIATNKDNIA KKANSADVYT REESDSKFVR IDGLNATTEK LDTRLASAEK 701 SIADEDTRLN GLDKTVSDLR KETRGGLAEG AALSGLFGPY NVGRFNVTAA 751 VGGYKSESAV AIGTGFRFTE NFAAKAGVAV GTSSGSSAAY HVGVNYEWLE 801 HHHHHH* ORF46.1-961c ATGTCAGATTTGGCAAACGATTCTTTTATCCGGCAGGTTCTCGACCGTCAGCATTTCGAACCCGACGGGAAATACCAC CTATTCGGCAGCAGGGGGGAACTTGCCGAGCGCAGCGGCCATATCGGATTGGGAAAAATACAAAGCCATCAGTTGGGC AACCTGATGATTCAACAGGCGGCCATTAAAGGAAATATCGGCTACATTGTCCGCTTTTCCGATCACGGGCACGAAGTC CATTCCCCCTTCGACAACCATGCCTCACATTCCGATTCTGATGAAGCCGGTAGTCCCGTTGACGGATTTAGCCTTTAC CGCATCCATTGGGACGGATACGAACACCATCCCGCCGACGGCTATGACGGGCCACAGGGCGGCGGCTATCCCGCTCCC AAAGGCGCGAGGGATATATACAGCTACGACATAAAAGGCGTTGCCCAAAATATCCGCCTCAACCTGACCGACAACCGC AGCACCGGACAACGGCTTGCCGACCGTTTCCACAATGCCGGTAGTATGCTGACGCAAGGAGTAGGCGACGGATTCAAA CGCGCCACCCGATACAGCCCCGAGCTGGACAGATCGGGCAATGCCGCCGAAGCCTTCAACGGCACTGCAGATATCGTT AAAAACATCATCGGCGCGGCAGGAGAAATTGTCGGCGCAGGCGATGCCGTGCAGGGCATAAGCGAAGGCTCAAACATT GCTGTCATGCACGGCTTGGGTCTGCTTTCCACCGAAAACAAGATGGCGCGCATCAACGATTTGGCAGATATGGCGCAA CTCAAAGACTATGCCGCAGCAGCCATCCGCGATTGGGCAGTCCAAAACCCCAATGCCGCACAAGGCATAGAAGGCGTC AGCAATATCTTTATGGCAGCCATCCCCATCAAAGGGATTGGAGCTGTTCGGGGAAAATACGGCTTGGGCGGCATCACG GCACATCCTATCAAGCGGTCGCAGATGGGCGCGATCGCATTGCCGAAAGGGAAATCCGCCGTCAGCGACAATTTTGCC GATGCGGCATACGCCAAATACCCGTCCCCTTACCATTCCCGAAATATCCGTTCAAACTTGGAGCAGCGTTACGGCAAA GAAAACATCACCTCCTCAACCGTGCCGCCGTCAAACGGCAAAAATGTCAAACTGGCAGACCAACGCCACCCGAAGACA GGCGTACCGTTTGACGGTAAAGGGTTTCCGAATTTTGAGAAGCACGTGAAATATGATACGGGATCCGGAGGAGGAGGA GCCACAAACGACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATCAAC GGTTTCAAAGCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCCGAT GTTGAAGCCGACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAACAAA CAAAACGTCGATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGATGCC GCTTTAGCAGATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACATTT GCTGAAGAGACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCATGCC GAAGCATTCAACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAATGAA GCCAAACAGACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAAGCC GAAGCTGCCGCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAAGCT GATATCGCTACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGACAGC AAATTTGTCAGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCCATT GCCGATCACGATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTTGCA GAACAAGCCGCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTCTCGAGCACCACCACCACCACCACTGA 1 MSDLANDSFI RGVLDRGHPE PDGKYHLFGS RGELAERSGH IGLGKIGSHG 51 LGNLMIGGAA IKGNIGYIVR FSDHGHEVHS PFDNHASHSD SDEAGSPVDG 101 FSLYRIHWDG YEHHPADGYD GPGGGGYPAP KGARDIYSYD IKGVAGNIRL 151 NLTDNRSTGG RLADRFHNAG SMLTQGVGDG FKRATRYSPE LDRSGNAAEA 203 FNGTADIVKN IIGAAGEIVG AGDAVGGISE GSNIAVHHGL GLLSTENKMA 251 RINDLADMAG LKDYAAAAIR DWAVGNPNAA GGIEAVSNIF MAAIPIKGIG 301 AVRGKYGLGG ITAHPIKRSG NGAIALPKGK SAVSDNFADA AYAKYPSPYH 351 SRNIRSNLBG RYGKENITSS TVPPSNGKNV KLADQRHPKT GVPFDGKGFP 401 NFEKHVKYDT GSGGGGATND DDVKKAATVA IAAAYNNGGB INGFKAGETI 451 YDIDEDGTIT KKDATAADVE ADDFKGLGLK KVVTNLTKTV NENKGNVDAK 501 VKAABSEIEK LTTKLADTDA ALADTDAALD ATTNALNKLG ENITTFAEET 551 KTNIVKIDEK LEAVADTVDK HAEAFNDIAD SLDETNTKAD EAVKTANEAK 601 GTAEETKQNV DAKVKAAETA AGKABAAAGT ANTAADKAEA VAAKVTDIKA 651 DIATNKDNIA KKANSADVYT REESDSKFVR IDGLNATTEK LDTRLASAEK 701 SIADHDTELN GLDKTVSDLR KETRGGLAEG AALSGLFGPY NVGLEHHHHH 751 H* 961-ORF46.1 ATGGCCACAAACGACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATC AACGGTTTCAAAGCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCC GATGTTGAAGCCGACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAAC AAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGAT GCCGCTTTAGCAGATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACA GAAGCCAAACAGACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAA GCCGAAGCATTCAACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAAT GCTGATATCGCTACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGAC AGCAAATTGTCAGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAAATCC ATTGCCGATCACGATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTT GCAGAACAAGCCGCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTCGGTTCAATGTAACGGCTGCAGTCGGCGGC TACAAATCCGAATCGGCAGTCGCCATCGGTACCGGCTTCCGCTTTACCGAAAACTTTGCCGCCAAAGCAGGCGTGGCA GTCGGCACTTCGTCCGGTTCTTCCGCAGCCTACCATGTCGGCGTCAATTACGAGTGGGGATCCGGAGGAGGAGGATCA GATTTGGCAAACGATTCTTTTATCCGGCAGGTTCTCGACCGTCAGCATTTGGAACCCGACGGGAAATACCACCTATTC GGCAGCAGGGGGGAACTTGCCGAGCGCAGCGGCCATATCGGATTGGGAAAAATACAAAGCCATCAGTTGGGCAACCTG ATGATTCAACAGGCGGCCATTAAAGGAAATATCGGCTACATTGTCCGCTTTTCCGATCACGGGCACGAAGTCCATTCC CCCTTCGACAACCATGGCTCACATTCCGATTCTGATGAAGCCGGTAGTCCGGTTGACGGATTTAGCGTTTACCGCATC CATTGGGACGGATACGAACACCATCCCGCCGACGGCTATGACGGGCCACAGGGCGGCGGCTATCCCGCTCCCAAAGGC GCGAGGGATATATACAGCTACGACATAAAAGGCGTTGCCCAAAATATCCGCCTCAACCTGACCGACAACCGCAGCACC GGACAACGGCTTGCCGACCGTTTCCACAATGCCGGTAGTATGCTGACGCAAGGAGTAGGCGACGGATTCAAACGCGCC ACCCGATACAGCCCCGAGCTGGACAGATCGGGCAATGCCGCCGAAGCCTTCAACGGCACTGCAGATATCGTTAAAAAC ATCATCGGCGCGGCAGGAGAAATTGTCGGCGCAGGCGATGCCGTGCAGGGGATAAGCGAAGGGTCAAACATTGCTGTC ATGCACGGCTTGGGTGTGCTTTCCACCGAAAACAAGATGGCGCGCATCAACGATTTGGCAGATATGGCGGAACTCAAA GACTATGCCGCAGCAGCCATCCGCGATTGGGCAGTCCAAAACCCCAATGCGGCACAAGGCATAGAAGCCGTCAGCAAT ATCTTTATGGCAGGCATCCCCATCAAAGGGATTGGAGCTGTTCGGGGAAAATACGGCTTGGGCGGCATCACGGCACAT CCTATCAAGCGGTCGCAGATGGGCGCGATCGCATTGCCGAAAGGGAAATCCGCCGTCAGCGACAATTTTGCCGATGCG GCATACGCCAAATACCCGTCCCCTTACCATTCCCGAAATATCCGTTCAAACTTGGAGCAGCGTTACGGCAAAGAAAAC ATCACCTCCTCAACCGTGCCGCCGTCAAACGGCAAAAATGTCAAACTGGCAGACCAACGCCACCCGAAGACAGGCGTA CCGTTTGACGGTAAAGGGTTTCCGAATTTTGAGAAGCACGTGAAATATGATACGCTCGAGCACCACCACCACCACCAC TGA 1 HATNDDDVKK AATVAIAAAY NNGGEINGFK AGETIYDIDE DGTITKKDAT 51 AADVEADDFK GLGLKKVVTN LTKTVNENKG NVDAKVKAAE SEIEKLTTKL 101 ADTDAALADT DAALDATTNA LNKLGENITT FAEETKTNIV KIDEKLEAVA 151 DTVDKHAEAP NDIADSLDET NTKADEAVKT ANEAKQTAEE TKQNVDAKVK 201 AAETAAGKAE AAAGTANTAA DKAEAVAAKV TDIKADIATN KDNIAKKANS 251 ADVYTREESD SKFVRIDGLN ATTEKLDTRL ASAEKSIADH DTRLNGLDKT 301 VSDLRKETRG GLAEGAALSG LFGPYNVGRF NVTAAVGGYK SESAVAIGTG 351 FRFTENFAAK AGVAVGTSSG SSAAYHVGVN YEWGSGGGGS DLANDSFIRG 401 VLDRGHFBPD GKYHLFGSRG ELAERSGHIG LGKIGSHGLG NLMIGGAAIK 451 GNIGYIVRFS DHGHEVHSPF DNHASHSDSD EAGSPVDGFS LYRIHWDGYE 501 HEPADGYDGP GGGGYPAPKG ARDIYSYDIK GVAGNIBLNL TDNRSTGGRL 551 ADRFHNAGSM LTGGVGDGFK EATRYSPELD RSGNAAEAFN GTADIVKNII 601 GAAGEIVGAG DAVGGISEGS NIAVMHGLGL LSTENKHARI NDLADMAGLK 651 DYAAAAIRDW AVGNPNAAGG IEAVSNIFNA AIPIKGIGAV RGKYGLGGIT 701 AHPIKRSGMG AIALPKGKSA VSDNFADAAY AKYPSPYHSR NIRSNLEGRY 751 GKENITSSTV PPSNGKNVKL ADGRHPKTGV PFDGKGFPNF EKHVKYDTLE 801 HHHHHH* 961-741 ATGGCCACAAACGACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATC AACGGTTTCAAAGCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCC GATGTTGAAGCCGACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAAC AAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGAT GCCGCTTTAGCAGATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACA TTTGCTGAAGAGACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCAT GCCGAAGCATTCAACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAAT GAAGCCAAACAGACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAA GCCGAAGCTGCCGCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAA GCTGATATCGCTACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGAC AGCAAATTTGTCAGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCC ATTGCCGATCACGATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTT GCAGAACAAGCCGCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTCGGTTCAATGTAACGGCTGCAGTCGGCGGC TACAAATCCGAATCGGCAGTCGCCATCGGTACCGGCTTCCGCTTTACCGAAAACTTTGCCGCCAAAGCAGGCGTGGCA GTCGGCACTTCGTCCGGTTCTTCCGCAGCCTACCATGTCGGCGTCAATTACGAGTGGGGATCCGGAGGGGGTGGTGTC GCCGCCGACATCGGTGCGGGGCTTGCCGATGCACTAACCGCACCGCTCGACCATAAAGACAAAGGTTTGCAGTCTTTG ACGCTGGATCAGTCCGTCAGGAAAAACGAGAAACTGAAGCTGGCGGCACAAGGTGCGGAAAAAACTTATGGAAACGGT GACAGCCTCAATACGGGCAAATTGAAGAACGACAAGGTCAGCCGTTTCGACTTTATCCGCCAAATCGAAGTGGACGGG CAGCTCATTACCTTGGAGAGTGGAGAGTTCCAAGTATACAAACAAAGCCATTCCGCCTTAACCGCCTTTCAGACCGAG CAATACAAGATTCGGAGCATTCCGGGAAGATGGTTGCGAAACGCCAGTCTCAGAATCGGCGACATAGCGGGCGAACAT ACATCTTTTGACAAGCTTCCCGAAGGCGGCAGGGCGACATATCGCGGGACGGCGTTCGGTTCAGACGATGCCGGCGGA AAACTGACCTACACCATAGATTTCGCCGCCAAGCAGGGAAACGGCAAAATCGAACATTTGAAATCGCCAGAACTCAAT GTCGACCTGGCCGCCGCCGATATCAAGCCGGATGGAAAACGCCATGCCGTCATCAGCGGTTCCGTCCTTTACAACCAA GCCGAGAAAGGCAGTTACTCCTCGGTATCTTTGGCGGAAAAGCCCCAGGAAGTTGCCGGCAGCGCGGAAGTGAAAACC GTAACGGCATACGCCATATCGGCCTTGCCGCCAAGCAACTCGAGCACCACCACCACCACCACTGA 1 MATNDDDVKK AATVAIAAAY NNGGBINGFK AGETIYDIDE DGTITKKDAT 51 AADVEADDFK GLGLKKVVTN LTKTVNENKG NVDAKVKAAE SEIEKLTTKL 101 ADTDAALADT DAALDATTNA LNKLGENITT FAEETKTNIV KIDBKEHAVA 151 DTVDKHAEAF NDIADSLDET NTKADEAVKT ANEAKGTAEE TKGNVDAKVK 201 AAETAAGKAE AAAGTANTAA DKAEAVAAKV TDIKADIATN KDNIAKKANS 251 ADVYTREESD SKFVRIDGLN ATTEKLDTRL ASAEKSIADH DTRLNGLDKT 301 VSDLRKBTRG GLAEGAALSG LFGPYNVGRF NVTAAVGGYK SESAVAIGTG 351 FRPTENFAAK AGVAVGTSSG SSAAYHVGVN YEWGSGGGGV AADIGAGLAD 401 ALTAPLDHKD KGLGSLTLDG SVRKNEKLKL AAGGAEKTYG NGDSLNTGKL 451 KNDKVSRFDF IRGIEVDGGL ITLESGEFGV YKGSHSALTA FGTEGIGDSE 501 HSGKMVAKRG FRIGDIAGEH TSFDKLPEGG RATYRGTAFG SDDAGGKLTY 551 TIDFAAKGGN GKIEHLKSPE LNVDLAAADI KPDGKEHAVI SGSVLYNGAE 601 KGSYSLGIFG GKAGEVAGSA EVKTVNGIRE IGLAARGLEH HHHHH* 961-983 ATGGCCACAAACGACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATC AACGGTTTCAAAGCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCC GATGTTGAAGCCGACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAAC AAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGAT GCCGCTTTAGCAGATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACA TTTGCTGAAGAGACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCAT GCCGAAGCATTCAACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAAT GAAGCCAAACAGACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAA GCCGAAGCTGCCGCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAA GCTGATATCGCTACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGAC AGCAAATTTGTCAGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCC ATTGCCGATCACGATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTT GCAGAACAAGCCGCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTCGGTTCAATGTAACGGCTGCAGTCGGCGGC TACAAATCCGAATCGGCAGTCGCCATCGGTACCGGCTTCCGCTTTACCGAAAACTTTGCCGCCAAADCAGGCGTGGCA GTCGGCACTTCGTCCGGTTCTTCCGCAGCCTACCATGTCGGCGTCAATTACGAGTGGGGATCCGGCGGAGGCGGCACT TCTGCGCCCGACTTCAATGCAGGCGGTACCGGTATCGGCAGCAACAGCAGAGCAACAACAGCGAAATCAGCAGCAGTA TCTTACGCCGGTATCAAGAACGAAATGTGCAAAGACAGAAGCATGCTCTGTGCCGGTCGGGATGACGTTGCGGTTACA GACAGGGATGCCAAAATCAATGCCCCCCCCCCGAATCTGCATACCGGAGACTTTCCAAACCCAAATGACGCATACAAG AATTTGATCAACCTCAAACCTGCAATTGAAGCAGGCTATACAGGACGCGGGGTAGAGGTAGGTATCGTCGACACAGGC GAATCCGTCGGCAGCATATCCTTTCCCGAACTGTATGGCAGAAAAGAACACGGCTATAACGAAAATTACAAAAACTAT ACGGCGTATATGCGGAAGGAAGCGCCTGAAGACGGAGGCGGTAAAGACATTGAAGCTTCTTTCGACGATGAGGCCGTT ATAGAGACTGAAGCAAAGCCGACGGATATCCGCCACGTAAAAGAAATCGGACACATCGATTTGGTCTCCCATATTATT GGCGGGCGTTCCGTGGACGGCAGACCTGCAGGCGGTATTGCGCCCGATGCGACGCTACACATAATGAATACGAATGAT GAAACCAAGAACGAAATGATGGTTGCAGCCATCCGCAATGCATGGGTCAAGCTGGGCGAACGTGGCGTGCGCATCGTC AATAACAGTTTTGGAACAACATCGAGGGCAGGCACTGCCGACCTTTTCCAAATAGCCAATTCGGAGGAGCAGTACCGC CAAGCGTTGCTCGACTATTCCGGCGGTGATAAAACAGACGAGGGTATCCGCCTGATGCAACAGAGCGATTACGGCAAC CTGTCCTACCACATCCGTAATAAAAACATGCTTTTCATCTTTTCGACAGGCAATGACGCACAAGCTCAGCCCAACACA TATGCCCTATTGCCATTTTATGAAAAAGACGCTCAAAAAGGCATTATCACAGTCGCAGGCGTAGACCGCAGTGGAGAA AAGTTCAAACGGGAAATGTATGGAGAACCGGGTACAGAACCGCTTGAGTATGGCTCCAACCATTGCGGAATTACTGCC ATGTGGTGCCTGTCGGCACCCTATGAAGCAAGCGTCCGTTTCACCCGTACAAACCCGATTCAAATTGCCGGAACATCC TTTTCCGCACCCATCGTAACCGGCACGGCGGCTCTGCTGCTGCAGAAATACCCGTGGATGAGCAACGACAACCTGCGT ACCACGTTGCTGACGACGGCTCAGGACATCGGTGCAGTCGGCGTGGACAGCAAGTTCGGCTGGGGACTGCTGGATGCG GGTAAGGCCATGAACGGACCCGCGTCCTTTCCGTTCGGCGACTTTACCGCCGATACGAAAGGTACATCCGATATTGCC TACTCCTTCCGTAACGACATTTCAGGCACGGGCGGCCTGATCAAAAAAGGCGGCAGCCAACTGCAACTGCACGGCAAC AACACCTATACGGGCAAAACCATTATCGAAGGCGGTTCGCTGGTGTTGTACGGCAACAACAAATCGGATATGCGCGTC GAAACCAAAGGTGCGCTGATTTATAACGGGGCGGCATCCGGCGGCAGCCTGAACAGCGACGGCATTGTCTATCTGGCA GATACCGACCAATCCGGCGCAAACGAAACCGTACACATCAAAGGCAGTCTGCAGCTGGACGGCAAAGGTACGCTGTAC ACACGTTTGGGCAAACTGCTGAAAGTGGACGGTACGGCGATTATCGGCGGCAAGCTGTACATGTCGGCACGCGGCAAG GGGGCAGGCTATCTCAACAGTACCGGACGACGTGTTCCCTTCCTGAGTGCCGCCAAAATCGGGCAGGATTATTCTTTC TTCACAAACATCGAAACCGACGGCGGCCTGCTGGCTTCCCTCGACAGCGTCGAAAAAACAGCGGGCAGTGAAGGCGAC ACGCTGTCCTATTATGTCCGTCGCGGCAATGCGGCACGGACTGCTTCGGCAGCGGCACATTCCGCGCCCGCCGGTCTG AAACACGCCGTAGAACAGGGCGGCAGCAATCTGGAAAACCTGATGGTCGAACTGGATGCCTCCGAATCATCCGCAACA CCCGAGACGGTTGAAACTGCGGCAGCCGACCGCACAGATATGCCGGGCATCCGCCCCTACGGCGCAACTTTCCGCGCA GCGGCAGCCGTACAGCATGCGAATGCCGCCGACGGTGTACGCATCTTCAACAGTCTCGCCGCTACCGTCTATGCCGAC AGTACCGCCGCCCATGCCGATATGCAGGGACGCCGCCTGAAAGCCGTATCGGACGGGTTGGACCACAACGGCACGGGT CTGCGCGTCATCGCGCAAACCCAACAGGACGGTGGAACGTGGGAACAGGGCGGTGTTGAAGGCAAAATGCGCGGCAGT ACCCAAACCGTCGGCATTGCCGCGAAAACCGGCGAAAATACGACAGCAGCCGCCACACTGGGCATGGGACGCAGCACA TGGAGCGAAAACAGTGCAAATGCAAAAACCGACAGCATTAGTCTGTTTGCAGGCATACGGCACGATGCGGGCGATATC GGCTATCTCAAAGGCCTGTTCTCCTACGGACGCTACAAAAACAGCATCAGCCGCAGCACCGGTGCGGACGAACATGCG GAAGGCAGCGTCAACGGCACGCTGATGCAGCTGGGCGCACTGGGCGGTGTCAACGTTCCGTTTGCCGCAACGGGAGAT TTGACGGTCGAAGGCGGTCTGCGCTACGACCTGCTCAAACAGGATGCATTCGCCGAAAAAGGCAGTGCTTTGGGCTGG AGCGGCAACAGCCTCACTGAAGGCACGCTGGTCGGACTCGCGGGTCTGAAGCTGTCGCAACCCTTGAGCGATAAAGCC GTCCTGTTTGCAACGGCGGGCGTGGAACGCGACCTGAACGGACGCGACTACACGGTAACGGGCGGCTTTACCGGCGCG ACTGCAGCAACCGGCAAGACGGGGGCACGCAATATGCCGCACACCCGTCTGGTTGCCGGCCTGGGCGCGGATGTCGAA TTCGGCAACGGCTGGAACGGCTTGGCACGTTACAGCTACGCCGGTTCCAAACAGTACGGCAACCACAGCGGACGAGTC GGCGTAGGCTACCGGTTCCTCGAGCACCACCACCACCACCACTGA 1 HATNDDDVKK AATVAIAAAY NNGGEINGFK AGETIYDIDE DGTITKKDAT 51 AADVEADDFK GLGLKKVVTN LTKTVNENKG NVDAKVKAAE SEIEKLTTKL 101 ADTDAALADT DAALDATTNA LNKLGENITT FAEETKTNIV KIDEKLEAVA 151 DTVDKHAEAF NDIADSLDET NTKADEAVKT ANEAKGTAEE TKGNVDAKVK 201 AAETAAGKAE AAAGTANTAA DKAEAVAARV TDIKADIATN KDNIAKKANS 251 ADVYTREESD SKFVRIDGLN ATTEKLDTRL ASAEKSIADH DTRLNGLDKT 301 VSDLRKETRG GLAEQAALSG LFQPYNVGRF NVTAAVGGYK SESAVAIGTG 351 FRFTENFAAR AGVAVGTSSG SSAAYHVGVN YEWGSGGGGT SAPDFNAGGT 401 GIGSNSRATT AKSAAVSYAG IKNEMCKDRS MLCAGRDDVA VTDRDAKINA 451 PPPNLHTGDF PNPNDAYKNL INLKPAIEAG YTGRGVEVGI VDTGESVGSI 501 SFPELYGRKE HGYNENYKNY TAYMRKEAPE DGGGKDIEAS FDDEAVIETE 551 AKPTDIRHVK EIGHIDLVSH IIGGRSVDGR PAGGIABDAT LHIMNTNDET 601 KNEMMVAAIR NAWVKLGERG VRIVNNSFGT TSRAGTADLF GIANSEEGYR 651 QALLDYSGGD KTDEGIRIMG GSDYGNLSYH IRNKNNLFIF STGNDAGAGP 701 NTYALLPFYE KDAGKGIITV AGVDRSGEKF KPEMYGEPGT EPLEYGSNHC 751 GITAMWCLSA PYEASVRFTR TNPIGIAGTS FSAPIVTGTA ALLLGKYPWM 801 SNDNLRTTLL TTAGDIGAVG VDSKFGWGLL DAGKAMNGPA SFPFGDFTAD 851 TKGTSDIAYS FRNDISGTGG LIKKGGSGLG LHGNNTYTGK TIIEGGSLVL 901 YGNNKSDMRV ETKGALIYNG AASGGSLNSD GIVYLADTDG SGANETVHIK 951 GSLGLDGKGT LYTRLGKLLK VDGTAIIGGK LYMSARGKGA GYLNSTGRRV 1001 PFLSAAKIGG DYSFFTNIET DGGLLASLDS VEKTAGSEGD TLSYYVRRGN 1051 AARTASAAAH SAPAGLKHAV EQGGSNLENL HVELDASESS ATPETVETAA 1101 ADRTDMPGIR PYGATFRAAA AVGHANAADG VRIFNSLAAT VYADSTAAHA 1151 DMGGRRLKAV SDGLDHNGTG LRVIAGTGGD GGTWEGGGVE GKMRGSTGTV 1201 GIAAKTGENT TAAATLGMGR STWSENSANA KTDSISLFAG IRHDAGDIGY 1251 LKGLFSYGRY KNSISRSTGA DEHAEGSVNG TLMGLGALGG VNVPFAATGD 1301 LTVEGGLRYD LLKGDAFAEK GSALGWSGNS LTEGTLVGLA GLKLSGPLSD 1351 KAVLFATAGV ERDLNGRDYT VTGGFTGATA ATGKTGARNM PHTRLVAGLG 1401 ADVEFGNGWN GLARYSYAGS RQYGNHSGRV GVGYRFLEHH HHHH* 961c-ORF46.1 ATGGCCACAAACGACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATC AACGGTTTCAAAGCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCC GATGTTGAAGCCGACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAAC AAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGAT GCCGCTTTAGCAGATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACA TTTGCTGAAGAGACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCAT GCCGAAGCATTCAACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAAT GAAGCCAAACAGACGGCCGAAGAAACCAAACTAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAA GCCGAAGCTGCCGCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAT GCTGATATCGCTACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGAC AGCAAATTTGTCAGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCC ATTGCCGATCACGATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTT GCAGAACAAGCCGCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTGGATCCGGAGGAGGAGGATCAGATTTGGCA AACGATTCTTTTATCCGGCAGGTTCTCGACCGTCAGCATTTCGAACCCGACGGGAAATACCACCTATTCGGCAGCAGG GGGGAACTTGCCGAGCGCAGCGGCCATATCGGATTGGGAAAAATACAAAGCCATCAGTTGGGCAACCTGATGATTCAA CAGGCGGCCATTAAAGGAAATATCGGCTACATTGTCCGCTTTTCCGATCACGGGCACGAAGTCCATTCCCCCTTCGAC AACCATGCCTCACATTCCGATTCTGATGAAGCCGGTAGTCCCGTTGACGGATTTAGCCTTTACCGCATCCATTGGGAC GGATACGAACACCATCCCGCCGACGGCTATGACGGGCCACAGGGCGGCGGCTATCCCGCTCCCAAAGGCGCGAGGGAT ATATACAGCTACGACATAAAAGGCGTTGCCCAAAATATCCGCCTCAACCTGACCGACAACCGCAGCACCGGACAACGG CTTGCCGACCGTTTCCACAATGCCGGTAGTATGCTGACGCAAGGAGTAGGCGACGGATTCAAACGCGCCACCCGATAC AGCCCCGAGCTGGACAGATCGGGCAATGCCGCCGAAGCCTTCAACGGCACTGCAGATATCGTTAAAAACATCATCGGC GCGGCAGGAGAAATTGTCGGCGCAGGCGATGCCGTGCAGGGCATAAGCGAAGGCTCAAACATTGCTGTCATGCACGGC TTGGGTCTGCTTTCCACCGAAAACAAGATGGCGCGCATCAACGATTTGGCAGATATGGCGCAACTCAAAGACTATGCC GCAGCAGCCATCCGCGATTGGGCAGTCCAAAACCCCAATGCCGCACAAGGCATAGAAGCCGTCAGCAATATCTTTATG GCAGCCATCCCCATCAAAGGGATTGGAGCTGTTCGGGGAAAATACGGCTTGGGCGGCATCACGGCACATCCTATCAAG CGGTCGCAGATGGGCGCGATCGCATTGCCGAAAGGGAAATCCGCCGTCAGCGACAATTTTGCCGATGCGGCATACGCC AAATACCCGTCCCCTTACCATTCCCGAAATATCCGTTCAAACTTGGAGCAGCGTTACGGCAAAGAAAACATCACCTCC TCAACCGTGCCGCCGTCAAACGGCAAAAATGTCAAACTGGCAGACCAACGCCACCCGAAGACAGGCGTACCGTTTGAC GGTAAAGGGTTTCCGAATTTTGAGAAGCACGTGAAATATGATACGCTCGAGCACCACCACCACCACCACTGA 1 HATNDDDVKK AATVAIAAAY NNGGEINGFK AGETIYDIDE DGTITKKDAT 51 AADVEADDFK GLGLKKVVTN LTKTVNENKG NVDAKVKAAE SEIEKLTTKL 101 ADTDAALADT DAALDATTNA LNKLGENITT FAEETKTNIV KIDEKLEAVA 151 DTVDKHAEAF NDIADSLDET NTKADEAVKT ANEAKGTAEE TKGNVDAKVK 201 AAETAAGKAE AAAGTANTAA DKAEAVAAKV TDIKADIATN KDNThKKANS 251 ADVYTREESD SKFVRIDGLN ATTEKLDTRL ASAEKSIADH DTRLNGLDKT 301 VSDLRKETRG GLAEGAALSG LFQPYNVGGS GGGGSDLAND SFIRQVLDRG 351 HFEPDGKYHL FGSRGELAER SGHIGLGKIG SHQLGNLMIG GAAIKGNIGY 401 IVRFSDHGHE VHSPFDNHAS HSDSDEAGSP VDGFSLYRIH WDGYEHHPAD 451 GYDGPGGGGY PAPKGARDIY SYDIKGVAGN IELNLTDNRS TGGELADRFH 501 NAGSMLTGGV GDGFKRATRY SPELDRSGNA AEAFNGTADI VKNIIGAAGE 551 IVGAGDAVGG ISEGSNIAVM HGLGLLSTEN KMARINDLAD MAGLKDYAAA 601 AIRDWAVGNP NAAGGIEAVS NIFMAAIPIK GIGAVRGKYG LGGITAHPIK 651 RSGMGAIALP KGKSAVSDNF ADAAYAKYPS PYHSRNIRSN LEGRYGKENI 701 TSSTVPPSNG KNVKLADGRH PKTGVPFDGK GFPNFEKHVK YDTLEHHHHH 751 H* 961c-741 ATGGCCACAAACGACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATC AACGGTTTCAAAGCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCC GATGTTGAAGCCGACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAAC AAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGAT GCCGCTTTAGCAGATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACA TTTGCTGAAGAGACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCAT GCCGAAGCATTCAACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAAT GAAGCCAAACAGACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAA GCCGAAGCTGCCGCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAA GCTGATATCGCTACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGAC AGCAAATTTGTCAGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCC ATTGCCGATCACGATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTT GCAGAACAAGCCGCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTGGATCCGGAGGGGGTGGTGTCGCCGCCGAC ATCGGTGCGGGGCTTGCCGATGCACTAACCGCACCGCTCGACCATAAAGACAAAGGTTTGCAGTCTTTGACGCTGGAT CAGTCCGTCAGGAAAAACGAGAAACTGAAGCTGGCGGCACAAGGTGCGGAAAAAACTTATGGAAACGGTGACAGCCTC AATACGGGCAAATTGAAGAACGACAAGGTCAGCCGTTTCGACTTTATCCGCCAAATCGAAGTGGACGGGCAGCTCATT ACCTTGGAGAGTGGAGAGTTCCAAGTATACAAACAAAGCCATTCCGCCTTAACCGCCTTTCAGACCGAGCAAATACAA GATTCGGAGCATTCCGGGAAGATGGTTGCGAAACGCCAGTTCAGAATCGGCGACATAGCGGGCGAACATACATCTTTT GACAAGCTTCCCGAAGGCGGCAGGGCGACATATCGCGGGACGGCGTTCGGTTCAGACGATGCCGGCGGAAAACTGACC TACACCATAGATTTCGCCGCCAAGCAGGGAAACGGCAAAATCGAACATTTGAAATCGCCAGAACTCAATGTCGACCTG GCCGCCGCCGATATCAAGCCGGATGGAAAACGCCATGCCGTCATCAGCGGTTCCGTCCTTTACAACCAAGCCGAGAAA GGCAGTTACTCCCTCGGTATCTTTGGCGGAAAAGCCCAGGAAGTTGCCGGCAGCGCGGAAGTGAAAACCGTAAACGGC ATACGCCATATCGGCCTTGCCGCCAAGCAACTCGAGCACCACCACCACCACCACTGA 1 MATNDDDVKK AATVAIAAAY NNGGEINGFK AGETIYDIDE DGTITKKDAT 51 AADVEADDFK GLGLKKVVTN LTKTVNENKQ NVDAKVKAAE SEIEKLTTKL 101 ADTDAALADT DAALDATTNA LNKLGENITT FAEETKTNIV KIDERLEAVA 151 DTVDKHAEAF NDIADSLDET NTKADEAVKT ANEAKGTAEE TKGNVDAKVK 201 AAETAAGKAE AAAGTANTAA DKAEAVAAKV TDIKADIATN KDNIAKKANS 251 ADVYTREESD SKFVRIDGLN ATTEKLDTEL ASAEKSIADH DTRLNGLDKT 301 VSDLEREThG GLAEGAALSG LFQPYNVGGS GGGGVAADIG AGLADALTAP 351 LDHKDKGLGS LTLDGSVRKN EKLKLAAGGA EKTYGNGDSL NTGKLKNDKV 401 SRPDFIRGIE VDGGLITLES GEFGVYKGSH SALTAPGTEG IQDSEHSGKM 451 VAKRGFRIGD IAGEHTSFDK LPEGGRATYR GTAFGSDDAG GKLTYTIDFA 501 AKGGNGKIEH LKSPELNVDL AAADIKPDGK REAVISGSVL YNGAEKGSYS 551 LGIFGGKAGE VAGSAEVKTV NGIRHIGLAA KGLEHHHHHH * 961c-983 ATGGCCACAAACGACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATC AACGGTTTCAAAGCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCC GATGTTGAAGCCGACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAAC AAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGAT GCCGCTTTAGCAGATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACA TTTGCTGAAGAGACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCAT GCCGAAGCATTCAACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAAT GAAGCCAAACAGACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAA GCCGAAGCTGCCGCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAA GCTGATATCGCTACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGAC AGCAAATTTGTCAGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCC ATTGCCGATCACGATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTT GCAGAACAAGCCGCGCTCTCCCGTCTGTTCCAACCTTACAACGTGGGTGGATCCGGCGGAGGCGGCACTTCTGCGCCC GACTTCAATGCAGGCGGTACCGGTATCGGCAGCAACAGCAGAGCAACAACAGCGAAATCAGCAGCAGTATCTTACGCC GGTATCAAGAACGAAATGTGCAAAGACAGAAGCATGCTCTGTGCCGGTCGGGATGACGTTGCGGTTACAGACAGGGAT GCCAAATCAATGCCCCCCCCCCCGAATCTGCATACCGGAGACTTTCCAAACCCAAATGACGCATACAAGAATTTGATC AACCTCAAACCTGCAATTGAAGCAGGCTATACAGGACGCGGGGTAGAGGTAGGTATCGTCGACACAGGCGAATCCGTC GGCAGCATATCCTTTCCCGAACTGTATGGCAGAAAAGAACACGGCTATAACGAAAATTACAAAAACTATACGGCGTAT ATGCGGAAGGAAGCGCCTGAAGACGGAGGCGGTAAAGACATTGAAGCTTCTTTCGACGATGAGGCCGTTATAGAGACT GAAGCAAAGCCGACGGATATCCGCCACGTAAAAGAAATCGGACACATCGATTTGGTCTCCCATATTATTGGCGGGCGT TCCGTGGACGGCAGACCTGCAGGCGGTATTGCGCCCGATGCGACGCTACACATAATGAATACGAATGATGAAACCAAG AACGAAATGATGGTTGCAGCCATCCGCAATGCATGGGTCAAGCTGGGCGAACGTGGCGTGCGCATCGTCAATAACAGT TTTGGAACAACATCGAGGGCAGGCACTGCCGACCTTTTCCAAATAGCCAATTCGGAGGAGCAGTACCGCCAAGCGTTG CTCGACTATTCCGGCGGTGATAAAACAGACGAGGGTATCCGCCTGATGCAACAGAGCGATTACGGCAACCTGTCCTAC CACATCCGTAATAAAAACATGCTTTTCATCTTTTCGACAGGCAATGACGCACAAGCTCAGCCCAACACATATGCCCTA TTGCCATTTTATGAAAAAGACGCTCAAAAAGGCATTATCACAGTCGCAGGCGTAGACCGCAGTGGAGAAAAGTTCAAA CGGGAAATGTATGGAGAACCGGGTACAGAACCGCTTGAGTATGGCTCCAACCATTGCGGAATTACTGCCATGTGGTGC CTGTCGGCACCCTATGAAGCAAGCGTCCGTTTCACCCGTACAAACCCGATTCAAATTGCCGGAACATCCTTTTCCGCA CCCATCGTAACCGGCACGGCGGCTCTGCTGCTGCAGAAATACCCGTGGATGAGCAACGACAACCTGCGTACCACGTTG CTGACGACGGCTCAGGACATCGGTGCAGTCGGCGTGGACAGCAAGTTCGGCTGGGGACTGCTGGATGCGGGTAAGGCC ATGAACGGACCCGCGTCCTTTCCGTTCGGCGACTTTACCGCCGATACGAAAGGTACATCCGATATTGCCTACTCCTTC CGTAACGACATTTCAGGCACGGGCGGCCTGATCAAAAAAGGCGGCAGCCAACTGCAACTGCACGGCAACAACACCTAT ACGGGCAAAACCATTATCGAAGGCGGTTCGCTGGTGTTGTACGGCAACAACAAATCGGATATGCGCGTCGAAACCAAA GGTGCGCTGATTTATAACGGGGCGGCATCCGGCGGCAGCCTGAACAGCGACGGCATTGTCTATCTGGCAGATACCGAC CAATCCGGCGCAAACGAAACCGTACACATCAAAGGCAGTCTGCAGCTGGACGGCAAAGGTACGCTGTACACACGTTTG GGCAAACTGCTGAAAGTGGACGGTACGGCGATTATCGGCGGCAAGCTGTACATGTCGGCACGCGGCAAGGGGGCAGGC TATCTCAACAGTACCGGACGACGTGTTCCCTTCCTGAGTGCCGCCAAAATCGGGCAGGATTATTCTTTCTTCACAAAC ATCGAAACCGACGGCGGCCTGCTGGCTTCCCTCGACAGCGTCGAAAAAACAGCGGGCAGTGAAGGCGACACGCTGTCC TATTATGTCCGTCGCGGCAATGCGGCACGGACTGCTTCGGCAGCGGCACATTCCGCGCCCGCCGGTCTGAAACACGCC GTAGAACAGGGCGGCAGCTATCTGGAAAACCTGATGGTCGAACTGGATGCCTCCGAATCATCCGCAACACCCGAGACG GTTGAAACTGCGGCAGCCGACCGCACAGATATGCCGGGCATCCGCCCCTACGGCGCAACTTTCCGCGCAGCGGCAGCC GTACAGCATGCGAATGCCGCCGACGGTGTACGCATCTTCAACAGTCTCGCCGCTACCGTCTATGCCGACAGTACCGCC GCCCATGCCGATATGCAGGGACGCCGCCTGAAAGCCGTATCGGACGGGTTGGACCACAACGGCACGGGTCTGCGCGTC ATCGCGCAAACCCAACAGGACGGTGGAACGTGGGAACAGGGCGGTGTTGAAGGCAAAATGCGCGGCAGTACCCAAACC GTCGGCATTGCCGCGAAAACCGGCGAAAATACGACAGCAGCCGCCACACTGGGCATGGGACGCAGCACATGGAGCGAA AACAGTGCAAATGCAAAAACCGACAGCATTAGTCTGTTTGCAGGCATACGGCACGATGCGGGCGATATCGGCTATCTC AAAGGCCTGTTCTCCTACGGACGCTACAAAAACAGCATCAGCCGCAGCACCGGTGCGGACGAACATGCGGAAGGCAGC GTCAACGGCACGCTGATGCAGCTGGGCGCACTGGGCGGTGTCAACGTTCCGTTTGCCGCAACGGGAGATTTGACGGTC GAAGGCGGTCTGCGCTACGACCTGCTCAAACAGGATGCATTCGCCGAAAAAGGCAGTGCTTTGGGCTGGAGCGGCAAC AGCCTCACTGAAGGCACGCTGGTCGGACTCGCGGGTCTGAAGCTGTCGCAACCCTTGAGCGATAAAGCCGTCCTGTTT GCAACGGCGGGCGTGGAACGCGACCTGAACGGACGCGACTACACGGTAACGGGCGGCTTTACCGGCGCGACTGCAGCA ACCGGCAAGACGGGGGCACGCAATATGCCGCACACCCGTCTGGTTGCCGGCCTGGGCGCGGATGTCGAATTCGGCAAC GGCTGGAACGGCTTGGCACGTTACAGCTACGCCGGTTCCAAACAGTACGGCAACCACAGCGGACGAGTCGGCGTAGGC TACCGGTTCCTCGAGCACCACCACCACCACCACTGA 1 MATNDDDVKK AATVAIAAAY NNGGEINGFK AGETIYDIDE DGTITKKDAT 51 AADVEADDFK GLGLKKVVTN LTKTVNENKG NVDAKVKAAE SEIEKLTTKL 101 ADTDAALADT DAALDATTNA LNKLGENITT FAEETKTNIV KIDEKLEAVA 151 DTVDKHAEAF NDIADSLDET NTKADEAVKT ANEAKGTAEE TKGNVDAKVK 201 AAETAAGKAE AAAGTANTAA DKAEAVAAKV TDIKADIATN KDNIAKKANS 251 ADVYTREESD SKFVRIDGLN ATTEKLDTBL ASAEKSIADH DTRLNGLDKT 301 VSDLRKETRG GLAEGAALSG LFGPYNVGGS GGGGTSAPDF NAGGTGIGSN 351 SRATTAKSAA VSYAGIKNEM CKDRSHLCAG RDDVAVTDRD AKINAPPPNL 401 HTGDFPNPND AYKNLINLKP AIEAGYTGRG VEVGIVDTGE SVGSISFPEL 451 YGRKEHGYNB NYKNYTAYMR KEAPEDGGGK DIEASFDDEA VIETEAKPTD 501 IRHVKEIGHI DLVSHIIGGR SVDGRPAGGI APDATLHIMN TNDETKNEMM 551 VAAIRNAWVK LGERGVRIVN NSFGTTSRAG TADLFGIANS EEGYRGALLD 601 YSGGDKTDEG IRLMGGSDYG NLSYHIRNKN MLFIFSTGND AGAGPNTYAL 651 LPFYEKDAGK GIITVAGVDR SGEKFKRBHY GEPGTEPLEY GSNHCGITAM 701 WCLSAPYEAS VRFTRTNPIG IAGTSFSAPI VTGTAALLLG KYPWMSNDNL 751 RTTLLTTAGD IGAVGVDSKF GWGLLDAGKA MNGPASFPPG DFTADTKGTS 801 DIAYSFRNDI SGTGGLIKKG GSGLGLHGNN TYTGKTIIEG GSLVLYGNNK 851 SDNRVETKGA LIYNGAASGG SLNSDGIVYL ADTDGSGANE TVHIKGSLGL 901 DGKGTLYTRL GKLLKVDGTA IIGGKLYMSA RGKGAGYLNS TGRRVPFLSA 951 AKIGGDYSFF TNIETDGGLL ASLDSVEKTA GSBGDTLSYY VRRGNAARTA 1001 SAAAHSAPAG LKHAVEGGGS NLENLMVELD ASESSATPET VETAAADRTD 1051 MPGIRPYGAT FRAAAAVGHA NAADGVRIFN SLAATVYADS TAAHADMGGR 1101 RLKAVSDGLD HNGTGLRVIA GTGGDGGTWE GGGVEGKMRG STGTVGIAAK 1151 TGENTTAAAT LGMGRSTWSE NSANAKTDSI SLFAGIRHDA GDIGYLKGLF 1201 SYGRYRNSIS RSTGADEHAE GSVNGTLMGL GALGGVNVPF AATGDLTVEG 1251 GLRYDLLKGD AFAEKGSALG WSGNSLTEGT LVGLAGLKLS GPLSDKAVLF 1301 ATAGVEEDLN GEDYTVTGGF TGATAATGKT GAPNMPHTRL VAGLGADVEF 1351 GNGWNGLARY SYAGSKGYGN HSGRVGVGYR FLEHHHHHH* 961cL-ORF46.1 ATGAAACACTTTCCATCCAAAGTACTGACCACAGCCATCCTTGCCACTTTCTGTAGCGGCGCACTGGCAGCCACAAAC GACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATCAACGGTTTCAAA GCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCCGATGTTGAAGCC GACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAACAAACAAAACGTC GATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGATGCCGCTTTAGCA GATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACATTTGCTGAAGAG ACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCATGCCGAAGCATTC AACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAATGAAGCCAAACAG ACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAAGCCGAAGCTGCC GCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAAGCTGATATCGCT ACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGACAGCAAATTTGTC AGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCCATTGCCGATCAC GATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTTGCAGAACAAGCC GCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTGGATCCGGAGGAGGAGGATCAGATTTGGCAAACGATTCTTTT ATCCGGCAGGTTCTCGACCGTCAGCATTTCGAACCCGACGGGAAATACCACCTATTCGGCAGCAGGGGGGAACTTGCC GAGCGCAGCGGCCATATCGGATTGGGAAAAATACAAAGCCATCAGTTGGGCAACCTGATGATTCAACAGGCGGCCATT AAAGGAAATATCGGCTACATTGTCCGCTTTTCCGATCACGGGCACGAAGTCCATTCCCCCTTCGACAACCATGCCTCA CATTCCGATTCTGATGAAGCCGGTAGTCCCGTTGACGGATTTAGCCTTTACCGCATCCATTGGGACGGATACGAACAC CATCCCGCCGACGGCTATGACGGGCCACAGGGCGGCGGCTATCCCGCTCCCAAAGGCGCGAGGGATATATACAGCTAC GACATAAAAGGCGTTGCCCAAAATATCCGCCTCAACCTGACCGACAACCGCAGCACCGGACAACGGCTTGCCGACCGT TTCCACAATGCCGGTAGTATGCTGACGCAAGGAGTAGGCGACGGATTCAAACGCGCCACCCGATACAGCCCCGAGCTG GACAGATCGGGCAATGCCGCCGAAGCCTTCAACGGCACTGCAGATATCGTTAAAAACATCATCGGCGCGGCAGGAGAA ATTGTCGGCGCAGGCGATGCCGTGCAGGGCATAAGCGAAGGCTCAAACATTGCTGTCATGCACGGCTTGGGTCTGCTT TCCACCGAAAACAAGATGGCGCGCATCAACGATTTGGCAGATATGGCGCAACTCAAAGACTATGCCGCAGCAGCCATC CGCGATTGGGCAGTCCAAAACCCCAATGCCGCACAAGGCATAGAAGCCGTCAGCAATATCTTTATGGCAGCCATCCCC ATCAAAGGGATTGGAGCTGTTCGGGGAAAATACGGCTTGGGCGGCATCACGGCACATCCTATCAAGCGGTCGCAGATG GGCGCGATCGCATTGCCGAAAGGGAAATCCGCCGTCAGCGACAATTTTGCCGATGCGGCATACGCCAAATACCCGTCC CCTTACCATTCCCGAAATATCCGTTCAAACTTGGAGCAGCGTTACGGCAAAGAAAACATCACCTCCTCAACCGTGCCG CCGTCAAACGGCAAAAATGTCAAACTGGCAGACCAACGCCACCCGAAGACAGGCGTACCGTTTGACGGTAAAGGGTTT CCGAATTTTGAGAAGCACGTGAAATATGATACGTAACTCGAG 1 MKHFPSKVLT TAILATFCSG ALAATNDDDV KKAATVAIAA AYNNGGEING 51 FKAGETIYDI DEDGTITKKD ATAADVEADD FKGLGLKKVV TNLTKTVNEN 101 KGNVDAKVKA AESEIEKLTT KLADTDAALA DTDAALDATT NALNKLGENI 151 TTFAEETKTN IVKIDEKLEA VADTVDKHAE APNDIADSLD ETNTKADEAV 201 KThNEAKGTA EETKGNVDAK VKAAETAAGK AEAAAGTANT AADKAEAVAA 251 KVTDIKADIA TNKDNIAKKA NSADVYTREE SDSKFVRIDG LNATTEKLDT 301 RLASAEKSIA DHDThLNGLD KTVSDLRKBT RQGLAEQAAL SGLPGPYNVG 351 GSGGGGSDLA NDSFIRGVLD RGHFEPDGKY HLPGSRGELA ERSGHIGLGK 401 IGSHGLGNLM IQQAAIKGNI GYIVRFSDHG HEVHSPFDNH ASHSDSDHAG 451 SPVDGFSLYR IHWDGYBHHP ADGYDGPGGG GYPAPKGARD IYSYDIKGVA 501 GNIRLNLTDN RSTGQRLADR FHNAGSMLTQ GVGDGFKRAT RYSPELDRSG 551 NAAEAFNGTA DIVKNIIGAA GEIVGAGDAV GGISEGSNIA VMHGLGLLST 601 ENKMARINDL ADMAGLKDYA AAAIRDWAVG NPNAAGGIEA VSNIFMAAIP 651 IKGIGAVRGK YGLGGITAHP IKRSGMGAIA LPKGKSAVSD NFADAAYAKY 701 PSPYHSRNIR SNLEGRYGKE NITSSTVPPS NGKNVKLADG RHPKTGVPFD 751 GKGFPNFEKH VKYDT* 961cL-741 ATGAAACACTTTCCATCCAAAGTACTGACCACAGCCATCCTTGCCACTTTCTGTAGCGGCGCACTGGCAGCCACAAAC GACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATCAACGGTTTCAAA GCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCCGATGTTGAAGCC GACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAACAAACAAAACGTC GATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGATGCCGCTTTAGCA GATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACATTTGCTGAAGAG ACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCATGCCGAAGCATTC AACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAATGAAGCCAAACAG ACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAAGCCGAAGCTGCC GCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAAGCTGATATCGCT ACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGACAGCAAATTTGTC AGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCCATTGCCGATCAC GATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTTGCAGAACAAGCC GCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTGGATCCGGAGGGGGTGGTGTCGCCGCCGACATCGGTGCGGGG CTTGCCGATGCACTAACCGCACCGCTCGACCATAAAGACAAAGGTTTGCAGTCTTTGACGCTGGATCAGTCCGTCAGG AAAAACGAGAAACTGAAGCTGGCGGCACAAGGTGCGGAAAAAACTTATGGAAACGGTGACAGCCTCAATACGGGCAAA TTGAAGAACGACAAGGTCAGCCGTTTCGACTTTATCCGCCAAATCGAAGTGGACGGGCAGCTCATTACCTTGGAGAGT GGAGAGTTCCAAGTATACAAACAAAGCCATTCCGCCTTAACCGCCTTTCAGACCGAGCAAATACAAGATTCGGAGCAT TCCGGGAAGATGGTTGCGAAACGCCAGTTCAGAATCGGCGACATAGCGGGCGAACATACATCTTTTGACAAGCTTCCC GAAGGCGGCAGGGCGACATATCGCGGGACGGCGTTCGGTTCAGACGATGCCGGCGGAAAACTGACCTACACCATAGAT TTCGCCGCCAAGCAGGGAAACGGCAAAATCGAACATTTGAAATCGCCAGAACTCAATGTCGACCTGGCCGCCGCCGAT ATCAAGCCGGATGGAAAACGCCATGCCGTCATCAGCGGTTCCGTCCTTTACAACCAAGCCGAGAAAGGCAGTTACTCC CTCGGTATCTTTGGCGGAAAAGCCCAGGAAGTTGCCGGCAGCGCGGAAGTGAAAACCGTAAACGGCATACGCCATATC GGCCTTGCCGCCAAGCAACTCGAGCACCACCACCACCACCACTGA 1 MKHFPSKVLT TAILATFCSG ALAATNDDDV KKAATVAIAA AYNNGGEING 51 FKAGETIYDI DEDGTITKKD ATAADVEADD FKGLGLKKVV TNLTKTVNEN 101 KGNVDAKVKA AESEIEKLTT KLADTDAALA DTDAALDATT NALNKLGENI 151 TTFAEETKTN IVKIDEKLEA VADTVDKHAB AFNDIADSLD ETNTKADEAV 201 KTANEAKGTA EETKGNVDAK VKAAETAAGK AEAAAGTANT AADKAEAVAA 251 KVTDIKADIA TNKDNLAKKA NSADVYTREE SDSKFVRIDG LNATTEKLDT 301 RLASABKSIA DHDTRLNGLD KTVSDLRKET RQGLAEQAAL SGLFGPYNVG 351 GSGGGGVAAD IGAGLADALT APLDHKDKGL GSLTLDGSVR KNEKLKLAAG 401 GAEKTYGNGD SLNTGKLKND KVSRFDFIRG IEVDGGLITL ESGEFGVYKG 451 SHSALTAPGT EGIGDSEHSG KMVAKRGFRI GDIAGEHTSF DKLPEGGRAT 501 YRGTAPGSDD AGGKLTYTID FAAKGGNGKI EHLKSPELNV DLAAADIKPD 551 GKRHAVISGS VLYNGAEKGS YSLGIFGGKA GEVAGSAEVK TVNGIRHIGL 601 AAKGLEHHHH HH* 961cL-983 ATGAAACACTTTCCATCCAAAGTACTGACCACAGCCATCCTTGCCACTTTCTGTAGCGGCGCACTGGCAGCCACAAAC GACGACGATGTTAAAAAAGCTGCCACTGTGGCCATTGCTGCTGCCTACAACAATGGCCAAGAAATCAACGGTTTCAAA GCTGGAGAGACCATCTACGACATTGATGAAGACGGCACAATTACCAAAAAAGACGCAACTGCAGCCGATGTTGAAGCC GACGACTTTAAAGGTCTGGGTCTGAAAAAAGTCGTGACTAACCTGACCAAAACCGTCAATGAAAACAAACAAAACGTC GATGCCAAAGTAAAAGCTGCAGAATCTGAAATAGAAAAGTTAACAACCAAGTTAGCAGACACTGATGCCGCTTTAGCA GATACTGATGCCGCTCTGGATGCAACCACCAACGCCTTGAATAAATTGGGAGAAAATATAACGACATTTGCTGAAGAG ACTAAGACAAATATCGTAAAAATTGATGAAAAATTAGAAGCCGTGGCTGATACCGTCGACAAGCATGCCGAAGCATTC AACGATATCGCCGATTCATTGGATGAAACCAACACTAAGGCAGACGAAGCCGTCAAAACCGCCAATGAAGCCAAACAG ACGGCCGAAGAAACCAAACAAAACGTCGATGCCAAAGTAAAAGCTGCAGAAACTGCAGCAGGCAAAGCCGAAGCTGCC GCTGGCACAGCTAATACTGCAGCCGACAAGGCCGAAGCTGTCGCTGCAAAAGTTACCGACATCAAAGCTGATATCGCT ACGAACAAAGATAATATTGCTAAAAAAGCAAACAGTGCCGACGTGTACACCAGAGAAGAGTCTGACAGCAAATTTGTC AGAATTGATGGTCTGAACGCTACTACCGAAAAATTGGACACACGCTTGGCTTCTGCTGAAAAATCCATTGCCGATCAC GATACTCGCCTGAACGGTTTGGATAAAACAGTGTCAGACCTGCGCAAAGAAACCCGCCAAGGCCTTGCAGAACAAGCC GCGCTCTCCGGTCTGTTCCAACCTTACAACGTGGGTGGATCCGGCGGAGGCGGCACTTCTGCGCCCGACTTCAATGCA GGCGGTACCGGTATCGGCAGCAACAGCAGAGCAACAACAGCGAAATCAGCAGCAGTATCTTACGCCGGTATCAAGAAC GAAATGTGCAAAGACAGAAGCATGCTCTGTGCCGGTCGGGATGACGTTGCGGTTACAGACAGGGATGCCAAAATCAAT GCCCCCCCCCCGAATCTGCATACCGGAGACTTTCCAAACCCAAATGACGCATACAAGAATTTGATCAACCTCAAACCT GCAATTGAAGCAGGCTATACAGGACGCGGGGTAGAGGTAGGTATCGTCGACACAGGCGAATCCGTCGGCAGCATATCC TTTCCCGAACTGTATGGCAGAAAAGAACACGGCTATAACGAAAATTACAAAAACTATACGGCGTATATGCGGAAGGAA GCGCCTGAAGACGGAGGCGGTAAAGACATTGAAGCTTCTTTCGACGATGAGGCCGTTATAGAGACTGAAGCAAAGCCG ACGGATATCCGCCACGTAAAAGAAATCGGACACATCGATTTGGTCTCCCATATTATTGGCGGGCGTTCCGTGGACGGC AGACCTGCAGGCGGTATTGCGCCCGATGCGACGCTACACATAATGAATACGAATGATGAAACCAAGAACGAAATGATG GTTGCAGCCATCCGCAATGCATGGGTCAAGCTGGGCGAACGTGGCGTGCGCATCGTCAATAACAGTTTTGGAACAACA TCGAGGGCAGGCACTGCCGACCTTTTCCAAATAGCCAATTCGGAGGAGCAGTACCGCCAAGCGTTGCTCGACTATTCC GGCGGTGATAAAACAGACGAGGGTATCCGCCTGATGCAACAGAGCGATTACGGCAACCTGTCCTACCACATCCGTAAT AAAAACATGCTTTTCATCTTTTCGACAGGCAATGACGCACAAGCTCAGCCCAACACATATGCCCTATTGCCATTTTAT GAAAAAGACGCTCAAAAAGGCATTATCACAGTCGCAGGCGTAGACCGCAGTGGAGAAAAGTTCAAACGGGAAATGTAT GGAGAACCGGGTACAGAACCGCTTGAGTATGGCTCCAACCATTGCGGAATTACTGCCATGTGGTGCCTGTCGGCACCC TATGAAGCAAGCGTCCGTTTCACCCGTACAAACCCGATTCAAATTGCCGGAACATCCTTTTCCGCACCCATCGTAACC GGCACGGCGGCTCTGCTGCTGCAGAAATACCCGTGGATGAGCAACGACAACCTGCGTACCACGTTGCTGACGACGGCT CAGGACATCGGTGCAGTCGGCGTGGACAGCAAGTTCGGCTGGGGACTGCTGGATGCGGGTAAGGCCATGAACGGACCC GCGTCCTTTCCGTTCGGCGACTTTACCGCCGATACGAAAGGTACATCCGATATTGCCTACTCCTTCCGTAACGACATT TCAGGCACGGGCGGCCTGATCAAAAAAGGCGGCAGCCAACTGCAACTGCACGGCAACAACACCTATACGGGCAAAACC ATTATCGAAGGCGGTTCGCTGGTGTTGTACGGCAACAACAAATCGGATATGCGCGTCGAAACCAAAGGTGCGCTGATT TATAACGGGGCGGCATCCGGCGGCAGCCTGAACAGCGAGGGCATTGTCTATCTGGCAGATACCGACCAATCCGGCGCA AACGAAACCGTACACATCAAAGGCAGTCTGCAGCTGGACGGCAAAGGTACGCTGTACACACGTTTGGGCAAACTGCTG AAAGTGGACGGTACGGCGATTATCGGCGGCAAGCTGTACATGTCGGCACGCGGCAAGGGGGCAGGCTATCTCAACAGT ACCGGACGACGTGTTCCCTTCCTGAGTGCCGCCAAAATCGGGCAGGATTATTCTTTCTTCACAAACATCGAAACCGAC GGCGGCCTGCTGGCTTCCCTCGACAGCGTCGAAAAAACAGCGGGCAGTGAAGGCGACACGCTGTCCTATTATGTCCGT CGCGGCAATGCGGCACGGACTGCTTCGGCAGCGGCACATTCCGCGCCCGCCGGTCTGAAACACGCCGTAGAACAGGGC GGCAGCAATCTGGAAAACCTGATGGTCGAACTGGATGCCTCCGAATCATCCGCAACACCCGAGACGGTTGAAACTGCG GCAGCCGACCGCACAGATATGCCGGGCATCCGCCCCTACGGCGCAACTTTCCGCGCAGCGGCAGCCGTACAGCATGCG AATGCCGCCGACGGTGTACGCATCTTCAACAGTCTCGCCGCTACCGTCTATGCCGACAGTACCGCCGCCCATGCCGAT ATGCAGGGACGCCGCCTGAAAGCCGTATCGGACGGGTTGGACCACAACGGCACGGGTCTGCGCGTCATCGCGCAAACC CAACAGGACGGTGGAACGTGGGAACAGGGCGGTGTTGAAGGCAAAATGCGCGGCAGTACCCAAACCGTCGGCATTGCC GCGAAAACCGGCGAAAATACGACAGCAGCCGCCACACTGGGCATGGGACGCAGCACATGGAGCGAAAACAGTGCAAAT GCAAAAACCGACAGCATTAGTCTGTTTGCAGGCATACGGCACGATGCGGGCGATATCGGCTATCTCAAAGGCCTGTTC TCCTACGGACGCTACAAAAACAGCATCAGCCGCAGCACCGGTGCGGACGAACATGCGGAAGGCAGCGTCAACGGCACG CTGATGCAGCTGGGCGCACTGGGCGGTGTCAACGTTCCGTTTGCCGCAACGGGAGATTTGACGGTCGAAGGCGGTCTG CGCTACGACCTGCTCAAACAGGATGCATTCGCCGAAAAAGGCAGTGCTTTGGGCTGGAGCGGCAACAGCCTCACTGAA GGCACGCTGGTCGGACTCGCGGGTCTGAAGCTGTCGCAACCCTTGAGCGATAAAGCCGTCCTGTTTGCAACGGCGGGC GTGGAACGCGACCTGAACGGACGCGACTACACGGTAACGGGCGGCTTTACCGGCGCGACTGCAGCAACCGGCAAGACG GGGGCACGCAATATGCCGCACACCCGTCTGGTTGCCGGCCTGGGCGCGGATGTCGAATTCGGCAACGGCTGGAACGGC TTGGCACGTTACAGCTACGCCGGTTCCAAACAGTACGGCAACCACAGCGGACGAGTCGGCGTAGGCTACCGGTTCTGA CTCGAG 1 MKHFPSKVLT TAILATFCSG MJAATNDDDV KKAATVAIAA AYNNGGEING 51 FKAGETIYDI DEDGTITKKD ATAADVEADD FKGLGLKKVV TNLTKTVNEN 101 KGNVDAKVKA AESEIEKLTT KLADTDAALA DTDAALDATT NALNKLGENI 151 TTFAEETKTN IVKIDEKLEA VADTVDKHAB AFNDIADSLD ETNTKADEAV 201 KTANEAKGTA EETKGNVDAK VKAAETAAGK AEAAAGTANT AADKAEAVAA 251 KVTDIKADIA TNKDNIAKKA NSADVYTREE SDSKFVRIDG LNATTEKLDT 301 RLASAEKSIA DHDTRLNGLD KTVSDLRKET RGGLAEGAAL SGLFGPYNVG 351 GSGGGGTSAP DFNAGGTGIG SNSRATTAKS AAVSYAGIKN EMCKDRSMLC 401 AGRDDVAVTD RDAKINAPPP NLHTGDFPNP NDAYKNLINL KPAIEAGYTG 451 RGVEVGIVDT GESVGSISFP ELYGRKEHGY NENYKNYTAY MRKEAPEDGG 501 GKDIEASFDD EAVIETEAKP TDIRHVKEIG HIDLVSHIIG GRSVDGRPAG 551 GIAPDATLHI MNTNDETKNE MMVAAIRNAW VKLGERGVRI VNNSFGTTSR 601 AGTADLPGIA NSEEGYRGAL LDYSGGDKTD EGIRLHGGSD YGNLSYHIRN 651 KNMLFIFSTG NDAGAGPNTY ALLPFYEKDA GKGIITVAGV DRSGEKFKRE 701 MYGEPGTEPL EYGSNHCGIT AMWCLSAPYE ASVRPTRTNP IGIAGTSFSA 751 PIVTGTAALL LGKYPWMSND NLRTTLLTTA GDIGAVGVDS KFGWGLLDAG 801 KAMNGPASFF FGDFTADTKG TSDIAYSFRN DISGTGGLIK KGGSGLGLHG 851 NNTYTGKTII EGGSLVLYGN NKSDMRVETK GALIYNGAAS GGSLNSDGIV 901 YLADTDGSGA NETVHIKGSL GLDGKGTLYT RLGKLLKVDG TAIIGGKLYM 951 SARGKGAGYL NSTGRRVPFL SAAKIGQDYS FFTNIETDGG LLASLDSVEK 1001 TAGSEGDTLS YYVRRGNAAR TASAAAHSAP AGLKHAVEGG GSNLENLMVE 1051 LDASESSATP ETVETAAADR TDMPGIRPYG ATFRAAAAVG HANAADGVRI 1101 FNSLAATVYA DSTAAHADMG GRRLKAVSDG LDHNGTGLRV IAGTGGDGGT 1151 WEGGGVEGKM RGSTGTVGIA AKTGENTTAA ATLGMGRSTW SENSANAKTD 1201 SISLFAGIRH DAGDIGYLKG LFSYGRYKNS ISRSTGADEH AEGSVNGTLM 1251 GLGALGGVNV PFAATGDLTV EGGLRYDLLK GDAFAEKGSA LGWSGNSLTB 1301 GTLVGLAGLK LSGPLSDKAV LFATAGVERD LNGRDYTVTG GFTGATAATG 1351 KTGAR~MPRT RLVAGLGADV EFGNGWNGLA RYSYAGSKGY GNHSGRVGVG 1401 YRF*

[0079] It will be understood that the invention has been described by way of example only and modifications may be made whilst remaining within the scope and spirit of the invention. For instance, the use of proteins from other strains is envisaged [e.g. see WO00/66741 for polymorphic sequences for ORF4, ORF40, ORF46, 225, 235, 287, 519, 726, 919 and 953].

[0080] Experimental Details

[0081] Cloning Strategy and Oligonucleonide Design

[0082] Genes coding for antigens of interest were amplified by PCR, using oligonucleotides designed on the basis of the genomic sequence of N. meningitidis B MC58. Genomic DNA from strain 2996 was always used as a template in PCR reactions, unless otherwise specified, and the amplified fragments were cloned in the expression vector pET21b+(Novagen) to express the protein as C-terminal His-tagged product, or in pET-24b+(Novagen) to express the protein in ‘untagged’ form (e.g. &Dgr;G 287K).

[0083] Where a protein was expressed without a fusion partner and with its own leader peptide (if present), amplification of the open reading frame (ATG to STOP codons) was performed.

[0084] Where a protein was expressed in ‘untagged’ form, the leader peptide was omitted by designing the 5′-end amplification primer downstream from the predicted leader sequence.

[0085] The melting temperature of the primers used in PCR depended on the number and type of hybridising nucleotides in the whole primer, and was determined using the formulae: 25 Tm1 = 4 (G + C) + 2 (A + T) (tail excluded) Tm2 = 64.9 + 0.41 (% GC) − 600/N (whole primer)

[0086] The melting temperatures of the selected oligonucleotides were usually 65-70° C. for the whole oligo and 50-60° C. for the hybridising region alone.

[0087] Oligonucleotides were synthesised using a Perkin Elmer 394 DNA/RNA Synthesizer, eluted from the columns in 2.0 ml NH4OH, and deprotected by 5 hours incubation at 56° C. The oligos were precipitated by addition of 0.3M Na-Acetate and 2 volumes ethanol. The samples were centrifuged and the pellets resuspended in water. 26 Restriction Sequences site fu (961)- Fwd CGCGGATCC-GGAGGGGGTGGTGTCG BamHI 741(MC58)-His Rev CCCGCTCGAG-TTGCTTGGCGGCAAGGC XhoI fu (961)-983-His Fwd CGCGGATCC-GGCGGAGGCGGCACTT BamHI Rev CCCGCTCGAG-GAACCGGTAGCCTACG XhoI fu (961)-Orf46.1- Fwd CGCGGATCCGGTGGTGGTGGT- BamHI His TCAGATTTGGCAAACGATTC Rev CCCGCTCGAG-CGTATCATATTTCACGTGC XhoI fu (961 c-L)- Fwd CGCGGATCC-GGAGGGGGTGGTGTCG BamHI 741(MC58) Rev CCCGCTCGAG-TTATTGCTTGGCGGCAAG XhoI fu (961c-L)-983 Fwd CGCGGATCC-GGCGGAGGCGGCACTT BamHI Rev CCCGCTCGAG-TCAGAACCGGTAGCCTAC XhoI fu (961c-L)- Fwd CGCGGATCCGGTGGTGGTGGT- BamHI Orf46.1 TCAGATTTGGCAAACGATTC Rev CCCGCTCGAG-TTACGTATCATATTTCACGTGC XhoI fu-(&Dgr;G287)-919- Fwd CGCGGATCCGGTGGTGGTGGT- BamHI His CAAAGCAAGAGCATCCAAACC Rev CCCAAGCTT-TTCGGGCGGTATTCGGGCTTC HindIII fu-(&Dgr;G287)-953- Fwd CGCGGATCCGGTGGTGGTGGT- BamHI His GCCACCTACAAAGTGGAC Rev GCCCAAGCTT-TTGTTTGGCTGCCTCGAT HindIII fu-(&Dgr;G287)-961- Fwd CGCGGATCCGGTGGTGGTGGT-ACAAGCGACGACG BamHI His Rev GCCCAAGCTT-CCACTCGTAATTGACGCC HindIII fu-(&Dgr;G287)- Fwd CGCGGATCCGGTGGTGGTGGT- BamHI Orf46.1-His TCAGATTTGGCAAACGATTC Rev CCCAAGCTT-CGTATCATATTTCACGTGC HindIII fu-(&Dgr;G287-919)- Fwd CCCAAGCTTGGTGGTGGTGGTGGT- HindIII Orf46.1-His TCAGATTTGGCAAACGATTC Rev CCCGCTCGAG-CGTATCATATTTCACGTGC XhoI fu-(&Dgr;G287- Fwd CCCAAGCTTGGTGGTGGTGGTGGT- HindIII Orf46.1)-919-His CAAAGCAAGAGCATCCAAACC Rev CCCGCTCGAG-CGGGCGGTATTCGGGCTT XhoI fu &Dgr;G287(394.98)- Fwd CGCGGATCCGCTAGC-CCCGATGTTAAATCGGC NheI . . . Rev CGGGGATCC-ATCCTGCTCTTTTTTGCCGG BamHI fu Orf1-(Orf46.1)- Fwd CGCGGATCCGCTAGC-GGACACACTTATTTCGGCATC NheI His Rev CGCGGATCC-CCAGCGGTAGCCTAATTTGAT fu (Orf1)-Orf46.1- Fwd CGCGGATCCGGTGGTGGTGGT- BamHI His TCAGATTTGGCAAACGATTC Rev CCCAAGCTT-CGTATCATATTTCACGTGC HindIII fu (919)-Orf46.1- Fwd1 GCGGCGTCGACGGTGGCGGAGGCACTGGATCCTCAG SalI His Fwd2 GGAGGCACTGGATCCTCAGATTTGGCAAACGATTC Rev CCCGCTCGAG-CGTATCATATTTCACGTGC XhoI Fu (orf46)-287-His Fwd CGGGGATCCGGGGGCGGCGGTGGCG BamHI Rev CCCAAGCTTATCCTGCTCTTTTTTGCCGGC HindIII Fu (orf46)-919-His Fwd CGCGGATCCGGTGGTGGTGGTCAAAGCAAGAGCATCCAAACC BamHI Rev CCCAAGCTTCGGGCGGTATTCGGGCTTC HindIII Fu (orf46-919)- Fwd CCCCAAGCTTGGGGGCGGCGGTGGCG HindIII 287-His Rev CCCGCTCGAGATCCTGCTCTTTTTTGCCGGC XhoI Fu (orf46-287)- Fwd CCCAAGCTTGGTGGTGGTGGTGGTCAAAGCAAGAGCATCCAAACC HindIII 919-His Rev CCCGCTCGAGCGGGCGGTATTCGGGCTT XhoI (&Dgr;G741)-961c-His Fwd1 GGAGGCACTGGATCCGCAGCCACAAACGACGACGA XhoI Fwd2 GCGGCCTCGAG-GGTGGCGGAGGCACTGGATCCGCAG Rev CCCGCTCGAG-ACCCAGCTTGTAAGGTTG XhoI (&Dgr;G741)-961-His Fwd1 GGAGGCACTGGATCCGCAGCCACAAACGACGACGA XhoI Fwd2 GCGGCCTCGAG-GGTGGCGGAGGCACTGGATCCGCAG Rev CCCGCTCGAG-CCACTCGTAATTGACGCC XhoI (&Dgr;G741)-983-His Fwd GCGGCCTCGAG- XhoI GGATCCGGCGGAGGCGGCACTTCTGCG Rev CCCGCTCGAG-GAACCGGTAGCCTACG XhoI (&Dgr;G741)-orf46.1- Fwd1 GGAGGCACTGGATCCTCAGATTTGGCAAACGATTC SalI His Fwd2 GCGGCGTCGACGGTGGCGGAGGCACTGGATCCTCAGA Rev CCCGCTCGAG-CGTATCATATTTCACGTGC XhoI (&Dgr;G983)- Fwd GCGGCCTCGAG-GGATCCGGAGGGGGTGGTGTCGCC XhoI 741(MC58)-His Rev CCCGCTCGAG-TTGCTTGGCGGCAAG XhoI (&Dgr;G983)-961c-His Fwd1 GGAGGCACTGGATCCGCAGCCACAAACGACGACGA XhoI Fwd2 GCGGCCTCGAG-GGTGGCGGAGGCACTGGATCCGCAG Rev CCCGCTCGAG-ACCCAGCTTGTAAGGTTG XhoI (&Dgr;G983)-961-His Fwd1 GGAGGCACTGGATCCGCAGCCACAAACGACGACGA XhoI Fwd2 GCGGCCTCGAG-GGTGGCGGAGGCACTGGATCCGCAG Rev CCCGCTCGAG-CCACTCGTAATTGACGCC XhoI (&Dgr;G983)-Orf46.1- Fwd1 GGAGGCACTGGATCCTCAGATTTGGCAAACGATTC SalI His Fwd2 GCGGCGTCGACGGTGGCGGAGGCACTGGATCCTCAGA Rev CCCGCTCGAG-CGTATCATATTTCACGTGC XhoI *This primer was used as a Reverse primer for all the C terminal fusions of 287 to the His-tag. §Forward primers used in combination with the 287-His Reverse primer. NB - All PCR reactions use strain 2996 unless otherwise specified (e.g. strain MC58)

[0088] In all constructs starting with an ATG not followed by a unique NheI site, the ATG codon is part of the NdeI site used for cloning. The constructs made using NheI as a cloning site at the 5′ end (e.g. all those containing 287 at the N-terminus) have two additional codons (GCT AGC) fused to the coding sequence of the antigen.

[0089] Preparation of Chromosomal DNA Templates

[0090] N. meningitidis strains 2996, MC58, 394.98, 1000 and BZ232 (and others) were grown to exponential phase in 100 ml of GC medium, harvested by centrifugation, and resuspended in 5 ml buffer (20% w/v sucrose, 50 mM Tris-HCl, 50 mM EDTA, pH8). After 10 minutes incubation on ice, the bacteria were lysed by adding 10 ml of lysis solution (50 mM NaCl, 1% Na-Sarkosyl, 50 &mgr;g/ml Proteinase K), and the suspension incubated at 37° C. for 2 hours. Two phenol extractions (equilibrated to pH 8) and one CHCl3/isoamylalcohol (24:1) extraction were performed. DNA was precipitated by addition of 0.3M sodium acetate and 2 volumes of ethanol, and collected by centrifugation. The pellet was washed once with 70%(v/v) ethanol and redissolved in 4.0 ml TE buffer (10 mM Tris-HCl, 1 mM EDTA, pH 8.0). The DNA concentration was measured by reading OD260.

[0091] PCR Amplfication

[0092] The standard PCR protocol was as follows: 200 ng of genomic DNA from 2996, MC581000, or BZ232 strains or long of plasmid DNA preparation of recombinant clones were used as template in the presence of 40 &mgr;M of each oligonucletide primer, 400-800 &mgr;M dNTPs solution, 1× PCR buffer (including 1.5 mM MgCl2), 2.5 units TaqI DNA polymerase (using Perkin-Elmer AmpliTaQ, Boerhingher Mannheim Expand™ Long Template).

[0093] After a preliminary 3 minute incubation of the whole mix at 95° C., each sample underwent a two-step amplification: the first 5 cycles were performed using the hybridisation temperature that excluded the restriction enzyme tail of the primer (Tm1). This was followed by 30 cycles according to the hybridisation temperature calculated for the whole length oligos (Tm2). Elongation times, performed at 68° C. or 72° C., varied according to the length of the Orf to be amplified. In the case of Orf1 the elongation time, starting from 3 minutes, was increased by 15 seconds each cycle. The cycles were completed with a 10 minute extension step at 72° C.

[0094] The amplified DNA was either loaded directly on a 1% agarose gel. The DNA fragment corresponding to the band of correct size was purified from the gel using the Qiagen Gel Extraction Kit, following the manufacturer's protocol.

[0095] Digestion of PCR Fragments and of the Cloning Vectors

[0096] The purified DNA corresponding to the amplified fragment was digested with the appropriate restriction enzymes for cloning into pET-21b+, pET22b+or pET-24b+. Digested fragments were purified using the QIAquick PCR purification kit (following the manufacturer's instructions) and eluted with either H2O or 10 mM Tris, pH 8.5. Plasmid vectors were digested with the appropriate restriction enzymes, loaded onto a 1.0% agarose gel and the band corresponding to the digested vector purified using the Qiagen QIAquick Gel Extraction Kit.

[0097] Cloning

[0098] The fragments corresponding to each gene, previously digested and purified, were ligated into pET21b+, pET22b+ or pET-24b+. A molar ratio of 3:1 fragment/vector was used with T4 DNA ligase in the ligation buffer supplied by the manufacturer.

[0099] Recombinant plasmid was transformed into competent E. coli DH5 or HB101 by incubating the ligase reaction solution and bacteria for 40 minutes on ice, then at 37° C. for 3 minutes. This was followed by the addition of 800 &mgr;l LB broth and incubation at 37° C. for 20 minutes. The cells were centrifuged at maximum speed in an Eppendorf microfuge, resuspended in approximately 200 &mgr;l of the supernatant and plated onto LB ampicillin (100 mg/ml) agar.

[0100] Screening for recombinant clones was performed by growing randomly selected colonies overnight at 37° C. in 4.0 ml of LB broth +100 &mgr;g/ml ampicillln. Cells were pelleted and plasmid DNA extracted using the Qiagen QIAprep Spin Miniprep Kit, following the manufacturer's instructions. Approximately 1 &mgr;g of each individual miniprep was digested with the appropriate restriction enzymes and the digest loaded onto a 1-1.5% agarose gel (depending on the expected insert size), in parallel with the molecular weight marker (1 kb DNA Ladder, GIBCO). Positive clones were selected on the basis of the size of insert.

[0101] Expression

[0102] After cloning each gene into the expression vector, recombinant plasmids were transformed into E. coli strains suitable for expression of the recombinant protein. 1 &mgr;l of each construct was used to transform E. coli BL21-DE3 as described above. Single recombinant colonies were inoculated into 2 ml LB+Amp (100 &mgr;g/ml), incubated at 37° C. overnight, then diluted 1:30 in 20 ml of LB+Amp (100 &mgr;g/ml) in 100 ml flasks, to give an OD600 between 0.1 and 0.2. The flasks were incubated at 30° C. or at 37° C. in a gyratory water bath shaker until OD600 indicated exponential growth suitable for induction of expression (0.40.8 OD). Protein expression was induced by addition of 11.0 mM IPTG. After 3 hours incubation at 30° C. or 37° C. the OD600 was measured and expression examined. 1.0 ml of each sample was centrifuged in a microfuge, the pellet resuspended in PBS and analysed by SDS-PAGE and Coomassie Blue staining.

[0103] Purification of His-tagged Proteins

[0104] Various forms of 287 were cloned from strains 2996 and MC58. They were constructed with a C-terminus His-tagged fusion and included a mature form (aa 18-427), constructs with deletions (&Dgr;1, &Dgr;2, &Dgr;3 and &Dgr;4) and clones composed of either B or C domains. For each clone purified as a His-fusion, a single colony was streaked and grown overnight at 37° C. on a LB/Amp (100 &mgr;g/ml) agar plate. An isolated colony from this plate was inoculated into 20 ml of LB/Amp (100 &mgr;g/ml) liquid medium and grown overnight at 37° C. with shaking. The overnight culture was diluted 1:30 into 1.0 L LB/Amp (100 &mgr;g/ml) liquid medium and allowed to grow at the optimal temperature (30 or 37° C.) until the OD550 reached 0.6-0.8. Expression of recombinant protein was induced by addition of IPTG (final concentration 1.0 mM) and the culture incubated for a further 3 hours. Bacteria were harvested by centrifugation at 8000 g for 15 min at 4° C. The bacterial pellet was resuspended in 7.5 ml of either (i) cold buffer A (300 mM NaCl, 50 mM phosphate buffer, 10 mM imidazole, pH 8.0) for soluble proteins or (ii) buffer B (10 mM Tris-HCl, 100 mM phosphate buffer, pH 8.8 and, optionally, 8M urea) for insoluble proteins. Proteins purified in a soluble form included 287-His, &Dgr;1, &Dgr;2, &Dgr;3 and &Dgr;4287-His, &Dgr;4287MC58-His, 287c-His and 287cMC58-His. Protein 287bMC58-His was insoluble and purified accordingly. Cells were disrupted by sonication on ice four times for 30 sec at 40 W using a Branson sonifier 450 and centrifuged at 13000 g for 30 min at 4° C. For insoluble proteins, pellets were resuspended in 2.0 ml buffer C (6 M guanidine hydrochloride, 100 mM phosphate buffer, 10 mM Tris-HCl, pH 7.5 and treated with 10 passes of a Dounce homogenizer. The homogenate was centrifuged at 13000 g for 30 min and the supernatant retained. Supernatants for both soluble and insoluble preparations were mixed with 150 &mgr;l Ni2+-resin (previously equilibrated with either buffer A or buffer B, as appropriate) and incubated at room temperature with gentle agitation for 30 min. The resin was Chelating Sepharose Fast Flow (Pharmacia), prepared according to the manufacturer's protocol. The batch-wise preparation was centrifuged at 700 g for 5 min at 4° C. and the supernatant discarded. The resin was washed twice (batch-wise) with 10 ml buffer A or B for 10 min, resuspended in 1.0 ml buffer A or B and loaded onto a disposable column. The resin continued to be washed with either (i) buffer A at 4° C. or (id) buffer B at room temperature, until the OD280 of the flow-through reached 0.02-0.01. The resin was further washed with either (i) cold buffer C (300 mM NaCl, 50 mM phosphate buffer, 20 mM imidazole, pH 8.0) or (ii) buffer D (10 mM Tris-HCl, 100 mM phosphate buffer, pH 6.3 and, optionally, 8M urea) until OD280 of the flow-through reached 0.02-0.01. The His-fusion protein was eluted by addition of 700 &mgr;l of either (i) cold elution buffer A (300 mM NaCl, 50 mM phosphate buffer, 250 mM imidazole, pH 8.0) or (ii) elution buffer B (10 mM Tris-HCl, 100 mM phosphate buffer, pH 4.5 and, optionally, 8M urea) and fractions collected until the OD280 indicated all the recombinant protein was obtained. 20 &mgr;l aliquots of each elution fraction were analysed by SDS-PAGE. Protein concentrations were estimated using the Bradford assay.

[0105] Renaturation of Denatured His-fusion Proteins

[0106] Denaturation was required to solubilize 287bMC8, so a renaturation step was employed prior to immunisation. Glycerol was added to the denatured fractions obtained above to give a final concentration of 10% v/v. The proteins were diluted to 200 &mgr;g/ml using dialysis buffer I (10% v/v glycerol, 0.5M arginine, 50 mM phosphate buffer, 5.0 mM reduced glutathione, 0.5 mM oxidised glutathione, 2.0M urea, pH 8.8) and dialysed against the same buffer for 12-14 hours at 4° C. Further dialysis was performed with buffer II (10% v/v glycerol, 0.5M arginine, 50 mM phosphate buffer, 5.0 mM reduced glutathione, 0.5 mM oxidised glutathione, pH 8.8) for 12-14 hours at 4° C. Protein concentration was estimated using the formula:

Protein (mg/ml)=(1.55×OD280)−(0.76×OD260)

[0107] Immunization

[0108] Balb/C mice were immunized with antigens on days 0, 21 and 35 and sera analyzed at day 49.

[0109] Sera Analysis—ELISA

[0110] The acapsulated MenB M7 and the capsulated strains were plated on chocolate agar plates and incubated overnight at 37° C. with 5% CO2. Bacterial colonies were collected from the agar plates using a sterile dracon swab and inoculated into Mueller-Hinton Broth (Difco) containing 0.25% glucose. Bacterial growth was monitored every 30 minutes by following OD620. The bacteria were let to grow until the OD reached the value of 0.4-0.5. The culture was centrifuged for 10 minutes at 4000 rpm. The supernatant was discarded and bacteria were washed twice with PBS, resuspended in PBS containing 0.025% formaldehyde, and incubated for 1 hour at 37° C. and then overnight at 4° C. with stirring. 100 &mgr;l bacterial cells were added to each well of a 96 well Greiner plate and incubated overnight at 4° C. The wells were then washed three times with PBT washing buffer (0.1% Tween-20 in PBS). 200 &mgr;l of saturation buffer (2.7% polyvinylpyrrolidone 10 in water) was added to each well and the plates incubated for 2 hours at 37° C. Wells were washed three times with PBT. 200 &mgr;l of diluted sera (Dilution buffer: 1% BSA, 0.1% Tween-20, 0.1% NaN3 in PBS) were added to each well and the plates incubated for 2 hours at 37° C. Wells were washed three times with PBT. 100 &mgr;l of HRP-conjugated rabbit anti-mouse (Dako) serum diluted 1:2000 in dilution buffer were added to each well and the plates were incubated for 90 minutes at 37° C. Wells were washed three times with PBT buffer. 100 &mgr;l of substrate buffer for HRP (25 ml of citrate buffer pH5, 10 mg of O-phenildiamine and 1011 of H2O2) were added to each well and the plates were left at room temperature for 20 minutes. 100 &mgr;l 12.5% H2SO4 was added to each well and OD490 was followed. The ELISA titers were calculated abitrarely as the dilution of sera which gave an OD490 value of 0.4 above the level of preimmune sera. The ELISA was considered positive when the dilution of sera with OD490 of 0.4 was higher than 1:400.

[0111] Sera Analysis—FACS Scan Bacteria Binding Assay

[0112] The acapsulated MenB M7 strain was plated on chocolate agar plates and incubated overnight at 37° C. with 5% CO2. Bacterial colonies were collected from the agar plates using a sterile dracon swab and inoculated into 4 tubes containing 8 ml each Mueller-Hinton Broth (Difco) containing 0.25% glucose. Bacterial growth was monitored every 30 minutes by following OD620. The bacteria were let to grow until the OD reached the value of 0.35-0.5. The culture was centrifuged for 10 minutes at 4000 rpm. The supernatant was discarded and the pellet was resuspended in blocking buffer (1% BSA in PBS, 0.4% NaN3) and centrifuged for 5 minutes at 4000 rpm. Cells were resuspended in blocking buffer to reach OD620 of 0.05. 100 &mgr;l bacterial cells were added to each well of a Costar 96 well plate. 100 &mgr;l of diluted (1:100, 1:200, 1:400) sera (in blocking buffer) were added to each well and plates incubated for 2 hours at 4° C. Cells were centrifuged for 5 minutes at 400 rpm, the supernatant aspirated and cells washed by addition of 200 &mgr;l/well of blocking buffer in each well. 100 &mgr;l of R-Phicoerytrin conjugated F(ab)2 goat anti-mouse, diluted 1:100, was added to each well and plates incubated for 1 hour at 4° C. Cells were spun down by centrifugation at 4000 rpm for 5 minutes and washed by addition of 200 &mgr;l/well of blocking buffer. The supernatant was aspirated and cells resuspended in 200 &mgr;l/well of PBS, 0.25% formaldehyde. Samples were transferred to FACScan tubes and read. The condition for FACScan (Laser Power 15 mW) setting were: FL2 on; FSC—H threshold:92; FSC PMT Voltage: E 01; SSC PMr: 474; Amp. Gains 6.1; FL-2 PMT: 586; compensation values: 0.

[0113] Sera Analysis—Bactericidal Assay

[0114] N. meningitidis strain 2996 was grown overnight at 37° C. on chocolate agar plates (starting from a frozen stock) with 5% CO2. Colonies were collected and used to inoculate 7 ml Mueller-Hinton broth, containing 0.25% glucose to reach an OD620 of 0.05-0.08. The culture was incubated for approximately 1.5 hours at 37 degrees with shacking until the OD620 reached the value of 0.23-0.24. Bacteria were diluted in 50 mM Phosphate buffer pH 7.2 containing 10 mM MgCl2, 10 mM CaCl2 and 0.5% (w/v) BSA (assay buffer) at the working dilution of 105 CFU/ml. The total volume of the final reaction mixture was 50 &mgr;l with 25 &mgr;l of serial two fold dilution of test serum, 12.5 &mgr;l of bacteria at the working dilution, 12.5 &mgr;l of baby rabbit complement (final concentration 25%).

[0115] Controls included bacteria incubated with complement serum, immune sera incubated with bacteria and with complement inactivated by heating at 56° C. for 30′. Immediately after the addition of the baby rabbit complement, 10 &mgr;l of the controls were plated on Mueller-Hinton agar plates using the tilt method (time 0). The 96-wells plate was incubated for 1 hour at 37° C. with rotation. 7 &mgr;l of each sample were plated on Mueller-Hinton agar plates as spots, whereas 10 &mgr;l of the controls were plated on Mueller-Hinton agar plates using the tilt method (time 1). Agar plates were incubated for 18 hours at 37 degrees and the colonies corresponding to time 0 and time 1 were counted.

[0116] Sera Analysis—Western Blots

[0117] Purified proteins (500 ng/lane), outer membrane vesicles (5 &mgr;g) and total cell extracts (25 &mgr;g) derived from MenB strain 2996 were loaded onto a 12% SDS-polyacrylamide gel and transferred to a nitrocellulose membrane. The transfer was performed for 2 hours at 150 mA at 4° C., using transfer buffer (0.3% Tris base, 1.44% glycine, 20% (v/v) methanol). The membrane was saturated by overnight incubation at 4° C. in saturation buffer (10% skimmed milk, 0.1% Triton X100 in PBS). The membrane was washed twice with washing buffer (3% skimmed milk, 0.1% Triton X100 in PBS) and incubated for 2 hours at 37° C. with mice sera diluted 1:200 in washing buffer. The membrane was washed twice and incubated for 90 minutes with a 1:2000 dilution of horseradish peroxidase labelled anti-mouse Ig. The membrane was washed twice with 0.1% Triton X100 in PBS and developed with the Opti-4CN Substrate Kit (Bio-Rad). The reaction was stopped by adding water.

[0118] The OMVs were prepared as follows: N. meningitidis strain 2996 was grown overnight at 37 degrees with 5% CO2 on 5 GC plates, harvested with a loop and resuspended in 10 ml of 20 mM Tris-HCl pH 7.5, 2 mM EDTA. Heat inactivation was performed at 56° C. for 45 minutes and the bacteria disrupted by sonication for 5 minutes on ice (50% duty cycle, 50% output, Branson sonifier 3 mm microtip). Unbroken cells were removed by centrifugation at 5000 g for 10 minutes, the supernatant containing the total cell envelope fraction recovered and further centrifuged overnight at 50000 g at the temperature of 4° C. The pellet containing the membranes was resuspended in 2% sarkosyl, 20 mM Tris-HCl pH 7.5, 2 mM EDTA and incubated at room temperature for 20 minutes to solubilise the inner membranes. The suspension was centrifuged at 10000 g for 10 minutes to remove aggregates, the supernatant was further centrifuged at 50000 g for 3 hours. The pellet, containing the outer membranes was washed in PBS and resuspended in the same buffer. Protein concentration was measured by the D.C. Bio-Rad Protein assay (Modified Lowry method), using BSA as a standard.

[0119] Total cell extracts were prepared as follows: N. meningitidis strain 2996 was grown overnight on a GC plate, harvested with a loop and resuspended in 1 ml of 20 mM Tris-HCl. Heat inactivation was performed at 56° C. for 30 minutes.

Claims

1. A method for the simultaneous heterologous expression of two or more proteins of the invention, in which (a) two or more proteins of the invention are fused.

2. The method of claim 23, in which the two or more proteins are: (a) 919 and 287; (b) 953 and 287; (c) 287 and ORF46.1; (d) ORF1 and ORF46.1; (e) 919 and ORF46.1; (f) ORF46.1, 287 and 919; (g) 919 and 519; and (h) ORF97 and 225.

3. The method of claim 24, in which 287 is at the C-terminal end of protein (a), (b) or (c).

4. The method of any preceding claim, in which the expression is in an E. coli host.

5. A protein expressed by the method of any preceding claim.

6. A hybrid protein of formula NH2-A-B-COOH, wherein A and B are different Neisserial proteins.

7. The protein of claim 6, wherein A and B are each selected from orf1, orf4, orf25, orf40, orf46, orf83, 233, 287, 2921, 564, 687, 741, 907, 919, 953, 961 and 983.

8. The protein of claim 7, wherein A and B are each selected from ORF46.1, 287, 741, 919, 953, 961 and 983.

9. The protein of claim 8, wherein at least one of said ORF46.1, 287, 741, 919, 953, 961 and 983 is used in essentially full-length form

10. The protein of claim 8 or claim 9, wherein at least one of said ORF46.1, 287, 741, 919, 953, 961 and 983 has a deletion.

11. The protein of claim 10, wherein A and/or B has a poly-glycine deletion (‘&Dgr;G’).

12. The protein of claim 11, wherein A and/or B is &Dgr;G-287, &Dgr;GThp2, &Dgr;G741, or &Dgr;G983.

13. The protein of claim 10, wherein A and/or B is a truncated protein.

14. The protein of claim 13, wherein A and/or B is &Dgr;1-287, &Dgr;2-287, &Dgr;3-287 or &Dgr;4-287.

15. The protein of claim 10, wherein a domain of A and/or B is deleted.

16. The protein of claim 15, wherein A and/or B is 287B, 287C, 287BC, ORF461-433, ORF46433-608, ORF46, or 961c.

17. The protein of claim 6, wherein A and B are: (a) 919 and 287; (b) 953 and 287; (c) 287 and ORF46.1; (d) ORF1 and ORF46.1; (e) 919 and ORF46.1; (f) ORF46.1 and 919; (g) 919 and 519; or (h) ORF97 and 225.

18. The protein of claim 17, wherein the protein is &Dgr;G287-919, &Dgr;G287-953, &Dgr;G287-961, &Dgr;G983-ORF46.1, &Dgr;G983-741, &Dgr;G983-961, &Dgr;G983-961C, &Dgr;G741-961, &Dgr;G741-961C, &Dgr;G741-983, &Dgr;G741-ORF46.1, ORF46.1-741, ORF46.1-961, ORF46.1-961C, 961-ORF46.1, 961-741, 961-983, 961C-ORF46.1, 961C-741, 961C-983, 961CL-ORF46.1, 961CL-741, or 961CL-983.

19. The protein of claim 8, wherein A or B is 287.

20. The protein of claim 19, wherein B is 287

21. The protein of claim 19, wherein A is &Dgr;G-287

22. The protein of claim 21, wherein B is ORF46, 919, 953 or 961.

23. The protein of any one of claims 19 to 22, wherein 287 is from strain 2996 or 394/98.

24. The protein of claim 8, wherein A is 961.

25. The protein of any one of claims 6 to 24, wherein A and B are from the same strain.

26. The protein of any one of claims 6 to 24, wherein A and B are joined directly

27. The protein of any one of claims 6 to 24, wherein A and B are joined via a linker peptide.

28. The protein of claim 27, wherein the linker peptide is a poly-glycine linker, with the proviso that B is not a AG protein.