人工重组的h7亚型流感病毒及其制备方法和应用的制作方法
专利名称：人工重组的h7亚型流感病毒及其制备方法和应用的制作方法
技术领域：
本发明涉及一株人工重组的流感病毒，该病毒的制备和应用。
背景技术：
禽流感病毒(avian influenza virus，AIV)是禽类及一些哺乳动物的呼吸系统疾病的重要病原。AIV在分类学上属于病毒界(Vira)----正粘病毒科(Orthomyxoviridae)----流感病毒属(Influenza Virus A and B)----禽流感病毒(avian influenza virus)。
根据表面蛋白抗原性不同，禽流感病毒(avian influenza virus，AIV)被分为15个血凝素(HA)和9个神经氨酸酶(NA)亚型。H7亚型AIV 1878年首次在意大利出现，该“真性鸡瘟”的病原是A/Chicken/Brescia/1902(H7N7)，其是禽流感历史上第一次文字记载的HPAIV，后来人们又分离到多株H7亚型AIV，神经氨酸酶亚型主要为N1、N2、N3、N7等，其中HPAIV占据一定的比例。在国内，虽然高致病性的H7亚型流感病毒还没有分离到，但是也曾分离到过一株自然分离株H7亚型LPAIV，而且对于高致病性的H7亚型病毒的高度危害，我们深信不疑。为了防止那可能发生的危害，做好高效力的H7亚型的流感疫苗，做好技术储备刻不容缓。但由于现地分离的野毒株鸡胚适应性较差，致使现行预储备的疫苗存在成本高、疫苗免疫期短等缺点。
发明内容
本发明的目的是提供一种人工重组的H7亚型流感病毒、该病毒的制备方法和应用，该病毒对我国流行的H7N2禽流感病毒具有特异免疫性，具有高滴度鸡胚适应性，对鸡及鸡胚无致病力。
本发明的目的是这样实现的一种人工重组的H7亚型流感病毒，其组成包括H7亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)的HA和NA基因，所述的病毒上还重组有流感病毒A/PR/8/34(H1N1)的6个内部基因PB2、PB1、PA、NP、M和NS，保藏号CCTCC-V200415。
上述的人工重组的H7亚型流感病毒，HA和NA基因具有序列表SEQ IDNo.4和SEQ ID No.6所述的序列，6个内部基因PB2、PB1、PA、NP、M和NS具有序列表SEQ ID No.1、SEQ ID No.2、SEQ ID No.3、SEQ ID No.5、SEQ ID No.7和SEQ ID No.8所述的序列，该病毒对我国流行的H7N2禽流感病毒具有特异免疫性，具有高滴度鸡胚适应性，对鸡及鸡胚无致病力。
上述的人工重组的H7亚型流感病毒的制备方法，包括以下步骤a.共感染-使两个病毒基因组发生重组流感病毒A/PR/8/34(PR8)(H1N1)与经紫外线处理过的H7N2亚型AIV国内分离株A/Chicken/HeBei/1/02(H7N2)共同接种10日龄鸡胚尿囊腔；b.筛选-除去表面基因来源于PR8病毒的病毒在上述特定的人工条件下收获的尿囊液，与倍比稀释的抗PR8的血清混合，室温下孵育之后接种鸡胚尿囊腔，收获后，对鸡胚尿囊液进行血凝活性检测，收获血清稀释度最低并且具有血凝活性的样品用于第二轮筛选；c.稀释纯化-除去内部基因来源于H7N2病毒同上方法筛选得到的病毒在鸡胚上进行3代有限克隆纯化，利用片段特异性引物RT-PCR鉴定并经过序列分析筛选HA和NA来自于H7N2、6个内部基因PB2、PB1、PA、NP、M及NS来自PR8的重组病毒。
上述的人工重组的H7亚型流感病毒，作为防制动物和人类H7亚型流感的疫苗株，应用于疫苗防制。
该病毒HA和NA基因来自H7N2亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)，6个内部基因来自PR8病毒；该病毒与我国H7N2亚型禽流感病毒流行株抗原性一致，并且具有良好的鸡胚适应性；作为防制H7亚型动物流感的疫苗株。
本发明的优点1.本发明Re-3以我国禽流感病毒H7N2为HA和NA基因供体，因此，该病毒与我国曾流行的H7N2亚型禽流感病毒抗原性一致。以此为疫苗株生产的疫苗，将对H7N2亚型禽流感病毒流行株产生良好的特异保护性。
2.本发明Re-3含有鸡胚高滴度适应株PR8的6个内部基因，病毒生长滴度与野毒相比，可提高10倍左右，以此为疫苗株生产疫苗，可大大降低疫苗生产成本，提高疫苗质量。
流感病毒A/PR/8/34(H1N1)(PR8)是实验室鸡胚适应株，即该病毒可在鸡胚内生长至很高滴度，这种鸡胚适应性由病毒的6个内部基因决定。另外，当两个流感病毒共同感染一个细胞或鸡胚时，可发生基因互换，从而产生新的基因重组病毒。因此，通过遗传重组方法，可将PR8病毒6个内部基因和流行株的HA、NA基因的进行重组而获得一个新的重组病毒，该重组病毒具有与HA和NA基因供体毒株一致的抗原性，同时还具有PR8病毒的鸡胚高滴度生长特性。
它是以H7N2亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)和A/PR/8/34(H1N1)(PR8)共同接种鸡胚，应用PR8的抗血清对鸡胚培养物进行2轮抗体筛选后、继续在鸡胚上进行有限稀释克隆纯化3代获得的、经测序确证含有H7N2的表面基因HA和NA，同时拥有PR8的6个内部基因PB2，PB1，PA，NP，M及NS的重组病毒。该病毒鸡胚生长滴度比自然分离株H7N2提高10倍左右，是理想的H7N2亚型禽流感疫苗株。
本发明的
具体实施例方式实施例1一种人工重组的H7亚型流感病毒，其组成包括H7亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)的HA和NA基因，所述的病毒上还重组有流感病毒A/PR/8/34(H1N1)的6个内部基因PB2、PB1、PA、NP、M和NS，保藏在中国典型培养物保藏中心，保藏号CCTCC-V200415，保藏日2004年9月6日。
实施例2实施例1所述的人工重组的H7亚型流感病毒，HA和NA基因具有序列表SEQ ID No.4和SEQ ID No.6所述的序列，6个内部基因PB2、PB1、PA、NP、M和NS具有序列表SEQ ID No.1、SEQ ID No.2、SEQ ID No.3、SEQ ID No.5、SEQ ID No.7和SEQ ID No.8所述的序列(附后)，该病毒对我国流行的H7N2禽流感病毒具有特异免疫性，具有高滴度鸡胚适应性，对鸡及鸡胚无致病力。]实施例3实施例1或2所述的人工重组的H7亚型流感病毒的制备方法，包括以下步骤a.共感染-使两个病毒基因组发生重组流感病毒A/PR/8/34(PR8)(H1N1)与经紫外线处理过的H7N2亚型AIV国内分离株A/Chicken/HeBei/1/02(H7N2)共同接种10日龄鸡胚尿囊腔；b.筛选-除去表面基因来源于PR8病毒的病毒在上述特定的人工条件下收获的尿囊液，与倍比稀释的抗PR8的血清混合，室温下孵育之后接种鸡胚尿囊腔，收获后，对鸡胚尿囊液进行血凝活性检测，收获血清稀释度最低并且具有血凝活性的样品用于第二轮筛选；c.稀释纯化-除去内部基因来源于H7N2病毒同上方法筛选得到的病毒在鸡胚上进行3代有限克隆纯化，利用片段特异性引物RT-PCR鉴定并经过序列分析筛选HA和NA来自于H7N2、6个内部基因PB2、PB1、PA、NP、M及NS来自PR8的重组病毒。
实施例4上述的人工重组的H7亚型流感病毒，作为防制动物和人类H7亚型流感的疫苗株，应用于疫苗防制。
基因序列附后。
重组流感病毒A/哈尔滨/Re-2/2003(H9N2)基因序列SEQ ID No.1ATG GAA AGA ATA AAA GAA CTA AGA AAT CTA ATG TCG CAG TCT CGC ACC CGC GAG ATA CTCMet Glu Arg Ile Lys Glu Leu Arg Asn Leu Met Ser Gln Ser Arg Thr Arg Glu Ile Leu5 10 15 20ACA AAA ACC ACC GTG GAC CAT ATG GCC ATA ATC AAG AAG TAC ACA TCA GGA AGA CAG GAGThr Lys Thr Thr Val Asp His Met Ala Ile Ile Lys Lys Tyr Thr Ser Gly Arg Gln Glu25 30 35 40AAG AAC CCA GCA CTT AGG ATG AAA TGG ATG ATG GCA ATG AAA TAT CCA ATT ACA GCA GACLys Asn Pro Ala Leu Arg Met Lys Trp Met Met Ala Met Lys Tyr Pro Ile Thr Ala Asp45 50 55 60AAG AGG ATA ACG GAA ATG ATT CCT GAG AGA AAT GAG CAA GGA CAA ACT TTA TGG AGT AAALys Arg Ile Thr Glu Met Ile Pro Glu Arg Asn Glu Gln Gly Gln Thr Leu Trp Ser Lys65 70 75 80ATG AAT GAT GCC GGA TCA GAC CGA GTG ATG GTA TCA CCT CTG GCT GTG ACA TGG TGG AATMet Asn Asp Ala Gly Ser Asp Arg Val Met Val Ser Pro Leu Ala Val Thr Trp Trp Asn85 90 95 100AGG AAT GGA CCA ATG ACA AAT ACA GTT CAT TAT CCA AAA ATC TAC AAA ACT TAT TTT GAAArg Asn Gly Pro Met Thr Asn Thr Val His Tyr Pro Lys Ile Tyr Lys Thr Tyr Phe Glu105 110 115 120AGA GTC GAA AGG CTA AAG CAT GGA ACC TTT GGC CCT GTC CAT TTT AGA AAC CAA GTC AAAArg Val Glu Arg Leu Lys His Gly Thr Phe Gly Pro Val His Phe Arg Asn Gln Val Lys125 130 135 140ATA CGT CGG AGA GTT GAC ATA AAT CCT GGT CAT GCA GAT CTC AGT GCC AAG GAG GCA CAGIle Arg Arg Arg Val Asp Ile Asn Pro Gly His Ala Asp Leu Ser Ala Lys Glu Ala Gln145 150 155 160GAT GTA ATC ATG GAA GTT GTT TTC CCT AAC GAA GTG GGA GCC AGG ATA CTA ACA TCG GAAAsp Val Ile Met Glu Val Val Phe Pro Asn Glu Val Gly Ala Arg Lle Leu Thr Ser Glu165 170 175 180TCG CAA CTA ACG ATA ACC AAA GAG AAG AAA GAA GAA CTC CAG GAT TGC AAA ATT TCT CCTSer Gln Leu Thr Ile Thr Lys Glu Lys Lys Glu Glu Leu Gln Asp Cys Lys Ile Ser Pro185 190 195 200TTG ATG GTT GCA TAC ATG TTG GAG AGA GAA CTG GTC CGC AAA ACG AGA TTC CTC CCA GTGLeu Met Val Ala Tyr Met Leu Glu Arg Glu Leu Val Arg Lys Thr Arg Phe Leu Pro Val205 210 215 220GCT GGT GGA ACA AGC AGT GTG TAC ATT GAA GTG TTG CAT TTG ACT CAA GGA ACA TGC TGGAla Gly Gly Thr Ser Ser Val Tyr Ile Glu Val Leu His Leu Thr Gln Gly Thr Cys Trp225 230 235 240GAA CAG ATG TAT ACT CCA GGA GGG GAA GTG AAG AAT GAT GAT GTT GAT CAA AGC TTG ATTGlu Gln Met Tyr Thr Pro Gly Gly Glu Val Lys Asn Asp Asp Val Asp Gln Ser Leu Ile245 250 255 260ATT GCT GCT AGG AAC ATA GTG AGA AGA GCT GCA GTA TCA GCA GAC CCA CTA GCA TCT TTAIle Ala Ala Arg Asn Ile Val Arg Arg Ala Ala Val Ser Ala Asp Pro Leu Ala Ser Leu265 270 275 280TTG GAG ATG TGC CAC AGC ACA CAG ATT GGT GGA ATT AGG ATG GTA GAC ATC CTT AAG CAGLeu Glu Met Cys His Ser Thr Gln Ile Gly Gly Ile Arg Met Val Asp Ile Leu Lys Gln285 290 295 300AAC CCA ACA GAA GAG CAA GCC GTG GGT ATA TGC AAG GCT GCA ATG GGA GTG AGA ATT AGCAsn Pro Thr Glu Glu Gln Ala Val Gly Ile Cys Lys Ala Ala Met Gly Leu Arg Ile Ser305 310 315 320TCA TCC TTC AGT TTT GGT GGA TTC ACA TTT AAG AGA ACA AGC GGA TCA TCA GTC AAG AGASer Ser Phe Ser Phe Gly Gly Phe Thr Phe Lys Arg Thr Ser Gly Ser Ser Val Lys Arg
325 330 335 340GAG GAA GAG GTG CTT ACG GGC AAT CTT CAA ACA TTG AAG ATA AGA GTG CAT GAG GGA TATGlu Glu Glu Val Leu Thr Gly Asn Leu Gln Thr Leu Lys Ile Arg Val His Glu Gly Tyr345 350 355 360GAA GAG TTC ACA ATG GTT GGG AGA AGA GCA ACA GCC ATA CTC AGA AAA GCA ACC AGG AGAGlu Glu Phe Thr Met Val Gly Arg Arg Ala Thr Ala Ile Leu Arg Lys Ala Thr Arg Arg365 370 375 380TTG ATT CAG CTG ATA GTG AGT GGG AGA GAC GAA CAG ICG ATT GCC GAA GCA ATA ATT GTGLeu Ile Gln Leu Ile Val Ser Gly Arg Asp Glu Gln Ser Ile Ala Glu Ala Ile Ile Val385 390 395 400GCC ATG GTA TTT TCA CAA GAG GAT TGT ATG ATA AAA GCA GTT AGA GGT GAT CTG AAT TTCAla Met Val Phe Ser Gln Glu Asp Cys Met Ile Lys Ala Val Arg Gly Asp Leu Asn Phe405 410 415 420GTC AAT AGG GCG AAT CAG CGA CTG AAT CCT ATG CAT CAA CTT TTA AGA CAT TTT CAG AAGVal Asn Arg Ala Asn Gln Arg Leu Asn Pro Met His Gln Leu Leu Arg His Phe Gln Lys425 430 435 440GAT GCG AAA GTG CTT TTT CAA AAT TGG GGA GTT GAA CCT ATC GAC AAT GTG ATG GGA ATGAsp Ala Lys Val Leu Phe Gln Asn Trp Gly Val Glu Pro Ile Asp Asn Val Met Gly Met445 450 455 460ATT GGG ATA TTG CCC GAC ATG ACT CCA AGC ATC GAG ATG TCA ATG AGA GGA GTG AGA ATCIle Gly Ile Leu Pro Asp Met Thr Pro Ser Ile Glu Met Ser Met Arg Gly Val Arg Ile465 470 475 480AGC AAA ATG GGT GTA GAT GAG TAC TCC AGC ACG GAG AGG GTA GTG GTG AGC ATT GAC CGGSer Lys Met Gly Val Asp Glu Tyr Ser Ser Thr Glu Arg Val Val Val Ser Ile Asp Arg485 490 495 500TTC TTG AGA GTC CGG GAC CAA CGA GGA AAT GTA CTA CTG TCT CCC GAG GAG GTC AGT GAAPhe Leu Arg Val Arg Asp Gln Arg Gly Asn Val Leu Leu Ser Pro Glu Glu Val Ser Glu505 510 515 520ACA CAG GGA ACA GAG AAA CTG ACA ATA ACT TAC TCA TCG TCA ATG ATG TGG GAG ATT AATThr Gln Gly Thr Glu Lys Leu Thr Ile Thr Tyr Ser Ser Ser Met Met Trp Glu Ile Asn525 530 535 540GGT CCT GAA TCA GTG TTG GTC AAT ACC TAT CAA TGG ATC ATC AGA AAC TGG GAA ACT GTTGly Pro Glu Ser Val Leu Val Asn Thr Tyr Gln Trp Ile Ile Arg Asn Trp Glu Thr Val545 550 555 560AAA ATT CAG TGG TCC CAG AAC CCT ACA ATG CTA TAC AAT AAA ATG GAA TTT GAA CCA TTTLys Ile Gln Trp Ser Gln Asn Pro Thr Met Leu Tyr Asn Lys Met Glu Phe Glu Pro Phe565 570 575 580CAG TCT TTA GTA CCT AAG GCC ATT AGA GGC CAA TAC AGT GGG TTT GTG AGA ACT CTG TTCGln Ser Leu Val Pro Lys Ala Ile Arg Gly Gln Tyr Ser Gly Phe Val Arg Thr Leu Phe585 590 595 600CAA CAA ATG AGG GAT GTG CTT GGG ACA TTT GAT ACC GCA CAG ATA ATA AAA CTT CTT CCCGln Gln Met Arg Asp Val Leu Gly Thr Phe Asp Thr Ala Gln Ile Ile Lys Leu Leu Pro605 610 615 620TTC GCA GCC GCT CCA CCA AAG CAA AGT AGA ATG CAG TTC TCC TCA TTT ACT GTG AAT GTGPhe Ala Ala Ala Pro Pro Lys Gln Ser Arg Met Gln Phe Ser Ser Phe Thr Val Asn Val625 630 635 640AGG GGA TCA GGA ATG AGA ATA CTT GTA AGG GGC AAT TCT CCT GTA TTC AAC TAC AAC AAGArg Gly Ser Gly Met Arg Ile Leu Val Arg Gly Asn Ser Pro Val Phe Asn Tyr Asn Lys645 650 655 660GCC ACG AAG AGA CTC ACA GTT CTC GGA AAG GAT GCT GGC ACT TTA ACC GAA GAC CCA GATAla Thr Lys Arg Leu Thr Val Leu Gly Lys Asp Ala Gly Thr Leu Thr Glu Asp Pro Asp665 670 675 680GAA GGC ACA GCT GGA GTG GAG TCC GCT GTT CTG AGG GGA TTC CTC ATT CTG GGC AAA GAA
Glu Gly Thr Ala Gly Val Glu Ser Ala Val Leu Arg Gly Phe Leu Ile Leu Gly Lys Glu685 690 695 700GAC AGG AGA TAT GGG CCA GCA TTA AGC ATC AAT GAA CTG AGC AAC CTT GCG AAA GGA GAGAsp Arg Arg Tyr Gly Pro Ala Leu Ser Ile Asn Glu Leu Ser Asn Leu Ala Lys Gly Glu705 710 715 720AAG GCT AAT GTG CTA ATT GGG CAA GGA GAC GTG GTG TTG GTA ATG AAA CGA AAA CGG GACLys Ala Asn Val Leu Ile Gly Gln Gly Asp Val Val Leu Val Met Lys Arg Lys Arg Asp725 730 735 740TCT AGC ATA CTT ACT GAC AGC CAG ACA GCG ACC AAA AGA ATT CGG ATG GCC ATC AATSer Ser Ile Leu Thr Asp Ser Gln Thr Ala Thr Lys Arg Ile Arg Met Ala Ile Asn745 750 755 759Met Glu Arg Ile Lys Glu Leu Arg Asn Leu Met Ser Gln Ser Arg Thr Arg Glu Ile Leu5 10 15 20Thr Lys Thr Thr Val Asp His Met Ala Ile Ile Lys Lys Tyr Thr Ser Gly Arg Gln Glu25 30 35 40Lys Asn Pro Ala Leu Arg Met Lys Trp Met Met Ala Met Lys Tyr Pro Ile Thr Ala Asp45 50 55 60Lys Arg Ile Thr Glu Met Ile Pro Glu Arg Asn Glu Gln Gly Gln Thr Leu Trp Ser Lys65 70 75 80Met Asn Asp Ala Gly Ser Asp Arg Val Met Val Ser Pro Leu Ala Val Thr Trp Trp Asn85 90 95 100Arg Asn Gly Pro Met Thr Asn Thr Val His Tyr Pro Lys Ile Tyr Lys Thr Tyr Phe Glu105 110 115 120Arg Val Glu Arg Leu Lys His Gly Thr Phe Gly Pro Val His Phe Arg Asn Gln Val Lys125 130 135 140Ile Arg Arg Arg Val Asp Ile Asn Pro Gly His Ala Asp Leu Ser Ala Lys Glu Ala Gln145 150 155 160Asp Val Ile Met Glu Val Val Phe Pro Asn Glu Val Gly Ala Arg Ile Leu Thr Ser Glu165 170 175 180Ser Gln Leu Thr Ile Thr Lys Glu Lys Lys Glu Glu Leu Gln Asp Cys Lys Ile Ser Pro185 190 195 200Leu Met Val Ala Tyr Met Leu Glu Arg Glu Leu Val Arg Lys Thr Arg Phe Leu Pro Val205 210 215 220Ala Gly Gly Thr Ser Ser Val Tyr Ile Glu Val Leu His Leu Thr Gln Gly Thr Cys Trp225 230 235 240Glu Gln Met Tyr Thr Pro Gly Gly Glu Val Lys Asn Asp Asp Val Asp Gln Ser Leu Ile245 250 255 260lle Ala Ala Arg Asn Ile Val Arg Arg Ala Ala Val Ser Ala Asp Pro Leu Ala Ser Leu265 270 275 280Leu Glu Met Cys His Ser Thr Gln Ile Gly Gly Ile Arg Met Val Asp Ile Leu Lys Gln285 290 295 300Asn Pro Thr Glu Glu Gln Ala Val Gly Ile Cys Lys Ala Ala Met Gly Leu Arg Ile Ser305 310 315 320Ser Ser Phe Ser Phe Gly Gly Phe Thr Phe Lys Arg Thr Ser Gly Ser Ser Val Lys Arg325 330 335 340Glu Glu Glu Val Leu Thr Gly Asn Leu Gln Thr Leu Lys Ile Arg Val His Glu Gly Tyr345 350 355 360Glu Glu Phe Thr Met Val Gly Arg Arg Ala Thr Ala Ile Leu Arg Lys Ala Thr Arg Arg365 370 375 380Leu Ile Gln Leu Ile Val Ser Gly Arg Asp Glu Gln Ser Ile Ala Glu Ala Ile Ile Val385 390 395 400
Ala Met Val Phe Ser Gln Glu Asp Cys Met Ile Lys Ala Val Arg Gly Asp Leu Asn Phe405 410 415 420Val Asn Arg Ala Asn Gln Arg Leu Asn Pro Met His Gln Leu Leu Arg His Phe Gln Lys425 430 435 440Asp Ala Lys Val Leu Phe Gln Asn Trp Gly Val Glu Pro Ile Asp Asn Val Met Gly Met445 450 455 460Ile Gly Ile Leu Pro Asp Met Thr Pro Ser Ile Glu Met Ser Met Arg Gly Val Arg Ile465 470 475 480Ser Lys Met Gly Val Asp Glu Tyr Ser Ser Thr Glu Arg Val Val Val Ser Ile Asp Arg485 490 495 500Phe Leu Arg Val Arg Asp Gln Arg Gly Asn Val Leu Leu Ser Pro Glu Glu Val Ser Glu505 510 515 520Thr Gln Gly Thr Glu Lys Leu Thr Ile Thr Tyr Ser Ser Ser Met Met Trp Glu Ile Asn525 530 535 540Gly Pro Glu Ser Val Leu Val Asn Thr Tyr Gln Trp Ile Ile Arg Asn Trp Glu Thr Val545 550 555 560Lys Ile Gln Trp Ser Gln Asn Pro Thr Met Leu Tyr Asn Lys Met Glu Phe Glu Pro Phe565 570 575 580Gln Ser Leu Val Pro Lys Ala lle Arg Gly Gln Tyr Ser Gly Phe Val Arg Thr Leu Phe585 590 595 600Gln Gln Met Arg Asp Val Leu Gly Thr Phe Asp Thr Ala Gln Ile Ile Lys Leu Leu Pro605 610 615 620Phe Ala Ala Ala Pro Pro Lys Gln Ser Arg Met Gln Phe Ser Ser Phe Thr Val Asn Val625 630 635 640Arg Gly Ser Gly Met Arg Ile Leu Val Arg Gly Asn Ser Pro Val Phe Asn Tyr Asn Lys645 650 655 660Ala Thr Lys Arg Leu Thr Val Leu Gly Lys Asp Ala Gly Thr Leu Thr Glu Asp Pro Asp665 670 675 680Glu Gly Thr Ala Gly Val Glu Ser Ala Val Leu Arg Gly Phe Leu Ile Leu Gly Lys Glu685 690 695 700Asp Arg Arg Tyr Gly Pro Ala Leu Ser Ile Asn Glu Leu Ser Asn Leu Ala Lys Gly Glu705 710 715 720Lys Ala Asn Val Leu Ile Gly Gln Gly Asp Val Val Leu Val Met Lys Arg Lys Arg Asp725 730 735 740Ser Ser Ile Leu Thr Asp Ser Gln Thr Ala Thr Lys Arg Ile Arg Met Ala lle Asn45 750 755 759SEQ ID No.2ATG GAT GTC AAT CCG ACC TTA CTT TTC TTA AAA GTG CCA GCA CAA AAT GCT ATA AGC ACAMet Asp Val Asn Pro Thr Leu Leu Phe Leu Lys Val Pro Ala Gln Asn Ala Ile Ser Thr5 10 15 20ACT TTC CCT TAT ACC GGA GAC CCT CCT TAC AGC CAT GGG ACA GGA ACA GGA TAC ACC ATGThr Phe Pro Tyr Thr Gly Asp Pro Pro Tyr Ser His Gly Thr Gly Thr Gly Tyr Thr Met25 30 35 40GAT ACT GTC AAC AGG ACA CAT CAG TAC TCA GAA AAG GCA AGA TGG ACA ACA AAC ACC GAAAsp Thr Val Asn Arg Thr His Gln Tyr Ser Glu Lys Ala Arg Trp Thr Thr Asn Thr Glu45 50 55 60ACT GGA GCA CCG CAA CTC AAC CCG ATT GAT GGG CCA CTG CCA GAA GAC AAT GAA CCA AGTThr Gly Ala Pro Gln Leu Asn Pro Ile Asp Gly Pro Leu Pro Glu Asp Asn Glu Pro Ser65 70 75 80GGT TAT GCC CAA ACA GAT TGT GTA TTG GAA GCA ATG GCT TTC CTT GAG GAA TCC CAT CCTGly Tyr Ala Gln Thr Asp Cys Val Leu Glu Ala Met Ala Phe Leu Glu Glu Ser His Pro85 90 95 100
GGT ATT TTT GAA AAC TCG TGT ATT GAA ACG ATG GAG GTT GTT CAG CAA ACA CGA GTA GACGly Ile Phe Glu Asn Ser Cys Ile Glu Thr Met Glu Val Val Gln Gln Thr Arg Val Asp105 110 115 120AAG CTG ACA CAA GGC CGA CAG ACC TAT GAC TGG ACT TTA AAT AGA AAC CAG CCT GCT GCALys Leu Thr Gln Gly Arg Gln Thr Tyr Asp Trp Thr Leu Asn Arg Asn Gln Pro Ala Ala125 130 135 140ACA GCA TTG GCC AAC ACA ATA GAA GTG TTC AGA TCA AAT GGC CTC ACG GCC AAT GAG TCTThr Ala Leu Ala Asn Thr Ile Glu Val Phe Arg Ser Asn Gly Leu Thr Ala Asn Glu Ser145 150 155 160GGA AGG CTC ATA GAC TTC CTT AAG GAT GTA ATG GAG TCA ATG AAA AAA GAA GAA ATG GGGGly Arg Leu Ile Asp Phe Leu Lys Asp Val Met Glu Ser Met Lys Lys Glu Glu Met Gly165 170 175 180ATC ACA ACT CAT TTT CAG AGA AAG AGA CGG GTG AGA GAC AAT ATG ACT AAG AAA ATG ATAIle Thr Thr His Phe Gln Arg Lys Arg Arg Val Arg Asp Asn Met Thr Lys Lys Met Ile185 190 195 200ACA CAG AGA ACA ATA GGT AAA AGG AAA CAG AGA TTG AAC AAA AGG AGT TAT CTA ATT AGAThr Gln Arg Thr Ile Gly Lys Arg Lys Gln Arg Leu Asn Lys Arg Ser Tyr Leu Ile Arg205 210 215 220GCA TTG ACC CTG AAC ACA ATG ACC AAA GAT GCT GAG AGA GGG AAG CTA AAA CGG AGA GCAAla Leu Thr Leu Asn Thr Met Thr Lys Asp Ala Glu Arg Gly Lys Leu Lys Arg Arg Ala225 230 235 240ATT GCA ACC CCA GGG ATG CAA ATA AGG GGG TTT GTA TAC TTT GTT GAG ACA CTG GCA AGGIle Ala Thr Pro Gly Met Gln Ile Arg Gly Phe Val Tyr Phe Val Glu Thr Leu Ala Arg245 250 255 260AGT ATA TGT GAG AAA CTT GAA CAA TCA GGG TTG CCA GTT GGA GGC AAT GAG AAG AAA GCASer Ile Cys Glu Lys Leu Glu Gln Ser Gly Leu Pro Val Gly Gly Asn Glu Lys Lys Ala265 270 275 280AAG TTG GCA AAT GTT GTA AGG AAG ATG ATG ACC AAT TCT CAG GAC ACC GAA CTT TCT TTGLys Leu Ala Asn Val Val Arg Lys Met Met Thr Asn Ser Gln Asp Thr Glu Leu Ser Leu285 290 295 300ACC ATC ACT GGA GAT AAC ACC AAA TGG AAC GAA AAT CAG AAT CCT CGG ATG TTT TTG GCCThr Ile Thr Gly Asp Asn Thr Lys Trp Asn Glu Asn Gln Asn Pro Arg Met Phe Leu Ala305 310 315 320ATG ATC ACA TAT ATG ACC AGA AAT CAG CCC GAA TGG TTC AGA AAT GTT CTA AGT ATT GCTMet Ile Thr Tyr Met Thr Arg Asn Gln Pro Glu Trp Phe Arg Asn Val Leu Ser Ile Ala325 330 335 340CCA ATA ATG TTC TCA AAC AAA ATG GCG AGA CTG GGA AAA GGG TAT ATG TTT GAG AGC AAGPro Ile Met Phe Ser Asn Lys Met Ala Arg Leu Gly Lys Gly Tyr Met Phe Glu Ser Lys345 350 355 360AGT ATG AAA CTT AGA ACT CAA ATA CCT GCA GAA ATG CTA GCA AGC ATT GAT TTG AAA TATSer Met Lys Leu Arg Thr Gln Ile Pro Ala Glu Met Leu Ala Ser Ile Asp Leu Lys Tyr365 370 375 380TTC AAT GAT TCA ACA AGA AAG AAG ATT GAA AAA ATC CGA CCG CTC TTA ATA GAG GGG ACTPhe Asn Asp Ser Thr Arg Lys Lys Ile Glu Lys Ile Arg Pro Leu Leu Ile Glu Gly Thr385 390 395 400GCA TCA TTG AGC CCT GGA ATG ATG ATG GGC ATG TTC AAT ATG TTA AGC ACT GTA TTA GGCAla Ser Leu Ser Pro Gly Met Met Met Gly Met Phe Asn Met Leu Ser Thr Val Leu Gly405 410 415 420GTC TCC ATC CTG AAT CTT GGA CAA AAG AGA TAC ACC AAG ACT ACT TAC TGG TGG GAT GGTVal Ser Ile Leu Asn Leu Gly Gln Lys Arg Tyr Thr Lys Thr Thr Tyr Trp Trp Asp Gly425 430 435 440CTT CAA TCC TCT GAC GAT TTT GCT CTG ATT GTG AAT GCA CCC AAT CAT GAA GGG ATT CAALeu Gln Ser Ser Asp Asp Phe Ala Leu Ile Val Asn Ala Pro Asn His Glu Gly Ile Gln
445 450 455 460GCC GGA GTC GAC AGG TTT TAT CGA ACC TGT AAG CTA CAT GGA ATC AAT ATG AGC AAG AAAAla Gly Val Asp Arg Phe Tyr Arg Thr Cys Lys Leu His Gly Ile Asn Met Ser Lys Lys465 470 475 480AAG TCT TAC ATA AAC AGA ACA GGT ACA TTT GAA TTC ACA AGT TTT TTC TAT CGT TAT GGGLys Ser Tyr Ile Asn Arg Thr Gly Thr Phe Glu Phe Thr Ser Phe Phe Tyr Arg Tyr Gly485 490 495 500TTT GTT GCC AAT TTC AGC ATG GAG CTT CCC AGT TTT GGT GTG TCT GGG AGC AAC GAG TCAPhe Val Ala Asn Phe Ser Met Glu Leu Pro Ser Phe Gly Val Ser Gly Ser Asn Glu Ser505 510 515 520GCG GAC ATG AGT ATT GGA GTT ACT GTC ATC AAA AAC AAT ATG ATA AAC AAT GAT CTT GGTAla Asp Met Ser Ile Gly Val Thr Val Ile Lys Asn Asn Met Ile Asn Asn Asp Leu Gly525 530 535 540CCA GCA ACA GCT CAA ATG GCC CTT CAG TTG TTC ATC AAA GAT TAC AGG TAC ACG TAC CGAPro Ala Thr Ala Gln Met Ala Leu Gln Leu Phe Ile Lys Asp Tyr Arg Tyr Thr Tyr Arg545 550 555 560TGC CAT AGA GGT GAC ACA CAA ATA CAA ACC CGA AGA TCA TTT GAA ATA AAG AAA CTG TGGCys His Arg Gly Asp Thr Gln Ile Gln Thr Arg Arg Ser Phe Glu Ile Lys Lys Leu Trp565 570 575 580GAG CAA ACC CGT TCC AAA GCT GGA CTG CTG GTC TCC GAC GGA GGC CCA AAT TTA TAC AACGlu Gln Thr Arg Ser Lys Ala Gly Leu Leu Val Ser Asp Gly Gly Pro Asn Leu Tyr Asn585 590 595 600ATT AGA AAT CTC CAC ATT CCT GAA GTC TGC CTA AAA TGG GAA TTG ATG GAT GAG GAT TACIle Arg Asn Leu His Ile Pro Glu Val Cys Leu Lys Trp Glu Leu Met Asp Glu Asp Tyr605 610 615 620CAG GGG CGT TTA TGC AAC CCA CTG AAC CCA TTT GTC AGC CAT AAA GAA ATT GAA TCA ATGGln Gly Arg Leu Cys Asn Pro Leu Asn Pro Phe Val Ser His Lys Glu Ile Glu Ser Met625 630 635 640AAC AAT GCA GTG ATG ATG CCA GCA CAT GGT CCA GCC AAA AAC ATG GAG TAT GAT GCT GTTAsn Asn Ala Val Met Met Pro Ala His Gly Pro Ala Lys Asn Met Glu Tyr Asp Ala Val645 650 655 660GCA ACA ACA CAC TCC TGG ATC CCC AAA AGA AAT CGA TCC ATC TTG AAT ACA AGT CAA AGAAla Thr Thr His Ser Trp Ile Pro Lys Arg Asn Arg Ser Ile Leu Asn Thr Ser Gln Arg665 670 675 680GGA GTA CTT GAA GAT GAA CAA ATG TAC CAA AGG TGC TGC AAT TTA TTT GAA AAA TTC TTCGly Val Leu Glu Asp Glu Gln Met Tyr Gln Arg Cys Cys Asn Leu Phe Glu Lys Phe Phe685 690 695 700CCC AGC AGT TCA TAC AGA AGA CCA GTC GGG ATA TCC AGT ATG GTG GAG GCT ATG GTT TCCPro Ser Ser Ser Tyr Arg Arg Pro Val Gly Ile Ser Ser Met Val Glu Ala Met Val Ser705 710 715 720AGA GCC CGA ATT GAT GCA CGG ATT GAT TTC GAA TCT GGA AGG ATA AAG AAA GAA GAG TTCArg Ala Arg Ile Asp Ala Arg Ile Asp Phe Glu Ser Gly Arg Ile Lys Lys Glu Glu Phe725 730 735 740ACT GAG ATC ATG AAG ATC TGT TCC ACC ATT GAA GAG CTC AGA CGG CAA AAA TAGThr Glu Ile Met Lys Ile Cys Ser Thr Ile Glu Glu Leu Arg Arg Gln Lys ***745 750 755Met Asp Val Asn Pro Thr Leu Leu Phe Leu Lys Val Pro Ala Gln Asn Ala Ile Ser Thr5 10 15 20Thr Phe Pro Tyr Thr Gly Asp Pro Pro Tyr Ser His Gly Thr Gly Thr Gly Tyr Thr Met25 30 35 40Asp Thr Val Asn Arg Thr His Gln Tyr Ser Glu Lys Ala Arg Trp Thr Thr Asn Thr Glu45 50 55 60
Thr Gly Ala Pro Gln Leu Asn Pro Ile Asp Gly Pro Leu Pro Glu Asp Asn Glu Pro Ser65 70 75 80Gly Tyr Ala Gln Thr Asp Cys Val Leu Glu Ala Met Ala Phe Leu Glu Glu Ser His Pro85 90 95 100Gly Ile Phe Glu Asn Ser Cys Ile Glu Thr Met Glu Val Val Gln Gln Thr Arg Val Asp105 110 115 120Lys Leu Thr Gln Gly Arg Gln Thr Tyr Asp Trp Thr Leu Asn Arg Asn Gln Pro Ala Ala125 130 135 140Thr Ala Leu Ala Asn Thr Ile Glu Val Phe Arg Ser Asn Gly Leu Thr Ala Asn Glu Ser145 150 155 160GGA AGG CTC ATA GAC TTC CTT AAG GAT GTA ATG GAG TCA ATG AAA AAA GAA GAA ATG GGGGly Arg Leu Ile Asp Phe Leu Lys Asp Val Met Glu Ser Met Lys Lys Glu Glu Met Gly165 170 175 180Ile Thr Thr His Phe Gln Arg Lys Arg Arg Val Arg Asp Asa Met Thr Lys Lys Met Ile185 190 195 200Thr Gln Arg Thr Ile Gly Lys Arg Lys Gln Arg Leu Asn Lys Arg Ser Tyr Leu Ile Arg205 210 215 220Ala Leu Thr Leu Asn Thr Met Thr Lys Asp Ala Glu Arg Gly Lys Leu Lys Arg Arg Ala225 230 235 240Ile Ala Thr Pro Gly Mer Gln Ile Arg Gly Phe Val Tyr Phe Val Glu Thr Leu Ala Arg245 250 255 260Ser Ile Cys Glu Lys Leu Glu Gln Ser Gly Leu Pro Val Gly Gly Asn Glu Lys Lys Ala265 270 275 280Lys Leu Ala Asn Val Val Arg Lys Met Met Thr Asn Ser Gln Asp Thr Glu Leu Ser Leu285 290 295 300Thr Ile Thr Gly Asp Asn Thr Lys Trp Asn Glu Asn Gln Asn Pro Arg Met Phe Leu Ala305 310 315 320Met Ile Thr Tyr Met Thr Arg Asn Gln Pro Glu Trp Phe Arg Asn Val Leu Ser Ile Ala325 330 335 340Pro Ile Met Phe Ser Asn Lys Met Ala Arg Leu Gly Lys Gly Tyr Met Phe Glu Ser Lys345 350 355 360Ser Met Lys Leu Arg Thr Gln Ile Pro Ala Glu Met Leu Ala Ser Ile Asp Leu Lys Tyr365 370 375 380Phe Asn Asp Ser Thr Arg Lys Lys Ile Glu Lys Ile Arg Pro Leu Leu Ile Glu Gly Thr385 390 395 400Ala Ser Leu Ser Pro Gly Met Met Met Gly Met Phe Asn Met Leu Ser Thr Val Leu Gly405 410 415 420Val Ser Ile Leu Asn Leu Gly Gln Lys Arg Tyr Thr Lys Thr Thr Tyr Trp Trp Asp Gly425 430 435 440Leu Gln Ser Ser Asp Asp Phe Ala Leu Ile Val Asn Ala Pro Asn His Glu Gly Ile Gln445 450 455 460Ala Gly Val Asp Arg Phe Tyr Arg Thr Cys Lys Leu His Gly Ile Asn Met Ser Lys Lys465 470 475 480Lys Ser Tyr Ile Asn Arg Thr Gly Thr Phe Glu Phe Thr Ser Phe Phe Tyr Arg Tyr Gly485 490 95 500Phe Val Ala Asn Phe Ser Met Glu Leu Pro Ser Phe Gly Val Ser Gly Ser Asn Glu Ser505 510 15 520Ala Asp Met Ser Ile Gly Val Thr Val Ile Lys Asn Asn Met Ile Asn Asn Asp Leu Gly525 530 535 540Pro Ala Thr Ala Gln Met Ala Leu Gln Leu Phe Ile Lys Asp Tyr Arg Tyr Thr Tyr Arg545 550 555 560Cys His Arg Gly Asp Thr Gln Ile Gln Thr Arg Arg Ser Phe Glu Ile Lys Lys Leu Trp565 570 575 580
Glu Gln Thr Arg Ser Lys Ala Gly Leu Leu Val Ser Asp Gly Gly Pro Asn Leu Tyr Asn585 590 595 600Ile Arg Asn Leu His Ile Pro Glu Val Cys Leu Lys Trp Glu Leu Met Asp Glu Asp Tyr605 610 615 620Gln Gly Arg Leu Cys Asn Pro Leu Asn Pro Phe Val Ser His Lys Glu Ile Glu Ser Met625 630 635 640Asn Asn Ala Val Met Met Pro Ala His Gly Pro Ala Lys Asn Met Glu Tyr Asp Ala Val645 650 655 660Ala Thr Thr His Ser Trp Ile Pro Lys Arg Asn Arg Ser Ile Leu Asn Thr Ser Gln Arg665 670 675 680Gly Val Leu Glu Asp Glu Gln Met Tyr Gln Arg Cys Cys Asn Leu Phe Glu Lys Phe Phe685 690 695 700Pro Ser Ser Ser Tyr Arg Arg Pro Val Gly Ile Ser Ser Met Val Glu Ala Met Val Ser705 710 715 720Arg Ala Arg Ile Asp Ala Arg Ile Asp Phe Glu Ser Gly Arg Ile Lys Lys Glu Glu Phe725 730 735 740Thr Glu Ile Met Lys Ile Cys Ser Thr Ile Glu Glu Leu Arg Arg Gln Lys ***745 750 755SEQ ID No.3ATG GAA GAT TTT GTG CGA CAA TGC TTC AAT CCG AGG ATT GTC GAG CTT GCG GAA AAA ACAMet Glu Asp Phe Val Arg Gln Cys Phe Asn Pro Met Ile Val Glu Leu Ala Glu Lys Thr5 10 15 20ATG AAA GAG TAT GGG GAG GAC CTG AAA ATC GAA ACA AAC AAA TTT GCA GCA ATA TGC ACTMet Lys Glu Tyr Gly Glu Asp Leu Lys Ile Glu Thr Asn Lys Phe Ala Ala Ile Cys Thr25 30 35 40CAC TTG GAA GTA TGC TTC ATG TAT TCA GAT TTC CAC TTC ATC AAT GAG CAA GGC GAG TCAHis Leu Glu Val Cys Phe Met Tyr Ser Asp Phe His Phe Ile Asn Glu Gln Gly GLu Ser45 50 55 60ATA ATC GTA GAA CTT GGT GAT CCT AAT GCA CTT TTG AAG CAC AGA TTT GAA ATA ATC GAGIle Ile Val Glu Leu Gly Asp Pro Asn Ala Leu Leu Lys His Arg Phe Glu Ile Ile Glu65 70 75 80GGA AGA GAT CGC ACA ATG GCC TGG ACA GTA GTA AAC AGT ATT TGC AAC ACT ACA GGG GCTGly Arg Asp Arg Thr Met Ala Trp Thr Val Val Asn Ser Ile Cys Asn Thr Thr Gly Ala85 90 95 100GAG AAA CCA AAG TTT CTA CCA GAT TTG TAT GAT TAC AAG GAA AAT AGA TTC ATC GAA ATTGlu Lys Pro Lys Phe Leu Pro Asp Leu Tyr Asp Tyr Lys Glu Asn Arg Phe Ile Glu Ile105 110 115 120GGA GTA ACA AGG AGA GAA GTT CAC ATA TAC TAT CTG GAA AAG GCC AAT AAA ATT AAA TCTGly Val Thr Arg Arg Glu Val His Ile Tyr Tyr Leu Glu Lys Ala Asn Lys Ile Lys Ser125 130 135 140GAG AAA ACA CAC ATC CAC ATT TTC TCG TTC ACT GGG GAA GAA ATG GCC ACA AAG GCC GACGlu Lys Thr His Ile His Ile Phe Ser Phe Thr Gly Glu Glu Met Ala Thr Lys Ala Asp145 150 155 160TAC ACT CTC GAT GAA GAA AGC AGG GCT AGG ATC AAA ACC AGG CTA TTC ACC ATA AGA CAATyr Thr Leu Asp Glu Glu Ser Arg Ala Arg Ile Lys Thr Arg Leu Phe Thr Ile Arg Gln165 170 175 180GAA ATG GCC AGC AGA GGC CTC TGG GAT TCC TTT CGT CAG TCC GAG AGA GGA GAA GAG ACAGlu Met Ala Ser Arg Gly Leu Trp Asp Ser Phe Arg Gln Ser Glu Arg Gly Glu Glu Thr185 190 195 200ATT GAA GAA AGG TTT GAA ATC ACA GGA ACA ATG CGC AAG CTT GCC GAC CAA AGT CTC CCG
Ile Glu Glu Arg Phe Glu Ile Thr Gly Thr Met Arg Lys Leu Ala Asp Gln Ser Leu Pro205 210 215 220CCG AAC TTC TCC AGC CTT GAA AAT TTT AGA GCC TAT GTG GAT GGA TTC GAA CCG AAC GGCPro Asn Phe Ser Ser Leu Glu Asn Phe Arg Ala Tyr Val Asp Gly Phe Glu Pro Asn Gly225 230 235 240TAC ATT GAG GGC AAG CTG TCT CAA ATG TCC AAA GAA GTA AAT GCT AGA ATT GAA CCT TTTTyr Ile Glu Gly Lys Leu Ser Gln Met Ser Lys Glu Val Asn Ala Arg Ile Glu Pro Phe245250 255 260TTG AAA ACA ACA CCA CGA CCA CTT AGA CTT CCG AAT GGG CCT CCC TGT TCT CAG CGG TCCLeu Lys Thr Thr Pro Arg Pro Leu Arg Leu Pro Asn Gly Pro Pro Cys Ser Gln Arg Ser265 270 275 280AAA TTC CTG CTG ATG GAT GCC TTA AAA TTA AGC ATT GAG GAC CCA AGT CAT GAA GGA GAGLys Phe Leu Leu Met Asp Ala Leu Lys Leu Ser Ile Glu Asp Pro Ser His Glu Gly Glu285 290 295 300GGA ATA CCG CTA TAT GAT GCA ATC AAA TGC ATG AGA ACA TTC TTT GGA TGG AAG GAA CCCGly Ile Pro Leu Tyr Asp Ala lle Lys Cys Met Arg Thr Phe Phe Gly Trp Lys Glu Pro305 310 315 320AAT GTT GTT AAA CCA CAC GAA AAG GGA ATA AAT CCA AAT TAT CTT CTG TCA TGG AAG CAAAsn Val Val Lys Pro His Glu Lys Gly Ile Asn Pro Asn Tyr Leu Leu Ser Trp Lys Gln325 330 335 340GTA CTG GCA GAA CTG CAG GAC ATT GAG AAT GAG GAG AAA ATT CCA AAG ACT AAA AAT ATGVal Leu Ala Glu Leu Gln Asp Ile Glu Asn Glu Glu Lys Ile Pro Lys Thr Lys Asn Met345 350 355 360AAA AAA ACA AGT CAG CTA AAG TGG GCA CTT GGT GAG AAC ATG GCA CCA GAA AAG GTA GACLys Lys Thr Ser Gln Leu Lys Trp Ala Leu Gly Glu Asn Met Ala Pro Glu Lys Val Asp365 370 375 380TTT GAC GAC TGT AAA GAT GTA GGT GAT TTG AAG CAA TAT GAT AGT GAT GAA CCA GAA TTGPhe Asp Asp Cys Lys Asp Val Gly Asp Leu Lys Gln Tyr Asp Ser Asp Glu Pro Glu Leu385 390 395 400AGG TCG CTT GCA AGT TGG ATT CAG AAT GAG TTC AAC AAG GCA TGC GAA CTG ACA GAT TCAArg Ser Leu Ala Ser Trp Ile Gln Asn Glu Phe Asn Lys Ala Cys Glu Leu Thr Asp Ser405 410 415 420AGC TGG ATA GAG CTT GAT GAG ATT GGA GAA GAT GTG GCT CCA ATT GAA CAC ATT GCA AGCSer Trp Ile Glu Leu Asp Glu Ile Gly Glu Asp Val Ala Pro Ile Glu His Ile Ala Ser425 430 435 440ATG AGA AGG AAT TAT TTC ACA TCA GAG GTG TCT CAC TGC AGA GCC ACA GAA TAC ATA ATGMet Arg Arg Asn Tyr Phe Thr Ser Glu Val Ser His Cys Arg Ala Thr Glu Tyr Ile Met445 450 455 460AAG GGG GTG TAC ATC AAT ACT GCC TTA CTT AAT GCA TCT TGT GCA GCA ATG GAT GAT TTCLys Gly Val Tyr Ile Asn Thr Ala Leu Leu Asn Ala Ser Cys Ala Ala Met Asp Asp Phe465 470 475 480CAA TTA ATT CCA ATG ATA AGC AAG TGT AGA ACT AAG GAG GGA AGG CGA AAG ACC AAC TTGGln Leu Ile Pro Met Ile Ser Lys Cys Arg Thr Lys Glu Gly Arg Arg Lys Thr Asn Leu485 490 495 500TAT GGT TTC ATC ATA AAA GGA AGA TCC CAC TTA AGG AAT GAC ACC GAC GTG GTA AAC TTTTyr Gly Phe Ile Ile Lys Gly Arg Ser His Leu Arg Asn Asp Thr Asp Val Val Asn Phe505 510 515 520GTG AGC ATG GAG TTT TCT CTC ACT GAC CCA AGA CTT GAA CCA CAC AAA TGG GAG AAG TACVal Ser Met Glu Phe Ser Leu Thr Asp Pro Arg Leu Glu Pro His Lys Trp Glu Lys Tyr525 530 535 540TGT GTT CTT GAG ATA GGA GAT ATG CTT CTA AGA AGT GCC ATA GGC CAG GTT TCA AGG CCCCys Val Leu Glu Ile Gly Asp Met Leu Leu Arg Ser Ala Ile Gly Gln Val Ser Arg Pro545 550 555 560
ATG TTC TTG TAT GTG AGG ACA AAT GGA ACC TCA AAA ATT AAA ATG AAA TGG GGA ATG GAGMet Phe Leu Tyr Val Arg Thr Asn Gly Thr Ser Lys Ile Lys Met Lys Trp Gly Met Glu565 570 575 580ATG AGG CGT TGT CTC CTC CAG TCA CTT CAA CAA ATT GAG AGT ATG ATT GAA GCT GAG TCCMet Arg Arg Cys Leu Leu Gln Ser Leu Gln Gln Ile Glu Ser Met Ile Glu Ala Glu Ser585 590 595 600TCT GTC AAA GAG AAA GAC ATG ACC AAA GAG TTC TTT GAG AAC AAA TCA GAA ACA TGG CCCSer Val Lys Glu Lys Asp Met Thr Lys Glu Phe Phe Glu Asn Lys Ser Glu Thr Trp Pro605 610 615 620ATT GGA GAG TCT CCC AAA GGA GTG GAG GAA AGT TCC ATT GGG AAG GTC TGC AGG ACT TTAIle Gly Glu Ser Pro Lys Gly Val Glu Glu Ser Ser Ile Gly Lys Val Cys Arg Thr Leu625 630 635 640TTA GCA AAG TCG GTA TTT AAC AGC TTG TAT GCA TCT CCA CAA CTA GAA GGA TTT TCA GCTLeu Ala Lys Ser Val Phe Asn Ser Leu Tyr Ala Ser Pro Gln Leu Glu Gly Phe Ser Ala645 650 655 660GAA TCA AGA AAA CTG CTT CTT ATC GTT CAG GCT CTT AGG GAC AAT CTG GAA CCT GGG ACCGlu Ser Arg Lys Leu Leu Leu Ile Val Gln Ala Leu Arg Asp Asn Leu Glu Pro Gly Thr665 670 675 680TTT GAT CTT GGG GGG CTA TAT GAA GCA ATT GAG GAG TGC CTA ATT AAT GAT CCC TGG GTTPhe Asp Leu Gly Gly Leu Tyr Glu Ala Ile Glu Glu Cys Leu Ile Asn Asp Pro Trp Val685 690 695 700TTG CTT AAT GCT TCT TGG TTC AAC TCC TTC CTT ACA CAT GCA TTG AGT TAGLeu Leu Asn Ala Ser Trp Phe Asn Ser Phe Leu Thr His Ala Leu Ser ***705 710 715Met Glu Asp Phe Val Arg Gln Cys Phe Asn Pro Met Ile Val Glu Leu Ala Glu Lys Thr5 10 15 20Met Lys Glu Tyr Gly Glu Asp Leu Lys Ile Glu Thr Asn Lys Phe Ala Ala Ile Cys Thr25 30 35 40His Leu Glu Val Cys Phe Met Tyr Ser Asp Phe His Phe Ile Asn Glu Gln Gly Glu Ser45 50 55 60Ile Ile Val Glu Leu Gly Asp Pro Asn Ala Leu Leu Lys His Arg Phe Glu Ile Ile Glu65 70 75 80Gly Arg Asp Arg Thr Met Ala Trp Thr Val Val Asn Ser Ile Cys Asn Thr Thr Gly Ala85 90 95 100Glu Lys Pro Lys Phe Leu Pro Asp Leu Tyr Asp Tyr Lys Glu Asn Arg Phe Ile Glu Ile105 110 115 120Gly Val Thr Arg Arg Glu Val His Ile Tyr Tyr Leu Glu Lys Ala Asn Lys Ile LVs Ser125 130 135 140Glu Lys Thr His Ile His Ile Phe Ser Phe Thr Gly Glu Glu Met Ala Thr Lys Ala Asp145 150 155 160Tyr Thr Leu Asp Glu Glu Ser Arg Ala Arg Ile Lys Thr Arg Leu Phe Thr Ile Arg Gln165 170 175 180Glu Met Ala Ser Arg Gly Leu Trp Asp Ser Phe Arg Gln Ser Glu Arg Gly Glu Glu Thr185 190 195 200Ile Glu Glu Arg Phe Glu Ile Thr Gly Thr Met Arg Lys Leu Ala Asp Gln Ser Leu Pro205 210 215 220Pro Asn Phe Ser Ser Leu Glu Asn Phe Arg Ala Tyr Val Asp Gly Phe Glu Pro Asn Gly225 230 235 240Tyr Ile Glu Gly Lys Leu Ser Gln Met Ser Lys Glu Val Asn Ala Arg Ile Glu Pro Phe245 250 255 260Leu Lys Thr Thr Pro Arg Pro Leu Arg Leu Pro Asn Gly Pro Pro Cys Ser Gln Arg Ser265 270 275 280
Lys Phe Leu Leu Met Asp Ala Leu Lys Leu Ser Ile Glu Asp Pro Ser His Glu Gly Glu285 290 295 300Gly Ile Pro Leu Tyr Asp Ala Ile Lys Cys Met Arg Thr Phe Phe Gly Trp Lys Glu Pro305 310 315 320Asn Val Val Lys Pro His Glu Lys Gly Ile Asn Pro Asn Tyr Leu Leu Ser Trp Lys Gln325 330 335 340Val Leu Ala Glu Leu Gln Asp Ile Glu Asn Glu Glu Lys Ile Pro Lys Thr Lys Asn Met345 350 355 360Lys Lys Thr Ser Gln Leu Lys Trp Ala Leu Gly Glu Asn Met Ala Pro Glu Lys Val Asp365 370 375 380Phe Asp Asp Cys Lys Asp Val Gly Asp Leu Lys Gln Tyr Asp Ser Asp Glu Pro Glu Leu385 390 395 400Arg Ser Leu Ala Ser Trp Ile Gln Asn Glu Phe Asn Lys Ala Cys Glu Leu Thr Asp Ser405 410 415 420Ser Trp Ile Glu Leu Asp Glu Ile Gly Glu Asp Val Ala Pro Ile Glu His Ile Ala Ser425 430 435 440Met Arg Arg Asn Tyr Phe Thr Ser Glu Val Ser His Cys Arg Ala Thr Glu Tyr Ile Met445 450 455 460Lys Gly Val Tyr Ile Asn Thr Ala Leu Leu Asn Ala Ser Cys Ala Ala Met Asp Asp Phe465 470 475 480Gln Leu Ile Pro Met Ile Ser Lys Cys Arg Thr Lys Glu Gly Arg Arg Lys Thr Asn Leu485 490 495 500Tyr Gly Phe Ile Ile Lys Gly Arg Ser His Leu Arg Asn Asp Thr Asp Val Val Asn Phe505 510 515 520Val Ser Met Glu Phe Ser Leu Thr Asp Pro Arg Leu Glu Pro His Lys Trp Glu Lys Tyr525 530 535 540Cys Val Leu Glu Ile Gly Asp Met Leu Leu Arg Ser Ala Ile Gly Gln Val Ser Arg Pro545 550 555 560Met Phe Leu Tyr Val Arg Thr Asn Gly Thr Ser Lys Ile Lys Met Lys Trp Gly Met Glu565 570 575 580Met Arg Arg Cys Leu Leu Gln Ser Leu Gln Gln Ile Glu Ser Met Ile Glu Ala Glu Ser585 590 595 600Ser Val Lys Glu Lys Asp Met Thr Lys Glu Phe Phe Glu Asn Lys Ser Glu Thr Trp Pro605 610 615 620Ile Gly Glu Ser Pro Lys Gly Val Glu Glu Ser Ser Ile Gly Lys Val Cys Arg Thr Leu625 630 635 640Leu Ala Lys Ser Val Phe Asn Ser Leu Tyr Ala Ser Pro Gln Leu Glu Gly Phe Ser Ala645 650 655 660Glu Ser Arg Lys Leu Leu Leu Ile Val Gln Ala Leu Arg Asp Asn Leu Glu Pro Gly Thr665 670 675 680Phe Asp Leu Gly Gly Leu Tyr Glu Ala Ile Glu Glu Cys Leu Ile Asn Asp Pro Trp Val685 690 695 700Leu Leu Asn Ala Ser Trp Phe Asn Ser Phe Leu Thr His Ala Leu Ser ***705 710 715SEQ ID No.4ATG AAC ACT CAA ATC CTG ATA CTC GCT CTT GTG GTG ATC ATC CCG ACA AAT GCA GAC AAAMet Asn Thr Gln Ile Leu Ile Leu Ala Leu Val Val Ile Ile Pro Thr Asn Ala Asp Lys5 10 15ATT TGC CTT GGG CAT CAT GCC GTG TCA AAC GGA ACT AAG GTA AAC ACA CTA ACT GAA AGAIle Cys Leu Gly His His Ala Val Ser Asn Gly Thr Lys Val Asn Thr Leu Thr Glu Arg25 30 35 40GGG GTG GAA GTT GTC AAT GCA ACT GAA ACA GTG GAG CGA ACA AAC ATT CCC AGG ATC TGC
Gly Val Glu Val Val Asn Ala Thr Glu Thr Val Glu Arg Thr Asn Ile Pro Arg Ile Cys45 50 55 60TCA CAA GGA AAA AGA ACA ATC GAC CTT GGC CAA TGT GGA CTG CTG GGA ACA ATC ACT GGASer Gln Gly Lys Arg Thr Ile Asp Leu Gly Gln Cys Gly Leu Leu Gly Thr Ile Thr Gly65 70 75 80CCA CCC CAA TGT GAC CAA TTC CTA GAA TTC TCA GCT GAT TTG ATT ATT GAA AGG CGG GAAPro Pro Gln Cys Asp Gln Phe Leu Glu Phe Ser Ala Asp Leu Ile Ile Glu Arg Arg Glu85 90 95 100GGA AAT GAT GTC TGT TAT CCT GGG AAA TTT GTA AAC GGA GAA GCT CTG AGA CAA ATT CTCGly Asn Asp Val Cys Tyr Pro Gly Lys Phe Val Asn Gly Glu Ala Leu Arg Gln Ile Leu105 110 115 120AGG GAA TCA GGC GGA ATT GAT AAG GAG ATA ATG GGG TTC ACA TAC AGC GAA ATA AGA ACTArg Glu Ser Gly Gly Ile Asp Lys Glu Ile Met Gly Phe Thr Tyr Ser Glu Ile Arg Thr125 130 135 140AAT GGA ACA ACC AGT GCA TGT AGA AGA TCA GGA TCT TCG TTC TAT GCA GAG ATG AAA TGGAsn Gly Thr Thr Ser Ala Cys Arg Arg Ser Gly Ser Ser Phe Tyr Ala Glu Met Lys Trp145 150 155 160CTT TTG TCA AAT ACA GAC AAT GCT GCT TTC CCG CAG ATG ACT AAG TCG TAC AAA AAC ACALeu Leu Ser Asn Thr Asp Asn Ala Ala Phe Pro Gln Met Thr Lys Ser Tyr Lys Asn Thr165 170 175 180AGG AAA GAC CCA GCT CTG ATA ATC TGG GGA ATC CAT CAT TCC GGA TCA AGC ACA GAA CAGArg Lys Asp Pro Ala Leu Ile Ile Trp Gly Ile His His Ser Gly Ser Ser Thr Glu Gln185 190 195 200ACC AAA TTA TAT GGG AGT GGA AGC AAA CTA ATA ACA GTT GGG AGT TCC AAT TAT CAA CAGThr Lys Leu Tyr Gly Ser Gly Ser Lys Leu Ile Thr Val Gly Ser Ser Asn Tyr Gln Gln205 210 215 220TCC TTT GTA CCG AGT CCG GGG GCG AGA CCA CAA GTG AAT GGC CAA TCT GGA AGG ATC GACSer Phe Val Pro Ser Pro Gly Ala Arg Pro Gln Val Asn Gly Gln Ser Gly Arg Ile Asp225 230 235 240TTC CAT TAC CTG ATG CTA AAT CCC AAT GAC ACA GTC ACT TTC AGT TTC AAT GGG GCT TTCPhe His Tyr Leu Met Leu Asn Pro Asn Asp Thr Val Thr Phe Ser Phe Asn Gly Ala Phe245 250 255 260ATA GCT CCA GAC CGT GCA AGT TTT CTG AGA GGG AAG TCT ATG GGG ATT CAG AGT GAT GTAIle Ala Pro Asp Arg Ala Ser Phe Leu Arg Gly Lys Ser Met Gly Ile Gln Ser Asp Val265 270 275 280CAA GTT GAT GCC AAT TGT GAG GGG GAT TGC TAT CAT AGT GGA GGA ACA ATA ATA AGT AATGln Val Asp Ala Asn Cys Glu Gly Asp Cys Tyr His Ser Gly Gly Thr Ile Ile Ser Asn285 290 295 300TTG CCT TTT CAG AAT ATC AAT AGC AGA GCA GTA GGA AAA TGT CCG AGG TAT GTT AAA CAALeu Pro Phe Gln Asn Ile Asn Ser Arg Ala Val Gly Lys Cys Pro Arg Tyr Val Lys Gln305 310 315 320GAG AGC CTG CTA CTG GCA ACA GGG ATG AAG AAC GTT CCC GAA ATT CCA AAA GGA AGA GGCGlu Ser Leu Leu Leu Ala Thr Gly Met Lys Asn Val Pro Glu Ile Pro Lys Gly Arg Gly325 330 335 340CTA TTT GGT GCC ATA GCG GGT TTC ATT GAA AAT GGA TGG GAA GGT CTG ATT GAT GGA TGGLeu Phe Gly Ala Ile Ala Gly Phe Ile Glu Asn Gly Trp Glu Gly Leu Ile Asp Gly Trp345 350 355 360TAT GGC TTC AGG CAT CAA AAT GCA CAA GGA GAG GGA ACT GCG GCA GAT TAC AAG AGC ACTTyr Gly Phe Arg His Gln Asn Ala Gln Gly Glu Gly Thr Ala Ala Asp Tyr Lys Ser Thr365 370 375 380CAA TCG GCA ATT GAT CAA ATA ACA GGA AAG TTA AAC CGG CTC ATA GAG AAA ACT AAC CAAGln Ser Ala Ile Asp Gln Ile Thr Gly Lys Leu Asn Arg Leu lle Glu Lys Thr Asn Gln385 390 395 400
CAG TTT GAG TTG ATA GAC AAT GAA TTC ACT GAG GTT GAA AAG CAA ATT GGC AAT GTG ATAGln Phe Glu Leu Ile Asp Asn Glu Phe Thr Glu Val Glu Lys Gln Ile Gly Asn Val Ile405 410 415 420AAT TGG ACC AGA GAT TCC ATA ACA GAA GTG TGG TCC TAC AAT GCC GAA CTC TTA GTG GCAAsn Trp Thr Arg Asp Ser Ile Thr Glu Val Trp Ser Tyr Asn Ala Glu Leu Leu Val Ala425 430 435 440ATG GAG AAT CAG CAC ACT ATC GAT CTG GCC GAT TCA GAA ATG AAC AAG CTG TAC GAA AGAMet Glu Asn Gln His Thr Ile Asp Leu Ala Asp Ser Glu Met Asn Lys Leu Tyr Glu Arg445 450 455 460GTG AGG AGA CAA TTG AGG GAG AAT GCT GAG GAA GAC GGC ACT GGT TGT TTC GAA ATA TTTVal Arg Arg Gln Leu Arg Glu Asn Ala Glu Glu Asp Gly Thr Gly Cys Phe Glu Ile Phe465 470 475 480CAC AAG TGT GAT GAC GAC TGT ATG GCC AGT ATT AGA AAC AAC ACT TAT GAT CAC AGC AAAHis Lys Cys Asp Asp Asp Cys Met Ala Ser Ile Arg Asn Asn Thr Tyr Asp His Ser Lys485 490 495 500TAC AGA GAA GAA GCA ATG CAA AAT AGG ATA CAG ATT GAC CCA GTC AAA CTG AGC AGC GGCTyr Arg Glu Glu Ala Met Gln Asn Arg Ile Gln Ile Asp Pro Val Lys Leu Ser Ser Gly505 510 515 520TAC AAA GAT GTG ATA CTT TGG TTT AGC TTC GGG GCA TCA TGT TTC ATA CTT CTG GCC ATTTyr Lys Asp Val Ile Leu Trp Phe Ser Phe Gly Ala Ser Cys Phe Ile Leu Leu Ala Ile525 530 535 540GCA ATG GGC CTT GTC TTC ATG TGT GTG AAG AAT GGA AAC ATG CGG TGC ACT ATT TGT ATAAla Met Gly Leu Val Phe Met Cys Val Lys Asn Gly Asn Met Arg Cys Thr Ile Cys Ile545 550 555 560TAA---565 570 575 580Met Asn Thr Gln Ile Leu Ile Leu Ala Leu Val Val Ile Ile Pro Thr Asn Ala Asp Lys5 10 15 20Ile Cys Leu Gly His His Ala Val Ser Asn Gly Thr Lys Val Asn Thr Leu Thr Glu Arg25 30 35 40Gly Val Glu Val Val Asn Ala Thr Glu Thr Val Glu Arg Thr Asn Ile Pro Arg Ile Cys45 50 55 60Ser Gln Gly Lys Arg Thr Ile Asp Leu Gly Gln Cys Gly Leu Leu Gly Thr Ile Thr Gly65 70 75 80Pro Pro Gln Cys Asp Gln Phe Leu Glu Phe Ser Ala Asp Leu Ile Ile Glu Arg Arg Glu85 90 95 100Gly Asn Asp Val Cys Tyr Pro Gly Lys Phe Val Asn Gly Glu Ala Leu Arg Gln Ile Leu105 110 115 120Arg Glu Ser Gly Gly Ile Asp Lys Glu Ile Met Gly Phe Thr Tyr Ser Glu Ile Arg Thr125 130 135 140Asn Gly Thr Thr Ser Ala Cys Arg Arg Ser Gly Ser Ser Phe Tyr Ala Glu Met Lys Trp145 150 155 160Leu Leu Ser Asn Thr Asp Asn Ala Ala Phe Pro Gln Met Thr Lys Ser Tyr Lys Asn Thr165 170 175 180Arg Lys Asp Pro Ala Leu Ile Ile Trp Gly Ile His His Ser Gly Ser Ser Thr Glu Gln185 190 195 200Thr Lys Leu Tyr Gly Ser Gly Ser Lys Leu Ile Thr Val Gly Ser Ser Asn Tyr Gln Gln205 210 215 220Ser Phe Val Pro Ser Pro Gly Ala Arg Pro Gln Val Asn Gly Gln Ser Gly Arg Ile Asp225 230 235 240Phe His Tyr Leu Met Leu Asn Pro Asn Asp Thr Val Thr Phe Ser Phe Asn Gly Ala Phe
245 250 255 260Ile Ala Pro Asp Arg Ala Ser Phe Leu Arg Gly Lys Ser Met Gly Ile Gln Ser Asp Val265 270 275 280Gln Val Asp Ala Asn Cys Glu Gly Asp Cys Tyr His Ser Gly Gly Thr Ile Ile Ser Asn285 290 295 300Leu Pro Phe Gln Asn Ile Asn Ser Arg Ala Val Gly Lys Cys Pro Arg Tyr Val Lys Gln305 310 315 320Glu Ser Leu Leu Leu Ala Thr Gly Met Lys Asn Val Pro Glu Ile Pro Lys Gly Arg Gly325 330 335 340Leu Phe Gly Ala Ile Ala Gly Phe Ile Glu Asn Gly Trp Glu Gly Leu Ile Asp Gly Trp345 350 355 360Tyr Gly Phe Arg His Gln Asn Ala Gln Gly Glu Gly Thr Ala Ala Asp Tyr Lys Ser Thr365 370 375 380Gln Ser Ala Ile Asp Gln Ile Thr Gly Lys Leu Asn Arg Leu Ile Glu Lys Thr Asn Gln385 390 395 400Gln Phe Glu Leu Ile Asp Asn Glu Phe Thr Glu Val Glu Lys Gln Ile Gly Asn Val Ile405 410 415 420Asn Trp Thr Arg Asp Ser Ile Thr Glu Val Trp Ser Tyr Asn Ala Glu Leu Leu Val Ala425 430 435 440Met Glu Asn Gln His Thr Ile Asp Leu Ala Asp Ser Glu Met Asn Lys Leu Tyr Glu Arg445 450 455 460Val Arg Arg Gln Leu Arg Glu Asn Ala Glu Glu Asp Gly Thr Gly Cys Phe Glu Ile Phe465 470 475 480His Lys Cys Asp Asp Asp Cys Met Ala Ser Ile Arg Asn Asn Thr Tyr Asp His Ser Lys485 490 495 500Tyr Arg Glu Glu Ala Met Gln Asn Arg Ile Gln Ile Asp Pro Val Lys Leu Ser Ser Gly505 510 515 520Tyr Lys Asp Val Ile Leu Trp Phe Ser Phe Gly Ala Ser Cys Phe Ile Leu Leu Ala Ile525 530 535 540Ala Met Gly Leu Val Phe Met Cys Val Lys Asn Gly Asn Met Arg Cys Thr Ile Cys Ile545 550 555 560SEQ ID No.5GCG TCC CAA GGC ACC AAA CGG TCT TAC GAA CAG ATG GAG ACT GAT GGA GAA CGC CAG 60Met Ala Ser Gln Gly Thr Lys Arg Ser Tyr Glu Gln Met Glu Thr Asp Gly Glu Arg Gln5 10 15 20AAT GCC ACT GAA ATC AGA GCA TCC GTC GGA AAA ATG ATT GGT GGA ATT GGA CGA TTC TACAsn Ala Thr Glu Ile Arg Ala Ser Val Gly Lys Met Ile Gly Gly Ile Gly Arg Phe Tyr25 30 35 40ATC CAA ATG TGC ACA GAA CTT AAA CTC AGT GAT TAT GAG GGA CGG TTG ATC CAA AAC AGCIle Gln Met Cys Thr Glu Leu Lys Leu Ser Asp Tyr Glu Gly Arg Leu Ile Gln Asn Ser45 50 55 60TTA ACA ATA GAG AGA ATG GTG CTC TCT GCT TTT GAC GAA AGG AGA AAT AAA TAC CTG GAALeu Thr Ile Glu Arg Met Val Leu Ser Ala Phe Asp Glu Arg Arg Asn Lys Tyr Leu Glu65 70 75 80GAA CAT CCC AGT GCG GGG AAG GAT CCT AAG AAA ACT GGA GGA CCT ATA TAC AGA AGA GTAGlu His Pro Ser Ala Gly Lys Asp Pro Lys Lys Thr Gly Gly Pro Ile Tyr Arg Arg Val85 90 95 100AAC GGA AAG TGG ATG AGA GAA CTC ATC CTT TAT GAC AAA GAA GAA ATA AGG CGA ATC TGGAsn Gly Lys Trp Met Arg Glu Leu Ile Leu Tyr Asp Lys Glu Glu Ile Arg Arg Ile Trp105 110 115 120CGC CAA GCT AAT AAT GGT GAC GAT GCA ACG GCT GGT CTG ACT CAC ATG ATG ATC TGG CAT
Arg Gln Ala Asn Asn Gly Asp Asp Ala Thr Ala Gly Leu Thr His Met Met Ile Trp His125 130 135 140TCC AAT TTG AAT GAT GCA ACT TAT CAG AGG ACA AGG GCT CTT GTT CGC ACC GGA ATG GATSer Asn Leu Asn Asp Ala Thr Tyr Gln Arg Thr Arg Ala Leu Val Arg Thr Gly Met Asp145 150 155 160CCC AGG ATG TGC TCT CTG ATG CAA GGT TCA ACT CTC CCT AGG AGG TCT GGA GCC GCA GGTPro Arg Met Cys Ser Leu Met Gln Gly Ser Thr Leu Pro Arg Arg Ser Gly Ala Ala Gly165 170 175 180GCT GCA GTC AAA GGA GTT GGA ACA ATG GTG ATG GAA TTG GTC AGG ATG ATC AAA CGT GGGAla Ala Val Lys Gly Val Gly Thr Met Val Met Glu Leu Val Arg Met Ile Lys Arg Gly185 190 195 200ATC AAT GAT CGG AAC TTC TGG AGG GGT GAG AAT GGA CGA AAA ACA AGA ATT GCT TAT GAAIle Asn Asp Arg Asn Phe Trp Arg Gly Glu Asn Gly Arg Lys Thr Arg lle Ala Tyr Glu205 210 215 220AGA ATG TGC AAC ATT CTC AAA GGG AAA TTT CAA ACT GCT GCA CAA AAA GCA ATG ATG GATArg Met Cys Asn Ile Leu Lys Gly Lys Phe Gln Thr Ala Ala Gln Lys Ala Met Met Asp225 230 235 240CAA GTG AGA GAG AGC CGG GAC CCA GGG AAT GCT GAG TTC GAA GAT CTC ACT TTT CTA GCAGln Val Arg Glu Ser Arg Asp Pro Gly Asn Ala Glu Phe Glu Asp Leu Thr Phe Leu Ala245 250 255 260CGG TCT GCA CTC ATA TTG AGA GGG TCG GTT GCT CAC AAG TCC TGC CTG CCT GCC TGT GTGArg Ser Ala Leu Ile Leu Arg Gly Ser Val Ala His Lys Ser Cys Leu Pro Ala Cys Val265 270 275 280TAT GGA CCT GCC GTA GCC AGT GGG TAC GAC TTT GAA AGA GAG GGA TAC TCT CTA GTC GGATyr Gly Pro Ala Val Ala Ser Gly Tyr Asp Phe Glu Arg Glu Gly Tyr Ser Leu Val Gly285 290 295 300ATA GAC CCT TTC AGA CTG CTT CAA AAC AGC CAA GTG TAC AGC CTA ATC AGA CCA AAT GAGlle Asp Pro Phe Arg Leu Leu Gln Asn Ser Gln Val Tyr Ser Leu Ile Arg Pro Asn Glu305 310 315 320AAT CCA GCA CAC AAG AGT CAA CTG GTG TGG ATG GCA TGC CAT TCT GCC GCA TTT GAA GATAsn Pro Ala His Lys Ser Gln Leu Val Trp Met Ala Cys His Ser Ala Ala Phe Glu Asp325 330 335 340CTA AGA GTA TTG AGC TTC ATC AAA GGG ACG AAG GTG GTC CCA AGA GGG AAG CTT TCC ACTLeu Arg Val Leu Ser Phe Ile Lys Gly Thr Lys Val Val Pro Arg Gly Lys Leu Ser Thr345 350 355 360AGA GGA GTT CAA ATT GCT TCC AAT GAA AAT ATG GAG ACT ATG GAA TCA AGT ACA CTT GAAArg Gly Val Gln Ile Ala Ser Asn Glu Asn Met Glu Thr Met Glu Ser Ser Thr Leu Glu365 370 375 380CTG AGA AGC AGG TAC TGG GCC ATA AGG ACC AGA AGT GGA GGA AAC ACC AAT CAA CAG AGGLeu Arg Ser Arg Tyr Trp Ala Ile Arg Thr Arg Ser Gly Gly Asn Thr Asn Gln Gln Arg385 390 395 400GCA TCT GCG GGC CAA ATC AGC ATA CAA CCT ACG TTC TCA GTA CAG AGA AAT CTC CCT TTTAla Ser Ala Gly Gln Ile Ser Ile Gln Pro Thr Phe Ser Val Gln Arg Asn Leu Pro Phe405 410 415 420GAC AGA ACA ACC GTT ATG GCA GCA TTC ACT GGG AAT ACA GAG GGG AGA ACA TCT GAC ATGAsp Arg Thr Thr Val Met Ala Ala Phe Thr Gly Asn Thr Glu Gly Arg Thr Ser Asp Met425 430 435 440AGG ACC GAA ATC ATA AGG ATG ATG GAA AGT GCA AGA CCA GAA GAT GTG TCT TTC CAG GGGArg Thr Glu Ile Ile Arg Met Met Glu Ser Ala Arg Pro Glu Asp Val Ser Phe Gln Gly445 450 455 460CGG GGA GTC TTC GAG CTC TCG GAC GAA AAG GCA GCG AGC CCG ATC GTG CCT TCC TTT GACArg Gly Val Phe Glu Leu Ser Asp Glu Lys Ala Ala Ser Pro Ser Val Pro Ser Phe Asp465 470 475 480
ATG AGT AAT GAA GGA TCT TAT TTC TTC GGA GAC AAT GCA GAG GAG TAC GAC AAT TAAMet Ser Asn Glu Gly Ser Tyr Phe Phe Gly Asp Asn Ala Glu Glu Tyr Asp Asn ***485 490 495Met Als Ser Gln Gly Thr Lys Arg Ser Tyr Glu Gln Met Glu Thr Asp Gly Glu Arg Gln5 10 15 20Asn Ala Thr Glu Ile Arg Ala Ser Val Gly Lys Met Ile Gly Gly Ile Gly Arg Phe Tyr25 30 35 40Ile Gln Met Cys Thr Glu Leu Lys Leu Ser Asp Tyr Glu Gly Arg Leu Ile Gln Asn Ser45 50 55 60Leu Thr Ile Glu Arg Met Val Leu Ser Ala Phe Asp Glu Arg Arg Asn Lys Tyr Leu Glu65 70 75 80Glu His Pro Ser Ala Gly Lys Asp Pro Lys Lys Thr Gly Gly Pro lle Tyr Arg Arg Val85 90 95 100Asn Gly Lys Trp Met Arg Glu Leu Ile Leu Tyr Asp Lys Glu Glu Ile Arg Arg Ile Trp105 110 115 120Arg Gln Ala Asn Asn Gly Asp Asp Ala Thr Ala Gly Leu Thr His Met Met Ile Trp His125 130 135 140Ser Asn Leu Asn Asp Ala Thr Tyr Gln Arg Thr Arg Ala Leu Val Arg Thr Gly Met Asp145 150 155 160Pro Arg Met Cys Ser Leu Met Gln Gly Ser Thr Leu Pro Arg Arg Ser Gly Ala Ala Gly165 170 175 180Ala Ala Val Lys Gly Val Gly Thr Met Val Met Glu Leu Val Arg Met Ile Lys Arg Gly185 190 195 200Ile Asn Asp Arg Asn Phe Trp Arg Gly Glu Asn Gly Arg Lys Thr Arg Ile Ala Tyr Glu205 210 215 220Arg Met Cys Asn Ile Leu Lys Gly Lys Phe Gln Thr Ala Ala Gln Lys Ala Met Met Asp225 230 235 240Gln Val Arg Glu Ser Arg Asp Pro Gly Asn Ala Glu Phe Glu Asp Leu Thr Phe Leu Ala245 250 255 260Arg Ser Ala Leu Ile Leu Arg Gly Ser Val Ala His Lys Ser Cys Leu Pro Ala Cys Val265 270 275 280Tyr Gly Pro Ala Val Ala Ser Gly Tyr Asp Phe Glu Arg Glu Gly Tyr Ser Leu Val Gly285 290 295 300ATA GAC CCT TTC AGA CTG CTT CAA AAC AGC CAA GTG TAC AGC CTA ATC AGA CCA AAT GAGIle Asp Pro Phe Arg Leu Leu Gln Asn Ser Gln Val Tyr Ser Leu Ile Arg Pro Asn Glu305 310 315 320Asn Pro Ala His Lys Ser Gln Leu Val Trp Met Ala Cys His Ser Ala Ala Phe Glu Asp325 330 335 340Leu Arg Val Leu Ser Phe Ile Lys Gly Thr Lys Val Val Pro Arg Gly Lys Leu Ser Thr345 350 355 360Arg Gly Val Gln Ile Ala Ser Asn Glu Asn Met Glu Thr Met Glu Ser Ser Thr Leu Glu365 370 375 380Leu Arg Ser Arg Tyr Trp Ala lle Arg Thr Arg Ser Gly Gly Asn Thr Asn Gln Gln Arg385 390 395 400Ala Ser Ala Gly Gln Ile Ser Ile Gln Pro Thr Phe Ser Val Gln Arg Asn Leu Pro Phe405 410 415 420Asp Arg Thr Thr Val Met Ala Ala Phe Thr Gly Asn Thr Glu Gly Arg Thr Ser Asp Met425 430 435 440Arg Thr Glu Ile Ile Arg Met Met Glu Ser Ala Arg Pro Glu Asp Val Ser Phe Gln Gly445 450 455 460Arg Gly Val Phe Glu Leu Ser Asp Glu Lys Ala Ala Ser Pro Ile Val Pro Ser Phe Asp465 470 475 480
Met Ser Asn Glu Gly Ser Tyr Phe Phe Gly Asp Asn Ala Glu Glu Tyr Asp Asn ***485 490 495SEQ ID No.6ATG AAT CCA AAT CAA AAG ATA ATA ACA ATT GGC TCT GTC TCA CTC ACC ATT GCA ACA ATAMet Asn Pro Asn Gln Lys Ile Ile Thr Ile Gly Ser Val Ser Leu Thr Ile Ala Thr Ile5 10 15 20TGC TTC CTC ATG CAG ATT GCT ATC CTG GCA ACT ACT GTG ACA CTG CAT TTT AAG CAA CATCys Phe Leu Met Gln Ile Ala Ile Leu Ala Thr Thr Val Thr Leu His Phe Lys Gln His25 30 35 40GAA TGC AAC CCC CCT GCA ATC AAC CAA GTG ACC CCA TGT GAA CCA ATC ATA GTA GAA AGGGlu Cys Asn Pro Pro Ala Ile Asn Gln Val Thr Pro Cys Glu Pro Ile Ile Val Glu Arg45 50 55 60AAC ATA ACA GAG ATA GTG TAT TTG AAC AGC ACC AGT ATA GAG AGA GAG ATT TGC CCC GAAAsn Ile Thr Glu Ile Val Tyr Leu Asn Ser Thr Ser Ile Glu Arg Glu Ile Cys Pro Glu65 70 75 80GCT GTG GAA TAC AGA AAT TGG TCA AAG CCG CAG TGT CAA ATT ACA GGA TTT GCT CCT TTCAla Val Glu Tyr Arg Asn Trp Ser Lys Pro Gln Cys Gln Ile Thr Gly Phe Ala Pro Phe85 90 95 100TCT AAG GAC AAT TCA ATT CGG CTT TCT GCT GGT GGG GAC ATT TGG GTG ACG AGA GAA CCTSer Lys Asp Asn Ser Ile Arg Leu Ser Ala Gly lys Asp Ile Trp Val Thr Arg Glu Pro105 110 115 120TAT GTG TCA TGC GAT CCT GGT AAA TGT TAT CAA TTT GCA CTT GGA CAG GGG ACC ACA CTATyr Val Ser Cys Asp Pro Gly Lys Cys Tyr Gln Phe Ala Leu Gly Gln Gly Thr Thr Leu125 130 135 140GAC AAC AAA CAT TCG AAT GGT ACA ATA CAT GAT AGA ATC CCT CAT CGG ACT CTC TTG ATGAsp Asn Lys His Ser Asn Gly Thr Ile His Asp Arg Ile Pro His Arg Thr Leu Leu Met145 150 155 160AAC GAA TTG GGT GTT CCG TTC CAT TTA GGG ACC AAA CAA GTA TGC ATA GCA TGG TCC AGCAsn Glu Leu Gly Val Pro Phe His Leu Gly Thr Lys Gln Val Cys Ile Ala Trp Ser Ser165 170 175 180TCA AGT TGT CAC GAT GGA AGA GCA TGG TTG CAT ATT TGT GTT ACT GGG GAT GAT AGA AATSer Ser Cys His Asp Gly Arg Ala Trp Leu His Ile Cys Val Thr Gly Asp Asp Arg Asn185 190 195 200GCG ACT GCC AGT TTC ATT TAT GAC GGG AGG CTT GTC GAC AGT ATT GGT TCA TGG TCT CAAAla Thr Ala Ser Phe Ile Tyr Asp Gly Arg Leu Val Asp Ser Ile Gly Ser Trp Ser Gln205 210 215 220AAT ATT CTC AGA ACA CAG GAG TCA GAA TGT GTT TGT ATC AAT GGG ACT TGC ACT GTA GTAAsn Ile Leu Arg Thr Gln Glu Ser Glu Cys Val Cys Ile Asn Gly Thr Cys Thr Val Val225 230 235 240ATG ACT GAT GGA AGT GCA TCA GGA AGA GCC GAT ACG AGA ATA CTA TTC ATT AAA GAG GGGMet Thr Asp Gly Ser Ala Ser Gly Arg Ala Asp Thr Arg Ile Leu Phe Ile Lys Glu Gly245 250 255 260AAA ATT GTC CAT ATT AGC CCA TTG TCA GGA AGT GCT CAG CAT ATA GAG GAA TGT TCC TGTLys Ile Val His Ile Ser Pro Leu Ser Gly Ser Ala Gln His Ile Glu Glu Cys Ser Cys265 270 275 280TAT CCT CGA TAT CCT GAC GTC AGG TGT ATC TGC AGA GAT AAC TGG AAA GGC TCT AAT AGGTyr Pro Arg Tyr Pro Asp Val Arg Cys Ile Cys Arg Asp Asn Trp Lys Gly Ser Asn Arg285 290 295 300CCC GTT ATA GAC ATA AAT ATG GCA GAT TAT AGC ATT GAT TCC AGT TAT GTA TGT TCA GGGPro Val Ile Asp Ile Asn Met Ala Asp Tyr Ser Ile Asp Ser Ser Tyr Val Cys Ser Gly305 310 315 320CTC GTT GGT GAC ACA CCC AGA AAT GAC GAC AGC TCC AGC AGT AGC AAA AGC AGG GAT CCT
Leu Val Gly Asp Thr Pro Arg Asn Asp Asp Ser Ser Ser Ser Ser Lys Ser Arg Asp Pro325 330 335 340AAT AAT GAG AGA GGG AAT CCA GGG GTG AAA GGC TGG GCT TTT GAT AAT GGA GAT GAT GTGAsn Asn Glu Arg Gly Asn Pro Gly Val Lys Gly Trp Ala Phe Asp Asn Gly Asp Asp Val345 350 355 360TGG ATG GGG AGA ACA ATC AGC AAA GAT TCA CGC TCA GGT TAC GAG ACT TTC AAA GTC ATTTrp Met Gly Arg Thr Ile Ser Lys Asp Ser Arg Ser Gly Tyr Glu Thr Phe Lys Val Ile365 370 375 380GGT GGT TGG TCC ACA CCT AAT TCC AAG TCG CAG GTC AAT AGA CAG GTC ATA GTT GAC AACGly Gly Trp Ser Thr Pro Asn Ser Lys Ser Gln Val Asn Arg Gln Val Ile Val Asp Asn385 390 395 400AAT AAC AGG TCT GGT TAC TCT GGT ATT TTC TCT GTT GAG GGC AAA AGC TGC ATC AAT AGGAsn Asn Arg Ser Gly Tyr Ser Gly Ile Phe Ser Val Glu Gly Lys Ser Cys Ile Asn Arg405 410 415 420TGT TTT TAT GTG GAG TTG ATA AGG GGA AGA CCA CAG GAA ACT AGA GTA TGG TGG ACT TCACys Phe Tyr Val Glu Leu Ile Arg Gly Arg Pro Gln Glu Thr Arg Val Trp Trp Thr Ser425 430 435 440AAC AGT ATT GTT GTA TTT TGT GGC ACT TCA GGT ACC TAT GGA ACA GGC TCA TGG CCT GATAsn Ser Ile Val Val Phe Cys Gly Thr Ser Gly Thr Tyr Gly Thr Gly Ser Trp Pro Asp445 450 455 460GGG GCG AAC ATC AAT TTT ATG CCT ATA TAA *Gly Ala Asn Ile Asn Phe Met Pro Ile --- *465 470 475 480Met Asn Pro Asn Gln Lys Ile Ile Thr Ile Gly Ser Val Ser Leu Thr Ile Ala Thr Ile5 10 15 20Cys Phe Leu Met Gln Ile Ala Ile Leu Ala Thr Thr Val Thr Leu His Phe Lys Gln His25 30 35 40Glu Cys Asn Pro Pro Ala Ile Asn Gln Val Thr Pro Cys Glu Pro Ile Ile Val Glu Arg45 50 55 60Asn Ile Thr Glu Ile Val Tyr Leu Asn Ser Thr Ser Ile Glu Arg Glu Ile Cys Pro Glu65 70 75 80Ala Val Glu Tyr Arg Asn Trp Ser Lys Pro Gln Cys Gln Ile Thr Gly Phe Ala Pro Phe85 90 95 100Ser Lys Asp Asn Ser Ile Arg Leu Ser Ala Gly lys Asp Ile Trp Val Thr Arg Glu Pro105 110 115 120Tyr Val Ser Cys Asp Pro Gly Lys Cys Tyr Gln Phe Ala Leu Gly Gln Gly Thr Thr Leu125 130 135 140Asp Asn Lys His Ser Asn Gly Thr Ile His Asp Arg Ile Pro His Arg Thr Leu Leu Met145 150 155 160Asn Glu Leu Gly Val Pro Phe His Leu Gly Thr Lys Gln Val Cys Ile Ala Trp Ser Ser165 170 175 180Ser Ser Cys His Asp Gly Arg Ala Trp Leu His Ile Cys Val Thr Gly Asp Asp Arg Asn185 190 195 200Ala Thr Ala Ser Phe Ile Tyr Asp Gly Arg Leu Val Asp Ser lle Gly Ser Trp Ser Gln205 210 215 220Asn lle Leu Arg Thr Gln Glu Ser Glu Cys Val Cys Ile Asn Gly Thr Cys Thr Val Val225 230 235 240Met Thr Asp Gly Ser Ala Ser Gly Arg Ala Asp Thr Arg Ile Leu Phe Ile Lys Glu Gly245 250 255 260Lys lle Val His Ile Ser Pro Leu Ser Gly Ser Ala Gln His Ile Glu Glu Cys Ser Cys265 270 275 280Tyr Pro Arg Tyr Pro Asp Val Arg Cys Ile Cys Arg Asp Asn Trp Lys Gly Ser Asn Arg
285 290 295 300Pro Val Ile Asp Ile Asn Met Ala Asp Tyr Ser Ile Asp Ser Ser Tyr Val Cys Ser Gly305 310 315 320Leu Val Gly Asp Thr Pro Arg Asn Asp Asp Ser Ser Ser Ser Ser Lys Ser Arg Asp Pro325 330 335 340Asn Asn Glu Arg Gly Asn Pro Gly Val Lys Gly Trp Ala Phe Asp Asn Gly Asp Asp Val345 350 355 360Trp Met Gly Arg Thr Ile Ser Lys Asp Ser Arg Ser Gly Tyr Glu Thr Phe Lys Val Ile365 370 375 380Gly Gly Trp Ser Thr Pro Asn Ser Lys Ser Gln Val Asn Arg Gln Val Ile Val Asp Asn385 390 395 400Asn Asn Arg Ser Gly Tyr Ser Gly Ile Phe Ser Val Glu Gly Lys Ser Cys Ile Asn Arg405 410 415 420Cys Phe Tyr Val Glu Leu Ile Arg Gly Arg Pro Gln Glu Thr Arg Val Trp Trp Thr Ser425 430 435 440Asn Ser Ile Val Val Phe Cys Gly Thr Ser Gly Thr Tyr Gly Thr Gly Ser Trp Pro Asp445 450 455 460Gly Ala Asn Ile Asn Phe Met Pro Ile --- *465 470 475 480SEQ ID No.7ATG AGT CTT CTA ACC GAG GTC GAA ACG TAC GTT CTC TCT ATC ATC CCG TCA GGC CCC CTCMet Ser Leu Leu Thr Glu Val Glu Thr Tyr Val Leu Ser Ile Ile Pro Ser Gly Pro Leu5 10 15 20AAA GCC GAG ATC GCA CAG AGA CTT GAA GAT GTC TTT GCA GGG AAG AAC ACC GAT CTT GAGLys Ala Glu Ile Ala Gln Arg Leu Glu Asp Val Phe Ala Gly Lys Asn Thr Asp Leu Glu25 30 35 40GTT CTC ATG GAA TGG CTA AAG ACA AGA CCA ATC CTG TCA CCT CTG ACT AAG GGG ATT TTAVal Leu Met Glu Trp Leu Lys Thr Arg Pro Ile Leu Ser Pro Leu Thr Lys Gly Ile Leu45 50 55 60GGA TTT GTG TTC ACG CTC ACC GTG CCC AGT GAG CGA GGA CTG CAG CGT AGA CGC TTT GTCGly Phe Val Phe Thr Leu Thr Val Pro Ser Glu Arg Gly Leu Gln Arg Arg Arg Phe Val65 70 75 80CAA AAT GCC CTT AAT GGG AAC GGG GAT CCA AAT AAC ATG GAC AAA GCA GTT AAA CTG TATGln Asn Ala Leu Asn Gly Asn Gly Asp Pro Asn Asn Met Asp Lys Ala Val Lys Leu Tyr85 90 95 100AGG AAG CTC AAG AGG GAG ATA ACA TTC CAT GGG GCC AAA GAA ATC TCA CTC AGT TAT TCTArg Lys Leu Lys Arg Glu Ile Thr Phe His Gly Ala Lys Glu Ile Ser Leu Ser Tyr Ser105 110 115 120GCT GGT GCA CTT GCC AGT TGT ATG GGC CTC ATA TAC AAC AGG ATG GGG GCT GTG ACC ACTAla Gly Ala Leu Ala Ser Cys Met Gly Leu Ile Tyr Asn Arg Met Gly Ala Val Thr Thr125 130 135 140GAA GTG GCA TTT GGC CTG GTA TGT GCA ACC TGT GAA CAG ATT GCT GAC TCC CAG CAT CGGGlu Val Ala Phe Gly Leu Val Cys Ala Thr Cys Glu Gln Ile Ala Asp Ser Gln His Arg145 150 155 160TCT CAT AGG CAA ATG GTG ACA ACA ACC AAC CCA CTA ATC AGA CAT GAG AAC AGA ATG GTTSer His Arg Gln Met Val Thr Thr Thr Asn Pro Leu Ile Arg His Glu Asn Arg Met Val165 170 175 180TTA GCC AGC ACT ACA GCT AAG GCT ATG GAG CAA ATG GCT GGA TCG AGT GAG CAA GCA GCALeu Ala Ser Thr Thr Ala Lys Ala Met Glu Gln Met Ala Gly Ser Ser Glu Gln Ala Ala185 190 195 200GAG GCC ATG GAG GTT GCT AGT CAG GCT AGG CAA ATG GTG CAA GCG ATG AGA ACC ATT GGG
Glu Ala Met Glu Val Ala Ser Gln Ala Arg Gln Met Val Gln Ala Met Arg Thr Ile Gly205 210 215 220ACT CAT CCT AGC TCC AGT GCT GGT CTG AAA AAT GAT CTT CTT GAA AAT TTG CAG GCC TATThr His Pro Ser Ser Ser Ala Gly Leu Lys Asn Asp Leu Leu Glu Asn Leu Gln Ala Tyr225 230 235 240CAG AAA CGA ATG GGG GTG CAG ATG CAA CGG TTC AAG TGAGln Lys Arg Met Gly Val Gln Met Gln Arg Phe Lys ***245 250Met Ser Leu Leu Thr Glu Val Glu Thr Tyr Val Leu Ser Ile Ile Pro Ser Gly Pro Leu5 10 15 20Lys Ala Glu Ile Ala Gln Arg Leu Glu Asp Val Phe Ala Gly Lys Asn Thr Asp Leu Glu25 30 35 40Val Leu Met Glu Trp Leu Lys Thr Arg Pro Ile Leu Ser Pro Leu Thr Lys Gly Ile Leu45 50 55 60Gly Phe Val Phe Thr Leu Thr Val Pro Ser Glu Arg Gly Leu Gln Arg Arg Arg Phe Val65 70 75 80Gln Asn Ala Leu Asn Gly Asn Gly Asp Pro Asn Asn Met Asp Lys Ala Val Lys Leu Tyr85 90 95 100Arg Lys Leu Lys Arg Glu Ile Thr Phe His Gly Ala Lys Glu Ile Ser Leu Ser Tyr Ser105 110 115 120Ala Gly Ala Leu Ala Ser Cys Met Gly Leu Ile Tyr Asn Arg Met Gly Ala Val Thr Thr125 130 135 140Glu Val Ala Phe Gly Leu Val Cys Ala Thr Cys Glu Gln Ile Ala Asp Ser Gln His Arg145 150 155 160Ser His Arg Gln Met Val Thr Thr Thr Asn Pro Leu Ile Arg His Glu Asn Arg Met Val165 170 175 180Leu Ala Ser Thr Thr Ala Lys Ala Met Glu Gln Met Ala Gly Ser Ser Glu Gln Ala Ala185 190 195 200Glu Ala Met Glu Val Ala Ser Gln Ala Arg Gln Met Val Gln Ala Met Arg Thr Ile Gly205 210 215 220Thr His Pro Ser Ser Ser Ala Gly Leu Lys Asn Asp Leu Leu Glu Asn Leu Gln Ala Tyr225 230 235 240Gln Lys Arg Met Gly Val Gln Met Gln Arg Phe Lys ***245 250SEQ ID No.8ATG GAT CCA AAC ACT GTG TCA AGC TTT CAG GTA GAT TGC TTT CTT TGG CAT GTC CGC AAAMet Asp Pro Asn Thr Val Ser Ser Phe Gln Val Asp Cys Phe Leu Trp His Val Arg Lys5 10 15 20CGA GTT GCA GAC CAA GAA CTA GGT GAT GCC CCA TTC CTT GAT CGG CTT CGC CGA GAT CAGArg Val Ala Asp Gln Glu Leu Gly Asp Ala Pro Phe Leu Asp Arg Leu Arg Arg Asp Gln25 30 35 40AAA TCC CTA AGA GGA AGG GGC AGC ACT CTT GGT CTG GAC ATC GAG ACA GCC ACA CGT GCTLys Ser Leu Arg Gly Arg Gly Ser Thr Leu Gly Leu Asp Ile Glu Thr Ala Thr Arg Ala45 50 55 60GGA AAG CAG ATA GTG GAG CGG ATT CTG AAA GAA GAA TCC GAT GAG GCA CTT AAA ATG ACCGly Lys Gln Ile Val Glu Arg Ile Leu Lys Glu Glu Ser Asp Glu Ala Leu Lys Met Thr65 70 75 80ATG GCC TCT GTA CCT GCG TCG CGT TAC CTA ACC GAC ATG ACT CTT GAG GAA ATG TCA AGGMet Ala Ser Val Pro Ala Ser Arg Tyr Leu Thr Asp Met Thr Leu Glu Gln Met Ser Arg85 90 95 100GAA TGG TCC ATG CTC ATA CCC AAG CAG AAA GTG GCA GGC CCT CTT TGT ATC AGA ATG GAC
Glu Trp Ser Met Leu Ile Pro Lys Gln Lys Val Ala Gly Pro Leu Cys Ile Arg Met Asp105 110 115 120CAG GCG ATC ATG GAT AAA AAC ATC ATA CTG AAA GCG AAC TTC AGT GTG ATT TTT GAC CGGGln Ala Ile Met Asp Lys Asn Ile Ile Leu Lys Ala Asn Phe Ser Val Ile Phe Asp Arg125 130 135 140CTG GAG ACT CTA ATA TTG CTA AGG GCT TTC ACC GAA GAG GGA GCA ATT GTT GGC GAA ATTLeu Glu Thr Leu Ile Leu Leu Arg Ala Phe Thr Glu Glu Gly Ala Ile Val Gly Glu Ile145 150 155 160TCA CCA TTG CCT TCT CTT CCA GGA CAT ACT GCT GAG GAT GTC AAA AAT GCA GTT GGA GTCSer Pro Leu Pro Ser Leu Pro Gly His Thr Ala Glu Asp Val Lys Asn Ala Val Gly Val165 170 175 180CTC ATC GGA GGA CTT GAA TGG AAT GAT AAC ACA GTT CGA GTC TCT GAA ACT CTA CAG AGALeu Ile Gly Gly Leu Glu Trp Asn Asp Asn Thr Val Arg Val Ser Glu Thr Leu Gln Arg185 190 195 200TTC GCT TGG AGA AGC AGT AAT GAG AAT GGG AGA CCT CCA CTC ACT CCA AAA CAG AAA CGAPhe Ala Trp Arg Ser Ser Asn Glu Asn Gly Arg Pro Pro Leu Thr Pro Lys Gln Lys Arg205 210 215 220GAA ATG GCG GGA ACA ATT AGG TCA GAA GTT TGAGlu Met Ala Gly Thr Ile Arg Ser Glu Val ***225 230Met Asp Pro Asn Thr Val Ser Ser Phe Gln Val Asp Cys Phe Leu Trp His Val Arg Lys5 10 15 20Arg Val Ala Asp Gln Glu Leu Gly Asp Ala Pro Phe Leu Asp Arg Leu Arg Arg Asp Gln25 30 35 40Lys Ser Leu Arg Gly Arg Gly Ser Thr Leu Gly Leu Asp Ile Glu Thr Ala Thr Arg Ala45 50 55 60Gly Lys Gln Ile Val Glu Arg Ile Leu Lys Glu Glu Ser Asp Glu Ala Leu Lys Met Thr65 70 75 80Met Ala Ser Val Pro Ala Ser Arg Tyr Leu Thr Asp Met Thr Leu Glu Glu Met Ser Arg85 90 95 100Glu Trp Ser Met Leu Ile Pro Lys Gln Lys Val Ala Gly Pro Leu Cys Ile Arg Met Asp105 110 115 120Gln Ala Ile Met Asp Lys Asn Ile Ile Leu Lys Ala Asn Phe Ser Val Ile Phe Asp Arg125 130 135 140Leu Glu Thr Leu Ile Leu Leu Arg Ala Phe Thr Glu Glu Gly Ala Ile Val Gly Glu Ile145 150 155 160Ser Pro Leu Pro Ser Leu Pro Gly His Thr Ala Glu Asp Val Lys Asn Ala Val Gly Val165 170 175 180Leu Ile Gly Gly Leu Glu Trp Asn Asp Asn Thr Val Arg Val Ser Glu Thr Leu Gln Arg185 190 195 200Phe Ala Trp Arg Ser Ser Asn Glu Asn Gly Arg Pro Pro Leu Thr Pro Lys Gln Lys Arg205 210 215 220Glu Met Ala Gly Thr Ile Arg Ser Glu Val ***225 230
权利要求
1一种人工重组的H7亚型流感病毒，其组成包括H7亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)的HA和NA基因，其特征是所述的病毒上还重组有流感病毒A/PR/8/34(H1N1)的6个内部基因PB2、PB1、PA、NP、M和NS，保藏号CCTCC-V200415。
2根据权利要求1所述的人工重组的H7亚型流感病毒，其特征是HA和NA基因具有序列表SEQ ID No.4和SEQ ID No.6所述的序列，6个内部基因PB2、PB1、PA、NP、M和NS具有序列表SEQ ID No.1、SEQ ID No.2、SEQID No.3、SEQ ID No.5、SEQ ID No.7和SEQ ID No.8所述的序列，该病毒对我国流行的H7N2禽流感病毒具有特异免疫性，具有高滴度鸡胚适应性，对鸡及鸡胚无致病力。
3一种权利要求1或2所述的人工重组的H7亚型流感病毒的制备方法，包括以下步骤a.共感染-使两个病毒基因组发生重组流感病毒A/PR/8/34(PR8)(H1N1)与经紫外线处理过的H7N2亚型AIV国内分离株A/Chicken/HeBei/1/02(H7N2)共同接种10日龄鸡胚尿囊腔；b.筛选-除去表面基因来源于PR8病毒的病毒在上述特定的人工条件下收获的尿囊液，与倍比稀释的抗PR8的血清混合，室温下孵育之后接种鸡胚尿囊腔，收获后，对鸡胚尿囊液进行血凝活性检测，收获血清稀释度最低并且具有血凝活性的样品用于第二轮筛选；c.稀释纯化-除去内部基因来源于H7N2病毒同上方法筛选得到的病毒在鸡胚上进行3代有限克隆纯化，利用片段特异性引物RT-PCR鉴定并经过序列分析筛选HA和NA来自于H7N2、6个内部基因PB2、PB1、PA、NP、M及NS来自PR8的重组病毒。
4.上述的人工重组的H7亚型流感病毒，作为防制动物和人类H7亚型流感的疫苗株，应用于疫苗防制。
全文摘要
人工重组的H7亚型流感病毒及其制备方法和应用，在国内，虽然高致病性的H7亚型流感病毒还没有分离到，但是也曾分离到过一株自然分离株H7亚型LPAIV。人工重组的H7亚型流感病毒，其组成包括H7亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)的HA和NA基因，所述的病毒上还重组有流感病毒A/PR/8/34(H1N1)的6个内部基因PB2、PB1、PA、NP、M和NS，保藏号CCTCC－V200415。该病毒对我国流行的H7N2禽流感病毒具有特异免疫性，具有高滴度鸡胚适应性，对鸡及鸡胚无致病力。
文档编号A61K39/00GK1746298SQ20041004386
公开日2006年3月15日 申请日期2004年9月10日 优先权日2004年9月10日
发明者陈化兰, 施建忠, 田国彬, 李雁冰 申请人:中国农业科学院哈尔滨兽医研究所
技术领域：
本发明涉及一株人工重组的流感病毒，该病毒的制备和应用。
背景技术：
禽流感病毒(avian influenza virus，AIV)是禽类及一些哺乳动物的呼吸系统疾病的重要病原。AIV在分类学上属于病毒界(Vira)----正粘病毒科(Orthomyxoviridae)----流感病毒属(Influenza Virus A and B)----禽流感病毒(avian influenza virus)。
根据表面蛋白抗原性不同，禽流感病毒(avian influenza virus，AIV)被分为15个血凝素(HA)和9个神经氨酸酶(NA)亚型。H7亚型AIV 1878年首次在意大利出现，该“真性鸡瘟”的病原是A/Chicken/Brescia/1902(H7N7)，其是禽流感历史上第一次文字记载的HPAIV，后来人们又分离到多株H7亚型AIV，神经氨酸酶亚型主要为N1、N2、N3、N7等，其中HPAIV占据一定的比例。在国内，虽然高致病性的H7亚型流感病毒还没有分离到，但是也曾分离到过一株自然分离株H7亚型LPAIV，而且对于高致病性的H7亚型病毒的高度危害，我们深信不疑。为了防止那可能发生的危害，做好高效力的H7亚型的流感疫苗，做好技术储备刻不容缓。但由于现地分离的野毒株鸡胚适应性较差，致使现行预储备的疫苗存在成本高、疫苗免疫期短等缺点。
发明内容
本发明的目的是提供一种人工重组的H7亚型流感病毒、该病毒的制备方法和应用，该病毒对我国流行的H7N2禽流感病毒具有特异免疫性，具有高滴度鸡胚适应性，对鸡及鸡胚无致病力。
本发明的目的是这样实现的一种人工重组的H7亚型流感病毒，其组成包括H7亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)的HA和NA基因，所述的病毒上还重组有流感病毒A/PR/8/34(H1N1)的6个内部基因PB2、PB1、PA、NP、M和NS，保藏号CCTCC-V200415。
上述的人工重组的H7亚型流感病毒，HA和NA基因具有序列表SEQ IDNo.4和SEQ ID No.6所述的序列，6个内部基因PB2、PB1、PA、NP、M和NS具有序列表SEQ ID No.1、SEQ ID No.2、SEQ ID No.3、SEQ ID No.5、SEQ ID No.7和SEQ ID No.8所述的序列，该病毒对我国流行的H7N2禽流感病毒具有特异免疫性，具有高滴度鸡胚适应性，对鸡及鸡胚无致病力。
上述的人工重组的H7亚型流感病毒的制备方法，包括以下步骤a.共感染-使两个病毒基因组发生重组流感病毒A/PR/8/34(PR8)(H1N1)与经紫外线处理过的H7N2亚型AIV国内分离株A/Chicken/HeBei/1/02(H7N2)共同接种10日龄鸡胚尿囊腔；b.筛选-除去表面基因来源于PR8病毒的病毒在上述特定的人工条件下收获的尿囊液，与倍比稀释的抗PR8的血清混合，室温下孵育之后接种鸡胚尿囊腔，收获后，对鸡胚尿囊液进行血凝活性检测，收获血清稀释度最低并且具有血凝活性的样品用于第二轮筛选；c.稀释纯化-除去内部基因来源于H7N2病毒同上方法筛选得到的病毒在鸡胚上进行3代有限克隆纯化，利用片段特异性引物RT-PCR鉴定并经过序列分析筛选HA和NA来自于H7N2、6个内部基因PB2、PB1、PA、NP、M及NS来自PR8的重组病毒。
上述的人工重组的H7亚型流感病毒，作为防制动物和人类H7亚型流感的疫苗株，应用于疫苗防制。
该病毒HA和NA基因来自H7N2亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)，6个内部基因来自PR8病毒；该病毒与我国H7N2亚型禽流感病毒流行株抗原性一致，并且具有良好的鸡胚适应性；作为防制H7亚型动物流感的疫苗株。
本发明的优点1.本发明Re-3以我国禽流感病毒H7N2为HA和NA基因供体，因此，该病毒与我国曾流行的H7N2亚型禽流感病毒抗原性一致。以此为疫苗株生产的疫苗，将对H7N2亚型禽流感病毒流行株产生良好的特异保护性。
2.本发明Re-3含有鸡胚高滴度适应株PR8的6个内部基因，病毒生长滴度与野毒相比，可提高10倍左右，以此为疫苗株生产疫苗，可大大降低疫苗生产成本，提高疫苗质量。
流感病毒A/PR/8/34(H1N1)(PR8)是实验室鸡胚适应株，即该病毒可在鸡胚内生长至很高滴度，这种鸡胚适应性由病毒的6个内部基因决定。另外，当两个流感病毒共同感染一个细胞或鸡胚时，可发生基因互换，从而产生新的基因重组病毒。因此，通过遗传重组方法，可将PR8病毒6个内部基因和流行株的HA、NA基因的进行重组而获得一个新的重组病毒，该重组病毒具有与HA和NA基因供体毒株一致的抗原性，同时还具有PR8病毒的鸡胚高滴度生长特性。
它是以H7N2亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)和A/PR/8/34(H1N1)(PR8)共同接种鸡胚，应用PR8的抗血清对鸡胚培养物进行2轮抗体筛选后、继续在鸡胚上进行有限稀释克隆纯化3代获得的、经测序确证含有H7N2的表面基因HA和NA，同时拥有PR8的6个内部基因PB2，PB1，PA，NP，M及NS的重组病毒。该病毒鸡胚生长滴度比自然分离株H7N2提高10倍左右，是理想的H7N2亚型禽流感疫苗株。
本发明的
具体实施例方式实施例1一种人工重组的H7亚型流感病毒，其组成包括H7亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)的HA和NA基因，所述的病毒上还重组有流感病毒A/PR/8/34(H1N1)的6个内部基因PB2、PB1、PA、NP、M和NS，保藏在中国典型培养物保藏中心，保藏号CCTCC-V200415，保藏日2004年9月6日。
实施例2实施例1所述的人工重组的H7亚型流感病毒，HA和NA基因具有序列表SEQ ID No.4和SEQ ID No.6所述的序列，6个内部基因PB2、PB1、PA、NP、M和NS具有序列表SEQ ID No.1、SEQ ID No.2、SEQ ID No.3、SEQ ID No.5、SEQ ID No.7和SEQ ID No.8所述的序列(附后)，该病毒对我国流行的H7N2禽流感病毒具有特异免疫性，具有高滴度鸡胚适应性，对鸡及鸡胚无致病力。]实施例3实施例1或2所述的人工重组的H7亚型流感病毒的制备方法，包括以下步骤a.共感染-使两个病毒基因组发生重组流感病毒A/PR/8/34(PR8)(H1N1)与经紫外线处理过的H7N2亚型AIV国内分离株A/Chicken/HeBei/1/02(H7N2)共同接种10日龄鸡胚尿囊腔；b.筛选-除去表面基因来源于PR8病毒的病毒在上述特定的人工条件下收获的尿囊液，与倍比稀释的抗PR8的血清混合，室温下孵育之后接种鸡胚尿囊腔，收获后，对鸡胚尿囊液进行血凝活性检测，收获血清稀释度最低并且具有血凝活性的样品用于第二轮筛选；c.稀释纯化-除去内部基因来源于H7N2病毒同上方法筛选得到的病毒在鸡胚上进行3代有限克隆纯化，利用片段特异性引物RT-PCR鉴定并经过序列分析筛选HA和NA来自于H7N2、6个内部基因PB2、PB1、PA、NP、M及NS来自PR8的重组病毒。
实施例4上述的人工重组的H7亚型流感病毒，作为防制动物和人类H7亚型流感的疫苗株，应用于疫苗防制。
基因序列附后。
重组流感病毒A/哈尔滨/Re-2/2003(H9N2)基因序列SEQ ID No.1ATG GAA AGA ATA AAA GAA CTA AGA AAT CTA ATG TCG CAG TCT CGC ACC CGC GAG ATA CTCMet Glu Arg Ile Lys Glu Leu Arg Asn Leu Met Ser Gln Ser Arg Thr Arg Glu Ile Leu5 10 15 20ACA AAA ACC ACC GTG GAC CAT ATG GCC ATA ATC AAG AAG TAC ACA TCA GGA AGA CAG GAGThr Lys Thr Thr Val Asp His Met Ala Ile Ile Lys Lys Tyr Thr Ser Gly Arg Gln Glu25 30 35 40AAG AAC CCA GCA CTT AGG ATG AAA TGG ATG ATG GCA ATG AAA TAT CCA ATT ACA GCA GACLys Asn Pro Ala Leu Arg Met Lys Trp Met Met Ala Met Lys Tyr Pro Ile Thr Ala Asp45 50 55 60AAG AGG ATA ACG GAA ATG ATT CCT GAG AGA AAT GAG CAA GGA CAA ACT TTA TGG AGT AAALys Arg Ile Thr Glu Met Ile Pro Glu Arg Asn Glu Gln Gly Gln Thr Leu Trp Ser Lys65 70 75 80ATG AAT GAT GCC GGA TCA GAC CGA GTG ATG GTA TCA CCT CTG GCT GTG ACA TGG TGG AATMet Asn Asp Ala Gly Ser Asp Arg Val Met Val Ser Pro Leu Ala Val Thr Trp Trp Asn85 90 95 100AGG AAT GGA CCA ATG ACA AAT ACA GTT CAT TAT CCA AAA ATC TAC AAA ACT TAT TTT GAAArg Asn Gly Pro Met Thr Asn Thr Val His Tyr Pro Lys Ile Tyr Lys Thr Tyr Phe Glu105 110 115 120AGA GTC GAA AGG CTA AAG CAT GGA ACC TTT GGC CCT GTC CAT TTT AGA AAC CAA GTC AAAArg Val Glu Arg Leu Lys His Gly Thr Phe Gly Pro Val His Phe Arg Asn Gln Val Lys125 130 135 140ATA CGT CGG AGA GTT GAC ATA AAT CCT GGT CAT GCA GAT CTC AGT GCC AAG GAG GCA CAGIle Arg Arg Arg Val Asp Ile Asn Pro Gly His Ala Asp Leu Ser Ala Lys Glu Ala Gln145 150 155 160GAT GTA ATC ATG GAA GTT GTT TTC CCT AAC GAA GTG GGA GCC AGG ATA CTA ACA TCG GAAAsp Val Ile Met Glu Val Val Phe Pro Asn Glu Val Gly Ala Arg Lle Leu Thr Ser Glu165 170 175 180TCG CAA CTA ACG ATA ACC AAA GAG AAG AAA GAA GAA CTC CAG GAT TGC AAA ATT TCT CCTSer Gln Leu Thr Ile Thr Lys Glu Lys Lys Glu Glu Leu Gln Asp Cys Lys Ile Ser Pro185 190 195 200TTG ATG GTT GCA TAC ATG TTG GAG AGA GAA CTG GTC CGC AAA ACG AGA TTC CTC CCA GTGLeu Met Val Ala Tyr Met Leu Glu Arg Glu Leu Val Arg Lys Thr Arg Phe Leu Pro Val205 210 215 220GCT GGT GGA ACA AGC AGT GTG TAC ATT GAA GTG TTG CAT TTG ACT CAA GGA ACA TGC TGGAla Gly Gly Thr Ser Ser Val Tyr Ile Glu Val Leu His Leu Thr Gln Gly Thr Cys Trp225 230 235 240GAA CAG ATG TAT ACT CCA GGA GGG GAA GTG AAG AAT GAT GAT GTT GAT CAA AGC TTG ATTGlu Gln Met Tyr Thr Pro Gly Gly Glu Val Lys Asn Asp Asp Val Asp Gln Ser Leu Ile245 250 255 260ATT GCT GCT AGG AAC ATA GTG AGA AGA GCT GCA GTA TCA GCA GAC CCA CTA GCA TCT TTAIle Ala Ala Arg Asn Ile Val Arg Arg Ala Ala Val Ser Ala Asp Pro Leu Ala Ser Leu265 270 275 280TTG GAG ATG TGC CAC AGC ACA CAG ATT GGT GGA ATT AGG ATG GTA GAC ATC CTT AAG CAGLeu Glu Met Cys His Ser Thr Gln Ile Gly Gly Ile Arg Met Val Asp Ile Leu Lys Gln285 290 295 300AAC CCA ACA GAA GAG CAA GCC GTG GGT ATA TGC AAG GCT GCA ATG GGA GTG AGA ATT AGCAsn Pro Thr Glu Glu Gln Ala Val Gly Ile Cys Lys Ala Ala Met Gly Leu Arg Ile Ser305 310 315 320TCA TCC TTC AGT TTT GGT GGA TTC ACA TTT AAG AGA ACA AGC GGA TCA TCA GTC AAG AGASer Ser Phe Ser Phe Gly Gly Phe Thr Phe Lys Arg Thr Ser Gly Ser Ser Val Lys Arg
325 330 335 340GAG GAA GAG GTG CTT ACG GGC AAT CTT CAA ACA TTG AAG ATA AGA GTG CAT GAG GGA TATGlu Glu Glu Val Leu Thr Gly Asn Leu Gln Thr Leu Lys Ile Arg Val His Glu Gly Tyr345 350 355 360GAA GAG TTC ACA ATG GTT GGG AGA AGA GCA ACA GCC ATA CTC AGA AAA GCA ACC AGG AGAGlu Glu Phe Thr Met Val Gly Arg Arg Ala Thr Ala Ile Leu Arg Lys Ala Thr Arg Arg365 370 375 380TTG ATT CAG CTG ATA GTG AGT GGG AGA GAC GAA CAG ICG ATT GCC GAA GCA ATA ATT GTGLeu Ile Gln Leu Ile Val Ser Gly Arg Asp Glu Gln Ser Ile Ala Glu Ala Ile Ile Val385 390 395 400GCC ATG GTA TTT TCA CAA GAG GAT TGT ATG ATA AAA GCA GTT AGA GGT GAT CTG AAT TTCAla Met Val Phe Ser Gln Glu Asp Cys Met Ile Lys Ala Val Arg Gly Asp Leu Asn Phe405 410 415 420GTC AAT AGG GCG AAT CAG CGA CTG AAT CCT ATG CAT CAA CTT TTA AGA CAT TTT CAG AAGVal Asn Arg Ala Asn Gln Arg Leu Asn Pro Met His Gln Leu Leu Arg His Phe Gln Lys425 430 435 440GAT GCG AAA GTG CTT TTT CAA AAT TGG GGA GTT GAA CCT ATC GAC AAT GTG ATG GGA ATGAsp Ala Lys Val Leu Phe Gln Asn Trp Gly Val Glu Pro Ile Asp Asn Val Met Gly Met445 450 455 460ATT GGG ATA TTG CCC GAC ATG ACT CCA AGC ATC GAG ATG TCA ATG AGA GGA GTG AGA ATCIle Gly Ile Leu Pro Asp Met Thr Pro Ser Ile Glu Met Ser Met Arg Gly Val Arg Ile465 470 475 480AGC AAA ATG GGT GTA GAT GAG TAC TCC AGC ACG GAG AGG GTA GTG GTG AGC ATT GAC CGGSer Lys Met Gly Val Asp Glu Tyr Ser Ser Thr Glu Arg Val Val Val Ser Ile Asp Arg485 490 495 500TTC TTG AGA GTC CGG GAC CAA CGA GGA AAT GTA CTA CTG TCT CCC GAG GAG GTC AGT GAAPhe Leu Arg Val Arg Asp Gln Arg Gly Asn Val Leu Leu Ser Pro Glu Glu Val Ser Glu505 510 515 520ACA CAG GGA ACA GAG AAA CTG ACA ATA ACT TAC TCA TCG TCA ATG ATG TGG GAG ATT AATThr Gln Gly Thr Glu Lys Leu Thr Ile Thr Tyr Ser Ser Ser Met Met Trp Glu Ile Asn525 530 535 540GGT CCT GAA TCA GTG TTG GTC AAT ACC TAT CAA TGG ATC ATC AGA AAC TGG GAA ACT GTTGly Pro Glu Ser Val Leu Val Asn Thr Tyr Gln Trp Ile Ile Arg Asn Trp Glu Thr Val545 550 555 560AAA ATT CAG TGG TCC CAG AAC CCT ACA ATG CTA TAC AAT AAA ATG GAA TTT GAA CCA TTTLys Ile Gln Trp Ser Gln Asn Pro Thr Met Leu Tyr Asn Lys Met Glu Phe Glu Pro Phe565 570 575 580CAG TCT TTA GTA CCT AAG GCC ATT AGA GGC CAA TAC AGT GGG TTT GTG AGA ACT CTG TTCGln Ser Leu Val Pro Lys Ala Ile Arg Gly Gln Tyr Ser Gly Phe Val Arg Thr Leu Phe585 590 595 600CAA CAA ATG AGG GAT GTG CTT GGG ACA TTT GAT ACC GCA CAG ATA ATA AAA CTT CTT CCCGln Gln Met Arg Asp Val Leu Gly Thr Phe Asp Thr Ala Gln Ile Ile Lys Leu Leu Pro605 610 615 620TTC GCA GCC GCT CCA CCA AAG CAA AGT AGA ATG CAG TTC TCC TCA TTT ACT GTG AAT GTGPhe Ala Ala Ala Pro Pro Lys Gln Ser Arg Met Gln Phe Ser Ser Phe Thr Val Asn Val625 630 635 640AGG GGA TCA GGA ATG AGA ATA CTT GTA AGG GGC AAT TCT CCT GTA TTC AAC TAC AAC AAGArg Gly Ser Gly Met Arg Ile Leu Val Arg Gly Asn Ser Pro Val Phe Asn Tyr Asn Lys645 650 655 660GCC ACG AAG AGA CTC ACA GTT CTC GGA AAG GAT GCT GGC ACT TTA ACC GAA GAC CCA GATAla Thr Lys Arg Leu Thr Val Leu Gly Lys Asp Ala Gly Thr Leu Thr Glu Asp Pro Asp665 670 675 680GAA GGC ACA GCT GGA GTG GAG TCC GCT GTT CTG AGG GGA TTC CTC ATT CTG GGC AAA GAA
Glu Gly Thr Ala Gly Val Glu Ser Ala Val Leu Arg Gly Phe Leu Ile Leu Gly Lys Glu685 690 695 700GAC AGG AGA TAT GGG CCA GCA TTA AGC ATC AAT GAA CTG AGC AAC CTT GCG AAA GGA GAGAsp Arg Arg Tyr Gly Pro Ala Leu Ser Ile Asn Glu Leu Ser Asn Leu Ala Lys Gly Glu705 710 715 720AAG GCT AAT GTG CTA ATT GGG CAA GGA GAC GTG GTG TTG GTA ATG AAA CGA AAA CGG GACLys Ala Asn Val Leu Ile Gly Gln Gly Asp Val Val Leu Val Met Lys Arg Lys Arg Asp725 730 735 740TCT AGC ATA CTT ACT GAC AGC CAG ACA GCG ACC AAA AGA ATT CGG ATG GCC ATC AATSer Ser Ile Leu Thr Asp Ser Gln Thr Ala Thr Lys Arg Ile Arg Met Ala Ile Asn745 750 755 759Met Glu Arg Ile Lys Glu Leu Arg Asn Leu Met Ser Gln Ser Arg Thr Arg Glu Ile Leu5 10 15 20Thr Lys Thr Thr Val Asp His Met Ala Ile Ile Lys Lys Tyr Thr Ser Gly Arg Gln Glu25 30 35 40Lys Asn Pro Ala Leu Arg Met Lys Trp Met Met Ala Met Lys Tyr Pro Ile Thr Ala Asp45 50 55 60Lys Arg Ile Thr Glu Met Ile Pro Glu Arg Asn Glu Gln Gly Gln Thr Leu Trp Ser Lys65 70 75 80Met Asn Asp Ala Gly Ser Asp Arg Val Met Val Ser Pro Leu Ala Val Thr Trp Trp Asn85 90 95 100Arg Asn Gly Pro Met Thr Asn Thr Val His Tyr Pro Lys Ile Tyr Lys Thr Tyr Phe Glu105 110 115 120Arg Val Glu Arg Leu Lys His Gly Thr Phe Gly Pro Val His Phe Arg Asn Gln Val Lys125 130 135 140Ile Arg Arg Arg Val Asp Ile Asn Pro Gly His Ala Asp Leu Ser Ala Lys Glu Ala Gln145 150 155 160Asp Val Ile Met Glu Val Val Phe Pro Asn Glu Val Gly Ala Arg Ile Leu Thr Ser Glu165 170 175 180Ser Gln Leu Thr Ile Thr Lys Glu Lys Lys Glu Glu Leu Gln Asp Cys Lys Ile Ser Pro185 190 195 200Leu Met Val Ala Tyr Met Leu Glu Arg Glu Leu Val Arg Lys Thr Arg Phe Leu Pro Val205 210 215 220Ala Gly Gly Thr Ser Ser Val Tyr Ile Glu Val Leu His Leu Thr Gln Gly Thr Cys Trp225 230 235 240Glu Gln Met Tyr Thr Pro Gly Gly Glu Val Lys Asn Asp Asp Val Asp Gln Ser Leu Ile245 250 255 260lle Ala Ala Arg Asn Ile Val Arg Arg Ala Ala Val Ser Ala Asp Pro Leu Ala Ser Leu265 270 275 280Leu Glu Met Cys His Ser Thr Gln Ile Gly Gly Ile Arg Met Val Asp Ile Leu Lys Gln285 290 295 300Asn Pro Thr Glu Glu Gln Ala Val Gly Ile Cys Lys Ala Ala Met Gly Leu Arg Ile Ser305 310 315 320Ser Ser Phe Ser Phe Gly Gly Phe Thr Phe Lys Arg Thr Ser Gly Ser Ser Val Lys Arg325 330 335 340Glu Glu Glu Val Leu Thr Gly Asn Leu Gln Thr Leu Lys Ile Arg Val His Glu Gly Tyr345 350 355 360Glu Glu Phe Thr Met Val Gly Arg Arg Ala Thr Ala Ile Leu Arg Lys Ala Thr Arg Arg365 370 375 380Leu Ile Gln Leu Ile Val Ser Gly Arg Asp Glu Gln Ser Ile Ala Glu Ala Ile Ile Val385 390 395 400
Ala Met Val Phe Ser Gln Glu Asp Cys Met Ile Lys Ala Val Arg Gly Asp Leu Asn Phe405 410 415 420Val Asn Arg Ala Asn Gln Arg Leu Asn Pro Met His Gln Leu Leu Arg His Phe Gln Lys425 430 435 440Asp Ala Lys Val Leu Phe Gln Asn Trp Gly Val Glu Pro Ile Asp Asn Val Met Gly Met445 450 455 460Ile Gly Ile Leu Pro Asp Met Thr Pro Ser Ile Glu Met Ser Met Arg Gly Val Arg Ile465 470 475 480Ser Lys Met Gly Val Asp Glu Tyr Ser Ser Thr Glu Arg Val Val Val Ser Ile Asp Arg485 490 495 500Phe Leu Arg Val Arg Asp Gln Arg Gly Asn Val Leu Leu Ser Pro Glu Glu Val Ser Glu505 510 515 520Thr Gln Gly Thr Glu Lys Leu Thr Ile Thr Tyr Ser Ser Ser Met Met Trp Glu Ile Asn525 530 535 540Gly Pro Glu Ser Val Leu Val Asn Thr Tyr Gln Trp Ile Ile Arg Asn Trp Glu Thr Val545 550 555 560Lys Ile Gln Trp Ser Gln Asn Pro Thr Met Leu Tyr Asn Lys Met Glu Phe Glu Pro Phe565 570 575 580Gln Ser Leu Val Pro Lys Ala lle Arg Gly Gln Tyr Ser Gly Phe Val Arg Thr Leu Phe585 590 595 600Gln Gln Met Arg Asp Val Leu Gly Thr Phe Asp Thr Ala Gln Ile Ile Lys Leu Leu Pro605 610 615 620Phe Ala Ala Ala Pro Pro Lys Gln Ser Arg Met Gln Phe Ser Ser Phe Thr Val Asn Val625 630 635 640Arg Gly Ser Gly Met Arg Ile Leu Val Arg Gly Asn Ser Pro Val Phe Asn Tyr Asn Lys645 650 655 660Ala Thr Lys Arg Leu Thr Val Leu Gly Lys Asp Ala Gly Thr Leu Thr Glu Asp Pro Asp665 670 675 680Glu Gly Thr Ala Gly Val Glu Ser Ala Val Leu Arg Gly Phe Leu Ile Leu Gly Lys Glu685 690 695 700Asp Arg Arg Tyr Gly Pro Ala Leu Ser Ile Asn Glu Leu Ser Asn Leu Ala Lys Gly Glu705 710 715 720Lys Ala Asn Val Leu Ile Gly Gln Gly Asp Val Val Leu Val Met Lys Arg Lys Arg Asp725 730 735 740Ser Ser Ile Leu Thr Asp Ser Gln Thr Ala Thr Lys Arg Ile Arg Met Ala lle Asn45 750 755 759SEQ ID No.2ATG GAT GTC AAT CCG ACC TTA CTT TTC TTA AAA GTG CCA GCA CAA AAT GCT ATA AGC ACAMet Asp Val Asn Pro Thr Leu Leu Phe Leu Lys Val Pro Ala Gln Asn Ala Ile Ser Thr5 10 15 20ACT TTC CCT TAT ACC GGA GAC CCT CCT TAC AGC CAT GGG ACA GGA ACA GGA TAC ACC ATGThr Phe Pro Tyr Thr Gly Asp Pro Pro Tyr Ser His Gly Thr Gly Thr Gly Tyr Thr Met25 30 35 40GAT ACT GTC AAC AGG ACA CAT CAG TAC TCA GAA AAG GCA AGA TGG ACA ACA AAC ACC GAAAsp Thr Val Asn Arg Thr His Gln Tyr Ser Glu Lys Ala Arg Trp Thr Thr Asn Thr Glu45 50 55 60ACT GGA GCA CCG CAA CTC AAC CCG ATT GAT GGG CCA CTG CCA GAA GAC AAT GAA CCA AGTThr Gly Ala Pro Gln Leu Asn Pro Ile Asp Gly Pro Leu Pro Glu Asp Asn Glu Pro Ser65 70 75 80GGT TAT GCC CAA ACA GAT TGT GTA TTG GAA GCA ATG GCT TTC CTT GAG GAA TCC CAT CCTGly Tyr Ala Gln Thr Asp Cys Val Leu Glu Ala Met Ala Phe Leu Glu Glu Ser His Pro85 90 95 100
GGT ATT TTT GAA AAC TCG TGT ATT GAA ACG ATG GAG GTT GTT CAG CAA ACA CGA GTA GACGly Ile Phe Glu Asn Ser Cys Ile Glu Thr Met Glu Val Val Gln Gln Thr Arg Val Asp105 110 115 120AAG CTG ACA CAA GGC CGA CAG ACC TAT GAC TGG ACT TTA AAT AGA AAC CAG CCT GCT GCALys Leu Thr Gln Gly Arg Gln Thr Tyr Asp Trp Thr Leu Asn Arg Asn Gln Pro Ala Ala125 130 135 140ACA GCA TTG GCC AAC ACA ATA GAA GTG TTC AGA TCA AAT GGC CTC ACG GCC AAT GAG TCTThr Ala Leu Ala Asn Thr Ile Glu Val Phe Arg Ser Asn Gly Leu Thr Ala Asn Glu Ser145 150 155 160GGA AGG CTC ATA GAC TTC CTT AAG GAT GTA ATG GAG TCA ATG AAA AAA GAA GAA ATG GGGGly Arg Leu Ile Asp Phe Leu Lys Asp Val Met Glu Ser Met Lys Lys Glu Glu Met Gly165 170 175 180ATC ACA ACT CAT TTT CAG AGA AAG AGA CGG GTG AGA GAC AAT ATG ACT AAG AAA ATG ATAIle Thr Thr His Phe Gln Arg Lys Arg Arg Val Arg Asp Asn Met Thr Lys Lys Met Ile185 190 195 200ACA CAG AGA ACA ATA GGT AAA AGG AAA CAG AGA TTG AAC AAA AGG AGT TAT CTA ATT AGAThr Gln Arg Thr Ile Gly Lys Arg Lys Gln Arg Leu Asn Lys Arg Ser Tyr Leu Ile Arg205 210 215 220GCA TTG ACC CTG AAC ACA ATG ACC AAA GAT GCT GAG AGA GGG AAG CTA AAA CGG AGA GCAAla Leu Thr Leu Asn Thr Met Thr Lys Asp Ala Glu Arg Gly Lys Leu Lys Arg Arg Ala225 230 235 240ATT GCA ACC CCA GGG ATG CAA ATA AGG GGG TTT GTA TAC TTT GTT GAG ACA CTG GCA AGGIle Ala Thr Pro Gly Met Gln Ile Arg Gly Phe Val Tyr Phe Val Glu Thr Leu Ala Arg245 250 255 260AGT ATA TGT GAG AAA CTT GAA CAA TCA GGG TTG CCA GTT GGA GGC AAT GAG AAG AAA GCASer Ile Cys Glu Lys Leu Glu Gln Ser Gly Leu Pro Val Gly Gly Asn Glu Lys Lys Ala265 270 275 280AAG TTG GCA AAT GTT GTA AGG AAG ATG ATG ACC AAT TCT CAG GAC ACC GAA CTT TCT TTGLys Leu Ala Asn Val Val Arg Lys Met Met Thr Asn Ser Gln Asp Thr Glu Leu Ser Leu285 290 295 300ACC ATC ACT GGA GAT AAC ACC AAA TGG AAC GAA AAT CAG AAT CCT CGG ATG TTT TTG GCCThr Ile Thr Gly Asp Asn Thr Lys Trp Asn Glu Asn Gln Asn Pro Arg Met Phe Leu Ala305 310 315 320ATG ATC ACA TAT ATG ACC AGA AAT CAG CCC GAA TGG TTC AGA AAT GTT CTA AGT ATT GCTMet Ile Thr Tyr Met Thr Arg Asn Gln Pro Glu Trp Phe Arg Asn Val Leu Ser Ile Ala325 330 335 340CCA ATA ATG TTC TCA AAC AAA ATG GCG AGA CTG GGA AAA GGG TAT ATG TTT GAG AGC AAGPro Ile Met Phe Ser Asn Lys Met Ala Arg Leu Gly Lys Gly Tyr Met Phe Glu Ser Lys345 350 355 360AGT ATG AAA CTT AGA ACT CAA ATA CCT GCA GAA ATG CTA GCA AGC ATT GAT TTG AAA TATSer Met Lys Leu Arg Thr Gln Ile Pro Ala Glu Met Leu Ala Ser Ile Asp Leu Lys Tyr365 370 375 380TTC AAT GAT TCA ACA AGA AAG AAG ATT GAA AAA ATC CGA CCG CTC TTA ATA GAG GGG ACTPhe Asn Asp Ser Thr Arg Lys Lys Ile Glu Lys Ile Arg Pro Leu Leu Ile Glu Gly Thr385 390 395 400GCA TCA TTG AGC CCT GGA ATG ATG ATG GGC ATG TTC AAT ATG TTA AGC ACT GTA TTA GGCAla Ser Leu Ser Pro Gly Met Met Met Gly Met Phe Asn Met Leu Ser Thr Val Leu Gly405 410 415 420GTC TCC ATC CTG AAT CTT GGA CAA AAG AGA TAC ACC AAG ACT ACT TAC TGG TGG GAT GGTVal Ser Ile Leu Asn Leu Gly Gln Lys Arg Tyr Thr Lys Thr Thr Tyr Trp Trp Asp Gly425 430 435 440CTT CAA TCC TCT GAC GAT TTT GCT CTG ATT GTG AAT GCA CCC AAT CAT GAA GGG ATT CAALeu Gln Ser Ser Asp Asp Phe Ala Leu Ile Val Asn Ala Pro Asn His Glu Gly Ile Gln
445 450 455 460GCC GGA GTC GAC AGG TTT TAT CGA ACC TGT AAG CTA CAT GGA ATC AAT ATG AGC AAG AAAAla Gly Val Asp Arg Phe Tyr Arg Thr Cys Lys Leu His Gly Ile Asn Met Ser Lys Lys465 470 475 480AAG TCT TAC ATA AAC AGA ACA GGT ACA TTT GAA TTC ACA AGT TTT TTC TAT CGT TAT GGGLys Ser Tyr Ile Asn Arg Thr Gly Thr Phe Glu Phe Thr Ser Phe Phe Tyr Arg Tyr Gly485 490 495 500TTT GTT GCC AAT TTC AGC ATG GAG CTT CCC AGT TTT GGT GTG TCT GGG AGC AAC GAG TCAPhe Val Ala Asn Phe Ser Met Glu Leu Pro Ser Phe Gly Val Ser Gly Ser Asn Glu Ser505 510 515 520GCG GAC ATG AGT ATT GGA GTT ACT GTC ATC AAA AAC AAT ATG ATA AAC AAT GAT CTT GGTAla Asp Met Ser Ile Gly Val Thr Val Ile Lys Asn Asn Met Ile Asn Asn Asp Leu Gly525 530 535 540CCA GCA ACA GCT CAA ATG GCC CTT CAG TTG TTC ATC AAA GAT TAC AGG TAC ACG TAC CGAPro Ala Thr Ala Gln Met Ala Leu Gln Leu Phe Ile Lys Asp Tyr Arg Tyr Thr Tyr Arg545 550 555 560TGC CAT AGA GGT GAC ACA CAA ATA CAA ACC CGA AGA TCA TTT GAA ATA AAG AAA CTG TGGCys His Arg Gly Asp Thr Gln Ile Gln Thr Arg Arg Ser Phe Glu Ile Lys Lys Leu Trp565 570 575 580GAG CAA ACC CGT TCC AAA GCT GGA CTG CTG GTC TCC GAC GGA GGC CCA AAT TTA TAC AACGlu Gln Thr Arg Ser Lys Ala Gly Leu Leu Val Ser Asp Gly Gly Pro Asn Leu Tyr Asn585 590 595 600ATT AGA AAT CTC CAC ATT CCT GAA GTC TGC CTA AAA TGG GAA TTG ATG GAT GAG GAT TACIle Arg Asn Leu His Ile Pro Glu Val Cys Leu Lys Trp Glu Leu Met Asp Glu Asp Tyr605 610 615 620CAG GGG CGT TTA TGC AAC CCA CTG AAC CCA TTT GTC AGC CAT AAA GAA ATT GAA TCA ATGGln Gly Arg Leu Cys Asn Pro Leu Asn Pro Phe Val Ser His Lys Glu Ile Glu Ser Met625 630 635 640AAC AAT GCA GTG ATG ATG CCA GCA CAT GGT CCA GCC AAA AAC ATG GAG TAT GAT GCT GTTAsn Asn Ala Val Met Met Pro Ala His Gly Pro Ala Lys Asn Met Glu Tyr Asp Ala Val645 650 655 660GCA ACA ACA CAC TCC TGG ATC CCC AAA AGA AAT CGA TCC ATC TTG AAT ACA AGT CAA AGAAla Thr Thr His Ser Trp Ile Pro Lys Arg Asn Arg Ser Ile Leu Asn Thr Ser Gln Arg665 670 675 680GGA GTA CTT GAA GAT GAA CAA ATG TAC CAA AGG TGC TGC AAT TTA TTT GAA AAA TTC TTCGly Val Leu Glu Asp Glu Gln Met Tyr Gln Arg Cys Cys Asn Leu Phe Glu Lys Phe Phe685 690 695 700CCC AGC AGT TCA TAC AGA AGA CCA GTC GGG ATA TCC AGT ATG GTG GAG GCT ATG GTT TCCPro Ser Ser Ser Tyr Arg Arg Pro Val Gly Ile Ser Ser Met Val Glu Ala Met Val Ser705 710 715 720AGA GCC CGA ATT GAT GCA CGG ATT GAT TTC GAA TCT GGA AGG ATA AAG AAA GAA GAG TTCArg Ala Arg Ile Asp Ala Arg Ile Asp Phe Glu Ser Gly Arg Ile Lys Lys Glu Glu Phe725 730 735 740ACT GAG ATC ATG AAG ATC TGT TCC ACC ATT GAA GAG CTC AGA CGG CAA AAA TAGThr Glu Ile Met Lys Ile Cys Ser Thr Ile Glu Glu Leu Arg Arg Gln Lys ***745 750 755Met Asp Val Asn Pro Thr Leu Leu Phe Leu Lys Val Pro Ala Gln Asn Ala Ile Ser Thr5 10 15 20Thr Phe Pro Tyr Thr Gly Asp Pro Pro Tyr Ser His Gly Thr Gly Thr Gly Tyr Thr Met25 30 35 40Asp Thr Val Asn Arg Thr His Gln Tyr Ser Glu Lys Ala Arg Trp Thr Thr Asn Thr Glu45 50 55 60
Thr Gly Ala Pro Gln Leu Asn Pro Ile Asp Gly Pro Leu Pro Glu Asp Asn Glu Pro Ser65 70 75 80Gly Tyr Ala Gln Thr Asp Cys Val Leu Glu Ala Met Ala Phe Leu Glu Glu Ser His Pro85 90 95 100Gly Ile Phe Glu Asn Ser Cys Ile Glu Thr Met Glu Val Val Gln Gln Thr Arg Val Asp105 110 115 120Lys Leu Thr Gln Gly Arg Gln Thr Tyr Asp Trp Thr Leu Asn Arg Asn Gln Pro Ala Ala125 130 135 140Thr Ala Leu Ala Asn Thr Ile Glu Val Phe Arg Ser Asn Gly Leu Thr Ala Asn Glu Ser145 150 155 160GGA AGG CTC ATA GAC TTC CTT AAG GAT GTA ATG GAG TCA ATG AAA AAA GAA GAA ATG GGGGly Arg Leu Ile Asp Phe Leu Lys Asp Val Met Glu Ser Met Lys Lys Glu Glu Met Gly165 170 175 180Ile Thr Thr His Phe Gln Arg Lys Arg Arg Val Arg Asp Asa Met Thr Lys Lys Met Ile185 190 195 200Thr Gln Arg Thr Ile Gly Lys Arg Lys Gln Arg Leu Asn Lys Arg Ser Tyr Leu Ile Arg205 210 215 220Ala Leu Thr Leu Asn Thr Met Thr Lys Asp Ala Glu Arg Gly Lys Leu Lys Arg Arg Ala225 230 235 240Ile Ala Thr Pro Gly Mer Gln Ile Arg Gly Phe Val Tyr Phe Val Glu Thr Leu Ala Arg245 250 255 260Ser Ile Cys Glu Lys Leu Glu Gln Ser Gly Leu Pro Val Gly Gly Asn Glu Lys Lys Ala265 270 275 280Lys Leu Ala Asn Val Val Arg Lys Met Met Thr Asn Ser Gln Asp Thr Glu Leu Ser Leu285 290 295 300Thr Ile Thr Gly Asp Asn Thr Lys Trp Asn Glu Asn Gln Asn Pro Arg Met Phe Leu Ala305 310 315 320Met Ile Thr Tyr Met Thr Arg Asn Gln Pro Glu Trp Phe Arg Asn Val Leu Ser Ile Ala325 330 335 340Pro Ile Met Phe Ser Asn Lys Met Ala Arg Leu Gly Lys Gly Tyr Met Phe Glu Ser Lys345 350 355 360Ser Met Lys Leu Arg Thr Gln Ile Pro Ala Glu Met Leu Ala Ser Ile Asp Leu Lys Tyr365 370 375 380Phe Asn Asp Ser Thr Arg Lys Lys Ile Glu Lys Ile Arg Pro Leu Leu Ile Glu Gly Thr385 390 395 400Ala Ser Leu Ser Pro Gly Met Met Met Gly Met Phe Asn Met Leu Ser Thr Val Leu Gly405 410 415 420Val Ser Ile Leu Asn Leu Gly Gln Lys Arg Tyr Thr Lys Thr Thr Tyr Trp Trp Asp Gly425 430 435 440Leu Gln Ser Ser Asp Asp Phe Ala Leu Ile Val Asn Ala Pro Asn His Glu Gly Ile Gln445 450 455 460Ala Gly Val Asp Arg Phe Tyr Arg Thr Cys Lys Leu His Gly Ile Asn Met Ser Lys Lys465 470 475 480Lys Ser Tyr Ile Asn Arg Thr Gly Thr Phe Glu Phe Thr Ser Phe Phe Tyr Arg Tyr Gly485 490 95 500Phe Val Ala Asn Phe Ser Met Glu Leu Pro Ser Phe Gly Val Ser Gly Ser Asn Glu Ser505 510 15 520Ala Asp Met Ser Ile Gly Val Thr Val Ile Lys Asn Asn Met Ile Asn Asn Asp Leu Gly525 530 535 540Pro Ala Thr Ala Gln Met Ala Leu Gln Leu Phe Ile Lys Asp Tyr Arg Tyr Thr Tyr Arg545 550 555 560Cys His Arg Gly Asp Thr Gln Ile Gln Thr Arg Arg Ser Phe Glu Ile Lys Lys Leu Trp565 570 575 580
Glu Gln Thr Arg Ser Lys Ala Gly Leu Leu Val Ser Asp Gly Gly Pro Asn Leu Tyr Asn585 590 595 600Ile Arg Asn Leu His Ile Pro Glu Val Cys Leu Lys Trp Glu Leu Met Asp Glu Asp Tyr605 610 615 620Gln Gly Arg Leu Cys Asn Pro Leu Asn Pro Phe Val Ser His Lys Glu Ile Glu Ser Met625 630 635 640Asn Asn Ala Val Met Met Pro Ala His Gly Pro Ala Lys Asn Met Glu Tyr Asp Ala Val645 650 655 660Ala Thr Thr His Ser Trp Ile Pro Lys Arg Asn Arg Ser Ile Leu Asn Thr Ser Gln Arg665 670 675 680Gly Val Leu Glu Asp Glu Gln Met Tyr Gln Arg Cys Cys Asn Leu Phe Glu Lys Phe Phe685 690 695 700Pro Ser Ser Ser Tyr Arg Arg Pro Val Gly Ile Ser Ser Met Val Glu Ala Met Val Ser705 710 715 720Arg Ala Arg Ile Asp Ala Arg Ile Asp Phe Glu Ser Gly Arg Ile Lys Lys Glu Glu Phe725 730 735 740Thr Glu Ile Met Lys Ile Cys Ser Thr Ile Glu Glu Leu Arg Arg Gln Lys ***745 750 755SEQ ID No.3ATG GAA GAT TTT GTG CGA CAA TGC TTC AAT CCG AGG ATT GTC GAG CTT GCG GAA AAA ACAMet Glu Asp Phe Val Arg Gln Cys Phe Asn Pro Met Ile Val Glu Leu Ala Glu Lys Thr5 10 15 20ATG AAA GAG TAT GGG GAG GAC CTG AAA ATC GAA ACA AAC AAA TTT GCA GCA ATA TGC ACTMet Lys Glu Tyr Gly Glu Asp Leu Lys Ile Glu Thr Asn Lys Phe Ala Ala Ile Cys Thr25 30 35 40CAC TTG GAA GTA TGC TTC ATG TAT TCA GAT TTC CAC TTC ATC AAT GAG CAA GGC GAG TCAHis Leu Glu Val Cys Phe Met Tyr Ser Asp Phe His Phe Ile Asn Glu Gln Gly GLu Ser45 50 55 60ATA ATC GTA GAA CTT GGT GAT CCT AAT GCA CTT TTG AAG CAC AGA TTT GAA ATA ATC GAGIle Ile Val Glu Leu Gly Asp Pro Asn Ala Leu Leu Lys His Arg Phe Glu Ile Ile Glu65 70 75 80GGA AGA GAT CGC ACA ATG GCC TGG ACA GTA GTA AAC AGT ATT TGC AAC ACT ACA GGG GCTGly Arg Asp Arg Thr Met Ala Trp Thr Val Val Asn Ser Ile Cys Asn Thr Thr Gly Ala85 90 95 100GAG AAA CCA AAG TTT CTA CCA GAT TTG TAT GAT TAC AAG GAA AAT AGA TTC ATC GAA ATTGlu Lys Pro Lys Phe Leu Pro Asp Leu Tyr Asp Tyr Lys Glu Asn Arg Phe Ile Glu Ile105 110 115 120GGA GTA ACA AGG AGA GAA GTT CAC ATA TAC TAT CTG GAA AAG GCC AAT AAA ATT AAA TCTGly Val Thr Arg Arg Glu Val His Ile Tyr Tyr Leu Glu Lys Ala Asn Lys Ile Lys Ser125 130 135 140GAG AAA ACA CAC ATC CAC ATT TTC TCG TTC ACT GGG GAA GAA ATG GCC ACA AAG GCC GACGlu Lys Thr His Ile His Ile Phe Ser Phe Thr Gly Glu Glu Met Ala Thr Lys Ala Asp145 150 155 160TAC ACT CTC GAT GAA GAA AGC AGG GCT AGG ATC AAA ACC AGG CTA TTC ACC ATA AGA CAATyr Thr Leu Asp Glu Glu Ser Arg Ala Arg Ile Lys Thr Arg Leu Phe Thr Ile Arg Gln165 170 175 180GAA ATG GCC AGC AGA GGC CTC TGG GAT TCC TTT CGT CAG TCC GAG AGA GGA GAA GAG ACAGlu Met Ala Ser Arg Gly Leu Trp Asp Ser Phe Arg Gln Ser Glu Arg Gly Glu Glu Thr185 190 195 200ATT GAA GAA AGG TTT GAA ATC ACA GGA ACA ATG CGC AAG CTT GCC GAC CAA AGT CTC CCG
Ile Glu Glu Arg Phe Glu Ile Thr Gly Thr Met Arg Lys Leu Ala Asp Gln Ser Leu Pro205 210 215 220CCG AAC TTC TCC AGC CTT GAA AAT TTT AGA GCC TAT GTG GAT GGA TTC GAA CCG AAC GGCPro Asn Phe Ser Ser Leu Glu Asn Phe Arg Ala Tyr Val Asp Gly Phe Glu Pro Asn Gly225 230 235 240TAC ATT GAG GGC AAG CTG TCT CAA ATG TCC AAA GAA GTA AAT GCT AGA ATT GAA CCT TTTTyr Ile Glu Gly Lys Leu Ser Gln Met Ser Lys Glu Val Asn Ala Arg Ile Glu Pro Phe245250 255 260TTG AAA ACA ACA CCA CGA CCA CTT AGA CTT CCG AAT GGG CCT CCC TGT TCT CAG CGG TCCLeu Lys Thr Thr Pro Arg Pro Leu Arg Leu Pro Asn Gly Pro Pro Cys Ser Gln Arg Ser265 270 275 280AAA TTC CTG CTG ATG GAT GCC TTA AAA TTA AGC ATT GAG GAC CCA AGT CAT GAA GGA GAGLys Phe Leu Leu Met Asp Ala Leu Lys Leu Ser Ile Glu Asp Pro Ser His Glu Gly Glu285 290 295 300GGA ATA CCG CTA TAT GAT GCA ATC AAA TGC ATG AGA ACA TTC TTT GGA TGG AAG GAA CCCGly Ile Pro Leu Tyr Asp Ala lle Lys Cys Met Arg Thr Phe Phe Gly Trp Lys Glu Pro305 310 315 320AAT GTT GTT AAA CCA CAC GAA AAG GGA ATA AAT CCA AAT TAT CTT CTG TCA TGG AAG CAAAsn Val Val Lys Pro His Glu Lys Gly Ile Asn Pro Asn Tyr Leu Leu Ser Trp Lys Gln325 330 335 340GTA CTG GCA GAA CTG CAG GAC ATT GAG AAT GAG GAG AAA ATT CCA AAG ACT AAA AAT ATGVal Leu Ala Glu Leu Gln Asp Ile Glu Asn Glu Glu Lys Ile Pro Lys Thr Lys Asn Met345 350 355 360AAA AAA ACA AGT CAG CTA AAG TGG GCA CTT GGT GAG AAC ATG GCA CCA GAA AAG GTA GACLys Lys Thr Ser Gln Leu Lys Trp Ala Leu Gly Glu Asn Met Ala Pro Glu Lys Val Asp365 370 375 380TTT GAC GAC TGT AAA GAT GTA GGT GAT TTG AAG CAA TAT GAT AGT GAT GAA CCA GAA TTGPhe Asp Asp Cys Lys Asp Val Gly Asp Leu Lys Gln Tyr Asp Ser Asp Glu Pro Glu Leu385 390 395 400AGG TCG CTT GCA AGT TGG ATT CAG AAT GAG TTC AAC AAG GCA TGC GAA CTG ACA GAT TCAArg Ser Leu Ala Ser Trp Ile Gln Asn Glu Phe Asn Lys Ala Cys Glu Leu Thr Asp Ser405 410 415 420AGC TGG ATA GAG CTT GAT GAG ATT GGA GAA GAT GTG GCT CCA ATT GAA CAC ATT GCA AGCSer Trp Ile Glu Leu Asp Glu Ile Gly Glu Asp Val Ala Pro Ile Glu His Ile Ala Ser425 430 435 440ATG AGA AGG AAT TAT TTC ACA TCA GAG GTG TCT CAC TGC AGA GCC ACA GAA TAC ATA ATGMet Arg Arg Asn Tyr Phe Thr Ser Glu Val Ser His Cys Arg Ala Thr Glu Tyr Ile Met445 450 455 460AAG GGG GTG TAC ATC AAT ACT GCC TTA CTT AAT GCA TCT TGT GCA GCA ATG GAT GAT TTCLys Gly Val Tyr Ile Asn Thr Ala Leu Leu Asn Ala Ser Cys Ala Ala Met Asp Asp Phe465 470 475 480CAA TTA ATT CCA ATG ATA AGC AAG TGT AGA ACT AAG GAG GGA AGG CGA AAG ACC AAC TTGGln Leu Ile Pro Met Ile Ser Lys Cys Arg Thr Lys Glu Gly Arg Arg Lys Thr Asn Leu485 490 495 500TAT GGT TTC ATC ATA AAA GGA AGA TCC CAC TTA AGG AAT GAC ACC GAC GTG GTA AAC TTTTyr Gly Phe Ile Ile Lys Gly Arg Ser His Leu Arg Asn Asp Thr Asp Val Val Asn Phe505 510 515 520GTG AGC ATG GAG TTT TCT CTC ACT GAC CCA AGA CTT GAA CCA CAC AAA TGG GAG AAG TACVal Ser Met Glu Phe Ser Leu Thr Asp Pro Arg Leu Glu Pro His Lys Trp Glu Lys Tyr525 530 535 540TGT GTT CTT GAG ATA GGA GAT ATG CTT CTA AGA AGT GCC ATA GGC CAG GTT TCA AGG CCCCys Val Leu Glu Ile Gly Asp Met Leu Leu Arg Ser Ala Ile Gly Gln Val Ser Arg Pro545 550 555 560
ATG TTC TTG TAT GTG AGG ACA AAT GGA ACC TCA AAA ATT AAA ATG AAA TGG GGA ATG GAGMet Phe Leu Tyr Val Arg Thr Asn Gly Thr Ser Lys Ile Lys Met Lys Trp Gly Met Glu565 570 575 580ATG AGG CGT TGT CTC CTC CAG TCA CTT CAA CAA ATT GAG AGT ATG ATT GAA GCT GAG TCCMet Arg Arg Cys Leu Leu Gln Ser Leu Gln Gln Ile Glu Ser Met Ile Glu Ala Glu Ser585 590 595 600TCT GTC AAA GAG AAA GAC ATG ACC AAA GAG TTC TTT GAG AAC AAA TCA GAA ACA TGG CCCSer Val Lys Glu Lys Asp Met Thr Lys Glu Phe Phe Glu Asn Lys Ser Glu Thr Trp Pro605 610 615 620ATT GGA GAG TCT CCC AAA GGA GTG GAG GAA AGT TCC ATT GGG AAG GTC TGC AGG ACT TTAIle Gly Glu Ser Pro Lys Gly Val Glu Glu Ser Ser Ile Gly Lys Val Cys Arg Thr Leu625 630 635 640TTA GCA AAG TCG GTA TTT AAC AGC TTG TAT GCA TCT CCA CAA CTA GAA GGA TTT TCA GCTLeu Ala Lys Ser Val Phe Asn Ser Leu Tyr Ala Ser Pro Gln Leu Glu Gly Phe Ser Ala645 650 655 660GAA TCA AGA AAA CTG CTT CTT ATC GTT CAG GCT CTT AGG GAC AAT CTG GAA CCT GGG ACCGlu Ser Arg Lys Leu Leu Leu Ile Val Gln Ala Leu Arg Asp Asn Leu Glu Pro Gly Thr665 670 675 680TTT GAT CTT GGG GGG CTA TAT GAA GCA ATT GAG GAG TGC CTA ATT AAT GAT CCC TGG GTTPhe Asp Leu Gly Gly Leu Tyr Glu Ala Ile Glu Glu Cys Leu Ile Asn Asp Pro Trp Val685 690 695 700TTG CTT AAT GCT TCT TGG TTC AAC TCC TTC CTT ACA CAT GCA TTG AGT TAGLeu Leu Asn Ala Ser Trp Phe Asn Ser Phe Leu Thr His Ala Leu Ser ***705 710 715Met Glu Asp Phe Val Arg Gln Cys Phe Asn Pro Met Ile Val Glu Leu Ala Glu Lys Thr5 10 15 20Met Lys Glu Tyr Gly Glu Asp Leu Lys Ile Glu Thr Asn Lys Phe Ala Ala Ile Cys Thr25 30 35 40His Leu Glu Val Cys Phe Met Tyr Ser Asp Phe His Phe Ile Asn Glu Gln Gly Glu Ser45 50 55 60Ile Ile Val Glu Leu Gly Asp Pro Asn Ala Leu Leu Lys His Arg Phe Glu Ile Ile Glu65 70 75 80Gly Arg Asp Arg Thr Met Ala Trp Thr Val Val Asn Ser Ile Cys Asn Thr Thr Gly Ala85 90 95 100Glu Lys Pro Lys Phe Leu Pro Asp Leu Tyr Asp Tyr Lys Glu Asn Arg Phe Ile Glu Ile105 110 115 120Gly Val Thr Arg Arg Glu Val His Ile Tyr Tyr Leu Glu Lys Ala Asn Lys Ile LVs Ser125 130 135 140Glu Lys Thr His Ile His Ile Phe Ser Phe Thr Gly Glu Glu Met Ala Thr Lys Ala Asp145 150 155 160Tyr Thr Leu Asp Glu Glu Ser Arg Ala Arg Ile Lys Thr Arg Leu Phe Thr Ile Arg Gln165 170 175 180Glu Met Ala Ser Arg Gly Leu Trp Asp Ser Phe Arg Gln Ser Glu Arg Gly Glu Glu Thr185 190 195 200Ile Glu Glu Arg Phe Glu Ile Thr Gly Thr Met Arg Lys Leu Ala Asp Gln Ser Leu Pro205 210 215 220Pro Asn Phe Ser Ser Leu Glu Asn Phe Arg Ala Tyr Val Asp Gly Phe Glu Pro Asn Gly225 230 235 240Tyr Ile Glu Gly Lys Leu Ser Gln Met Ser Lys Glu Val Asn Ala Arg Ile Glu Pro Phe245 250 255 260Leu Lys Thr Thr Pro Arg Pro Leu Arg Leu Pro Asn Gly Pro Pro Cys Ser Gln Arg Ser265 270 275 280
Lys Phe Leu Leu Met Asp Ala Leu Lys Leu Ser Ile Glu Asp Pro Ser His Glu Gly Glu285 290 295 300Gly Ile Pro Leu Tyr Asp Ala Ile Lys Cys Met Arg Thr Phe Phe Gly Trp Lys Glu Pro305 310 315 320Asn Val Val Lys Pro His Glu Lys Gly Ile Asn Pro Asn Tyr Leu Leu Ser Trp Lys Gln325 330 335 340Val Leu Ala Glu Leu Gln Asp Ile Glu Asn Glu Glu Lys Ile Pro Lys Thr Lys Asn Met345 350 355 360Lys Lys Thr Ser Gln Leu Lys Trp Ala Leu Gly Glu Asn Met Ala Pro Glu Lys Val Asp365 370 375 380Phe Asp Asp Cys Lys Asp Val Gly Asp Leu Lys Gln Tyr Asp Ser Asp Glu Pro Glu Leu385 390 395 400Arg Ser Leu Ala Ser Trp Ile Gln Asn Glu Phe Asn Lys Ala Cys Glu Leu Thr Asp Ser405 410 415 420Ser Trp Ile Glu Leu Asp Glu Ile Gly Glu Asp Val Ala Pro Ile Glu His Ile Ala Ser425 430 435 440Met Arg Arg Asn Tyr Phe Thr Ser Glu Val Ser His Cys Arg Ala Thr Glu Tyr Ile Met445 450 455 460Lys Gly Val Tyr Ile Asn Thr Ala Leu Leu Asn Ala Ser Cys Ala Ala Met Asp Asp Phe465 470 475 480Gln Leu Ile Pro Met Ile Ser Lys Cys Arg Thr Lys Glu Gly Arg Arg Lys Thr Asn Leu485 490 495 500Tyr Gly Phe Ile Ile Lys Gly Arg Ser His Leu Arg Asn Asp Thr Asp Val Val Asn Phe505 510 515 520Val Ser Met Glu Phe Ser Leu Thr Asp Pro Arg Leu Glu Pro His Lys Trp Glu Lys Tyr525 530 535 540Cys Val Leu Glu Ile Gly Asp Met Leu Leu Arg Ser Ala Ile Gly Gln Val Ser Arg Pro545 550 555 560Met Phe Leu Tyr Val Arg Thr Asn Gly Thr Ser Lys Ile Lys Met Lys Trp Gly Met Glu565 570 575 580Met Arg Arg Cys Leu Leu Gln Ser Leu Gln Gln Ile Glu Ser Met Ile Glu Ala Glu Ser585 590 595 600Ser Val Lys Glu Lys Asp Met Thr Lys Glu Phe Phe Glu Asn Lys Ser Glu Thr Trp Pro605 610 615 620Ile Gly Glu Ser Pro Lys Gly Val Glu Glu Ser Ser Ile Gly Lys Val Cys Arg Thr Leu625 630 635 640Leu Ala Lys Ser Val Phe Asn Ser Leu Tyr Ala Ser Pro Gln Leu Glu Gly Phe Ser Ala645 650 655 660Glu Ser Arg Lys Leu Leu Leu Ile Val Gln Ala Leu Arg Asp Asn Leu Glu Pro Gly Thr665 670 675 680Phe Asp Leu Gly Gly Leu Tyr Glu Ala Ile Glu Glu Cys Leu Ile Asn Asp Pro Trp Val685 690 695 700Leu Leu Asn Ala Ser Trp Phe Asn Ser Phe Leu Thr His Ala Leu Ser ***705 710 715SEQ ID No.4ATG AAC ACT CAA ATC CTG ATA CTC GCT CTT GTG GTG ATC ATC CCG ACA AAT GCA GAC AAAMet Asn Thr Gln Ile Leu Ile Leu Ala Leu Val Val Ile Ile Pro Thr Asn Ala Asp Lys5 10 15ATT TGC CTT GGG CAT CAT GCC GTG TCA AAC GGA ACT AAG GTA AAC ACA CTA ACT GAA AGAIle Cys Leu Gly His His Ala Val Ser Asn Gly Thr Lys Val Asn Thr Leu Thr Glu Arg25 30 35 40GGG GTG GAA GTT GTC AAT GCA ACT GAA ACA GTG GAG CGA ACA AAC ATT CCC AGG ATC TGC
Gly Val Glu Val Val Asn Ala Thr Glu Thr Val Glu Arg Thr Asn Ile Pro Arg Ile Cys45 50 55 60TCA CAA GGA AAA AGA ACA ATC GAC CTT GGC CAA TGT GGA CTG CTG GGA ACA ATC ACT GGASer Gln Gly Lys Arg Thr Ile Asp Leu Gly Gln Cys Gly Leu Leu Gly Thr Ile Thr Gly65 70 75 80CCA CCC CAA TGT GAC CAA TTC CTA GAA TTC TCA GCT GAT TTG ATT ATT GAA AGG CGG GAAPro Pro Gln Cys Asp Gln Phe Leu Glu Phe Ser Ala Asp Leu Ile Ile Glu Arg Arg Glu85 90 95 100GGA AAT GAT GTC TGT TAT CCT GGG AAA TTT GTA AAC GGA GAA GCT CTG AGA CAA ATT CTCGly Asn Asp Val Cys Tyr Pro Gly Lys Phe Val Asn Gly Glu Ala Leu Arg Gln Ile Leu105 110 115 120AGG GAA TCA GGC GGA ATT GAT AAG GAG ATA ATG GGG TTC ACA TAC AGC GAA ATA AGA ACTArg Glu Ser Gly Gly Ile Asp Lys Glu Ile Met Gly Phe Thr Tyr Ser Glu Ile Arg Thr125 130 135 140AAT GGA ACA ACC AGT GCA TGT AGA AGA TCA GGA TCT TCG TTC TAT GCA GAG ATG AAA TGGAsn Gly Thr Thr Ser Ala Cys Arg Arg Ser Gly Ser Ser Phe Tyr Ala Glu Met Lys Trp145 150 155 160CTT TTG TCA AAT ACA GAC AAT GCT GCT TTC CCG CAG ATG ACT AAG TCG TAC AAA AAC ACALeu Leu Ser Asn Thr Asp Asn Ala Ala Phe Pro Gln Met Thr Lys Ser Tyr Lys Asn Thr165 170 175 180AGG AAA GAC CCA GCT CTG ATA ATC TGG GGA ATC CAT CAT TCC GGA TCA AGC ACA GAA CAGArg Lys Asp Pro Ala Leu Ile Ile Trp Gly Ile His His Ser Gly Ser Ser Thr Glu Gln185 190 195 200ACC AAA TTA TAT GGG AGT GGA AGC AAA CTA ATA ACA GTT GGG AGT TCC AAT TAT CAA CAGThr Lys Leu Tyr Gly Ser Gly Ser Lys Leu Ile Thr Val Gly Ser Ser Asn Tyr Gln Gln205 210 215 220TCC TTT GTA CCG AGT CCG GGG GCG AGA CCA CAA GTG AAT GGC CAA TCT GGA AGG ATC GACSer Phe Val Pro Ser Pro Gly Ala Arg Pro Gln Val Asn Gly Gln Ser Gly Arg Ile Asp225 230 235 240TTC CAT TAC CTG ATG CTA AAT CCC AAT GAC ACA GTC ACT TTC AGT TTC AAT GGG GCT TTCPhe His Tyr Leu Met Leu Asn Pro Asn Asp Thr Val Thr Phe Ser Phe Asn Gly Ala Phe245 250 255 260ATA GCT CCA GAC CGT GCA AGT TTT CTG AGA GGG AAG TCT ATG GGG ATT CAG AGT GAT GTAIle Ala Pro Asp Arg Ala Ser Phe Leu Arg Gly Lys Ser Met Gly Ile Gln Ser Asp Val265 270 275 280CAA GTT GAT GCC AAT TGT GAG GGG GAT TGC TAT CAT AGT GGA GGA ACA ATA ATA AGT AATGln Val Asp Ala Asn Cys Glu Gly Asp Cys Tyr His Ser Gly Gly Thr Ile Ile Ser Asn285 290 295 300TTG CCT TTT CAG AAT ATC AAT AGC AGA GCA GTA GGA AAA TGT CCG AGG TAT GTT AAA CAALeu Pro Phe Gln Asn Ile Asn Ser Arg Ala Val Gly Lys Cys Pro Arg Tyr Val Lys Gln305 310 315 320GAG AGC CTG CTA CTG GCA ACA GGG ATG AAG AAC GTT CCC GAA ATT CCA AAA GGA AGA GGCGlu Ser Leu Leu Leu Ala Thr Gly Met Lys Asn Val Pro Glu Ile Pro Lys Gly Arg Gly325 330 335 340CTA TTT GGT GCC ATA GCG GGT TTC ATT GAA AAT GGA TGG GAA GGT CTG ATT GAT GGA TGGLeu Phe Gly Ala Ile Ala Gly Phe Ile Glu Asn Gly Trp Glu Gly Leu Ile Asp Gly Trp345 350 355 360TAT GGC TTC AGG CAT CAA AAT GCA CAA GGA GAG GGA ACT GCG GCA GAT TAC AAG AGC ACTTyr Gly Phe Arg His Gln Asn Ala Gln Gly Glu Gly Thr Ala Ala Asp Tyr Lys Ser Thr365 370 375 380CAA TCG GCA ATT GAT CAA ATA ACA GGA AAG TTA AAC CGG CTC ATA GAG AAA ACT AAC CAAGln Ser Ala Ile Asp Gln Ile Thr Gly Lys Leu Asn Arg Leu lle Glu Lys Thr Asn Gln385 390 395 400
CAG TTT GAG TTG ATA GAC AAT GAA TTC ACT GAG GTT GAA AAG CAA ATT GGC AAT GTG ATAGln Phe Glu Leu Ile Asp Asn Glu Phe Thr Glu Val Glu Lys Gln Ile Gly Asn Val Ile405 410 415 420AAT TGG ACC AGA GAT TCC ATA ACA GAA GTG TGG TCC TAC AAT GCC GAA CTC TTA GTG GCAAsn Trp Thr Arg Asp Ser Ile Thr Glu Val Trp Ser Tyr Asn Ala Glu Leu Leu Val Ala425 430 435 440ATG GAG AAT CAG CAC ACT ATC GAT CTG GCC GAT TCA GAA ATG AAC AAG CTG TAC GAA AGAMet Glu Asn Gln His Thr Ile Asp Leu Ala Asp Ser Glu Met Asn Lys Leu Tyr Glu Arg445 450 455 460GTG AGG AGA CAA TTG AGG GAG AAT GCT GAG GAA GAC GGC ACT GGT TGT TTC GAA ATA TTTVal Arg Arg Gln Leu Arg Glu Asn Ala Glu Glu Asp Gly Thr Gly Cys Phe Glu Ile Phe465 470 475 480CAC AAG TGT GAT GAC GAC TGT ATG GCC AGT ATT AGA AAC AAC ACT TAT GAT CAC AGC AAAHis Lys Cys Asp Asp Asp Cys Met Ala Ser Ile Arg Asn Asn Thr Tyr Asp His Ser Lys485 490 495 500TAC AGA GAA GAA GCA ATG CAA AAT AGG ATA CAG ATT GAC CCA GTC AAA CTG AGC AGC GGCTyr Arg Glu Glu Ala Met Gln Asn Arg Ile Gln Ile Asp Pro Val Lys Leu Ser Ser Gly505 510 515 520TAC AAA GAT GTG ATA CTT TGG TTT AGC TTC GGG GCA TCA TGT TTC ATA CTT CTG GCC ATTTyr Lys Asp Val Ile Leu Trp Phe Ser Phe Gly Ala Ser Cys Phe Ile Leu Leu Ala Ile525 530 535 540GCA ATG GGC CTT GTC TTC ATG TGT GTG AAG AAT GGA AAC ATG CGG TGC ACT ATT TGT ATAAla Met Gly Leu Val Phe Met Cys Val Lys Asn Gly Asn Met Arg Cys Thr Ile Cys Ile545 550 555 560TAA---565 570 575 580Met Asn Thr Gln Ile Leu Ile Leu Ala Leu Val Val Ile Ile Pro Thr Asn Ala Asp Lys5 10 15 20Ile Cys Leu Gly His His Ala Val Ser Asn Gly Thr Lys Val Asn Thr Leu Thr Glu Arg25 30 35 40Gly Val Glu Val Val Asn Ala Thr Glu Thr Val Glu Arg Thr Asn Ile Pro Arg Ile Cys45 50 55 60Ser Gln Gly Lys Arg Thr Ile Asp Leu Gly Gln Cys Gly Leu Leu Gly Thr Ile Thr Gly65 70 75 80Pro Pro Gln Cys Asp Gln Phe Leu Glu Phe Ser Ala Asp Leu Ile Ile Glu Arg Arg Glu85 90 95 100Gly Asn Asp Val Cys Tyr Pro Gly Lys Phe Val Asn Gly Glu Ala Leu Arg Gln Ile Leu105 110 115 120Arg Glu Ser Gly Gly Ile Asp Lys Glu Ile Met Gly Phe Thr Tyr Ser Glu Ile Arg Thr125 130 135 140Asn Gly Thr Thr Ser Ala Cys Arg Arg Ser Gly Ser Ser Phe Tyr Ala Glu Met Lys Trp145 150 155 160Leu Leu Ser Asn Thr Asp Asn Ala Ala Phe Pro Gln Met Thr Lys Ser Tyr Lys Asn Thr165 170 175 180Arg Lys Asp Pro Ala Leu Ile Ile Trp Gly Ile His His Ser Gly Ser Ser Thr Glu Gln185 190 195 200Thr Lys Leu Tyr Gly Ser Gly Ser Lys Leu Ile Thr Val Gly Ser Ser Asn Tyr Gln Gln205 210 215 220Ser Phe Val Pro Ser Pro Gly Ala Arg Pro Gln Val Asn Gly Gln Ser Gly Arg Ile Asp225 230 235 240Phe His Tyr Leu Met Leu Asn Pro Asn Asp Thr Val Thr Phe Ser Phe Asn Gly Ala Phe
245 250 255 260Ile Ala Pro Asp Arg Ala Ser Phe Leu Arg Gly Lys Ser Met Gly Ile Gln Ser Asp Val265 270 275 280Gln Val Asp Ala Asn Cys Glu Gly Asp Cys Tyr His Ser Gly Gly Thr Ile Ile Ser Asn285 290 295 300Leu Pro Phe Gln Asn Ile Asn Ser Arg Ala Val Gly Lys Cys Pro Arg Tyr Val Lys Gln305 310 315 320Glu Ser Leu Leu Leu Ala Thr Gly Met Lys Asn Val Pro Glu Ile Pro Lys Gly Arg Gly325 330 335 340Leu Phe Gly Ala Ile Ala Gly Phe Ile Glu Asn Gly Trp Glu Gly Leu Ile Asp Gly Trp345 350 355 360Tyr Gly Phe Arg His Gln Asn Ala Gln Gly Glu Gly Thr Ala Ala Asp Tyr Lys Ser Thr365 370 375 380Gln Ser Ala Ile Asp Gln Ile Thr Gly Lys Leu Asn Arg Leu Ile Glu Lys Thr Asn Gln385 390 395 400Gln Phe Glu Leu Ile Asp Asn Glu Phe Thr Glu Val Glu Lys Gln Ile Gly Asn Val Ile405 410 415 420Asn Trp Thr Arg Asp Ser Ile Thr Glu Val Trp Ser Tyr Asn Ala Glu Leu Leu Val Ala425 430 435 440Met Glu Asn Gln His Thr Ile Asp Leu Ala Asp Ser Glu Met Asn Lys Leu Tyr Glu Arg445 450 455 460Val Arg Arg Gln Leu Arg Glu Asn Ala Glu Glu Asp Gly Thr Gly Cys Phe Glu Ile Phe465 470 475 480His Lys Cys Asp Asp Asp Cys Met Ala Ser Ile Arg Asn Asn Thr Tyr Asp His Ser Lys485 490 495 500Tyr Arg Glu Glu Ala Met Gln Asn Arg Ile Gln Ile Asp Pro Val Lys Leu Ser Ser Gly505 510 515 520Tyr Lys Asp Val Ile Leu Trp Phe Ser Phe Gly Ala Ser Cys Phe Ile Leu Leu Ala Ile525 530 535 540Ala Met Gly Leu Val Phe Met Cys Val Lys Asn Gly Asn Met Arg Cys Thr Ile Cys Ile545 550 555 560SEQ ID No.5GCG TCC CAA GGC ACC AAA CGG TCT TAC GAA CAG ATG GAG ACT GAT GGA GAA CGC CAG 60Met Ala Ser Gln Gly Thr Lys Arg Ser Tyr Glu Gln Met Glu Thr Asp Gly Glu Arg Gln5 10 15 20AAT GCC ACT GAA ATC AGA GCA TCC GTC GGA AAA ATG ATT GGT GGA ATT GGA CGA TTC TACAsn Ala Thr Glu Ile Arg Ala Ser Val Gly Lys Met Ile Gly Gly Ile Gly Arg Phe Tyr25 30 35 40ATC CAA ATG TGC ACA GAA CTT AAA CTC AGT GAT TAT GAG GGA CGG TTG ATC CAA AAC AGCIle Gln Met Cys Thr Glu Leu Lys Leu Ser Asp Tyr Glu Gly Arg Leu Ile Gln Asn Ser45 50 55 60TTA ACA ATA GAG AGA ATG GTG CTC TCT GCT TTT GAC GAA AGG AGA AAT AAA TAC CTG GAALeu Thr Ile Glu Arg Met Val Leu Ser Ala Phe Asp Glu Arg Arg Asn Lys Tyr Leu Glu65 70 75 80GAA CAT CCC AGT GCG GGG AAG GAT CCT AAG AAA ACT GGA GGA CCT ATA TAC AGA AGA GTAGlu His Pro Ser Ala Gly Lys Asp Pro Lys Lys Thr Gly Gly Pro Ile Tyr Arg Arg Val85 90 95 100AAC GGA AAG TGG ATG AGA GAA CTC ATC CTT TAT GAC AAA GAA GAA ATA AGG CGA ATC TGGAsn Gly Lys Trp Met Arg Glu Leu Ile Leu Tyr Asp Lys Glu Glu Ile Arg Arg Ile Trp105 110 115 120CGC CAA GCT AAT AAT GGT GAC GAT GCA ACG GCT GGT CTG ACT CAC ATG ATG ATC TGG CAT
Arg Gln Ala Asn Asn Gly Asp Asp Ala Thr Ala Gly Leu Thr His Met Met Ile Trp His125 130 135 140TCC AAT TTG AAT GAT GCA ACT TAT CAG AGG ACA AGG GCT CTT GTT CGC ACC GGA ATG GATSer Asn Leu Asn Asp Ala Thr Tyr Gln Arg Thr Arg Ala Leu Val Arg Thr Gly Met Asp145 150 155 160CCC AGG ATG TGC TCT CTG ATG CAA GGT TCA ACT CTC CCT AGG AGG TCT GGA GCC GCA GGTPro Arg Met Cys Ser Leu Met Gln Gly Ser Thr Leu Pro Arg Arg Ser Gly Ala Ala Gly165 170 175 180GCT GCA GTC AAA GGA GTT GGA ACA ATG GTG ATG GAA TTG GTC AGG ATG ATC AAA CGT GGGAla Ala Val Lys Gly Val Gly Thr Met Val Met Glu Leu Val Arg Met Ile Lys Arg Gly185 190 195 200ATC AAT GAT CGG AAC TTC TGG AGG GGT GAG AAT GGA CGA AAA ACA AGA ATT GCT TAT GAAIle Asn Asp Arg Asn Phe Trp Arg Gly Glu Asn Gly Arg Lys Thr Arg lle Ala Tyr Glu205 210 215 220AGA ATG TGC AAC ATT CTC AAA GGG AAA TTT CAA ACT GCT GCA CAA AAA GCA ATG ATG GATArg Met Cys Asn Ile Leu Lys Gly Lys Phe Gln Thr Ala Ala Gln Lys Ala Met Met Asp225 230 235 240CAA GTG AGA GAG AGC CGG GAC CCA GGG AAT GCT GAG TTC GAA GAT CTC ACT TTT CTA GCAGln Val Arg Glu Ser Arg Asp Pro Gly Asn Ala Glu Phe Glu Asp Leu Thr Phe Leu Ala245 250 255 260CGG TCT GCA CTC ATA TTG AGA GGG TCG GTT GCT CAC AAG TCC TGC CTG CCT GCC TGT GTGArg Ser Ala Leu Ile Leu Arg Gly Ser Val Ala His Lys Ser Cys Leu Pro Ala Cys Val265 270 275 280TAT GGA CCT GCC GTA GCC AGT GGG TAC GAC TTT GAA AGA GAG GGA TAC TCT CTA GTC GGATyr Gly Pro Ala Val Ala Ser Gly Tyr Asp Phe Glu Arg Glu Gly Tyr Ser Leu Val Gly285 290 295 300ATA GAC CCT TTC AGA CTG CTT CAA AAC AGC CAA GTG TAC AGC CTA ATC AGA CCA AAT GAGlle Asp Pro Phe Arg Leu Leu Gln Asn Ser Gln Val Tyr Ser Leu Ile Arg Pro Asn Glu305 310 315 320AAT CCA GCA CAC AAG AGT CAA CTG GTG TGG ATG GCA TGC CAT TCT GCC GCA TTT GAA GATAsn Pro Ala His Lys Ser Gln Leu Val Trp Met Ala Cys His Ser Ala Ala Phe Glu Asp325 330 335 340CTA AGA GTA TTG AGC TTC ATC AAA GGG ACG AAG GTG GTC CCA AGA GGG AAG CTT TCC ACTLeu Arg Val Leu Ser Phe Ile Lys Gly Thr Lys Val Val Pro Arg Gly Lys Leu Ser Thr345 350 355 360AGA GGA GTT CAA ATT GCT TCC AAT GAA AAT ATG GAG ACT ATG GAA TCA AGT ACA CTT GAAArg Gly Val Gln Ile Ala Ser Asn Glu Asn Met Glu Thr Met Glu Ser Ser Thr Leu Glu365 370 375 380CTG AGA AGC AGG TAC TGG GCC ATA AGG ACC AGA AGT GGA GGA AAC ACC AAT CAA CAG AGGLeu Arg Ser Arg Tyr Trp Ala Ile Arg Thr Arg Ser Gly Gly Asn Thr Asn Gln Gln Arg385 390 395 400GCA TCT GCG GGC CAA ATC AGC ATA CAA CCT ACG TTC TCA GTA CAG AGA AAT CTC CCT TTTAla Ser Ala Gly Gln Ile Ser Ile Gln Pro Thr Phe Ser Val Gln Arg Asn Leu Pro Phe405 410 415 420GAC AGA ACA ACC GTT ATG GCA GCA TTC ACT GGG AAT ACA GAG GGG AGA ACA TCT GAC ATGAsp Arg Thr Thr Val Met Ala Ala Phe Thr Gly Asn Thr Glu Gly Arg Thr Ser Asp Met425 430 435 440AGG ACC GAA ATC ATA AGG ATG ATG GAA AGT GCA AGA CCA GAA GAT GTG TCT TTC CAG GGGArg Thr Glu Ile Ile Arg Met Met Glu Ser Ala Arg Pro Glu Asp Val Ser Phe Gln Gly445 450 455 460CGG GGA GTC TTC GAG CTC TCG GAC GAA AAG GCA GCG AGC CCG ATC GTG CCT TCC TTT GACArg Gly Val Phe Glu Leu Ser Asp Glu Lys Ala Ala Ser Pro Ser Val Pro Ser Phe Asp465 470 475 480
ATG AGT AAT GAA GGA TCT TAT TTC TTC GGA GAC AAT GCA GAG GAG TAC GAC AAT TAAMet Ser Asn Glu Gly Ser Tyr Phe Phe Gly Asp Asn Ala Glu Glu Tyr Asp Asn ***485 490 495Met Als Ser Gln Gly Thr Lys Arg Ser Tyr Glu Gln Met Glu Thr Asp Gly Glu Arg Gln5 10 15 20Asn Ala Thr Glu Ile Arg Ala Ser Val Gly Lys Met Ile Gly Gly Ile Gly Arg Phe Tyr25 30 35 40Ile Gln Met Cys Thr Glu Leu Lys Leu Ser Asp Tyr Glu Gly Arg Leu Ile Gln Asn Ser45 50 55 60Leu Thr Ile Glu Arg Met Val Leu Ser Ala Phe Asp Glu Arg Arg Asn Lys Tyr Leu Glu65 70 75 80Glu His Pro Ser Ala Gly Lys Asp Pro Lys Lys Thr Gly Gly Pro lle Tyr Arg Arg Val85 90 95 100Asn Gly Lys Trp Met Arg Glu Leu Ile Leu Tyr Asp Lys Glu Glu Ile Arg Arg Ile Trp105 110 115 120Arg Gln Ala Asn Asn Gly Asp Asp Ala Thr Ala Gly Leu Thr His Met Met Ile Trp His125 130 135 140Ser Asn Leu Asn Asp Ala Thr Tyr Gln Arg Thr Arg Ala Leu Val Arg Thr Gly Met Asp145 150 155 160Pro Arg Met Cys Ser Leu Met Gln Gly Ser Thr Leu Pro Arg Arg Ser Gly Ala Ala Gly165 170 175 180Ala Ala Val Lys Gly Val Gly Thr Met Val Met Glu Leu Val Arg Met Ile Lys Arg Gly185 190 195 200Ile Asn Asp Arg Asn Phe Trp Arg Gly Glu Asn Gly Arg Lys Thr Arg Ile Ala Tyr Glu205 210 215 220Arg Met Cys Asn Ile Leu Lys Gly Lys Phe Gln Thr Ala Ala Gln Lys Ala Met Met Asp225 230 235 240Gln Val Arg Glu Ser Arg Asp Pro Gly Asn Ala Glu Phe Glu Asp Leu Thr Phe Leu Ala245 250 255 260Arg Ser Ala Leu Ile Leu Arg Gly Ser Val Ala His Lys Ser Cys Leu Pro Ala Cys Val265 270 275 280Tyr Gly Pro Ala Val Ala Ser Gly Tyr Asp Phe Glu Arg Glu Gly Tyr Ser Leu Val Gly285 290 295 300ATA GAC CCT TTC AGA CTG CTT CAA AAC AGC CAA GTG TAC AGC CTA ATC AGA CCA AAT GAGIle Asp Pro Phe Arg Leu Leu Gln Asn Ser Gln Val Tyr Ser Leu Ile Arg Pro Asn Glu305 310 315 320Asn Pro Ala His Lys Ser Gln Leu Val Trp Met Ala Cys His Ser Ala Ala Phe Glu Asp325 330 335 340Leu Arg Val Leu Ser Phe Ile Lys Gly Thr Lys Val Val Pro Arg Gly Lys Leu Ser Thr345 350 355 360Arg Gly Val Gln Ile Ala Ser Asn Glu Asn Met Glu Thr Met Glu Ser Ser Thr Leu Glu365 370 375 380Leu Arg Ser Arg Tyr Trp Ala lle Arg Thr Arg Ser Gly Gly Asn Thr Asn Gln Gln Arg385 390 395 400Ala Ser Ala Gly Gln Ile Ser Ile Gln Pro Thr Phe Ser Val Gln Arg Asn Leu Pro Phe405 410 415 420Asp Arg Thr Thr Val Met Ala Ala Phe Thr Gly Asn Thr Glu Gly Arg Thr Ser Asp Met425 430 435 440Arg Thr Glu Ile Ile Arg Met Met Glu Ser Ala Arg Pro Glu Asp Val Ser Phe Gln Gly445 450 455 460Arg Gly Val Phe Glu Leu Ser Asp Glu Lys Ala Ala Ser Pro Ile Val Pro Ser Phe Asp465 470 475 480
Met Ser Asn Glu Gly Ser Tyr Phe Phe Gly Asp Asn Ala Glu Glu Tyr Asp Asn ***485 490 495SEQ ID No.6ATG AAT CCA AAT CAA AAG ATA ATA ACA ATT GGC TCT GTC TCA CTC ACC ATT GCA ACA ATAMet Asn Pro Asn Gln Lys Ile Ile Thr Ile Gly Ser Val Ser Leu Thr Ile Ala Thr Ile5 10 15 20TGC TTC CTC ATG CAG ATT GCT ATC CTG GCA ACT ACT GTG ACA CTG CAT TTT AAG CAA CATCys Phe Leu Met Gln Ile Ala Ile Leu Ala Thr Thr Val Thr Leu His Phe Lys Gln His25 30 35 40GAA TGC AAC CCC CCT GCA ATC AAC CAA GTG ACC CCA TGT GAA CCA ATC ATA GTA GAA AGGGlu Cys Asn Pro Pro Ala Ile Asn Gln Val Thr Pro Cys Glu Pro Ile Ile Val Glu Arg45 50 55 60AAC ATA ACA GAG ATA GTG TAT TTG AAC AGC ACC AGT ATA GAG AGA GAG ATT TGC CCC GAAAsn Ile Thr Glu Ile Val Tyr Leu Asn Ser Thr Ser Ile Glu Arg Glu Ile Cys Pro Glu65 70 75 80GCT GTG GAA TAC AGA AAT TGG TCA AAG CCG CAG TGT CAA ATT ACA GGA TTT GCT CCT TTCAla Val Glu Tyr Arg Asn Trp Ser Lys Pro Gln Cys Gln Ile Thr Gly Phe Ala Pro Phe85 90 95 100TCT AAG GAC AAT TCA ATT CGG CTT TCT GCT GGT GGG GAC ATT TGG GTG ACG AGA GAA CCTSer Lys Asp Asn Ser Ile Arg Leu Ser Ala Gly lys Asp Ile Trp Val Thr Arg Glu Pro105 110 115 120TAT GTG TCA TGC GAT CCT GGT AAA TGT TAT CAA TTT GCA CTT GGA CAG GGG ACC ACA CTATyr Val Ser Cys Asp Pro Gly Lys Cys Tyr Gln Phe Ala Leu Gly Gln Gly Thr Thr Leu125 130 135 140GAC AAC AAA CAT TCG AAT GGT ACA ATA CAT GAT AGA ATC CCT CAT CGG ACT CTC TTG ATGAsp Asn Lys His Ser Asn Gly Thr Ile His Asp Arg Ile Pro His Arg Thr Leu Leu Met145 150 155 160AAC GAA TTG GGT GTT CCG TTC CAT TTA GGG ACC AAA CAA GTA TGC ATA GCA TGG TCC AGCAsn Glu Leu Gly Val Pro Phe His Leu Gly Thr Lys Gln Val Cys Ile Ala Trp Ser Ser165 170 175 180TCA AGT TGT CAC GAT GGA AGA GCA TGG TTG CAT ATT TGT GTT ACT GGG GAT GAT AGA AATSer Ser Cys His Asp Gly Arg Ala Trp Leu His Ile Cys Val Thr Gly Asp Asp Arg Asn185 190 195 200GCG ACT GCC AGT TTC ATT TAT GAC GGG AGG CTT GTC GAC AGT ATT GGT TCA TGG TCT CAAAla Thr Ala Ser Phe Ile Tyr Asp Gly Arg Leu Val Asp Ser Ile Gly Ser Trp Ser Gln205 210 215 220AAT ATT CTC AGA ACA CAG GAG TCA GAA TGT GTT TGT ATC AAT GGG ACT TGC ACT GTA GTAAsn Ile Leu Arg Thr Gln Glu Ser Glu Cys Val Cys Ile Asn Gly Thr Cys Thr Val Val225 230 235 240ATG ACT GAT GGA AGT GCA TCA GGA AGA GCC GAT ACG AGA ATA CTA TTC ATT AAA GAG GGGMet Thr Asp Gly Ser Ala Ser Gly Arg Ala Asp Thr Arg Ile Leu Phe Ile Lys Glu Gly245 250 255 260AAA ATT GTC CAT ATT AGC CCA TTG TCA GGA AGT GCT CAG CAT ATA GAG GAA TGT TCC TGTLys Ile Val His Ile Ser Pro Leu Ser Gly Ser Ala Gln His Ile Glu Glu Cys Ser Cys265 270 275 280TAT CCT CGA TAT CCT GAC GTC AGG TGT ATC TGC AGA GAT AAC TGG AAA GGC TCT AAT AGGTyr Pro Arg Tyr Pro Asp Val Arg Cys Ile Cys Arg Asp Asn Trp Lys Gly Ser Asn Arg285 290 295 300CCC GTT ATA GAC ATA AAT ATG GCA GAT TAT AGC ATT GAT TCC AGT TAT GTA TGT TCA GGGPro Val Ile Asp Ile Asn Met Ala Asp Tyr Ser Ile Asp Ser Ser Tyr Val Cys Ser Gly305 310 315 320CTC GTT GGT GAC ACA CCC AGA AAT GAC GAC AGC TCC AGC AGT AGC AAA AGC AGG GAT CCT
Leu Val Gly Asp Thr Pro Arg Asn Asp Asp Ser Ser Ser Ser Ser Lys Ser Arg Asp Pro325 330 335 340AAT AAT GAG AGA GGG AAT CCA GGG GTG AAA GGC TGG GCT TTT GAT AAT GGA GAT GAT GTGAsn Asn Glu Arg Gly Asn Pro Gly Val Lys Gly Trp Ala Phe Asp Asn Gly Asp Asp Val345 350 355 360TGG ATG GGG AGA ACA ATC AGC AAA GAT TCA CGC TCA GGT TAC GAG ACT TTC AAA GTC ATTTrp Met Gly Arg Thr Ile Ser Lys Asp Ser Arg Ser Gly Tyr Glu Thr Phe Lys Val Ile365 370 375 380GGT GGT TGG TCC ACA CCT AAT TCC AAG TCG CAG GTC AAT AGA CAG GTC ATA GTT GAC AACGly Gly Trp Ser Thr Pro Asn Ser Lys Ser Gln Val Asn Arg Gln Val Ile Val Asp Asn385 390 395 400AAT AAC AGG TCT GGT TAC TCT GGT ATT TTC TCT GTT GAG GGC AAA AGC TGC ATC AAT AGGAsn Asn Arg Ser Gly Tyr Ser Gly Ile Phe Ser Val Glu Gly Lys Ser Cys Ile Asn Arg405 410 415 420TGT TTT TAT GTG GAG TTG ATA AGG GGA AGA CCA CAG GAA ACT AGA GTA TGG TGG ACT TCACys Phe Tyr Val Glu Leu Ile Arg Gly Arg Pro Gln Glu Thr Arg Val Trp Trp Thr Ser425 430 435 440AAC AGT ATT GTT GTA TTT TGT GGC ACT TCA GGT ACC TAT GGA ACA GGC TCA TGG CCT GATAsn Ser Ile Val Val Phe Cys Gly Thr Ser Gly Thr Tyr Gly Thr Gly Ser Trp Pro Asp445 450 455 460GGG GCG AAC ATC AAT TTT ATG CCT ATA TAA *Gly Ala Asn Ile Asn Phe Met Pro Ile --- *465 470 475 480Met Asn Pro Asn Gln Lys Ile Ile Thr Ile Gly Ser Val Ser Leu Thr Ile Ala Thr Ile5 10 15 20Cys Phe Leu Met Gln Ile Ala Ile Leu Ala Thr Thr Val Thr Leu His Phe Lys Gln His25 30 35 40Glu Cys Asn Pro Pro Ala Ile Asn Gln Val Thr Pro Cys Glu Pro Ile Ile Val Glu Arg45 50 55 60Asn Ile Thr Glu Ile Val Tyr Leu Asn Ser Thr Ser Ile Glu Arg Glu Ile Cys Pro Glu65 70 75 80Ala Val Glu Tyr Arg Asn Trp Ser Lys Pro Gln Cys Gln Ile Thr Gly Phe Ala Pro Phe85 90 95 100Ser Lys Asp Asn Ser Ile Arg Leu Ser Ala Gly lys Asp Ile Trp Val Thr Arg Glu Pro105 110 115 120Tyr Val Ser Cys Asp Pro Gly Lys Cys Tyr Gln Phe Ala Leu Gly Gln Gly Thr Thr Leu125 130 135 140Asp Asn Lys His Ser Asn Gly Thr Ile His Asp Arg Ile Pro His Arg Thr Leu Leu Met145 150 155 160Asn Glu Leu Gly Val Pro Phe His Leu Gly Thr Lys Gln Val Cys Ile Ala Trp Ser Ser165 170 175 180Ser Ser Cys His Asp Gly Arg Ala Trp Leu His Ile Cys Val Thr Gly Asp Asp Arg Asn185 190 195 200Ala Thr Ala Ser Phe Ile Tyr Asp Gly Arg Leu Val Asp Ser lle Gly Ser Trp Ser Gln205 210 215 220Asn lle Leu Arg Thr Gln Glu Ser Glu Cys Val Cys Ile Asn Gly Thr Cys Thr Val Val225 230 235 240Met Thr Asp Gly Ser Ala Ser Gly Arg Ala Asp Thr Arg Ile Leu Phe Ile Lys Glu Gly245 250 255 260Lys lle Val His Ile Ser Pro Leu Ser Gly Ser Ala Gln His Ile Glu Glu Cys Ser Cys265 270 275 280Tyr Pro Arg Tyr Pro Asp Val Arg Cys Ile Cys Arg Asp Asn Trp Lys Gly Ser Asn Arg
285 290 295 300Pro Val Ile Asp Ile Asn Met Ala Asp Tyr Ser Ile Asp Ser Ser Tyr Val Cys Ser Gly305 310 315 320Leu Val Gly Asp Thr Pro Arg Asn Asp Asp Ser Ser Ser Ser Ser Lys Ser Arg Asp Pro325 330 335 340Asn Asn Glu Arg Gly Asn Pro Gly Val Lys Gly Trp Ala Phe Asp Asn Gly Asp Asp Val345 350 355 360Trp Met Gly Arg Thr Ile Ser Lys Asp Ser Arg Ser Gly Tyr Glu Thr Phe Lys Val Ile365 370 375 380Gly Gly Trp Ser Thr Pro Asn Ser Lys Ser Gln Val Asn Arg Gln Val Ile Val Asp Asn385 390 395 400Asn Asn Arg Ser Gly Tyr Ser Gly Ile Phe Ser Val Glu Gly Lys Ser Cys Ile Asn Arg405 410 415 420Cys Phe Tyr Val Glu Leu Ile Arg Gly Arg Pro Gln Glu Thr Arg Val Trp Trp Thr Ser425 430 435 440Asn Ser Ile Val Val Phe Cys Gly Thr Ser Gly Thr Tyr Gly Thr Gly Ser Trp Pro Asp445 450 455 460Gly Ala Asn Ile Asn Phe Met Pro Ile --- *465 470 475 480SEQ ID No.7ATG AGT CTT CTA ACC GAG GTC GAA ACG TAC GTT CTC TCT ATC ATC CCG TCA GGC CCC CTCMet Ser Leu Leu Thr Glu Val Glu Thr Tyr Val Leu Ser Ile Ile Pro Ser Gly Pro Leu5 10 15 20AAA GCC GAG ATC GCA CAG AGA CTT GAA GAT GTC TTT GCA GGG AAG AAC ACC GAT CTT GAGLys Ala Glu Ile Ala Gln Arg Leu Glu Asp Val Phe Ala Gly Lys Asn Thr Asp Leu Glu25 30 35 40GTT CTC ATG GAA TGG CTA AAG ACA AGA CCA ATC CTG TCA CCT CTG ACT AAG GGG ATT TTAVal Leu Met Glu Trp Leu Lys Thr Arg Pro Ile Leu Ser Pro Leu Thr Lys Gly Ile Leu45 50 55 60GGA TTT GTG TTC ACG CTC ACC GTG CCC AGT GAG CGA GGA CTG CAG CGT AGA CGC TTT GTCGly Phe Val Phe Thr Leu Thr Val Pro Ser Glu Arg Gly Leu Gln Arg Arg Arg Phe Val65 70 75 80CAA AAT GCC CTT AAT GGG AAC GGG GAT CCA AAT AAC ATG GAC AAA GCA GTT AAA CTG TATGln Asn Ala Leu Asn Gly Asn Gly Asp Pro Asn Asn Met Asp Lys Ala Val Lys Leu Tyr85 90 95 100AGG AAG CTC AAG AGG GAG ATA ACA TTC CAT GGG GCC AAA GAA ATC TCA CTC AGT TAT TCTArg Lys Leu Lys Arg Glu Ile Thr Phe His Gly Ala Lys Glu Ile Ser Leu Ser Tyr Ser105 110 115 120GCT GGT GCA CTT GCC AGT TGT ATG GGC CTC ATA TAC AAC AGG ATG GGG GCT GTG ACC ACTAla Gly Ala Leu Ala Ser Cys Met Gly Leu Ile Tyr Asn Arg Met Gly Ala Val Thr Thr125 130 135 140GAA GTG GCA TTT GGC CTG GTA TGT GCA ACC TGT GAA CAG ATT GCT GAC TCC CAG CAT CGGGlu Val Ala Phe Gly Leu Val Cys Ala Thr Cys Glu Gln Ile Ala Asp Ser Gln His Arg145 150 155 160TCT CAT AGG CAA ATG GTG ACA ACA ACC AAC CCA CTA ATC AGA CAT GAG AAC AGA ATG GTTSer His Arg Gln Met Val Thr Thr Thr Asn Pro Leu Ile Arg His Glu Asn Arg Met Val165 170 175 180TTA GCC AGC ACT ACA GCT AAG GCT ATG GAG CAA ATG GCT GGA TCG AGT GAG CAA GCA GCALeu Ala Ser Thr Thr Ala Lys Ala Met Glu Gln Met Ala Gly Ser Ser Glu Gln Ala Ala185 190 195 200GAG GCC ATG GAG GTT GCT AGT CAG GCT AGG CAA ATG GTG CAA GCG ATG AGA ACC ATT GGG
Glu Ala Met Glu Val Ala Ser Gln Ala Arg Gln Met Val Gln Ala Met Arg Thr Ile Gly205 210 215 220ACT CAT CCT AGC TCC AGT GCT GGT CTG AAA AAT GAT CTT CTT GAA AAT TTG CAG GCC TATThr His Pro Ser Ser Ser Ala Gly Leu Lys Asn Asp Leu Leu Glu Asn Leu Gln Ala Tyr225 230 235 240CAG AAA CGA ATG GGG GTG CAG ATG CAA CGG TTC AAG TGAGln Lys Arg Met Gly Val Gln Met Gln Arg Phe Lys ***245 250Met Ser Leu Leu Thr Glu Val Glu Thr Tyr Val Leu Ser Ile Ile Pro Ser Gly Pro Leu5 10 15 20Lys Ala Glu Ile Ala Gln Arg Leu Glu Asp Val Phe Ala Gly Lys Asn Thr Asp Leu Glu25 30 35 40Val Leu Met Glu Trp Leu Lys Thr Arg Pro Ile Leu Ser Pro Leu Thr Lys Gly Ile Leu45 50 55 60Gly Phe Val Phe Thr Leu Thr Val Pro Ser Glu Arg Gly Leu Gln Arg Arg Arg Phe Val65 70 75 80Gln Asn Ala Leu Asn Gly Asn Gly Asp Pro Asn Asn Met Asp Lys Ala Val Lys Leu Tyr85 90 95 100Arg Lys Leu Lys Arg Glu Ile Thr Phe His Gly Ala Lys Glu Ile Ser Leu Ser Tyr Ser105 110 115 120Ala Gly Ala Leu Ala Ser Cys Met Gly Leu Ile Tyr Asn Arg Met Gly Ala Val Thr Thr125 130 135 140Glu Val Ala Phe Gly Leu Val Cys Ala Thr Cys Glu Gln Ile Ala Asp Ser Gln His Arg145 150 155 160Ser His Arg Gln Met Val Thr Thr Thr Asn Pro Leu Ile Arg His Glu Asn Arg Met Val165 170 175 180Leu Ala Ser Thr Thr Ala Lys Ala Met Glu Gln Met Ala Gly Ser Ser Glu Gln Ala Ala185 190 195 200Glu Ala Met Glu Val Ala Ser Gln Ala Arg Gln Met Val Gln Ala Met Arg Thr Ile Gly205 210 215 220Thr His Pro Ser Ser Ser Ala Gly Leu Lys Asn Asp Leu Leu Glu Asn Leu Gln Ala Tyr225 230 235 240Gln Lys Arg Met Gly Val Gln Met Gln Arg Phe Lys ***245 250SEQ ID No.8ATG GAT CCA AAC ACT GTG TCA AGC TTT CAG GTA GAT TGC TTT CTT TGG CAT GTC CGC AAAMet Asp Pro Asn Thr Val Ser Ser Phe Gln Val Asp Cys Phe Leu Trp His Val Arg Lys5 10 15 20CGA GTT GCA GAC CAA GAA CTA GGT GAT GCC CCA TTC CTT GAT CGG CTT CGC CGA GAT CAGArg Val Ala Asp Gln Glu Leu Gly Asp Ala Pro Phe Leu Asp Arg Leu Arg Arg Asp Gln25 30 35 40AAA TCC CTA AGA GGA AGG GGC AGC ACT CTT GGT CTG GAC ATC GAG ACA GCC ACA CGT GCTLys Ser Leu Arg Gly Arg Gly Ser Thr Leu Gly Leu Asp Ile Glu Thr Ala Thr Arg Ala45 50 55 60GGA AAG CAG ATA GTG GAG CGG ATT CTG AAA GAA GAA TCC GAT GAG GCA CTT AAA ATG ACCGly Lys Gln Ile Val Glu Arg Ile Leu Lys Glu Glu Ser Asp Glu Ala Leu Lys Met Thr65 70 75 80ATG GCC TCT GTA CCT GCG TCG CGT TAC CTA ACC GAC ATG ACT CTT GAG GAA ATG TCA AGGMet Ala Ser Val Pro Ala Ser Arg Tyr Leu Thr Asp Met Thr Leu Glu Gln Met Ser Arg85 90 95 100GAA TGG TCC ATG CTC ATA CCC AAG CAG AAA GTG GCA GGC CCT CTT TGT ATC AGA ATG GAC
Glu Trp Ser Met Leu Ile Pro Lys Gln Lys Val Ala Gly Pro Leu Cys Ile Arg Met Asp105 110 115 120CAG GCG ATC ATG GAT AAA AAC ATC ATA CTG AAA GCG AAC TTC AGT GTG ATT TTT GAC CGGGln Ala Ile Met Asp Lys Asn Ile Ile Leu Lys Ala Asn Phe Ser Val Ile Phe Asp Arg125 130 135 140CTG GAG ACT CTA ATA TTG CTA AGG GCT TTC ACC GAA GAG GGA GCA ATT GTT GGC GAA ATTLeu Glu Thr Leu Ile Leu Leu Arg Ala Phe Thr Glu Glu Gly Ala Ile Val Gly Glu Ile145 150 155 160TCA CCA TTG CCT TCT CTT CCA GGA CAT ACT GCT GAG GAT GTC AAA AAT GCA GTT GGA GTCSer Pro Leu Pro Ser Leu Pro Gly His Thr Ala Glu Asp Val Lys Asn Ala Val Gly Val165 170 175 180CTC ATC GGA GGA CTT GAA TGG AAT GAT AAC ACA GTT CGA GTC TCT GAA ACT CTA CAG AGALeu Ile Gly Gly Leu Glu Trp Asn Asp Asn Thr Val Arg Val Ser Glu Thr Leu Gln Arg185 190 195 200TTC GCT TGG AGA AGC AGT AAT GAG AAT GGG AGA CCT CCA CTC ACT CCA AAA CAG AAA CGAPhe Ala Trp Arg Ser Ser Asn Glu Asn Gly Arg Pro Pro Leu Thr Pro Lys Gln Lys Arg205 210 215 220GAA ATG GCG GGA ACA ATT AGG TCA GAA GTT TGAGlu Met Ala Gly Thr Ile Arg Ser Glu Val ***225 230Met Asp Pro Asn Thr Val Ser Ser Phe Gln Val Asp Cys Phe Leu Trp His Val Arg Lys5 10 15 20Arg Val Ala Asp Gln Glu Leu Gly Asp Ala Pro Phe Leu Asp Arg Leu Arg Arg Asp Gln25 30 35 40Lys Ser Leu Arg Gly Arg Gly Ser Thr Leu Gly Leu Asp Ile Glu Thr Ala Thr Arg Ala45 50 55 60Gly Lys Gln Ile Val Glu Arg Ile Leu Lys Glu Glu Ser Asp Glu Ala Leu Lys Met Thr65 70 75 80Met Ala Ser Val Pro Ala Ser Arg Tyr Leu Thr Asp Met Thr Leu Glu Glu Met Ser Arg85 90 95 100Glu Trp Ser Met Leu Ile Pro Lys Gln Lys Val Ala Gly Pro Leu Cys Ile Arg Met Asp105 110 115 120Gln Ala Ile Met Asp Lys Asn Ile Ile Leu Lys Ala Asn Phe Ser Val Ile Phe Asp Arg125 130 135 140Leu Glu Thr Leu Ile Leu Leu Arg Ala Phe Thr Glu Glu Gly Ala Ile Val Gly Glu Ile145 150 155 160Ser Pro Leu Pro Ser Leu Pro Gly His Thr Ala Glu Asp Val Lys Asn Ala Val Gly Val165 170 175 180Leu Ile Gly Gly Leu Glu Trp Asn Asp Asn Thr Val Arg Val Ser Glu Thr Leu Gln Arg185 190 195 200Phe Ala Trp Arg Ser Ser Asn Glu Asn Gly Arg Pro Pro Leu Thr Pro Lys Gln Lys Arg205 210 215 220Glu Met Ala Gly Thr Ile Arg Ser Glu Val ***225 230
权利要求
1一种人工重组的H7亚型流感病毒，其组成包括H7亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)的HA和NA基因，其特征是所述的病毒上还重组有流感病毒A/PR/8/34(H1N1)的6个内部基因PB2、PB1、PA、NP、M和NS，保藏号CCTCC-V200415。
2根据权利要求1所述的人工重组的H7亚型流感病毒，其特征是HA和NA基因具有序列表SEQ ID No.4和SEQ ID No.6所述的序列，6个内部基因PB2、PB1、PA、NP、M和NS具有序列表SEQ ID No.1、SEQ ID No.2、SEQID No.3、SEQ ID No.5、SEQ ID No.7和SEQ ID No.8所述的序列，该病毒对我国流行的H7N2禽流感病毒具有特异免疫性，具有高滴度鸡胚适应性，对鸡及鸡胚无致病力。
3一种权利要求1或2所述的人工重组的H7亚型流感病毒的制备方法，包括以下步骤a.共感染-使两个病毒基因组发生重组流感病毒A/PR/8/34(PR8)(H1N1)与经紫外线处理过的H7N2亚型AIV国内分离株A/Chicken/HeBei/1/02(H7N2)共同接种10日龄鸡胚尿囊腔；b.筛选-除去表面基因来源于PR8病毒的病毒在上述特定的人工条件下收获的尿囊液，与倍比稀释的抗PR8的血清混合，室温下孵育之后接种鸡胚尿囊腔，收获后，对鸡胚尿囊液进行血凝活性检测，收获血清稀释度最低并且具有血凝活性的样品用于第二轮筛选；c.稀释纯化-除去内部基因来源于H7N2病毒同上方法筛选得到的病毒在鸡胚上进行3代有限克隆纯化，利用片段特异性引物RT-PCR鉴定并经过序列分析筛选HA和NA来自于H7N2、6个内部基因PB2、PB1、PA、NP、M及NS来自PR8的重组病毒。
4.上述的人工重组的H7亚型流感病毒，作为防制动物和人类H7亚型流感的疫苗株，应用于疫苗防制。
全文摘要
人工重组的H7亚型流感病毒及其制备方法和应用，在国内，虽然高致病性的H7亚型流感病毒还没有分离到，但是也曾分离到过一株自然分离株H7亚型LPAIV。人工重组的H7亚型流感病毒，其组成包括H7亚型禽流感病毒A/Chicken/HeBei/1/02(H7N2)的HA和NA基因，所述的病毒上还重组有流感病毒A/PR/8/34(H1N1)的6个内部基因PB2、PB1、PA、NP、M和NS，保藏号CCTCC－V200415。该病毒对我国流行的H7N2禽流感病毒具有特异免疫性，具有高滴度鸡胚适应性，对鸡及鸡胚无致病力。
文档编号A61K39/00GK1746298SQ20041004386
公开日2006年3月15日 申请日期2004年9月10日 优先权日2004年9月10日
发明者陈化兰, 施建忠, 田国彬, 李雁冰 申请人:中国农业科学院哈尔滨兽医研究所
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