CN1209374C - 具有促进3t3细胞转化功能的新的人蛋白及其编码序列 - Google Patents
具有促进3t3细胞转化功能的新的人蛋白及其编码序列 Download PDFInfo
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Abstract
本发明公开了一类新的具有促进3T3细胞转化功能的人蛋白,编码此多肽的多核苷酸和经重组技术产生该多肽的方法。本发明还公开了抗此多肽的拮抗剂及其治疗作用。本发明还公开了编码这类新的具有促进3T3细胞转化功能的人蛋白的多核苷酸的用途。
Description
技术领域
本发明属于生物技术领域,具体地说,本发明涉及新的编码具有促进3T3细胞转化功能的人蛋白的多核苷酸,以及此多核苷酸编码的多肽。本发明还涉及此多核苷酸和多肽的用途和制备。
背景技术
人基因组学研究目前是国际上的热点,除人染色体DNA大规模测序,表达序列测序(EST)的方法外,还缺少从功能开始的筛选具有功能基因的高通量的方法。
癌症是危害人类健康的主要疾病之一。为了有效地治疗和预防肿瘤,目前人们已越来越关注肿瘤的基因治疗。因此,本领域迫切需要开发研究与癌细胞生长相关的人蛋白及其激动剂/抑制剂。
发明内容
本发明的目的是提供一类新的具有促进3T3细胞转化功能的人蛋白多肽以及其片段、类似物和衍生物。
本发明的另一目的是提供编码这些多肽的多核苷酸。
本发明的另一目的是提供生产这些多肽的方法以及该多肽和编码序列的用途。
在本发明的第一方面,提供新颖的分离出的具有促进3T3细胞转化功能的蛋白多肽,它包含具有选自下组的氨基酸序列的多肽:SEQ ID NO:2、5、8、11、14、17、20、23;或其保守性变异多肽、或其活性片段、或其活性衍生物。
较佳地,该多肽是具有选自下组的氨基酸序列的多肽:SEQ ID NO:2、5、8、11、14、17、20、23。
在本发明的第二方面,提供了一种分离的多核苷酸,它包含一核苷酸序列,该核苷酸序列与选自下组的一种核苷酸序列有至少85%相同性:(a)编码上述的具有促进3T3细胞转化功能的蛋白多肽的多核苷酸;(b)与多核苷酸(a)互补的多核苷酸。较佳地,该多核苷酸编码的多肽具有选自下组的氨基酸序列:SEQ ID NO:2、5、8、11、14、17、20、23。更佳地,该多核苷酸的序列选自下组:SEQ ID NO:3、6、9、12、15、18、21、24的编码区序列或全长序列。
在本发明的第三方面,提供了含有上述多核苷酸的载体,以及被该载体转化或转导的宿主细胞或者被上述多核苷酸直接转化或转导的宿主细胞。
在本发明的第四方面,提供了制备具有促进3T3细胞转化功能的蛋白活性的多肽的制备方法,该方法包含:(a)在适合表达具有促进3T3细胞转化功能的蛋白的条件下,培养上述被转化或转导的宿主细胞;(b)从培养物中分离出具有促进3T3细胞转化功能的蛋白活性的多肽。
在本发明的第五方面,提供了与上述的具有促进3T3细胞转化功能的蛋白多肽特异性结合的抗体。还提供了可用于检测的核酸分子,它含有上述的多核苷酸中连续10个核苷酸至全长核苷酸,较佳地它含有连续的约10-800个核苷酸。
在本发明的第六方面,提供了一种药物组合物,它含有安全有效量的本发明的具有促进3T3细胞转化功能的蛋白多肽以及药学上可接受的载体。这些药物组合物可用于促进细胞的生长。本发明还提供了一种药物组合物,它含有安全有效量的针对本发明的具有促进3T3细胞转化功能的蛋白多肽的拮抗剂(如抗体)以及药学上可接受的载体。该药物组合物可治疗癌症以及细胞异常增殖等病症。
本发明的其它方面由于本文的技术的公开,对本领域的技术人员而言是显而易见的。
3T3细胞是一种小鼠成纤维细胞(J.Cell.Biol.,17:299,1963)。在癌症研究领域中,常将外源基因(尤其是人基因)引入3T3细胞,观察其对3T3细胞生长的影响情况。通常认为,对3T3细胞生长(或恶性转化)有影响的基因是癌症相关基因,其中对3T3细胞生长或转化有抑制作用的基因大多是抑癌基因,而对3T3细胞生长或转化有促进作用的基因大多是(原)癌基因。
本发明采用大规模cDNA克隆转染小鼠胚胎成纤维细胞3T3,在获得具有促进生长作用的基础上,经测序证明为新的基因,进一步得到全长cDNA克隆。DNA转染试验证明,本发明的具有促进3T3细胞转化功能的蛋白对3T3细胞具有促进克隆形成的作用,其促进率≥50%。
如本文所用,“分离的”是指物质从其原始环境中分离出来(如果是天然的物质,原始环境即是天然环境)。如活体细胞内的天然状态下的多聚核苷酸和多肽是没有分离纯化的,但同样的多聚核苷酸或多肽如从天然状态中同存在的其他物质中分开,则为分离纯化的。
如本文所用,“分离的具有促进3T3细胞转化功能的蛋白或多肽”是指具有促进3T3细胞转化功能的蛋白多肽基本上不含天然与其相关的其它蛋白、脂类、糖类或其它物质。本领域的技术人员能用标准的蛋白质纯化技术纯化具有促进3T3细胞转化功能的蛋白。基本上纯的多肽在非还原聚丙烯酰胺凝胶上能产生单一的主带。
本发明的多肽可以是重组多肽、天然多肽、合成多肽,优选重组多肽。本发明的多肽可以是天然纯化的产物,或是化学合成的产物,或使用重组技术从原核或真核宿主(例如,细菌、酵母、高等植物、昆虫和哺乳动物细胞)中产生。根据重组生产方案所用的宿主,本发明的多肽可以是糖基化的,或可以是非糖基化的。本发明的多肽还可包括或不包括起始的甲硫氨酸残基。
本发明还包括具有促进3T3细胞转化功能的人蛋白的片段、衍生物和类似物。如本文所用,术语“片段”、“衍生物”和“类似物”是指基本上保持本发明的天然具有促进3T3细胞转化功能的人蛋白相同的生物学功能或活性的多肽。本发明的多肽片段、衍生物或类似物可以是(i)有一个或多个保守或非保守性氨基酸残基(优选保守性氨基酸残基)被取代的多肽,而这样的取代的氨基酸残基可以是也可以不是由遗传密码编码的,或(ii)在一个或多个氨基酸残基中具有取代基团的多肽,或(iii)成熟多肽与另一个化合物(比如延长多肽半衰期的化合物,例如聚乙二醇)融合所形成的多肽,或(iv)附加的氨基酸序列融合到此多肽序列而形成的多肽(如前导序列或分泌序列或用来纯化此多肽的序列或蛋白原序列)。根据本文的教导,这些片段、衍生物和类似物属于本领域熟练技术人员公知的范围。
本发明的多核苷酸可以是DNA形式或RNA形式。DNA形式包括cDNA、基因组DNA或人工合成的DNA。DNA可以是单链的或是双链的。DNA可以是编码链或非编码链。以PP12719蛋白(在本申请中,蛋白质的命名采用其克隆编号)为例,编码成熟多肽的编码区序列可以与SEQ ID NO:3所示的编码区序列相同或者是简并的变异体。如本文所用,“简并的变异体”在本发明中是指编码具有SEQ ID NO:2的蛋白质,但与SEQ ID NO:3所示的编码区序列有差别的核酸序列。再以PP13181蛋白(在本申请中,蛋白质的命名采用其克隆编号)为例,编码成熟多肽的编码区序列可以与SEQ ID NO:6所示的编码区序列相同或者是简并的变异体。对于其他具有促进3T3细胞转化功能的蛋白,依此类推。
编码成熟多肽的多核苷酸包括:只编码成熟多肽的编码序列;成熟多肽的编码序列和各种附加编码序列;成熟多肽的编码序列(和任选的附加编码序列)以及非编码序列。
术语“编码多肽的多核苷酸”可以是包括编码此多肽的多核苷酸,也可以是还包括附加编码和/或非编码序列的多核苷酸。
本发明还涉及上述多核苷酸的变异体,其编码与本发明有相同的氨基酸序列的多肽或多肽的片段、类似物和衍生物。此多核苷酸的变异体可以是天然发生的等位变异体或非天然发生的变异体。这些核苷酸变异体包括取代变异体、缺失变异体和插入变异体。如本领域所知的,等位变异体是一个多核苷酸的替换形式,它可能是一个或多个核苷酸的取代、缺失或插入,但不会从实质上改变其编码的多肽的功能。
本发明还涉及与上述的序列杂交且两个序列之间具有至少50%,较佳地至少70%,更佳地至少80%相同性的多核苷酸。本发明特别涉及在严格条件下与本发明所述多核苷酸可杂交的多核苷酸。在本发明中,“严格条件”是指:(1)在较低离子强度和较高温度下的杂交和洗脱,如0.2×SSC,0.1%SDS,60℃;或(2)杂交时加有变性剂,如50%(v/v)甲酰胺,0.1%小牛血清/0.1%Ficoll,42℃等;或(3)仅在两条序列之间的相同性至少在95%以上,更好是97%以上时才发生杂交。并且,可杂交的多核苷酸编码的多肽与SEQ IDNO:2所示的成熟多肽(以PP12719蛋白为例)有相同的生物学功能和活性。
本发明还涉及与上述的序列杂交的核酸片段。如本文所用,“核酸片段”的长度至少含15个核苷酸,较好是至少30个核苷酸,更好是至少50个核苷酸,最好是至少100个核苷酸以上。核酸片段可用于核酸的扩增技术(如PCR)以确定和/或分离编码具有促进3T3细胞转化功能的蛋白的多聚核苷酸。
本发明中的多肽和多核苷酸优选以分离的形式提供,更佳地被纯化至均质。
本发明的DNA序列能用几种方法获得。例如,用本领域熟知的杂交技术分离DNA。这些技术包括但不局限于:1)用探针与基因组或cDNA文库杂交以检出同源性核苷酸序列,和2)表达文库的抗体筛选以检出具有共同结构特征的克隆的DNA片段。
编码具有促进3T3细胞转化功能的蛋白的特异DNA片段序列产生也能用下列方法获得:1)从基因组DNA分离双链DNA序列;2)化学合成DNA序列以获得所需多肽的双链DNA。
当需要的多肽产物的整个氨基酸序列已知时,DNA序列的直接化学合成是经常选用的方法。如果所需的氨基酸的整个序列不清楚时,DNA序列的直接化学合成是不可能的,选用的方法是cDNA序列的分离。分离感兴趣的cDNA的标准方法是从高表达该基因的供体细胞分离mRNA并进行逆转录,形成质粒或噬菌体cDNA文库。提取mRNA的方法已有多种成熟的技术,试剂盒也可从商业途径获得(Qiagene)。而构建cDNA文库也是通常的方法(Sambrook,et al.,Molecular Cloning,A Laboratory Manual,Cold Spring HarborLaboratory.New York,1989)。还可得到商业供应的cDNA文库,如Clontech公司的不同cDNA文库。当结合使用聚合酶反应技术时,即使极少的表达产物也能克隆。
可用常规方法从这些cDNA文库中筛选本发明的基因。这些方法包括(但不限于):(1)DNA-DNA或DNA-RNA杂交;(2)标志基因的功能出现或丧失;(3)测定具有促进3T3细胞转化功能的蛋白的转录本的水平;(4)通过免疫学技术或测定生物学活性,来检测基因表达的蛋白产物。上述方法可单用,也可多种方法联合应用。
在第(1)种方法中,杂交所用的探针是与本发明的多核苷酸的任何一部分同源,其长度至少15个核苷酸,较好是至少30个核苷酸,更好是至少50个核苷酸,最好是至少100个核苷酸。此外,探针的长度通常在2kb之内,较佳地为1kb之内。此处所用的探针通常是在本发明的基因DNA序列信息的基础上化学合成的DNA序列。本发明的基因本身或者片段当然可以用作探针。DNA探针的标记可用放射性同位素,荧光素或酶(如碱性磷酸酶)等。
在第(4)种方法中,检测具有促进3T3细胞转化功能的蛋白基因表达的蛋白产物可用免疫学技术如Western印迹法,放射免疫沉淀法,酶联免疫吸附法(ELISA)等。
应用PCR技术扩增DNA/RNA的方法(Saiki,et al.Science 1985;230:1350-1354)被优选用于获得本发明的基因。特别是很难从文库中得到全长的cDNA时,可优选使用RACE法(RACE-cDNA末端快速扩增法),用于PCR的引物可根据本文所公开的本发明的序列信息适当地选择,并可用常规方法合成。可用常规方法如通过凝胶电泳分离和纯化扩增的DNA/RNA片段。
如上所述得到的本发明的基因,或者各种DNA片段等的核苷酸序列的测定可用常规方法如双脱氧链终止法(Sanger et al.PNAS,1977,74:5463-5467)。这类核苷酸序列测定也可用商业测序试剂盒等。为了获得全长的cDNA序列,测序需反复进行。有时需要测定多个克隆的cDNA序列,才能拼接成全长的cDNA序列。
本发明也涉及包含本发明多核苷酸的载体,以及用本发明的载体或具有促进3T3细胞转化功能的蛋白编码序列经基因工程产生的宿主细胞,以及经重组技术产生本发明所述多肽的方法。
通过常规的重组DNA技术(Science,1984;224:1431),可利用本发明的多聚核苷酸序列可用来表达或生产重组的具有促进3T3细胞转化功能的蛋白多肽。一般来说有以下步骤:
(1).用本发明的编码具有促进3T3细胞转化功能的人蛋白的多核苷酸(或变异体),或用含有该多核苷酸的重组表达载体转化或转导合适的宿主细胞;
(2).在合适的培养基中培养的宿主细胞;
(3).从培养基或细胞中分离、纯化蛋白质。
本发明中,具有促进3T3细胞转化功能的人蛋白多核苷酸序列可插入到重组表达载体中。术语“重组表达载体”指本领域熟知的细菌质粒、噬菌体、酵母质粒、植物细胞病毒、哺乳动物细胞病毒如腺病毒、逆转录病毒或其他载体。在本发明中适用的载体包括但不限于:在细菌中表达的基于T7的表达载体(Rosenberg,et al.Gene,1987,56:125);在哺乳动物细胞中表达的pMSXND表达载体(Lee and Nathans,J Bio Chem.263:3521,1988)和在昆虫细胞中表达的来源于杆状病毒的载体。总之,只要能在宿主体内复制和稳定,任何质粒和载体都可以用。表达载体的一个重要特征是通常含有复制起点、启动子、标记基因和翻译控制元件。
本领域的技术人员熟知的方法能用于构建含具有促进3T3细胞转化功能的人蛋白编码DNA序列和合适的转录/翻译控制信号的表达载体。这些方法包括体外重组DNA技术、DNA合成技术、体内重组技术等(Sambroook,et al)。所述的DNA序列可有效连接到表达载体中的适当启动子上,以指导mRNA合成。这些启动子的代表性例子有:大肠杆菌的lac或trp启动子;λ噬菌体PL启动子;真核启动子包括CMV立即早期启动子、早期和晚期SV40启动子和其他一些已知的可控制基因在原核或真核细胞或其病毒中表达的启动子。表达载体还包括翻译起始用的核糖体结合位点和转录终止子。
此外,表达载体优选地包含一个或多个选择性标记基因,以提供用于选择转化的宿主细胞的表型性状,如真核细胞培养用的二氢叶酸还原酶、新霉素抗性以及绿色荧光蛋白(GFP),或用于大肠杆菌的四环素或氨苄青霉素抗性。
包含上述的适当DNA序列以及适当启动子或者控制序列的载体,可以用于转化适当的宿主细胞,以使其能够表达蛋白质。
宿主细胞可以是原核细胞,如细菌细胞;或是低等真核细胞,如酵母细胞;或是高等真核细胞,如哺乳动物细胞。代表性例子有:大肠杆菌,链霉菌属;鼠伤寒沙门氏菌的细菌细胞;真菌细胞如酵母;植物细胞;果蝇S2或Sf9的昆虫细胞;CHO、COS或Bowes黑素瘤细胞的动物细胞等。
本发明的多核苷酸在高等真核细胞中表达时,如果在载体中插入增强子序列时将会使转录得到增强。增强子是DNA的顺式作用因子,通常大约有10到300个碱基对,作用于启动子以增强基因的转录。可举的例子包括在复制起始点晚期一侧的100到270个碱基对的SV40增强子、在复制起始点晚期一侧的多瘤增强子以及腺病毒增强子等。
本领域一般技术人员都清楚如何选择适当的载体、启动子、增强子和宿主细胞。
用重组DNA转化宿主细胞可用本领域技术人员熟知的常规技术进行。当宿主为原核生物如大肠杆菌时,能吸收DNA的感受态细胞可在指数生长期后收获,用CaCl2法处理,所用的步骤在本领域众所周知。可供选择的是用MgCl2。如果需要,转化也可用电穿孔的方法进行。当宿主是真核生物,可选用如下的DNA转染方法:磷酸钙共沉淀法,常规机械方法如显微注射、电穿孔、脂质体包装等。
获得的转化子可以用常规方法培养,表达本发明的基因所编码的多肽。根据所用的宿主细胞,培养中所用的培养基可选自各种常规培养基。在适于宿主细胞生长的条件下进行培养。当宿主细胞生长到适当的细胞密度后,用合适的方法(如温度转换或化学诱导)诱导选择的启动子,将细胞再培养一段时间。
在上面的方法中的重组多肽可包被于细胞内、细胞外或在细胞膜上表达或分泌到细胞外。如果需要,可利用其物理的、化学的和其它特性通过各种分离方法分离和纯化重组的蛋白。这些方法是本领域技术人员所熟知的。这些方法的例子包括但并不限于:常规的复性处理、用蛋白沉淀剂处理(盐析方法)、离心、渗透破菌、超处理、超离心、分子筛层析(凝胶过滤)、吸附层析、离子交换层析、高效液相层析(HPLC)和其它各种液相层析技术及这些方法的结合。
重组的具有促进3T3细胞转化功能的人蛋白或多肽有多方面的用途。这些用途包括(但不限于):直接做为药物治疗具有促进3T3细胞转化功能的蛋白功能低下或丧失所致的疾病,和用于筛选促进或对抗具有促进3T3细胞转化功能的蛋白功能的抗体、多肽或其它配体。例如,该抗体可用于治疗癌症或细胞异常增殖。用重组表达的本发明蛋白筛选多肽库可用于寻找有治疗价值的能抑制或刺激具有促进3T3细胞转化功能的人蛋白功能的多肽分子。
本发明也提供了筛选药物以鉴定提高(激动剂)或阻遏(拮抗剂)具有促进3T3细胞转化功能的人蛋白的药剂的方法。激动剂提高具有促进3T3细胞转化功能的人蛋白刺激细胞增殖等生物功能,而拮抗剂阻止和治疗与细胞过度增殖有关的紊乱如各种癌症。
具有促进3T3细胞转化功能的人蛋白的拮抗剂包括筛选出的抗体、化合物、受体缺失物和类似物等。具有促进3T3细胞转化功能的人蛋白的拮抗剂可以与具有促进3T3细胞转化功能的人蛋白结合并消除其功能,或是抑制具有促进3T3细胞转化功能的人蛋白的产生,或是与多肽的活性位点结合使多肽不能发挥生物学功能。具有促进3T3细胞转化功能的人蛋白的拮抗剂可用于治疗用途。
在筛选作为拮抗剂的化合物时,可以将具有促进3T3细胞转化功能的蛋白加入生物分析测定中,通过测定化合物影响具有促进3T3细胞转化功能的蛋白和其受体之间的相互作用来确定化合物是否是拮抗剂。用上述筛选化合物的同样方法,可以筛选出起拮抗剂作用的受体缺失物和类似物。
本发明蛋白的拮抗剂可直接用于疾病治疗,例如,各种恶性肿瘤、和细胞异常增殖等。
本发明的多肽,及其片段、衍生物、类似物或它们的细胞可以用来作为抗原以生产抗体。这些抗体可以是多克隆或单克隆抗体。多克隆抗体可以通过将此多肽直接注射动物的方法得到。制备单克隆抗体的技术包括杂交瘤技术,三瘤技术,人B-细胞杂交瘤技术,EBV-杂交瘤技术等。
可以将本发明的多肽和拮抗剂与合适的药物载体组合后使用。这些载体可以是水、葡萄糖、乙醇、盐类、缓冲液、甘油以及它们的组合。组合物包含安全有效量的多肽或拮抗剂以及不影响药物效果的载体和赋形剂。这些组合物可以作为药物用于疾病治疗。
本发明还提供含有一种或多种容器的药盒或试剂盒,容器中装有一种或多种本发明的药用组合物成分。与这些容器一起,可以有由制造、使用或销售药品或生物制品的政府管理机构所给出的指示性提示,该提示反映出生产、使用或销售的政府管理机构许可其在人体上施用。此外,本发明的多肽可以与其它的治疗化合物结合使用。
药物组合物可以以方便的方式给药,如通过局部、静脉内、腹膜内、肌内、皮下、鼻内或皮内的给药途径。具有促进3T3细胞转化功能的蛋白或其特异性抗体,可按有效地治疗和/或预防具体的适应症的量来给药。施用于患者的具有促进3T3细胞转化功能的蛋白的量和剂量范围将取决于许多因素,如给药方式、待治疗者的健康条件和诊断医生的判断。
具有促进3T3细胞转化功能的人蛋白的多聚核苷酸也可用于多种治疗目的。基因治疗技术可用于治疗由于具有促进3T3细胞转化功能的蛋白的无表达或异常/无活性的具有促进3T3细胞转化功能的蛋白的表达所致的细胞发育或代谢异常。重组的基因治疗载体(如病毒载体)可设计成表达变异的具有促进3T3细胞转化功能的蛋白,以抑制内源性的具有促进3T3细胞转化功能的蛋白活性。例如,一种变异的具有促进3T3细胞转化功能的蛋白可以是缩短的、缺失了信号传导功能域的具有促进3T3细胞转化功能的蛋白,虽可与下游的底物结合,但缺乏信号传导活性。因此重组的基因治疗载体可用于治疗具有促进3T3细胞转化功能的蛋白表达或活性异常所致的疾病。来源于病毒的表达载体如逆转录病毒、腺病毒、腺病毒相关病毒、单纯疱疹病毒、细小病毒等可用于将具有促进3T3细胞转化功能的蛋白基因转移至细胞内。构建携带具有促进3T3细胞转化功能的蛋白基因的重组病毒载体的方法可见于已有文献(Sambrook,et al.)。另外重组具有促进3T3细胞转化功能的人蛋白基因可包装到脂质体中转移至细胞内。
抑制具有促进3T3细胞转化功能的人蛋白mRNA的寡聚核苷酸(包括反义RNA和DNA)以及核酶也在本发明的范围之内。核酶是一种能特异性分解特定RNA的酶样RNA分子,其作用机制是核酶分子与互补的靶RNA特异性杂交后进行核酸内切作用。反义的RNA和DNA及核酶可用已有的任何RNA或DNA合成技术获得,如固相磷酸酰胺化学合成法合成寡核苷酸的技术已广泛应用。反义RNA分子可通过编码该RNA的DNA序列在体外或体内转录获得。这种DNA序列已整合到载体的RNA聚合酶启动子的下游。为了增加核酸分子的稳定性,可用多种方法对其进行修饰,如增加两侧的序列长度,核糖核苷之间的连接应用磷酸硫酯键或肽键而非磷酸二酯键。
多聚核苷酸导入组织或细胞内的方法包括:将多聚核苷酸直接注入到体内组织中;或在体外通过载体(如病毒、噬菌体或质粒等)先将多聚核苷酸导入细胞中,再将细胞移植到体内等。由于本发明蛋白具有促进3T3细胞转化的功能,因此本发明蛋白编码序列的反义序列,可被引入细胞以抑制细胞的异常增殖(如癌变)。
本发明还提供了针对具有促进3T3细胞转化功能的人蛋白抗原决定簇的抗体。这些抗体包括(但不限于):多克隆抗体、单克隆抗体、嵌合抗体、单链抗体、Fab片段和Fab表达文库产生的片段。
抗具有促进3T3细胞转化功能的人蛋白的抗体可用于免疫组织化学技术中,检测活检标本中的具有促进3T3细胞转化功能的人蛋白。
与具有促进3T3细胞转化功能的人蛋白结合的单克隆抗体也可用放射性同位素标记,注入体内可跟踪其位置和分布。这种放射性标记的抗体可作为一种非创伤性诊断方法用于肿瘤细胞的定位和判断是否有转移。
本发明中的抗体可用于治疗或预防与具有促进3T3细胞转化功能的人蛋白相关的疾病。给予适当剂量的抗体可以阻断具有促进3T3细胞转化功能的人蛋白的产生或活性,从而抑制癌细胞的生长和/或细胞的异常增殖。
抗体也可用于设计针对体内某一特殊部位的免疫毒素。如具有促进3T3细胞转化功能的人蛋白高亲和性的单克隆抗体可与细菌或植物毒素(如白喉毒素,蓖麻蛋白,红豆碱等)共价结合。一种通常的方法是用巯基交联剂如SPDP,攻击抗体的氨基,通过二硫键的交换,将毒素结合于抗体上,这种杂交抗体可用于杀灭有关的阳性细胞(如癌细胞)。
多克隆抗体的生产可用具有促进3T3细胞转化功能的人蛋白或多肽免疫动物,如家兔,小鼠,大鼠等。多种佐剂可用于增强免疫反应,包括但不限于弗氏佐剂等。
具有促进3T3细胞转化功能的人蛋白单克隆抗体可用杂交瘤技术生产(Kohler andMilstein。Nature,1975,256:495-497)。将人恒定区和非人源的可变区结合的嵌合抗体可用已有的技术生产(Morrison et al,PNAS,1985,81:6851)。而已有的生产单链抗体的技术(U.S.Pat No.4946778)也可用于生产抗具有促进3T3细胞转化功能的人蛋白的单链抗体。
能与具有促进3T3细胞转化功能的人蛋白结合的多肽分子可通过筛选由各种可能组合的氨基酸结合于固相物组成的随机多肽库而获得。筛选时,必须对具有促进3T3细胞转化功能的人蛋白分子进行标记。
本发明还涉及定量和定位检测具有促进3T3细胞转化功能的人蛋白水平的诊断试验方法。这些试验为本领域所熟知,且包括FISH测定和放射免疫测定。试验中所检测的具有促进3T3细胞转化功能的蛋白水平,可以用作解释具有促进3T3细胞转化功能的蛋白在各种疾病中的重要性和用于诊断具有促进3T3细胞转化功能的蛋白起作用的疾病。
具有促进3T3细胞转化功能的蛋白的多聚核苷酸可用于具有促进3T3细胞转化功能的蛋白相关疾病的诊断和治疗。在诊断方面,具有促进3T3细胞转化功能的蛋白的多聚核苷酸可用于检测具有促进3T3细胞转化功能的蛋白的表达与否或在疾病状态下具有促进3T3细胞转化功能的蛋白的异常表达。如具有促进3T3细胞转化功能的蛋白DNA序列可用于对活检标本的杂交以判断具有促进3T3细胞转化功能的蛋白的表达异常。杂交技术包括Southern印迹法,Northern印迹法、原位杂交等。这些技术方法都是公开的成熟技术,相关的试剂盒都可从商业途径得到。本发明的多核苷酸的一部分或全部可作为探针固定在微阵列(Microarray)或DNA芯片(即基因芯片)上,用于分析组织中基因的差异表达分析和基因诊断。用具有促进3T3细胞转化功能的蛋白特异的引物进行RNA-聚合酶链反应(RT-PCR)体外扩增也可检测具有促进3T3细胞转化功能的蛋白的转录产物。
检测具有促进3T3细胞转化功能的蛋白基因的突变也可用于诊断具有促进3T3细胞转化功能的蛋白相关的疾病。具有促进3T3细胞转化功能的蛋白突变的形式包括与正常野生型具有促进3T3细胞转化功能的蛋白DNA序列相比的点突变、易位、缺失、重组和其它任何异常等。可用已有的技术如Southern印迹法、DNA序列分析、PCR和原位杂交检测突变。另外,突变有可能影响蛋白的表达,因此用Northern印迹法、Western印迹法可间接判断基因有无突变。
本发明的序列对染色体鉴定也是有价值的。这些序列会特异性地针对某条人染色体具体位置且并可以与其杂交。目前,需要鉴定染色体上的各基因的具体位点。然而现在只有很少的基于实际序列数据(重复多态性)的染色体标记物可用于标记染色体位置。为了将这些序列与疾病相关基因相关联。第一步就是将本发明DNA序列定位于染色体上。
简而言之,根据cDNA制备PCR引物(优选15-35bp),可以将序列定位于染色体上。然后,将这些引物用于PCR筛选含各条人染色体的体细胞杂合细胞。只有那些含有相应于引物的人基因的杂合细胞会产生扩增的片段。
体细胞杂合细胞的PCR定位法,是将DNA定位到具体染色体的快捷方法。使用本发明的的寡核苷酸引物,通过类似方法,可利用一组来自特定染色体的片段或大量基因组克隆而实现亚定位。可用于染色体定位的其它类似策略包括原位杂交、用标记的流式分选的染色体预筛选和杂交预选,从而构建染色体特异的cDNA库。
将cDNA克隆与中期染色体进行荧光原位杂交(FISH),可以在一个步骤中精确地进行染色体定位。此技术的综述,参见Verma等,Human Chromosomes:a Manual of BasicTechniques,Pergamon Press,New York(1988)。
一旦序列被定位到准确的染色体位置,此序列在染色体上的物理位置就可以与基因图数据相关联。这些数据可见于例如,V.Mckusick,Mendelian Inheritance in Man(可通过与Johns Hopkins University Welch Medical Library联机获得)。然后可通过连锁分析,确定基因与业已定位到染色体区域上的疾病之间的关系。
接着,需要测定患病和未患病个体间的cDNA或基因组序列差异。如果在一些或所有的患病个体中观察到某突变,而该突变在任何正常个体中未观察到,则该突变可能是疾病的病因。比较患病和未患病个体,通常涉及首先寻找染色体中结构的变化,如从染色体水平可见的或用基于cDNA序列的PCR可检测的缺失或易位。
本发明的具有促进3T3细胞转化功能的蛋白核苷酸全长序列或其片段通常可以用PCR扩增法、重组法或人工合成的方法获得。对于PCR扩增法,可根据本发明所公开的有关核苷酸序列,尤其是开放阅读框序列来设计引物,并用市售的cDNA库或按本领域技术人员已知的常规方法所制备的cDNA库作为模板,扩增而得有关序列。当序列较长时,常常需要进行两次或多次PCR扩增,然后再将各次扩增出的片段按正确次序拼接在一起。
一旦获得了有关的序列,就可以用重组法来大批量地获得有关序列。这通常是将其克隆入载体,再转入细胞,然后通过常规方法从增殖后的宿主细胞中分离得到有关序列。
此外,还可用人工合成的方法来合成有关序列,尤其是片段长度较短时。通常,通过先合成多个小片段,然后再进行连接可获得序列很长的片段。
目前,已经可以完全通过化学合成来编码本发明蛋白(或其片段,或其衍生物)的DNA序列。然后可将该DNA序列引入本领域中的各种DNA分子(如载体)和细胞中。此外,还可通过化学合成将突变引入本发明蛋白序列中。
此外,由于本发明的具有促进3T3细胞转化功能的蛋白具有源自人的天然氨基酸序列,因此,与来源于其他物种的同族蛋白相比,预计在施用于人时将具有更高的活性和/或更低的副作用(例如在人体内的免疫原性更低或没有)。
下面结合具体实施例,进一步阐述本发明。应理解,这些实施例仅用于说明本发明而不用于限制本发明的范围。下列实施例中未注明具体条件的实验方法,通常按照常规条件如Sambrook等人,分子克隆:实验室手册(New York:Cold Spring Harbor LaboratoryPress,1989)中所述的条件,或按照制造厂商所建议的条件。注意,在核苷酸和氨基酸组合序列中,(1)给出的是起始和终止编码子第一个核苷酸的位置,(2)分子量单位是道尔顿。
具体实施方式
实施例1:cDNA基因的获得及对3T3细胞克隆形成的促进作用
PP12719、PP13181、PP13191、PP13479、PP13439、PP13842、PP14673和PP14776是通过用常规方法构建人胎盘cDNA文库获得的。取3、6、10月龄的胎盘组织,用Trizol试剂(GIBCO BRL公司)按厂方说明书提取总RNA,用mRNA提纯试剂盒(Pharmacia公司)提取mRNA。用pCMV-script TMXR cDNA文库构建试剂盒(Stratagene公司)构建上述mRNA的cDNA文库。其中反转录酶改用MMLV-RT-Superscript II(GIBCOBRL),反转录反应在42℃进行。转化XL 10-Gold感受细胞,获得了1×106cfu/μg滴度的cDNA文库。第一轮随机挑取cDNA克隆,其后以高丰度cDNA克隆和已证明有抑癌细胞生长功能的cDNA克隆为探针,杂交筛选cDNA文库,挑取弱阳性及阴性克隆。用Qiagen 96孔板质粒抽提试剂盒,按厂家说明书进行质粒DNA的提取。质粒DNA和空载体同时转染3T3细胞系。100ng DNA酒精沉淀干燥后,加6μl H2O溶解,待转染。每份DNA样品中加0.74μl脂质体及9.3μl无血清培液,混匀后,室温放置10分钟。每管中加150μl无血清培液,均分加入3孔生长于96孔板的3T3细胞中,37℃放置2小时,每孔再加50μl无血清培液,37℃ 24小时。每孔换100μl全培液,37℃ 24小时,换含G418的全培液100μl,37℃ 24-48小时,边观察,边换G418浓度不等的培液。约2-3次后,直到镜检细胞有克隆形成,计数。发现以上克隆有促进细胞克隆形成作用,结果如下表所示。
cDNA克隆转染细胞(3T3)的克隆形成情况
cDNA克隆名称 | cDNA克隆数(三个重复) | 空载体克隆数(三个重复) |
PP12719PP13181PP13191PP13439PP13479PP13842PP14673PP14776 | 52 51 5344 46 4343 43 4069 56 6244 49 4042 38 2458 57 6246 43 41 | 27 29 3027 29 3027 29 3027 29 3027 29 3027 29 3027 29 3027 29 30 |
对cDNA克隆采用双脱氧终止法,在ABI377 DNA自动测序仪上测定其一端近500bp的核苷酸序列。分析后,确定为新基因克隆,进行另一端测序。对于仍未获得全长cDNA序列的,设计引物,再次进行测序,直到获得全长序列(SEQ ID NO:1、4、7、10、13、16、19、22)。
实施例2:从胎盘cDNA中PCR获得全长基因:
取3、6、10月龄的胎盘组织,用Trizol试剂(GIBCO BRL公司)按厂方说明书提取总RNA,用mRNA提纯试剂盒(Pharmacia公司)提取mRNA。用MMLV-RT-SuperscriptII(GIBCO BRL),反转录酶在42℃进行反转录反应,获得胎盘cDNA。利用各个基因的转异引物(如下表所示),按97℃ 3分钟、1个循环;94℃ 30秒→60℃ 30秒→72℃ 1分钟,共35个循环;72℃ 10分钟,1个循环,进行PCR扩增,获得含有完整开放阅读框序列的各蛋白基因的扩增产物。扩增产物经测序验证,与实施例1测得的序列相符,随后用常规技术将扩增产物转入宿主细胞,获得重组蛋白(SEQ ID NO:2、5、8、11、14、17、20、23)。
基因特异引物
克隆名称 | 特异引物1(5′→3′) | 特异引物2(5′→3′) |
PP12719PP13181PP13191PP13439PP13479PP13842PP14673PP14776 | CTGGAAACCCTGAAGCTGAGGAGATGCCGTGGACTTCCTCCATGGAGAAGGAGCAGTGACCATAGCTGGTGTCCTCCTGCACCGTGTCCTTCCCTCCCTCAGAGGGATGGGATTGGGGTACGTGAAGCTGAAAGCCACTGCCATCCCACTGAAACT | CCAATCAGAGCCAACCTAGCACCCGACTCGTAGGTGAGCCACCAGGACTGGCTGCCTGGGCCAGTTCCATCCTCTTAGCCTCTGTGATCCTTTCCCTGGCCAAAAAGAAAAGGGAGAGATAACAGCTCCTGGAAGGCCCTCAAGCAATACACCCACC |
实施例3:cDNA克隆序列分析
1.PP12719
A:核苷酸序列(SEQ ID NO:1)长度:1179
1 GTGGGATTAC AGGCGTGAGC CACCACCACA CCCAGCCCTG GGAATGGAGT CTTGATTCTT
61 CTCTGCCCCT CACTGATTCT TCCAAACTGG AAACCCTGAA GCTGAGAGCC CAGCATGGTT
121 CCTGGCAAAC AGCAGGCACT CAAATATTGA TTGGTTTACT GTATGACTAG TAGAGACCCC
181 AACGAGCAAA ACTGTGGCCT AATAAAATTC TGGCTCCTCT CCCAGACTTC CCCTCCCTTT
241 GAGAAATGCC AGAAGCTTCT TAGGGAGGCT CTTGCCAACC TAGACATCAC AGGCACTCAT
301 GGGGCAGCTC CAGCCTCTTC CTCCTGTCAT CACCATAATG CATCCATATC TACAATATGG
361 CAAATTTCAT ATCCTTCCAA CCTCTTTCCT GCATTATTGA TGGGCTGTGT GCACTTTTTA
421 AAAAATCAAT TAGATCAGGG CGTGGAGCTG GAGTTCAAAG AAGCCTTTAA AAGTCTGCTC
481 TTCTGTTTTG CTGTTTTGAA TAGGCACAGA TAAAGCTTTC CCTCTGGTTT GAATAAGCCA
541 AGCTCAGTGC TAGGTTGGCT CTGATTGGCC AGGACTAGGA AAATGCGGTT AAGATGCAAA
601 CACAAGCAAA TATAACCCAG TATCTCTGTG GCCATTACTA AGCTAAGGCA GCAGGACCTG
661 GAGCCTCCTG CTTTGGAGTG GTTCTTCAAT ACTGCTGCTG CTTACGCGCC GGGAAACTGG
721 GAAGGCTGGT GAGCGAGAAG GCAAGGTAAG GTCTCTGATT TACGGGGGCA TGCCAGTTAA
781 TCCTCCTGAA TGAGGAAGAA ATGAAAGGAA GAGGAGCTTG AGAGTCCCTT GGCTTTGTCT
841 TCTGAGATGA GGCTTTTAAA ATGAACCAGG AGTCTGGCTG GCCAGTTTTG CAAACTTCTT
901 GTTCAGAAGA ACCCCTTAGA GGCAGCTGAA CATATAGGGC ACATTTTCAA AAGCTGGGAA
961 AGACAGACCC CTTTCACCTA TCCCTAGAGA AAGATGGTAT GGAGCAAGGC AGGAGGAGTT
1021 AAAACCAGCT TCCTGGCCCA GAGAGGTGGC TTACACCTGT AATCCCAACA CTTTGGGAAG
1081 CCAGGGCATG AGGATTGTTT GAGCCCAGGA GTTTGTAACT TGTGACCATC CTGGGCAACA
1141 TAGTGAGACC CTGTCTCTAC AAAAAAAAAA AAAAAAAAA
B:氨基酸序列(SEQ ID NO:2) 长度:116
1 MTSRDPNEQN CGLIKFWLLS QTSPPFEKCQ KLLREALANL DITGTHGAAP ASSSCHHHNA
61 SISTIWQISY PSNLFPALLM GCVHFLKNQL DQGVELEFKE AFKSLLFCFA VLNRHR
C.核苷酸及氨基酸组合序列(SEQ ID NO:3) 克隆号:PP12719
起始编码子:163 ATG 终止编码子:511 TAA 蛋白质分子量:13057.34
1 GTG GGA TTA CAG GCG TGA GCC ACC ACC ACA CCC AGC CCT GGG AAT GGA 48
49 GTC TTG ATT CTT CTC TGC CCC TCA CTG ATT CTT CCA AAC TGG AAA CCC 96
97 TGA AGC TGA GAG CCC AGC ATG GTT CCT GGC AAA CAG CAG GCA CTC AAA 144
145 TAT TGA TTG GTT TAC TGT ATG ACT AGT AGA GAC CCC AAC GAG CAA AAC 192
1 Met Thr Ser Arg Asp Pro Asn Glu Gln Asn 10
193 TGT GGC CTA ATA AAA TTC TGG CTC CTC TCC CAG ACT TCC CCT CCC TTT 240
11 Cys Gly Leu Ile Lys Phe Trp Leu Leu Ser Gln Thr Ser Pro Pro Phe 26
241 GAG AAA TGC CAG AAG CTT CTT AGG GAG GCT CTT GCC AAC CTA GAC ATC 288
27 Glu Lys Cys Gln Lys Leu Leu Arg Glu Ala Leu Ala Asn Leu Asp Ile 42
289 ACA GGC ACT CAT GGG GCA GCT CCA GCC TCT TCC TCC TGT CAT CAC CAT 336
43 Thr Gly Thr His Gly Ala Ala Pro Ala Ser Ser Ser Cys His His His 58
337 AAT GCA TCC ATA TCT ACA ATA TGG CAA ATT TCA TAT CCT TCC AAC CTC 384
59 Asn Ala Ser Ile Ser Thr Ile Trp Gln Ile Ser Tyr Pro Ser Asn Leu 74
385 TTT CCT GCA TTA TTG ATG GGC TGT GTG CAC TTT TTA AAA AAT CAA TTA 432
75 Phe Pro Ala Leu Leu Met Gly Cys Val His Phe Leu Lys Asn Gln Leu 90
433 GAT CAG GGC GTG GAG CTG GAG TTC AAA GAA GCC TTT AAA AGT CTG CTC 480
91 Asp Gln Gly Val Glu Leu Glu Phe Lys Glu Ala Phe Lys Ser Leu Leu 106
481 TTC TGT TTT GCT GTT TTG AAT AGG CAC AGA TAA AGC TTT CCC TCT GGT 528
107 Phe Cys Phe Ala Val Leu Asn Arg His Arg *** 117
529 TTG AAT AAG CCA AGC TCA GTG CTA GGT TGG CTC TGA TTG GCC AGG ACT 576
577 AGG AAA ATG CGG TTA AGA TGC AAA CAC AAG CAA ATA TAA CCC AGT ATC 624
625 TCT GTG GCC ATT ACT AAG CTA AGG CAG CAG GAC CTG GAG CCT CCT GCT 672
673 TTG GAG TGG TTC TTC AAT ACT GCT GCT GCT TAC GCG CCG GGA AAC TGG 720
721 GAA GGC TGG TGA GCG AGA AGG CAA GGT AAG GTC TCT GAT TTA CGG GGG 768
769 CAT GCC AGT TAA TCC TCC TGA ATG AGG AAG AAA TGA AAG GAA GAG GAG 816
817 CTT GAG AGT CCC TTG GCT TTG TCT TCT GAG ATG AGG CTT TTA AAA TGA 864
865 ACC AGG AGT CTG GCT GGC CAG TTT TGC AAA CTT CTT GTT CAG AAG AAC 912
913 CCC TTA GAG GCA GCT GAA CAT ATA GGG CAC ATT TTC AAA AGC TGG GAA 960
961 AGA CAG ACC CCT TTC ACC TAT CCC TAG AGA AAG ATG GTA TGG AGC AAG 1008
1009 GCA GGA GGA GTT AAA ACC AGC TTC CTG GCC CAG AGA GGT GGC TTA CAC 1056
1057 CTG TAA TCC CAA CAC TTT GGG AAC CCA GGG CAT GAG GAT TCT TTG AGC 1104
1105 CCA GGA GTT TGT AAC TTG TGA CCA TCC TGG GCA ACA TAG TGA GAC CCT 1152
1153 GTC TCT ACA AAA AAA AAA AAA AAA AAA 1179
2.PP13181
A:核苷酸序列(SEQ ID NO:4)长度:1652
1 GCGGCCCCCC TCTGAGGGCG AGTTCATCGA CTGCTTCCAC AAAATCAAGC TGGCGATTAA
61 CTTGCTGGTG GGTCCGGTGG CCCCAGCCCT GCCCCACTGT CTGTGCTGAG GGGAGGGTGG
121 AGGCCCCGCC CCGCCCCGGC ACCTGCTCAC TTGTTCCCAC CCCCAGGCAA AGCTGCAGAA
181 GCACATCCAG AACCCCAGCG CCGCGGAGCT CGTGCACTTC CTCTTCGGGC CTCTGGACCT
241 GGTGCCTGGG GCCGGGCGGC AGGGGCGCGC AGGGTGGGGG CCCAGAGGCC TCTGCAGCAT
301 CTCCCCGGGG TCGGGGTTGG GGCAGCAGGT GCCCGCCTTG GGCAGCCCGG TTCACGCTGT
361 GTGGCCACTC TCCTGGGGTC CAAAGTCCCT TCCCGAGGGC CAGCCTGTGG AGCTACGGGG
421 GTGCTGGGCC AGGGTCTGTG GGCCTCAGTC CCCTCTGAAC CTCACTGTGC CCCAGATCGT
481 CAACACCTGC AGTGGCCCAG ACATCGCACG CTCCGTCTCC TGCCCACTGC TCTCCCGAGA
541 TGCCGTGGAC TTCCTGCGCG GCCACCTGGT CCCTAAGGAG ATGTCGCTGT GGGAGTCACT
601 GGGAGAGAGC TGGATGCGGC CCCGTTCCGA GTGGCCGCGG GAGCCACAGG TGCCCCTCTA
661 CGTGCCCAAG TTCCACAGCG GCTGGGAGCC TCCTGTGGAT GTGCTGCAGG AGGCCCCCTG
721 GGAGGTGGAG GGGCTGGCGT CTGCCCCCAT CGAGGAGGTG AGTCCAGTGA GCCGACAGTC
781 CATAAGAAAC TCCCAGAAGC ACAGCCCCAC TTCAGAGCCC ACCCCCCCGG GGGATGCCCT
841 ACCACCAGTC AGCTCCCCAC ATACTCACAG GGGCTACCAG CCAACACCAG CCATGGCCAA
901 GTACGTCAAG ATCCTGTATG ACTTCACAGC CCGAAATGCC AACGAGCTAT CGGTGCTCAA
961 GGATGAGGTC CTAGAGGTGC TGGAGGACGG CCGGCAGTGG TGGAAGCTGC GCAGCCGCAG
1021 CGGCCAGGCG GGGTACGTGC CCTGCAACAT CCTAGGCGAG GCGCGACCGG AGGACGCCGG
1081 CGCCCCGTTC GAGCAGGCCG GTCAGAAAAG TACTGGGGCC CCGCCAGCCC GACCCACAAG
1141 CTACCCCCAA GCTTCCCGGG GAACAAAGAC GAGCTCATGC AGCACATGGA CGAGGTCAAC
1201 GACGAGCTCA TCCGGAAAAT CAGCAACATC AGGGCGCAGC CACAGAGGCA CTTCCGCGTG
1261 GAGCGCAGCC AGCCCGTGAG CCAGCCGCTC ACCTACGAGT CGGGTCCGGA CGAGGTCCGC
1321 GCCTGGCTGG AAGCCAAGGC CTTCAGCCCG CGGATCGTGG AGAACCTGGG CATCCTGACC
1381 GGGCCGCAGC TCTTCTCCCT CAACAAGGAG GAGCTGAAGA AAGTGTGCGG CGAGGAGGGC
1441 GTCCGCGTGT ACAGCCAGCT CACCATGCAG AAGGCCTTCC TGGAGAAGCA GCAAAGTGGG
1501 TCGGAGCTGG AAGAACTCAT GAACAAGTTT CATTCCATGA ATCAGAGGAG GGGGGAGGAC
1561 AGCTAGGCCC AGCTGCCTTG GGCTGGGGCC TGCGGAGGGG AAGCCCACCC ACAATGCATG
1621 GAGTATTATT TTTAAAAAAA AAAAAAAAAA AA
B:氨基酸序列(SEQ ID NO:5) 长度:232
1 MSLWESLGES WMRPRSEWPR EPQVPLYVPK FHSGWEPPVD VLQEAPWEVE GLASAPIEEV
61 SPVSRQSIRN SQKHSPTSEP TPPGDALPPV SSPHTHRGYQ PTPAAAKYVK ILYDFTARNA
121 NELSVLKDEV LEVLEDGRQW WKLRSRSGQA GYVPCNILGE ARPEDAGAPF EQAGQKSTGA
181 PPARPTSYPQ ASRGTKTSSC STWTRSTTSS SGKSATSGRS HRGTSAWSAA SP
C.核苷酸及氨基酸组合序列(SEQ ID NO:6) 克隆号:PP13181
起始编码子:581 ATG 终止编码子:1277 TGA 蛋白质分子量:25362.69
1 G CGG CCC CCC TCT GAG GGC GAG TTC ATC GAC TGC TTC CAG AAA ATC 46
47 AAG CTG GCG ATT AAC TTG CTG GTG GGT CCG GTG GCC CCA GCC CTG CCC 94
95 CAC TGT CTG TGC TGA GGG GAG GGT GGA GGC CCC GCC CCG CCC CGG CAC 142
143 CTG CTC ACT TGT TCC CAC CCC CAG GCA AAG CTG CAG AAG CAC ATC CAG 190
191 AAC CCC AGC GCC GCG GAG CTC GTG CAC TTC CTC TTC GGG CCT CTG GAC 238
239 CTG GTG CCT GGG GCC GGG CGG CAG GGG CGC GCA GGG TGG GGG CCC AGA 286
287 GGC CTC TGC AGC ATC TCC CCG GGG TCG GGG TTG GGG CAG CAG GTG CCC 334
335 GCC TTG GGC AGC CCG GTT CAC GCT GTG TGG CCA CTC TCC TGG GGT CCA 382
383 AAG TCC CTT CCC GAG GGC CAG CCT GTG GAG CTA CGG GGG TGC TGG GCC 430
431 AGG GTC TGT GGG CCT CAG TCC CCT CTG AAC CTC ACT GTG CCC CAG ATC 478
479 GTC AAC ACC TGC AGT GGC CCA GAC ATC GCA CGC TCC GTC TCC TGC CCA 526
527 CTG CTC TCC CGA GAT GCC GTG GAC TTC CTG CGC GGC CAC CTG GTC CCT 574
575 AAG GAG ATG TCG CTG TGG GAG TCA CTG GGA GAG AGC TGG ATG CGG CCC 622
1 Met Ser Leu Trp Glu Ser Leu Gly Glu Ser Trp Met Arg Pro 14
623 CGT TCC GAG TGG CCG CGG GAG CCA CAG GTG CCC CTC TAC GTG CCC AAG 670
15 Arg Ser Glu Trp Pro Arg Glu Pro Gln Val Pro Leu Tyr Val Pro Lys 30
671 TTC CAC AGC GGC TGG GAG CCT CCT GTG GAT GTG CTG CAG GAG GCC CCC 718
31 Phe His Ser Gly Trp Glu Pro Pro Val Asp Val Leu Gln Glu Ala Pro 46
719 TGG GAG GTG GAG GGG CTG GCG TCT GCC CCC ATC GAG GAG GTG AGT CCA 766
47 Trp Glu Val Glu Gly Leu Ala Ser Ala Pro Ile Glu Glu Val Ser Pro 62
767 GTG AGC CGA CAG TCC ATA AGA AAC TCC CAG AAG CAC AGC CCC ACT TCA 814
63 Val Ser Arg Gln Ser Ile Arg Asn Ser Gln Lys His Ser Pro Thr Ser 78
815 GAG CCC ACC CCC CCG GGG GAT GCC CTA CCA CCA GTC AGC TCC CCA CAT 862
79 Glu Pro Thr Pro Pro Gly Asp Ala Leu Pro Pro Val Ser Ser Pro His 94
863 ACT CAC AGG GCC TAC CAG CCA ACA CCA GCC ATG GCC AAG TAC GTC AAG 910
95 Thr His Arg Gly Tyr Gln Pro Thr Pro Ala Met Ala Lys Tyr Val Lys 110
911 ATC CTG TAT GAC TTC ACA GCC CGA AAT GCC AAC GAG CTA TCG GTG CTC 958
111 Ile Leu Tyr Asp Phe Thr Ala Arg Asn Ala Asn Glu Leu Ser Val Leu 126
959 AAG GAT GAG GTC CTA GAG GTG CTG GAG GAC GGC CGG CAG TGG TGG AAG 1006
127 Lys Asp Glu Val Leu Glu Val Leu Glu Asp Gly Arg Gln Trp Trp Lys 142
1007 CTG CGC AGC CGC AGC GGC CAG GCG GGG TAC GTG CCC TGC AAC ATC CTA 1054
143 Leu Arg Ser Arg Ser Gly Gln Ala Gly Tyr Val Pro Cys Asn Ile Leu 158
1055 GGC GAG GCG CGA CCG GAG GAC GCC GGC GCC CCG TTC GAG CAG GCC GGT 1102
159 Gly Glu Ala Arg Pro Glu Asp Ala Gly Ala Pro Phe Glu Gln Ala Gly 174
1103 CAG AAA AGT ACT GGG GCC CCG CCA GCC CGA CCC ACA AGC TAC CCC CAA 1150
175 Gln Lys Ser Thr Gly Ala Pro Pro Ala Arg Pro Thr Ser Tyr Pro Gln 190
1151 GCT TCC CGG GGA ACA AAG ACG AGC TCA TGC AGC ACA TGG ACG AGG TCA 1198
191 Ala Ser Arg Gly Thr Lys Thr Ser Ser Cys Ser Thr Trp Thr Arg Ser 206
1199 ACG ACG AGC TCA TCC GGA AAA TCA GCA ACA TCA GGG CGC AGC CAC AGA 1246
207 Thr Thr Ser Ser Ser Gly Lys Ser Ala Thr Ser Gly Arg Ser His Arg 222
1247 GGC ACT TCC GCG TGG AGC GCA GCC AGC CCG TGA GCC AGC CGC TCA CCT 1294
223 Gly Thr Ser Ala Trp Ser Ala Ala Ser Pro *** 233
1295 ACG AGT CGG GTC CGG ACG AGG TCC GCG CCT GGC TGG AAG CCA AGG CCT 1342
1343 TCA GCC CGC GGA TCG TGG AGA ACC TGG GCA TCC TGA CCG GGC CGC AGC 1390
1391 TCT TCT CCC TCA ACA AGG AGG AGC TGA AGA AAG TGT GCG GCG AGG AGG 1438
1439 GCG TCC GCG TGT ACA GCC AGC TCA CCA TGC AGA AGG CCT TCC TGG AGA 1486
1487 AGC AGC AAA GTG GGT CGG AGC TGG AAG AAC TCA TGA ACA AGT TTC ATT 1534
1535 CCA TGA ATC AGA GGA GGG GGG AGG ACA GCT AGG CCC AGC TGC CTT GGG 1582
1583 CTG GGG CCT GCG GAG GGG AAG CCC ACC CAC AAT GCA TGG AGT ATT ATT 1630
1631 TTT AAA AAA AAA AAA AAA AAA A 1652
3.PP13191
A:核苷酸序列(SEQ ID NO:7)长度:1489
1 GCCTGTCCAC ACCTCTGCCC CAGAGCTGCC TCCTGCCTGG CACTGCCGCC ACACTCCCCT
61 CCTGGGATGG GGCTTCTGCT CCCGGGCTCA CTCAAGGAGA CTGCGGCATG TTGACCACAC
121 CAGACTGGGT TTCAGGGAAT GGGCATGCCA GGTGCCAAGG AGCCAAACAG ATGGCTTTCC
181 AGGCAGCAAG GTCCTTGGGG CCTTCTTGGA GGAGCTTGGG TGACAGCCAG GTGAGCACCC
241 AGACCCCAGA CCCTCATGTG CTGTGTGCCT GGCCCCTTCT GTACTGGCCA TTTGTGGCCA
301 GGGCCAAGCC TGTGACTCAA CTCCAGGGGC AAGATGGGGA GTGAGCTGAT GGCTCCGAGA
361 CTGGTCAGGA GCCCAGGCCA GTGAGATGGG GCCTGGAGCC TTGTCTGTGT CACATTAGGT
421 ACCATGGGAG CTGCTGAGAC CTGACATTTT GTCCCCTGCC TACATGGCTT GGCCCATGGA
481 GAAGGAGCAG TGAATGGGAT CGTCGGGGAA GCCCCTCTTC CTGCTCTGCT CCCCTGGAAA
541 CTGTTGCAAA ACTCCCACCG CCTCATGGCA AATGCCCAAA GCATGTTCCG CACCCAGGCG
601 GGGGCCCCTG CTAATGAGAA CCTTGGTGCA GCTGCAGCCA GGAGGGGAGC GGGCCCAGGA
661 GCCAGGCTCA GGTCCAGCTG GTTCCTCTCT GGCGCCTTCT GAACCCGTCT CAGCAGGTCC
721 ACAGCACCTG GGCAGAGGTC AGAGACCAGG GGAGGCCGGG CCTTGCCCTC CCTTCTGCCC
781 AGGGCCCAGT GTTCTTGATA GAAGACCCTT CTGGGGAGCC AGGGAGCTCA GGGGACAGAT
841 AAGGGAAGGA CGCCCCCTGA CTCCAGGCCC CTGAGCCTGG CGGGAAGTGG CTGCGGCCCA
901 GGCAGCCAGT CCTGGTGGTG TTCTCCCTGC ATGCCCTCCG TGGCTGGGCT GCCACCCCAC
961 CCGGCCCGAA TCTGTCTTGA CCTGCAGGAA TACACGGGCG GCGCCAGGCA TTACCTCACA
1021 GCGGGACTAC ACAGTTGCTG GCTTTGCTCC TGGGCAAGGA GGAGCAGGCC AGAGCCTCTT
1081 TTGCTTCCTT TTCTTGCCCA TGCCGCTTCT AGAAGCCAGG CACAGGTTGC CAAGAGGTGA
1141 CACGAAACAG GAGGAAACTC AGTGACCTCT GCCTCTCCCA CATTCCTCCC CGCGGGGGAG
1201 GACCTCGCCG CTCTGAAGAG CACCGTGCAC ATGTGGGTGC ACAAACGTGG GTGTTGGTGT
1261 GGACGGGGCG CAGATCTCCG TGGATGAACT GCGTCTGGAC TCTTAGATTC ATAAAATATT
1321 CGAGGGTTTG GGAGTCACAG ACCCTCCCCT CTCCTCAGTG CACTTTAGCA TTTGCACGGT
1381 GTCTTCCCCG GACAGCACAG CAATAAATGG TGTGATTGCG TGGAAAAAAA AAAAAAAAAA
1441 AAAAAAAAAA TAAAAAAAAA AAAAAAAAAA AAAAAAAAAA AAAAAAAAA
B:氨基酸序列(SEQ ID NO:8) 长度:126
1 MGSSGKPLFL LCSPGNCCKT PTASWQMPKA CSAPRRGPLL MRTLVQLQPG GERAQEPGSG
61 PAGSSLAPSE PVSAGPQHLG RGQRPGEAGP CPPFCPGPSV LDRRPFWGAR ELRGQIREGR
121 PLTPGP
C.核苷酸及氨基酸组合序列(SEQ ID NO:9) 克隆号:PP13191
起始编码子:494 ATG 终止编码子:872 TGA 蛋白质分子量:13131.41
1 G CCT GTC CAC ACC TCT GCC CCA GAG CTG CCT CCT GCC TGG CAC TGC 46
47 CGC CAC ACT CCC CTC CTG GGA TGG GGC TTC TGC TCC CGG GCT CAC TCA 94
95 AGG AGA CTG CGG CAT GTT GAC CAC ACC AGA CTG GGT TTC AGG GAA TGG 142
143 GCA TGC CAG GTG CCA AGG AGC CAA ACA GAT GGC TTT CCA GGC AGC AAG 190
191 GTC CTT GGG GCC TTC TTG GAG GAG CTT GGG TGA CAG CCA GGT GAG CAC 238
239 CCA GAC CCC AGA CCC TCA TGT GCT GTG TGC CTG GCC CCT TCT GTA CTG 286
287 GCC ATT TGT GGC CAG GGC CAA GCC TGT GAG TCA ACT CCA GGG GCA AGA 334
335 TGG GGA GTG AGC TGA TGG CTC CGA GAC TGG TCA GGA GCC CAG GCC AGT 382
383 GAG ATG GGG CCT GGA GCC TTG TCT GTG TCA CAT TAG GTA CCA TGG GAG 430
431 CTG CTG AGA CCT GAC ATT TTG TCC CCC GCC TAC ATG GCT TGG CCC ATG 478
479 GAG AAG GAG CAG TGA ATG GGA TCG TCG GGG AAG CCC CTC TTC CTG CTC 526
1 Met Gly Ser Ser Gly Lys Pro Leu Phe Leu Leu 11
527 TGC TCC CCT GGA AAC TGT TGC AAA ACT CCC ACC GCC TCA TGG CAA ATG 674
12 Cys Ser Pro Gly Asn Cys Cys Lys Thr Pro Thr Ala Ser Trp Gln Met 27
575 CCC AAA GCA TGT TCC GCA CCC AGG CGG GGG CCC CTG CTA ATG AGA ACC 622
28 Pro Lys Ala Cys Ser Ala Pro Arg Arg Gly Pro Leu Leu Met Arg Thr 43
623 TTG GTG CAG CTG CAG CCA GGA GGG GAG CGG GCC CAG GAG CCA GGC TCA 670
44 Leu Val Gln Leu Gln Pro Gly Gly Glu Arg Ala Gln Glu Pro Gly Ser 59
671 GGT CCA GCT GGT TCC TCT CTG GCG CCT TCT GAA CCC GTC TCA GCA GGT 718
60 Gly Pro Ala Gly Ser Ser Leu Ala Pro Ser Glu Pro Val Ser Ala Gly 75
719 CCA CAG CAC CTG GGC AGA GGT CAG AGA CCA GGG GAG GCC GGG CCT TGC 766
76 Pro Gln His Leu Gly Arg Gly Gln Arg Pro Gly Glu Ala Gly Pro Cys 91
767 CCT CCC TTC TGC CCA GGG CCC AGT GTT CTT GAT AGA AGA CCC TTC TGG 814
92 Pro Pro Phe Cys Pro Gly Pro Ser Val Leu Asp Arg Arg Pro Phe Trp 107
815 GGA GCC AGG GAG CTC AGG GGA CAG ATA AGG GAA GGA CGC CCC CTG ACT 862
108 Gly Ala Arg Glu Leu Arg Gly Gln Ile Arg Glu Gly Arg Pro Leu Thr 123
863 CCA GGC CCC TGA GCC TGG CGG GAA GTG GCT GCG GCC CAG GCA GCC AGT 910
124 Pro Gly Pro *** 127
911 CCT GGT GGT GTT CTC CCT GCA TGC CCT CCG TGG CTG GGC TGC CAC CCC 958
959 ACC CGG CCC GAA TCT GTC TTG ACC TGC AGG AAT ACA CGG GCG GCG CCA 1006
1007 GGC ATT ACC TCA CAG CGG GAC TAC ACA GTT GCT GGC TTT GCT CCT GGG 1054
1055 CAA GGA GGA GCA GGC CAG AGC CTC TTT TGC TTC CTT TTC TTG CCC ATG 1102
1103 CCG CTT CTA GAA GCC AGG CAC AGG TTG CCA AGA GGT GAC ACG AAA CAG 1150
1151 GAG GAA ACT CAG TGA CCT CTG CCT CTC CCA CAT TCC TCC CCG CGG GGG 1198
1199 AGG ACC TCG CCG CTC TGA AGA GCA CCG TGC ACA TGT GGG TGC ACA AAC 1246
1247 GTG GGT GTT GGT GTG GAC GGG GCG CAG ATC TCC GTG GAT GAA CTG CGT 1294
1295 CTG GAC TCT TAG ATT CAT AAA ATA TTC GAG GGT TTG GGA GTC ACA GAC 1342
1343 CCT CCC CTC TCC TCA GTG CAC TTT AGC ATT TGC ACG GTG TCT TCC CCG 1390
1391 GAC AGC ACA GCA ATA AAT GGT GTG ATT GCG TGG AAA AAA AAA AAA AAA 1438
1439 AAA AAA AAA AAA TAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA 1486
1487 AAA 1489
4.PP13439
A:核苷酸序列(SEQ ID NO:10) 长度:2935
1 GCCAGCCCGC AGAGCACGGT CCTGGGGGTG TGATCACAGC TGCTGACCGC GGCTCCCGCC
61 GTGTGATGTG ACCACTCGGA GGTGGGGCGA GGGGGCCGGG GGCCAGCAGC AGGGAGTGTG
121 TAAAAGGTCT TCTTCCTCAC AATACGATTG AAGGTGTTTG GGACGTAGGG CTCAGATCCA
181 GGAGTGCCGT CCTCCAAGCC TTCCATAGGA TTTTGTGACG CCCTTGAGAA CCGCCCTTGT
241 AGACTCAGTC CTGAACTACC GCCACGCCTG TATTTGCTCC TCCTCATCTC GGATTATTGC
301 TTTCTTTACT TCCTCTGTGA AGCACTAAAT ACTAGTAGCT TTTTAAGAAC AGACCACAAC
361 AGTTTCTTCG TTTGACCATT ACTAAGTCTG CCTCCAAAAT GGGGAGTTCC GTTAACTTTA
421 TCAGCCAGCC CTGCCAATTT TAGTTAAGAT AAGCCTTCGT GGGCCTCCAA AGGCCTCTGA
481 GAGACTAGGT ACCCAGAGAC CTCTGAGGTA TGTTAAATGG TTTAGTTCGT GGTGGTGCAA
541 TGAAAACTTA AAGAAGGAAA GGAATCTGCA ATTTGAACTG TTAACAGACC CCACATTTAC
601 ACACTGCCTA TTTGAATAAG AATCTCTTTC AGTATTAAAT GTTTTGTGAC ACACAAATGA
661 ATTTGTACAC AGTCTTTTAG CTTGAATTTG TAGACAGCCT TTAAATGTTG TTTGGGGATC
721 TTTGGACTAA TTGTATAAGC GCTCAGTCTG GTCTCAAACT CATGATCTCA GGTTATCCAT
781 GCGCCTCGGC CTCCGAAAAT GCTGCGATTA TAGGCGTGAG CCACCACTCC CAGCCCCTTC
841 TTGTGATGAT GTGAGATGAT AAAATGCCTG TGTGAAGATG AAGTGAGGCG AATAACCTAG
901 GCAGTGTGAC ATAAGTGGAA AACGCCAGAA ATAAACAATT CATGTTATAA ATTGTGCACC
961 ATTCTGAGTA GTGTGATGAA ATCTTGCACT CCTCCATTCC ATCCTGCCAG CATTGCCAGT
1021 CACTCGGTGG CCATCTCTGT GATCAGATTA ACAGTGGTTT TATCAGTATC ATAGGGCTTA
1081 TGTTCAAGTA ACTGTATTTT ACTTAATAAT GGCTACAAAG TGCAAAAGTA GCGATATAAT
1141 ATTTTTGGAC CATGGTTGAC TGCAGGTAAC TGAAATTTTA GTTACATGAA ACCACAGATA
1201 AGGGGGGACT GCACTACACA TAAAATAGGC ATAAGTAACC TCAAGACAAT AGACATTATT
1261 AAAATATAGA AAAGTAATAA AGAAGTGATA AATGGAGAAC TTTTGCTTTA ATATAATTTA
1321 TTGTAAGTTT ATATAATTTA ATTTTTAGTA ACTTCAATTT TTAATATTTT AATTTTTAAT
1381 GATTTATGTG TTTAACAACT GACTCACAAA ATTGCTGAAA ATTTCCCAAT CTTGATGAAA
1441 ATTTCCCAGT CTGCTCTCAG GAGCCTATAT GAGCCAGCCC AGAATACCAC TGGAGTTATA
1501 ATTTTAGAAG ATTACCTTGG TAGCAATCTG TAGAAATGTT TTGTAGGGGA GCTACAGTAG
1561 GGATCAAAAG TGGAGACAGA TTACGAAATG TCTTCAGACC CTGCCAGCAT GAGGGTTCAA
1621 ATGATGTCTA GTCAGTGGAG ATCACGCAAA TTTCACTGGT TTGTGTTCCC ACGTGGATCC
1681 GTAGGACCTT CAATGCAAGT CTCCTTCAGG TATTCCTGGT GGTATCTTGG CATTGTCTCA
1741 CATGTGTTTT GAGAGCCCCA CTTCCTTGGG GAGTCTTCCG CTTTGGCTTC CTTGTTGATG
1801 ACCGTGGGCC AAAGGGCTTT GTGTGTGAGG CTCCACCTCA GCCATAGCTG GTGTCCTCCT
1861 GAAGAGGATT TATGCCAAAA GGAGTCACAT GCCAGGACTT GTTTTTTTCA AGTTTGCAAA
1921 TCTGGGGGGA GGGTGGGGCT CTAGAAAGAT GCTGGAACCT GGTATGTGCC CTGCAGAAAT
1981 GTCCTTGACT CCTTTTGTCA CCAGGAGCCT AAATTTACTG TGTTTCTCTG TGACTCACAG
2041 AGGCAGGGGG AAAGACAATC CTCTCCACTT CTTCCTGCAG CTACTTGCTC CATCAGCCTC
2101 AGCCAGTCTG GGGCTGGGCA GGGAAACAGG CCCTCTTCCT TCTCCTGGTT TGTCCCCTGG
2161 TTTGCTCAGC GAGACCTTCG TGTCTGAGGG CCCCCTTCTC ACCTTGCAGA TCAGCTGGTG
2221 TGGTACTGAA TGCCTGATGG ATGGCCAGGG GCCCCTGATC CATCTGCATT CCCAGAATAT
2281 AAGAATCTCA TTTGGGGCCA CCCAGCCCCC ACTACTCTTA GTATATGAAA TAAGGTGGAA
2341 TAGATAAGAG GATGGAACTG GCCCCTGGGT GCATGATGGG GCCAATAACG GAGGTACAGG
2401 GTGCATATGT GGCTGGGCTG TGAGACAGGG AAAGAGGATG GAATTGAGAT GCAAGGTAGA
2461 AAAGCATGAC TATGTTCAGT TTGGGACTTG ATGAAATAGA TACTGGTAGG ACAGTTCACT
2521 AGGCACTGGG CCTTCAGGAG AAAGAGAAGA AAGGAATGGT GGCATTGGGA GTAGTAAAGC
2581 TGTGGGTATG AATGAGTATC TATGAGACTC ACCTCCCTGT GAGGACATAG AATGAGGAGA
2641 CACAGTCTAC AGGTAGACTA GGTCGAGTTC AGAAGCCTGA AGAACACCAG TGTTTAAGGG
2701 ATGGTGTTGG GAAAGGGAGC CAGGCCAGCC AGAGAGGAAT GTTCCGGAAC TCTGAAAGGA
2761 AGGAAATGGC AGGAACAAAG GAGCTGGGGC ACAACAGTGG TGACTCACAC TGGGAACACT
2821 TCGGCCCATT GTGTTTATGC TTATAGTTTC CCAAGTTTAT CTTTTCAGGG TTATGATTAC
2881 GTTAACCTCA CCTCCCTCCC TCCCTCTAAA ACAAAACAAA AAAAAAAAAA AAAAA
B:氨基酸序列(SEQ ID NO:11) 长度:152
1 MPGLVFFKFA NLGGGWGSRK MLEPGMCPAE MSLTPFVTRS LNLLCFSVTH RGRGKDNPLH
61 FFLQLLAPSA SASLGLGRET GPLPSPGLSP GLLSETFVSE GPLLTLQISW CGTECLMDGQ
121 GPLIHLHSQN IRISFGATQP PLLLVYEIRW NR
C.核苷酸及氨基酸组合序列(SEQ ID NO:12) 克隆号:PP13439
起始编码子:1889 ATG 终止编码子:2345 TAA 蛋白质分子量:16504.36
1 G CCA GCC CGC AGA GCA CGG TCC TGG GGG TGT GAT CAC AGC TGC TGA 46
47 CCG CGG CTC CCG CCG TGT GAT GTG ACC ACT CGG AGG TGG GGC GAG GGG 94
95 GCC GGG GGC CAG CAG CAG GGA GTG TGT AAA AGG TCT TCT TCC TCA CAA 142
143 TAC GAT TGA AGG TGT TTG GGA CGT AGG GCT CAG ATC CAG GAG TGC CGT 190
191 CCT CCA AGC CTT CCA TAG GAT TTT GTG ACG CCC TTG AGA ACC GCC CTT 238
239 GTA GAC TCA GTC CTG AAC TAC CGC CAC GCC TGT ATT TGC TCC TCC TCA 286
287 TCT CGG ATT ATT GCT TTC TTT ACT TCC TCT GTG AAG CAC TAA ATA CTA 334
335 GTA GCT TTT TAA GAA CAG ACC ACA ACA GTT TCT TCG TTT GAC CAT TAC 382
383 TAA GTC TGC CTC CAA AAT GGG GAG TTC CGT TAA CTT TAT CAG CCA GCC 430
431 CTG CCA ATT TTA GTT AAG ATA AGC CTT CGT GGG CCT CCA AAG GCC TCT 478
479 GAG AGA CTA GGT ACC CAG AGA CCT CTG AGG TAT GTT AAA TGG TTT AGT 526
527 TCG TGG TGG TGC AAT GAA AAC TTA AAG AAG GAA AGG AAT CTG CAA TTT 574
575 GAA CTG TTA ACA GAC CCC ACA TTT ACA CAC TGC CTA TTT GAA TAA GAA 622
623 TCT CTT TCA GTA TTA AAT GTT TTG TGA CAC ACA AAT GAA TTT GTA CAC 670
671 AGT CTT GCA GCT TGA ATT TGT AGA CAG CCT TTA AAT GTT GTT TGG GGA 718
719 TCT TTG GAC TAA TTG TAT AAG CGC TCA GTC TGG TCT CAA ACT CAT GAT 766
767 CTC AGG TTA TCC ATG CGC CTC GGC CTC CGA AAA TGC TGC GAT TAT AGG 814
815 CGT GAG CCA CCA CTC CCA GCC CCT TCT TGT GAT GAT GTG AGA TGA TAA 862
863 AAT GCC TGT GTG AAG ATG AAG TGA GGC GAA TAA CCT AGG CAG TGT GAC 910
911 ATA AGT GGA AAA CGC CAG AAA TAA ACA ATT CAT GTT ATA AAT TGT GCA 958
959 CCA TTC TGA GTA GTG TGA TGA AAT CTT GCA CTC CTC CAT TCC ATC CTG 1006
1007 CCA GCA TTG CCA GTC ACT CGG TGG CCA TCT CTG TGA TCA GAT TAA CAG 1054
1055 TGG TTT TAT CAG TAT CAT AGG GCT TAT GTT CAA GTA ACT GTA TTT TAC 1102
1103 TTA ATA ATG GCT ACA AAG TGC AAA AGT AGC GAT ATA ATA TTT TTG GAC 1150
1151 CAT GGT TGA CTG CAG GTA ACT GAA ATT TTA GTT ACA TGA AAC CAC AGA 1198
1199 TAA GGG GGG ACT GCA CTA CAC ATA AAA TAG GCA TAA GTA ACC TCA AGA 1246
1247 CAA TAG ACA TTA TTA AAA TAT AGA AAA GTA ATA AAG AAG TGA TAA ATG 1294
1295 GAG AAC TTT TGC TTT AAT ATA ATT TAT TGT AAG TTT ATA TAA TTT AAT 1342
1343 TTT TAG TAA CTT CAA TTT TTA ATA TTT TAA TTT TTA ATG ATT TAT GTG 1390
1391 TTT AAC AAC TGA CTC ACA AAA TTG CTG AAA ATT TCC CAA TCT TGA TGA 1438
1439 AAA TTT CCC AGT CTG CTC TCA GGA GCC TAT ATG AGC CAG CCC AGA ATA 1486
1487 CCA CTG GAG TTA TAA TTT TAG AAG ATT ACC TTG GTA GCA ATC TGT AGA 1534
1535 AAT GTT TTG TAG GGG AGC TAC AGT AGG GAT CAA AAG TGG AGA CAG ATT 1582
1583 ACG AAA TGT CTT CAG ACC CTG CCA GCA TGA GGG TTC AAA TGA TGT CTA 1630
1631 GTC AGT GGA GAT CAC GCA AAT TTC ACT GGT TTG TGT TCC CAC GTG GAT 1678
1679 CCG TAG GAC CTT CAA TGC AAG TCT CCT TCA GGT ATT CCT GGT GGT ATC 1726
1727 TTG GCA TTG TCT CAC ATG TGT TTT GAG AGC CCC ACT TCC TTG GGG AGT 1774
1775 CTT CCG CTT TGG CTT CCT TGT TGA TGA CCG TGG GCC AAA GGG CTT TGT 1822
1823 GTG TGA GGC TCC ACC TCA GCC ATA GCT GGT GTC CTC CTG AAG AGG ATT 1870
1871 TAT GCC AAA AGG AGT CAC ATG CCA GGA CTT GTT TTT TTC AAG TTT GCA 1918
1 Met Pro Gly Leu Val Phe Phe Lys Phe Ala 10
1919 AAT CTG GGG GGA GGG TGG GGC TCT AGA AAG ATG CTG GAA CCT GGT ATG 1966
11 Asn Leu Gly Gly Gly Trp Gly Ser Arg Lys Met Leu Glu Pro Gly Met 26
1967 TGC CCT GCA GAA ATG TCC TTG ACT CCT TTT GTC ACC AGG AGC CTA AAT 2014
27 Cys Pro Ala Glu Met Ser Leu Thr Pro Phe Val Thr Arg Ser Leu Asn 42
2015 TTA CTG TGT TTC TCT GTG ACT CAC AGA GGC AGG GGG AAA GAC AAT CCT 2062
43 Leu Leu Cys Phe Ser Val Thr His Arg Gly Arg Gly Lys Asp Asn Pro 58
2063 CTC CAC TTC TTC CTG CAG CTA CTT GCT CCA TCA GCC TCA GCC AGT CTG 2110
59 Leu His Phe Phe Leu Gln Leu Leu Ala Pro Ser Ala Ser Ala Ser Leu 74
2111 GGG CTG GGC AGG GAA ACA GGC CCT CTT CCT TCT CCT GGT TTG TCC CCT 2158
75 Gly Leu Gly Arg Glu Thr Gly Pro Leu Pro Ser Pro Gly Leu Ser Pro 90
2159 GGT TTG CTC AGC GAG ACC TTC GTG TCT GAG GGC CCC CTT CTC ACC TTG 2206
91 Gly Leu Leu Ser Glu Thr Phe Val Ser Glu Gly Pro Leu Leu Thr Leu 106
2207 CAG ATC AGC TGG TGT GGT ACT GAA TGC CTG ATG GAT GGC CAG GGG CCC 2254
107 Gln Ile Ser Trp Cys Gly Thr Glu Cys Leu Met Asp Gly Gln Gly Pro 122
2255 CTG ATC CAT CTG CAT TCC CAG AAT ATA AGA ATC TCA TTT GGG GCC ACC 2302
123 Leu Ile His Leu His Ser Gln Asn Ile Arg Ile Ser Phe Gly Ala Thr 138
2303 CAG CCC CCA CTA CTC TTA GTA TAT GAA ATA AGG TGG AAT AGA TAA GAG 2350
139 Gln Pro Pro Leu Leu Leu Val Tyr Glu Ile Arg Trp Asn Arg *** 153
2351 GAT GGA ACT GGC CCC TGG GTG CAT GAT GGG GCC AAT AAC GGA GGT ACA 2398
2399 GGG TGC ATA TGT GGC TGG GCT GTG AGA CAG GGA AAG AGG ATG GAA TTG 2446
2447 AGA TGC AAG GTA GAA AAG CAT GAC TAT GTT CAG TTT GGG ACT TGA TGA 2494
2495 AAT AGA TAC TGG TAG GAC AGT TCA CTA GGC ACT GGG CCT TCA GGA GAA 2542
2543 AGA GAA GAA AGG AAT GGT GGC ATT GGG AGT AGT AAA GCT GTG GGT ATG 2590
2591 AAT GAG TAT CTA TGA GAC TCA CCT CCC TGT GAG GAC ATA GAA TGA GGA 2638
2639 GAC ACA GTC TAC AGG TAG ACT AGG TCG AGT TCA GAA GCC TGA AGA ACA 2686
2687 CCA GTG TTT AAG GGA TGG TGT TGG GAA AGG GAG CCA GGC CAG CCA GAG 2734
2735 AGG AAT GTT CCG GAA CTC TGA AAG GAA GGA AAT GGC AGG AAC AAA GGA 2782
2783 GCT GGG GCA CAA CAG TGG TGA CTC ACA CTG GGA ACA CTT CGG CCC ATT 2830
2831 GTG TTT ATG CTT ATA GTT TCC CAA GTT TAT CTT TTC AGG GTT ATG ATT 2878
2879 ACG TTA ACC TCA CCT CCC TCC CTC CCT CTA AAA CAA AAC AAA AAA AAA 2926
2927 AAA AAA AAA 2935
5.PP13479
A:核苷酸序列(SEQ ID NO:13)长度:2323
1 GGCCGGATTC CCAGTGGTGG CGAGGAGGTG GTATTTTTTT AAGTGTCAGT GTGGCATTGT
61 GGTTGCCTCA ATAAAACAGA GTCTTTTTTT GTCTTTTTTT TTTGAGACAG GGTCTCGCTC
121 TCACCCAGGC TGGAGTGCAG TGACACAGTT GCAGCTCACT GCAGCCTCGA CCTCCTGGGC
181 TCAAGCAATC CTCCCGCCTC AGCCTCCCGA GTAGCTGAGT CTACAGATCT CCACAATGGA
241 AGTTCGAAGC AAGCAAAAGC CACGCAAACC ACAGGCCGAT CTGTCTGAGC CCTAGGATTT
301 GCCCCGGTTC TGCTTCAGCC ACCAGCACCG TCTGCTCCTC CTCAGAATCC TTCCTCCCCC
361 GTGGCCCGCC CGCCGTGTCC CTCCTCCTCC ACGGGCCGCC CACCGTGTCC TTCCCTCCCC
421 CGTGGCCCAC CCACCATGTC CTTCCCTCCC CTGTGGCCCA CCCGCCATGT CCCTGCCTCC
481 CACCCGACAT GCCCCTTGAA GCTGCCTGGG CCCTGCTGTT GTCCCCACTG CCTGTGTGAC
541 TCTGCGCCCC CTTCCCTACC CTGCCCCACC CTGGTTCAGG GAGCGTCCAG GCCCATTCTC
601 ATCCTCAGGG CCTTCCCTGG CCCTTGCCAC TCTGTGCCGT GTCATGACCT GAAGCTGCAG
661 GTGGGCGCCT CCCCCTTCCG TCATGGCTGT CCCCCTTCTG TGAGGTGTCC CAGCCGCCTG
721 ATTGCCGGAG TCCCAGGGTG CTCGGTGCTG TCGTGGAGCC TGGGACATTC ACTGTCTGGG
781 ATTGATTCCA GGGTTGGAGC CACACCTGGT CTGGGGCATT CGCTGTCCTG GGTCAGAGCC
841 CCTCCTGGTC TGGGACATTC GCTGTCTGGG GTTGGAGCCA CACCTGGTCT GGGGCATTTG
901 CTGTCCGGGG TCGGAGCCTC ACCTGGTGAA GATACAGAAC ATGCTGCTGC CCTAACCCCG
961 TGTGGTGTGC CCCCTGTCCC CGGGTGTCGT TCCCATAGCC AGCCCTTGTC TCATCTCGTC
1021 TCATCCTCTA GATGCTGTGG GCCCTGAGGG AAAGGATCAC AGAGGCGCTG AGCCGGGATG
1081 GCTACGTGTA CAAGTACGAC CTCTCCCTCC CTGTGGAGCG GCTCTACGAC ATCGTGACTG
1141 ACCTGCGCGC CCGCCTCGGC CCGCACGCCA AGCACGTGGT GGGCTATGGC CACCTTGGAG
1201 ATGGTAACCT GCACCTCAAT GTGACGGCGG AGGCCTTCAG CCCCTCGCTC CTGGCTGCCC
1261 TGGAGGCCCA GGTGTACGAG TGGACGGCCG GGCAGCAGGG CAGCGTCAGC GCGGAGCACG
1321 GAGTGGGCTT CAGGAAGAGG GACGTCCTGG GCTACAGCAA GCCACCGGGG GCCCTGCAGC
1381 TCATGCAGCA GCTCAAGGCC CTGCTGGACC CCAAGGGCAT CCTCAACCCC TACAAGACGC
1441 TGCCCAGCCA GGCCTGACGG CCACTCCTGC TGCTGCCAAG GCCCACTGGG GGTCGGCGGG
1501 TGGCTCTCGG GCGGGGGTGT TGCGGTGGCT CTGAGGGATG AGCCGGCAGT GGGCAGGGGA
1561 CCAGGCACCT GGTTGAAGGG ACTGGGAGCC CGCACTGGGG AACTGCCGGA CGCAGGCCCT
1621 CGGGCAGGAG CATCTGGCAG AGTGGGGGGC GTGGCAGGCA CCCTCCTTTG CAGGGCGAGG
1681 TGGGGCCTCT GCAGCCATCC TGGACAGGCC GGGGTGGCGG CAGCTTTGCC CACGTGGAAG
1741 CGGGGTGGGT CTCACTTGCG TGGTGGCCCC TGGCCCCATC TTGCCTGCTG CGGCCTGGGG
1801 AGCAGGCGCT GGGTGGTGGT TCTGCCTGCT TGCTGCTCGT TCCCCGGGCA TGCGTGGGCA
1861 GCGGGGGGCA TGCGTGGGCA GCAGGGGGCG TGGGCAGCGG GGGCATGGGC AGGACCACGT
1921 GGGCCGTGAT CGTGGGTTGC CGAGAGGACC TGAGCGCTGC GGCTCTGCTG AATGGAGCCG
1981 GGTCCCTCAG GCCGTGGACG CCCTCGGGAG GGGGGTGACT GTGGCTTGTG TCTGGACAGG
2041 AATGTGTCAT TTCCCACATC TTCTAGAGGG CTGCCAGCTG GGAAGACAGT TATCAGGGCA
2101 AGCTGTGCTC TGAGTTTCGG GTTCTGCTCC TACAAAGAAC GTGCGGTGCT GCGGGCGAGG
2161 GCCCCGGCAC GGACAAGGGC CACTGCAGAG TGTGTTTCTG CTCGTCAGCT GCCCTGGGCA
2221 GCGGATGGGC TGGGCGATGC AGCTGGATGC ACATCTCATT CTGTCATGAA TGTCCAGTAA
2281 AAATCTGAAT TGGTTGCAAA AAAAAAAAAA AAAAAAAAAA AAA
B:氨基酸序列(SEQ ID NO:14) 长度:203
1 MSFPPLWPTR HVPASHPTCP LKLPGPCCCP HCLCDSAPPS LPCPTLVQGA SRPILILRAF
61 PGPCHSVPCH DLKLQVGASP FRHGCPPSVR CPSRLIAGVP GCSVLSWSLG HSLSGIDSRV
121 GATPGLGHSL SWVRAPPGLG HSLSGVGATP GLGHLLSGVG ASPGEDTEHA AALTPCGVPP
181 VPGCRSHSQP LSHLVSSSRC CGP
C.核苷酸及氨基酸组合序列(SEQ ID NO:15) 克隆号:PP13479
起始编码子:436 ATG 终止编码子:1045 TGA 蛋白质分子量:17771.81
1 GGC CGG ATT CCC AGT GGT GGC GAG GAG GTG GTA TTT TTT TAA GTG TCA 48
49 GTG TGG CAT TGT GGT TGC CTC AAT AAA ACA GAG TCT TTT TTT GTC TTT 96
97 TTT TTT TGA GAC AGG GTC TCG CTC TCA CCC AGG CTG GAG TGC AGT GAC 144
145 ACA GTT GCA GCT CAC TGC AGC CTC GAC CTC CTG GGC TCA AGC AAT CCT 192
193 CCC GCC TCA GCC TCC CGA GTA GCT GAG TCT ACA GAT CTC CAC AAT GGA 240
241 AGT TCG AAG CAA GCA AAA GCC ACG CAA ACC ACA GGC CGA TCT GTC TGA 288
289 GCC CTA GGA TTT GGC CCG GTT CTG CTT CAG CCA CCA GCA CCG TCT GCT 336
337 CCT CCT CAG AAT CCT TCC TCC CCC GTG GCC CGC CCG CCG TGT CCC TCC 384
385 TCC TCC ACG GGC CGC CCA CCG TGT CCT TCC CTC CCC CGT GGC CCA CCC 432
433 ACC ATG TCC TTC CCT CCC CTG TGG CCC ACC CGC CAT GTC CCT GCC TCC 480
1 Met Ser Phe Pro Pro Leu Trp Pro Thr Arg His Val Pro Ala Ser 15
481 CAC CCG ACA TGC CCC TTG AAG CTG CCT GGG CCC TGC TGT TGT CCC CAC 528
16 His Pro Thr Cys Pro Leu Lys Leu Pro Gly Pro Cys Cys Cys Pro His 31
529 TGC CTG TGT GAC TCT GCG CCC CCT TCC CTA CCC TGC CCC ACC CTG GTT 576
32 Cys Leu Cys Asp Ser Ala Pro Pro Ser Leu Pro Cys Pro Thr Leu Val 47
577 CAG GGA GCG TCC AGG CCC ATT CTC ATC CTC AGG GCC TTC CCT GGC CCT 624
48 Gln Gly Ala Ser Arg Pro Ile Leu Ile Leu Arg Ala Phe Pro Gly Pro 63
625 TGC CAC TCT GTG CCG TGT CAT GAC CTG AAG CTG CAG GTG GGC GCC TCC 672
64 Cys His Ser Val Pro Cys His Asp Leu Lys Leu Gln Val Gly Ala Ser 79
673 CCC TTC CGT CAT GGC TGT CCC CCT TCT GTG AGG TGT CCC AGC CGC CTG 720
80 Pro Phe Arg His Gly Cys Pro Pro Ser Val Arg Cys Pro Ser Arg Leu 95
721 ATT GCC GGA GTC CCA GGG TGC TCG GTG CTG TCG TGG AGC CTG GGA CAT 768
96 Ile Ala Gly Val Pro Gly Cys Ser Val Leu Ser Trp Ser Leu Gly His 111
769 TCA CTG TCT GGG ATT GAT TCC AGG GTT GGA GCC ACA CCT GGT CTG GGG 816
112 Ser Leu Ser Gly Ile Asp Ser Arg Val Gly Ala Thr Pro Gly Leu Gly 127
817 CAT TCG CTG TCC TGG GTC AGA GCC CCT CCT GGT CTG GGA CAT TCG CTG 864
128 His Ser Leu Ser Trp Val Arg Ala Pro Pro Gly Leu Gly His Ser Leu 143
865 TCT GGG GTT GGA GCC ACA CCT GGT CTG GGG CAT TTG CTG TCC GGG GTC 912
144 Ser Gly Val Gly Ala Thr Pro Gly Leu Gly His Leu Leu Ser Gly Val 159
913 GGA GCC TCA CCT GGT GAA GAT ACA GAA CAT GCT GCT GCC CTA ACC CCG 960
160 Gly Ala Ser Pro Gly Glu Asp Thr Glu His Ala Ala Ala Leu Thr Pro 175
961 TGT GGT GTG CCC CCT GTC CCC GGG TGT CGT TCC CAT AGC CAG CCC TTG 1008
176 Cys Gly Val Pro Pro Val Pro Gly Cys Arg Ser His Ser Gln Pro Leu 191
1009 TCT CAT CTC GTC TCA TCC TCT AGA TGC TGT GGG CCC TGA GGG AAA GGA 1056
192 Ser His Leu Val Ser Ser Ser Arg Cys Cys Gly Pro *** 204
1057 TCA CAG AGG CGC TGA GCC GGG ATG GCT ACG TGT ACA AGT ACG ACC TCT 1104
1105 CCC TCC CTG TGG AGC GGC TCT ACG ACA TCG TGA CTG ACC TGC GCG CCC 1152
1153 GCC TCG GCC CGC ACG CCA AGC ACG TGG TGG GCT ATG GCC ACC TTG GAG 1200
1201 ATG GTA ACC TGC ACC TCA ATG TGA CGG CGG AGG CCT TCA GCC CCT CGC 1248
1249 TCC TGG CTG CCC TGG AGC CCC ACG TGT ACG AGT GGA CGG CCG GGC AGC 1296
1297 AGG GCA GCG TCA GCG CGG AGC ACG GAG TGG GCT TCA GGA AGA GGG ACG 1344
1345 TCC TGG GCT ACA GCA AGC CAC CGG GGG CCC TGC AGC TCA TGC AGC AGC 1392
1393 TCA AGG CCC TGC TGG ACC CCA AGG GCA TCC TCA ACC CCT ACA AGA CGC 1440
1441 TGC CCA GCC AGG CCT GAC GGC CAC TCC TGC TGC TGC CAA GGC CCA CTG 1488
1489 GGG GTC GGC GGG TGG CTC TCG GGC GGG GGT GTT GCG GTG GCT CTG AGG 1536
1537 GAT GAG CCG GCA GTG GGC AGG GGA CCA GGC ACC TGG TTG AAG GGA CTG 1584
1585 GGA GCC CGC ACT GGG GAA CTG CCG GAC GCA GGC CCT CGG GCA GGA GCA 1632
1633 TCT GGC AGA GTG GGG GGC GTG GCA GGC ACC CTC CTT TGC AGG GCG AGG 1680
1681 TGG GGC CTC TGC AGC CAT CCT GGA CAG GCC GGG GTG GCG GCA GCT TTG 1728
1729 CCC ACG TGG AAG CGG GGT GGG TCT CAC TTG CGT GGT GGC CCC TGG CCC 1776
1777 CAT CTT GCC TGC TGC GGC CTG GGG AGC AGG CGC TGG GTG GTG GTT CTG 1824
1825 CCT GCT TGC TGC TCG TTC CCC GGG CAT GCG TGG GCA GCG GGG GGC ATG 1872
1873 CGT GGG CAG CAG GGG GCG TGG GCA GCG GGG GCA TGG GCA GGA CCA CGT 1920
1921 GGG CCG TGA TCG TGG GTT GCC GAG AGG ACC TGA GCG CTG CGG CTC TGC 1968
1969 TGA ATG GAG CCG GGT CCC TCA GGC CGT GGA CGC CCT CGG GAG GGG GGT 2016
2017 GAC TGT GGC TTG TGT CTG GAC AGG AAT GTG TCA TTT CCC ACA TCT TCT 2064
2065 AGA GGG CTG CCA GCT GGG AAG ACA GTT ATC AGG GCA AGC TGT GCT CTG 2112
2113 AGT TTC GGG TTC TGC TCC TAC AAA GAA CGT GCG GTG CTG CGG GCG AGG 2160
2161 GCC CCG GCA CGG ACA AGG GCC ACT GCA GAG TGT GTT TCT GCT CGT CAG 2208
2209 CTG CCC TGG GCA GCG GAT GGG CTG GGC GAT GCA GCT GGA TGC ACA TCT 2256
2257 CAT TCT GTC ATG AAT GTC CAG TAA AAA TCT GAA TTG GTT GCA AAA AAA 2304
2305 AAA AAA AAA AAA AAA AAA A 2323
6.PP13842
A:核苷酸序列(SEQ ID NO:16)长度:1259
1 GCTGCAGCCC CTGCCCCCGC CCCTCCTCGC TGGGTGCTCA GAAGGCTGAC AGCTGCGCCA
61 GGCTGAGGCG GCAGTCGATG CTGGAGTTGT CCGGGCCCGT GTAGGCCAGG CCCAGGGGCT
121 CTAGGAAGGC CCGGCAGGCC CCAGCGCTGC CTTTGCGGAT TCTGTTTTTG AGCCGTGGAC
181 TTGGGTTGTA AATTTATTTG TGGGGAGTGC GCTCCAGGAA GAGCCACCAT CCCTGCCCCC
241 GTTTTCCCAC CGGGGAGTCT GTACAGAGAT TTTTCTACGT TTTTATTTTT TGCCTCAGAG
301 GGATGGGATT GGGGAGGAGG GGATGGGCAG CGGAGGGTTG GGGGCATGGT CTGCAGGCTC
361 ATCTGTGTCC GCTTTCACTC CACTAATGCT GTCTCAGTGT TTTCTCTCTC TCTCTTTCGA
421 GCTTGCACTC CGGTACCCGA CCCGGCGCCC TGGCCCATCC CATGCCGGGG GGCCAGTGGA
481 AAGAAGACAG GCCGTCCAGC CCGTGCCCGC CTGCGGCGGG G(CACCCAGC AAGCCCGCCC
541 ACCGCCCGCT GCCTCACCTG CTTCGCCACA GACTCTTGTT CCCAGCCCCT TGGGGCCTCC
601 GTGTTTGGGG TGGGGGAGCT GCTTAGAGAC TGTGCCCGTC CTCGGCCCCC CACCCTGAAG
661 TGCCAGCACC ACCAGCACCA GATCCTCCGC CGCCACACCG CACTGAGGAC ACGCCGGCCG
721 GGCCGCCTCG TCTCAAGTTG TATAAAGTTG TCTCCGTGTC CCCTCCTCCC TCTGCCCCCA
781 GTGTTTCTTC TGATTTTTTT TTCCCCTTTC CCTCCCTCCC CCTCCGCATT CTTCCCTTGG
841 TTCAGCACAG GTAAAACGGT TCCCCTCCCT CCCTGCCTTC ATGGATCACC AGCTCACGTC
901 ATGTTGCCTT CTCTTTTCTT TGTGTGTGTG TTTATTTAAG TTATTTTTCT TCCTCCTCTC
961 CCTTTTCTTT TTGGCCCTCC CTCCCTCCCT CTTCTGCCAT GTAACTGGAG GATGTGCTAT
1021 GAGTTTGCAA ACAGCTGGAC TGTCAGGCTG CTTTTTTTCC AGATGTTCCT CCTCTGCCTC
1081 CCCTTCCCCT CCTCTCCCCT CCTTTTCCTT CCTTCCTTCC TTTCCTTGGA GCACTGAGCA
1141 CCATTTGGAA GCTTGAGAGA AACCAAAATT AAAGAGAGAA AGAGAGAAAA AAAAAAAAAA
1201 AAAAAAAAAA AAAAAAAAAA AAAAAAAAAA AAAAAAAAAA AAAAAAAAAA AACCTCGGG
B:氨基酸序列(SEQ ID NO:17) 长度:197
1 MVCRLTCVRF HSTNAVSVFS LSLFRACTPV PDPAPWPIPC RGASGKKTGR PARARLRRGH
61 PASPPTARCL TCFATDSCSQ PLGASVFGVG ELLRDCARPR PPTLKCQHHQ HQILRRHTAL
121 RTRRPGRLVS SCIKLSPCPL LPLPPVFLLI FFSPFPPSPS AFFPWFSTGK TVPLPPCLHG
181 SPAHVMLPSL FFVCVFI
C.核苷酸及氨基酸组合序列(SEQ ID NO:18) 克隆号:PP13842
起始编码子:346 ATG 终止编码子:937 TAA 蛋白质分子量:21622.56
1 GCT GCA GCC CCT GCC CCC GCC CCT CCT CGC TGG GTG CTC AGA AGG CTG 48
49 ACA GCT GCG CCA GGC TGA GGC GGC AGT CGA TGC TGG AGT TGT CCG GGC 96
97 CCG TGT AGG CCA GGC CCA GGG GCT CTA GGA AGG CCC GGC AGG CCC CAG 144
145 CGC TGC CTT TGC GGA TTC TGT TTT TGA GCC GTG GAC TTG GGT TGT AAA 192
193 TTT ATT TGT GGG GAG TGC GCT CCA GGA AGA GCC ACC ATC CCT GCC CCC 240
241 GTT TTC CCA CCG GGG AGT CTG TAC AGA GAT TTT TCT ACG TTT TTA TTT 288
289 TTT GCC TCA GAG GGA TGG GAT TGG GGA GGA GGG GAT GGG CAG CGG AGG 336
337 GTT GGG GGC ATG GTC TGC AGG CTC ATC TGT GTC CGC TTT CAC TCC ACT 384
1 Met Val Cys Arg Leu Ile Cys Val Arg Phe His Ser Thr 13
385 AAT GCT GTC TCA GTG TTT TCT CTC TCT CTC TTT CGA GCT TGC ACT CCG 432
14 Asn Ala Val Ser Val Phe Ser Leu Ser Leu Phe Arg Ala Cys Thr Pro 29
433 GTA CCC GAC CCG GCG CCC TGG CCC ATC CCA TGC CGG GGG GCC AGT GGA 480
30 Val Pro Asp Pro Ala Pro Trp Pro Ile Pro Cys Arg Gly Ala Ser Gly 45
481 AAG AAG ACA GGC CGT CCA GCC CGT GCC CGC CTG CGG CGG GGG CAC CCA 528
46 Lys Lys Thr Gly Arg Pro Ala Arg Ala Arg Leu Arg Arg Gly His Pro 61
529 GCA AGC CCG CCC ACC GCC CGC TGC CTC ACC TGC TTC GCC ACA GAC TCT 576
62 Ala Ser Pro Pro Thr Ala Arg Cys Leu Thr Cys Phe Ala Thr Asp Ser 77
577 TGT TCC CAG CCC CTT GGG GCC TCC GTG TTT GGG GTG GGG GAG CTG CTT 624
78 Cys Ser Gln Pro Leu Gly Ala Ser Val Phe Gly Val Gly Glu Leu Leu 93
625 AGA GAC TGT GCC CGT CCT CGG CCC CCC ACC CTG AAG TGC CAG CAC CAC 672
94 Arg Asp Cys Ala Arg Pro Arg Pro Pro Thr Leu Lys Cys Gln His His 109
673 CAG CAC CAG ATC CTC CGC CGC CAC ACC GCA CTG AGG ACA CGC CGG CCG 720
110 Gln His Gln Ile Leu Arg Arg His Thr Ala Leu Arg Thr Arg Arg Pro 125
721 GGC CGC CTC GTC TCA AGT TGT ATA AAG TTG TCT CCG TGT CCC CTC CTC 768
126 Gly Arg Leu Val Ser Ser Cys Ile Lys Leu Ser Pro Cys Pro Leu Leu 141
769 CCT CTG CCC CCA GTG TTT CTT CTG ATT TTT TTT TCC CCT TTC CCT CCC 816
142 Pro Leu Pro Pro Val Phe Leu Leu Ile Phe Phe Ser Pro Phe Pro Pro 157
817 TCC CCC TCC GCA TTC TTC CCT TGG TTC AGC ACA GGT AAA ACG GTT CCC 864
158 Ser Pro Ser Ala Phe Phe Pro Trp Phe Ser Thr Gly Lys Thr Val Pro 173
865 CTC CCT CCC TGC CTT CAT GGA TCA CCA GCT CAC GTC ATG TTG CCT TCT 912
174 Leu Pro Pro Cys Leu His Gly Ser Pro Ala His Val Met Leu Pro Ser 189
913 CTT TTC TTT GTG TGT GTG TTT ATT TAA GTT ATT TTT CTT CCT CCT CTC 960
190 Leu Phe Phe Val Cys Val Phe Ile *** 198
961 CCT TTT CTT TTT GGC CCT CCC TCC CTC CCT CTT CTG CCA TGT AAC TGG 1008
1009 AGG ATG TGC TAT GAG TTT GCA AAC AGC TGG ACT GTC AGG CTG CTT TTT 1056
1057 TTC CAG ATG TTC CTC CTC TGC CTC CCC TTC CCC TCC TCT CCC CTC CTT 1104
1105 TTC CTT CCT TCC TTC CTT TCC TTG GAG CAC TGA GCA CCA TTT GGA AGC 1152
1153 TTG AGA GAA ACC AAA ATT AAA GAG AGA AAG AGA GAA AAA AAA AAA AAA 1200
1201 AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA 1248
1249 AAA ACC TCG GG 1259
7.PP14673
A:核苷酸序列(SEQ ID NO:19)长度:2333
1 GCGGCCGTAG CGGCCGGGGC TGCGGTAGCC ACTTTAGATT TGGGCAAGGA CTTTAGATTC
61 GGGCTCTGTT CTGTTTCCGC CGTCCTGCTT CCTGCCGAGG CTGGCCCAGG CAGCCGCGCT
121 TCGAAGGACG CCGCCGGGAG CTGCGGAGCA TGCGTGGAGT GGCAGTGCTA ACGGCTGGTG
181 TCTCGCACTG TTGGCCTGTG AAGGTACGTG AAGCTGAAAG CCTGGAATGG CTGGAAAGGG
241 GTCATCAGGC AGGCGGCCCC TGCTGCTGGG GCTGCTGGTG GCCGTAGCCA CTGTCCACCT
301 GGTCATCTGT CCCTACACCA AAGTGGAGGA GAGCTTCAAC CTGCAGGCCA CACATGACCT
361 GCTCTACCAC TGGCAAGACC TGGAGCAGTA CGACCATCTT GAGTTCCCCG GAGTCGTCCC
421 CAGGACGTTC CTCGGGCCAG TGGTGATCGC AGTGTTCTCC AGCCCCGCGG TTTACGTGCT
481 TTCGCTGTTA GAAATGTCCA AGTTTTACTC TCAGCTAATA GTTAGAGGAG TGCTTGGACT
541 CGGCGTGATT TTTGGACTCT GGACGTTACA AAAGGAAGTG AGACGGCACT TCGGGGCCAT
601 GGTGGCCACC ATGTTCTGCT GGGTGACGGC CATGCAGTTC CACCTGATGT TCTACTGCAC
661 GCGGACACTG CCCAATGTGC TGGCCCTGCC TGTAGTCCTG CTGGCCCTCG CGGCCTGGCT
721 GCGGCACGAG TGGGCCCGCT TCATCTGGCT GTCAGCCTTC GCCATCATCG TGTTCAGGGT
781 GGAGCTGTGC CTGTTCCTGG GCCTCCTGCT GCTGCTGGCC TTGGGCAACC GAAAGGTTTC
841 TGTAGTCAGA GCCCTTCGCC ACGCCGTCCC GGCAGGGATC CTCTGTTTAG GACTGACGGT
901 TGCTGTGGAC TCTTATTTTT GGCGGCAGCT CACTTGGCCG GAAGGAAAGG TGCTTTGGTA
961 CAACACTGTC CTGAACAAAA GCTCCAACTG GGGGACCTCC CCGCTGCTGT GGTACTTCTA
1021 CTCAGCCCTG CCCCGCGGCC TGGGCTGCAG CCTGCTCTTC ATCCCCCTGG GCTTGGTAGA
1081 CAGAAGGACG CACGCGCCGA CGGTGCTGGC ACTGGGCTTC ATGGCACTCT ACTCCCTCCT
1141 GCCACACAAG GAGCTACGCT TCATCATCTA TGCCTTCCCC ATGCTCAACA TCACGGCTGC
1201 CAGAGGCTGC TCCTACCTGC TGAATAACTA TAAAAAGTCT TGGCTGTACA AAGCGGGGTC
1261 TCTGCTTGTG ATCGGACACC TCGTGGTGAA TGCCGCCTAC TCAGCCACGG CCCTGTATGT
1321 GTCCCATTTC AACTACCCAG GTGGCGTCGC AATGCAGAGG CTGCACCAGC TGGTGCCCCC
1381 CCAGACAGAC GTCCTTCTGC ACATTGACGT GGCAGCCGCC CAGACAGGTG TGTCTCGGTT
1441 TCTCCAAGTC AACAGCGCCT GGAGGTACGA CAAGAGGGAG GATGTGCAGC CGGGGACAGG
1501 CATGCTGGCA TACACACACA TCCTCATGGA GGCGGCCCCT GGGCTCCTGG CCCTCTACAG
1561 GGACACACAC CGGGTCCTGG CCAGCGTCGT GGGGACCACA GGTGTGAGTC TGAACCTGAC
1621 CCAACTGCCC CCCTTCAACG TCCACCTGCA GACAAAGCTG GTGCTTCTGG AGAGGCTCCC
1681 CCGGCCGTCC TGAGGGGGAC CAGGCAGCCC TCAGCAGCCA CAGGCCTTCC AGGAGCTGTT
1741 ATCACTACCA GTTTCTGGCA CAATTCCAGC ACAATTATGA CAATTCAGAG AAGCAAGTCA
1801 AAGGACTGGG CACCTGCCTC TGACAGACAC CAGACCAGGT CCAGGGCCTC CTCCACAGCC
1861 TCAGCTGGGG CTCTCAGCAC CAAAGAACGA GGGGCCCAGG TCTTGTTGGC ACCCCGGGAG
1921 CCACTGCCCA GGGTGATTGG TGGCCAGCTC AGGGCTTCCT GCGGGTGACT GTCGCCCAGA
1981 CCAGGTGCCA TTCATGACTA ATCAGGAGCA GCGGGCTCAC CCAGGCACCT GTCTGCCAGG
2041 AGGCCACCGT GTGTCCTGCC CACCCAGGGG GAGCTGTATT TTGGCAGCAC CCCACGCTTG
2101 CTGCCCGAGG GCCTCTTGGG GCACCTAAGA CAGCACCCCC TCTCAGGGGA GACCATGGTG
2161 GCCCCGGCCG CACCCCCCCA CCCTGGTGCC ACCACTGCAA CTTTTGTATT CACAGGCATC
2221 CCATCTCCAT CACAGATAAA ATCTTAGGAG ATAAACACAT TCAAAAAGGA ATGAGATAAA
2281 AAGAATAAGG CAATAAATGT TGATTGGAAC CTCTCAAAAA AAAAAAAAAA AAA
B:氨基酸序列(SEQ ID NO:20) 长度:488
1 MAGKGSSGRR PLLLGLLVAV ATVHLVICPY TKVEESFNLQ ATHDLLYHWQ DLEQYDHLEF
61 PGVVPRTFLG PVVIAVFSSP AVYVLSLLEM SKFYSQLIVR GVLGLGVIFG LWTLQKEVRR
121 HFGAMVATMF CWVTAMQFHL MFYCTRTLPN VLALPVVLLA LAAWLRHEWA RFIWLSAFAI
181 IVFRVELCLF LGLLLLLALG NRKVSVVRAL RHAVPAGILC LGLTVAVDSY FWRQLTWPEG
241 KVLWYNTVLN KSSNWGTSPL LWYFYSALPR GLGCSLLFIP LGLVDRRTHA PTVLALGFMA
301 LYSLLPHKEL RFIIYAFPML NITAARGCSY LLNNYKKSWL YKAGSLLVIG HLVVNAAYSA
361 TALYVSHFNY PGGVAMQRLH QLVPPQTDVL LHIDVAAAQT GVSRFLQVNS AWRYDKREDV
421 QPGTGMLAYT HILMEAAPGL LALYRDTHRV LASVVGTTGV SLNLTQLPPF NVHLQTKLVL
481 LERLPRPS
C.核苷酸及氨基酸组合序列(SEQ ID NO:21) 克隆号:PP14673
起始编码子:227 ATG 终止编码子:1691 TGA 蛋白质分子量:54651.70
1 G CGG CCG TAG CGG CCG GGG CTG CGG TAG CCA CTT TAG ATT TGG GCA 46
47 AGG ACT TTA GAT TCG GGC TCT GTT CTG TTT CCG CCG TCC TGC TTC CTG 94
95 CCG AGG CTG GCC CAG GCA GCC GCG CTT CGA AGG ACG CCG CCG GGA GCT 142
143 GCG GAG CAT GCG TGG AGT GGC AGT GCT AAC GGC TGG TGT CTC GCA CTG 190
191 TTG GCC TGT GAA GGT ACG TGA AGC TGA AAG CCT GGA ATG GCT GGA AAG 238
1 Met Ala Gly Lys 4
239 GGG TCA TCA GGC AGG CGG CCC CTG CTG CTG GGG CTG CTG GTG GCC GTA 286
5 Gly Ser Ser Gly Arg Arg Pro Leu Leu Leu Gly Leu Leu Val Ala Val 20
287 GCC ACT GTC CAC CTG GTC ATC TGT CCC TAC ACC AAA GTG GAG GAG AGC 334
21 Ala Thr Val His Leu Val Ile Cys Pro Tyr Thr Lys Val Glu Glu Ser 36
335 TTC AAC CTG CAG GCC ACA CAT GAC CTG CTC TAC CAC TGG CAA GAC CTG 382
37 Phe Asn Leu Gln Ala Thr His Asp Leu Leu Tyr His Trp Gln Asp Leu 52
383 GAG CAG TAC GAC CAT CTT GAG TTC CCC GGA GTC GTC CCC AGG ACG TTC 430
53 Glu Gln Tyr Asp His Leu Glu Phe Pro Gly Val Val Pro Arg Thr Phe 68
431 CTC GGG CCA GTG GTG ATC GCA GTG TTC TCC AGC CCC GCG GTT TAC GTG 478
69 Leu Gly Pro Val Val Ile Ala Val Phe Ser Ser Pro Ala Val Tyr Val 84
479 CTT TCG CTG TTA GAA ATG TCC AAG TTT TAC TCT CAG CTA ATA GTT AGA 526
85 Leu Ser Leu Leu Glu Met Ser Lys Phe Tyr Ser Gln Leu Ile Val Arg 100
527 GGA GTG CTT GGA CTC GGC GTG ATT TTT GGA CTC TGG ACG TTA CAA AAG 574
101 Gly Val Leu Gly Leu Gly Val Ile Phe Gly Leu Trp Thr Leu Gln Lys 116
575 GAA GTG AGA CGG CAC TTC GGG GCC ATG GTG GCC ACC ATG TTC TGC TGG 622
117 Glu Val Arg Arg His Phe Gly Ala Met Val Ala Thr Met Phe Cys Trp 132
623 GTG ACG GCC ATG CAG TTC CAC CTG ATG TTC TAC TGC ACG CGG ACA CTG 670
133 Val Thr Ala Met Gln Phe His Leu Met Phe Tyr Cys Thr Arg Thr Leu 148
671 CCC AAT GTG CTG GCC CTG CCT GTA GTC CTG CTG GCC CTC GCG GCC TGG 718
149 Pro Asn Val Leu Ala Leu Pro Val Val Leu Leu Ala Leu Ala Ala Trp 164
719 CTG CGG CAC GAG TGG GCC CGC TTC ATC TGG CTG TCA GCC TTC GCC ATC 766
165 Leu Arg His Glu Trp Ala Arg Phe Ile Trp Leu Ser Ala Phe Ala Ile 180
767 ATC GTG TTC AGG GTG GAG CTG TGC CTG TTC CTG GGC CTC CTG CTG CTG 814
181 Ile Val Phe Arg Val Glu Leu Cys Leu Phe Leu Gly Leu Leu Leu Leu 196
815 CTG GCC TTG GGC AAC CGA AAG GTT TCT GTA GTC AGA GCC CTT CGC CAC 862
197 Leu Ala Leu Gly Asn Arg Lys Val Ser Val Val Arg Ala Leu Arg His 212
863 GCC GTC CCG GCA GGG ATC CTC TGT TTA GGA CTG ACG GTT GCT GTG GAC 910
213 Ala Val Pro Ala Gly Ile Leu Cys Leu Gly Leu Thr Val Ala Val Asp 228
911 TCT TAT TTT TGG CGG CAG CTC ACT TGG CCG GAA GGA AAG GTG CTT TGG 958
229 Ser Tyr Phe Trp Arg Gln Leu Thr Trp Pro Glu Gly Lys Val Leu Trp 244
959 TAC AAC ACT GTC CTG AAC AAA AGC TCC AAC TGC GGG ACC TCC CCG CTG 1006
245 Tyr Asn Thr Val Leu Asn Lys Ser Ser Asn Trp Gly Thr Ser Pro Leu 260
1007 CTG TGG TAC TTC TAC TCA GCC CTG CCC CGC GGC CTG GGC TGC AGC CTG 1054
261 Leu Trp Tyr Phe Tyr Ser Ala Leu Pro Arg Gly Leu Gly Cys Ser Leu 276
1055 CTC TTC ATC CCC CTG GGC TTG GTA GAC AGA AGG ACG CAC GCG CCG ACG 1102
277 Leu Phe Ile Pro Leu Gly Leu Val Asp Arg Arg Thr His Ala Pro Thr 292
1103 GTG CTG GCA CTG GGC TTC ATG GCA CTC TAC TCC CTC CTG CCA CAC AAG 1150
293 Val Leu Ala Leu Gly Phe Met Ala Leu Tyr Ser Leu Leu Pro His Lys 308
1151 GAG CTA CGC TTC ATC ATC TAT GCC TTC CCC ATG CTC AAC ATC ACG GCT 1198
309 Glu Leu Arg Phe Ile Ile Tyr Ala Phe Pro Met Leu Asn Ile Thr Ala 324
1199 GCC AGA GGC TGC TCC TAC CTG CTG AAT AAC TAT AAA AAG TCT TGG CTG 1246
325 Ala Arg Gly Cys Ser Tyr Leu Leu Asn Asn Tyr Lys Lys Ser Trp Leu 340
1247 TAC AAA GCG GGG TCT CTG CTT GTG ATC GGA CAC CTC GTG GTG AAT GCC 1294
341 Tyr Lys Ala Gly Ser Leu Leu Val Ile Gly His Leu Val Val Asn Ala 356
1295 GCC TAC TCA GCC ACG GCC CTG TAT GTG TCC CAT TTC AAC TAC CCA GGT 1342
357 Ala Tyr Ser Ala Thr Ala Leu Tyr Val Ser His Phe Asn Tyr Pro Gly 372
1343 GGC GTC GCA ATG CAG AGG CTG CAC CAG CTG GTG CCC CCC CAG ACA GAC 1390
373 Gly Val Ala Met Gln Arg Leu His Gln Leu Val Pro Pro Gln Thr Asp 388
1391 GTC CTT CTG CAC ATT GAC GTG GCA GCC GCC CAG ACA GGT GTG TCT CGG 1438
389 Val Leu Leu His Ile Asp Val Ala Ala Ala Gln Thr Gly Val Ser Arg 404
1439 TTT CTC CAA GTC AAC AGC GCC TGG AGG TAC GAC AAG AGG GAG GAT GTG 1486
405 Phe Leu Gln Val Asn Ser Ala Trp Arg Tyr Asp Lys Arg Glu Asp Val 420
1487 CAG CCG GGG ACA GGC ATG CTG GCA TAC ACA CAC ATC CTC ATG GAG GCG 1534
421 Gln Pro Gly Thr Gly Met Leu Ala Tyr Thr His Ile Leu Met Glu Ala 436
1535 GCC CCT GGG CTC CTG GCC CTC TAC AGG GAC ACA CAC CGG GTC CTG GCC 1582
437 Ala Pro Gly Leu Leu Ala Leu Tyr Arg Asp Thr His Arg Val Leu Ala 452
1583 AGC GTC GTG GGG ACC ACA GGT GTG AGT CTG AAC CTG ACC CAA CTG CCC 1630
453 Ser Val Val Gly Thr Thr Gly Val Ser Leu Asn Leu Thr Gln Leu Pro 468
1631 CCC TTC AAC GTC CAC CTG CAG ACA AAG CTG GTG CTT CTG GAG AGG CTC 1678
469 Pro Phe Asn Val His Leu Gln Thr Lys Leu Val Leu Leu Glu Arg Leu 484
1679 CCC CGG CCG TCC TGA GGG GGA CCA GGC AGC CCT CAG CAG CCA CAG GCC 1726
485 Pro Arg Pro Ser *** 489
1727 TTC CAG GAG CTG TTA TCA CTA CCA GTT TCT GGC ACA ATT CCA GCA CAA 1774
1775 TTA TGA CAA TTC AGA GAA GCA AGT CAA AGG ACT GGG CAC CTG CCT CTG 1822
1823 ACA GAC ACC AGA CCA GGT CCA GGG CCT CCT CCA CAG CCT CAG CTG GGG 1870
1871 CTC TCA GCA CCA AAG AAC GAG GGG CCC AGG TCT TGT TGG CAC CCC GGG 1918
1919 AGC CAC TGC CCA GGG TGA TTG GTG GCC AGC TCA GGG CTT CCT GCG GGT 1966
1967 GAC TGT CGC CCA GAC CAG GTG CCA TTC ATG ACT AAT CAG GAG CAG CGG 2014
2015 GCT CAC CCA GGC ACC TGT CTG CCA GGA GGC CAC CGT GTG TCC TGC CCA 2062
2063 CCC AGG GGG AGC TGT ATT TTG GCA GCA CCC CAC GCT TGC TGC CCG AGG 2110
2111 GCC TCT TGG GGC ACC TAA GAC AGC ACC CCC TCT CAG GGG AGA CCA TGG 2158
2159 TGG CCC CGG CCG CAC CCC CCC ACC CTG GTG CCA CCA CTG CTT CTT TTG 2206
2207 TAT TCA CAG GCA TCC CAT CTC CAT CAC AGA TAA AAT CTT AGG AGA TAA 2254
2255 ACA CAT TCA AAA AGG AAT GAG ATA AAA AGA ATA AGG CAA TAA ATG TTG 2302
2303 ATT GGA ACC TCT CAA AAA AAA AAA AAA AAA A 2333
8.PP14776
A:核苷酸序列(SEQ ID NO:22) 长度:1405
1 GTTTGGGTAG ATTTATAGGT GTTTAATAGC CATAGTCAAA GCACATTTAT TAATGATAAA
61 TGGAAATGGA GACCCCTAAG GTATGCAGTA AAGACAGGTC CTCGGTTTGG GATTCGCCAC
121 CGTTTCCTCC CACGCATTTT AATGAAAGAT TTAAATGAAA ACACAGATTT GCTTTCCACA
181 TTTTAAAAAT TATTCAAAAC TAGAGTCAAA GGGGAGAATC TGATACACTG GATAACAGAA
241 TTGAAATTCA ATAAGGCATT CAGAAGCAAA AGCAGTTATG CTGTAACAAG TCAGCTGCAC
301 TGCCATCCCA CTGAAACTAT GATCGATACC TGGCATTTAT GAGAACTTAT GATACACCAG
361 GAACTTCGAT GTGCATCATC CCAGATGATC CTCACAGAAA TCCTAGGAGG TTCTACGATG
421 AAATCAGGAC CCCGCTCCTC TCTGACATCC GCATCGATTA TCCCCCCAGC TCAGTGGTGC
481 AGGCCACCAA GACCCTGTTC CCCAACTACT TCAACGGCTC GGAGATCATC ATTGCGGGGA
541 AGCTGGTGGA CAGGAAGCTG GATCACCTGC ACGTGGAGGT CACCGCCAGC AACAGTAAGA
601 AATTCATCAT CCTGAAGACA GATGTGCCTG TGCGGCCTCA GAAGGCAGGG AAAGATGTCA
661 CAGGAAGCCC CAGGCCTGGA GGCGATGGAG AGGGGGACCC CAACCACATC GAGCGTCTCT
721 GGAGCTACCT CACCACAAAG GAGCTGCTGA GCTCCTGGCT GCAAAGTGAC GATGAACCGG
781 AGAAGGAGCG GCTGCGGCAG CGGGCCCAGG CCCTGGCTGT GAGCTACCGC TTCCTCACTC
841 CCTTCACCTC CATGAAGCTG AGGGGGCCGG TCCCACGCAT GGACGGCCTG GAGGAGGCCC
901 ACGGCATGTC GGCTGCCATG GGACCCGAAC CGGTGGTGCA GAGCGTGCGA GGAGCTGGCA
961 CGCAGCCAGG ACCTTTGCTC AAGAAGCCAT ACCAGCCAAG AATTAAAATC TCTAAAACAT
1021 CAGGTAAAGC AAAGGATGCG GTTGTGTGTG GATTGAGAGT AAGAGATGTT TAAAAAGTGT
1081 AAAACCAGGC CAGGTGTACT GGCTCACGCC TGTAATCCCA GCACTTTGGG AGGCCGAGGT
1141 GGGTGTATTG CTTGAGGTCA GGAGTTCAAG ACCAGCCTGG CCAACATGGT GCAACCTCGT
1201 CTCTACTAAG AATACAAAAA TTAGCCAGGC ATGGTGGCAG GGGCCTGTAG TTCCAGCTAC
1261 TGGGGAGGCT GAGGCAGGAG AATCACTTGA ACCTGGGAAG TTGAGGTTAC AGTGAGCCGA
1321 GATTGTGCCA CCACACTCTA GCCTGGACTA CAAGAGTGAA ACTCTGTCTC AAAAAAAAAA
1381 AAAAAAAAAA AAAAAAAAAA AAAAA
B:氨基酸序列(SEQ ID NO:23) 长度:244
1 MRTYDTPGTS MCIIPDDPHR NPRRFYDEIR TPLLSDIRID YPPSSVVQAT KTLFPNYFNG
61 SEIIIAGKLV DRKLDHLHVE VTASNSKKFI ILKTDVPVRP QKAGKDVTGS PRPGGDGEGD
121 PNHIERLWSY LTTKELLSSW LQSDDEPEKE RLRQRAQALA VSYRFLTPFT SMKLRGPVPR
181 MDGLEEAHGM SAAMGPEPVV QSVRGAGTQP GPLLKKPYQP RIKISKTSGK AKDAVVCGLR
241 VRDV
C.核苷酸及氨基酸组合序列(SEQ ID NO:24) 克隆号:PP14776
起始编码子:339 ATG 终止编码子:1071 TAA 蛋白质分子量:27185.72
1 GT TTG GGT AGA TTT ATA GGT GTT TAA TAG CCA TAG TCA AAG CAC ATT 47
48 TAT TAA TGA TAA ATG GAA ATG GAG ACC CCT AAG GTA TGC AGT AAA GAC 95
96 AGG TCC TCG GTT TGG GAT TCG CCA CCG TTT CCT CCC ACG CAT TTT AAT 143
144 GAA AGA TTT AAA TGA AAA CAC AGA TTT GCT TTC CAC ATT TTA AAA ATT 191
192 ATT CAA AAC TAG AGT CAA AGG GGA GAA TCT GAT ACA CTG GAT AAC AGA 239
240 ATT GAA ATT CAA TAA GGC ATT CAG AAG CAA AAG CAG TTA TGC TGT AAC 287
288 AAG TCA GCT GCA CTG CCA TCC CAC TGA AAC TAT GAT CGA TAC CTG GCA 335
336 TTT ATG AGA ACT TAT GAT ACA CCA GGA ACT TCG ATG TGC ATC ATC CCA 383
1 Met Arg Thr Tyr Asp Thr Pro Gly Thr Ser Met Cys Ile Ile Pro 15
384 GAT GAT CCT CAC AGA AAT CCT AGG AGG TTC TAC GAT GAA ATC AGG ACC 431
16 Asp Asp Pro His Arg Asn Pro Arg Arg Phe Tyr Asp Glu Ile Arg Thr 31
432 CCG CTC CTC TCT GAC ATC CGC ATC GAT TAT CCC CCC AGC TCA GTG GTG 479
32 Pro Leu Leu Ser Asp Ile Arg Ile Asp Tyr Pro Pro Ser Ser Val Val 47
480 CAG GCC ACC AAG ACC CTG TTC CCC AAC TAC TTC AAC GGC TCG GAG ATC 527
48 Gln Ala Thr Lys Thr Leu Phe Pro Asn Tyr Phe Asn Gly Ser Glu Ile 63
528 ATC ATT GCG GGG AAG CTG GTG GAC AGG AAG CTG GAT CAC CTG CAC GTG 575
64 Ile Ile Ala Gly Lys Leu Val Asp Arg Lys Leu Asp His Leu His Val 79
576 GAG GTC ACC CCC AGC AAC AGT AAG AAA TTC ATC ATC CTG AAG ACA GAT 623
80 Glu Val Thr Ala Ser Asn Ser Lys Lys Phe Ile Ile Leu Lys Thr Asp 95
624 GTG CCT GTG CGG CCT CAG AAG GCA GGG AAA GAT GTC ACA GGA AGC CCC 671
96 Val Pro Val Arg Pro Gln Lys Ala Gly Lys Asp Val Thr Gly Ser Pro 111
672 AGG CCT GGA GGC GAT GGA GAG GGG GAC CCC AAC CAC ATC GAG CGT CTC 719
112 Arg Pro Gly Gly Asp Gly Glu Gly Asp Pro Asn His Ile Glu Arg Leu 127
720 TGG AGC TAC CTC ACC ACA AAG GAG CTG CTG AGC TCC TGG CTG CAA AGT 767
128 Trp Ser Tyr Leu Thr Thr Lys Glu Leu Leu Ser Ser Trp Leu Gln Ser 143
768 GAC GAT GAA CCG GAG AAG GAG CGG CTG CGG CAG CGG GCC CAG GCC CTG 815
144 Asp Asp Glu Pro Glu Lys Glu Arg Leu Arg Gln Arg Ala Gln Ala Leu 159
816 GCT GTG AGC TAC CGC TTC CTC ACT CCC TTC ACC TCC ATG AAG CTG AGG 863
160 Ala Val Ser Tyr Arg Phe Leu Thr Pro Phe Thr Ser Met Lys Leu Arg 175
864 GGG CCG GTC CCA CGC ATG GAC GGC CTG GAG GAG GCC CAC GGC ATG TCG 911
176 Gly Pro Val Pro Arg Met Asp Gly Leu Glu Glu Ala His Gly Met Ser 191
912 GCT GCC ATG GGA CCC GAA CCG GTG GTG CAG AGC GTG CGA GGA GCT GGC 959
192 Ala Ala Met Gly Pro Glu Pro Val Val Gln Ser Val Arg Gly Ala Gly 207
960 ACG CAG CCA GGA CCT TTG CTC AAG AAG CCA TAC CAG CCA AGA ATT AAA 1007
208 Thr Gln Pro Gly Pro Leu Leu Lys Lys Pro Tyr Gln Pro Arg Ile Lys 223
1008 ATC TCT AAA ACA TCA GGT AAA GCA AAG GAT GCG GTT GTG TGT GGA TTG 1055
224 Ile Ser Lys Thr Ser Gly Lys Ala Lys Asp Ala Val Val Cys Gly Leu 239
1056 AGA GTA AGA GAT GTT TAA AAA GTG TAA AAC CAG GCC AGG TGT ACT GGC 1103
240 Arg Val Arg Asp Val *** 245
1104 TCA CGC CTG TAA TCC CAG CAC TTT GGG AGG CCG AGG TGG GTG TAT TGC 1151
1152 TTG AGG TCA GGA GTT CAA GAC CAG CCT GGC CAA CAT GGT GCA ACC TCG 1199
1200 TCT CTA CTA AGA ATA CAA AAA TTA GCC AGG CAT GGT GGC AGG GGC CTG 1247
1248 TAG TTC CAG CTA CTG GGG AGG CTG AGG CAG GAG AAT CAC TTG AAC CTG 1295
1296 GGA AGT TGA GGT TAC AGT GAG CCG AGA TTG TGC CAC CAC ACT CTA GCC 1343
1344 TGG ACT ACA AGA GTG AAA CTC TGT CTC AAA AAA AAA AAA AAA AAA AAA 1391
1392 AAA AAA AAA AAA AA 1405
Claims (9)
1.一种分离的具有促进3T3细胞转化功能的人蛋白多肽,其特征在于,它是具有选自下组的氨基酸序列的多肽:SEQ ID NO:2、5、8、11、14、17、20、23。
2.如权利要求1所述的多肽,其特征在于,该多肽的氨基酸序列选自下组:SEQ ID NO:2、11、20。
3.一种分离的多核苷酸,其特征在于,选自下组:
(a)编码如权利要求1所述多肽的多核苷酸;
(b)与多核苷酸(a)完全互补的多核苷酸。
4.如权利要求3所述的多核苷酸,其特征在于,该多核苷酸编码的多肽具有选自下组的氨基酸序列:SEQ ID NO:2、5、8、11、14、17、20、23。
5.如权利要求3所述的多核苷酸,其特征在于,该多核苷酸的序列选自下组:
SEQ ID NO:3、6、9、12、15、18、21、24的编码区序列或全长序列。
6.一种载体,其特征在于,它含有权利要求3所述的多核苷酸。
7.一种遗传工程化的宿主细胞,其特征在于,它是选自下组的一种宿主细胞:
(a)用权利要求6所述的载体转化或转导的宿主细胞;
(b)用权利要求3所述的多核苷酸转化或转导的宿主细胞。
8.一种具有促进3T3细胞转化功能的人蛋白活性的多肽的制备方法,其特征在于,该方法包含:
(a)在适合表达具有促进3T3细胞转化功能的人蛋白的条件下,培养权利要求7所述的宿主细胞;
(b)从培养物中分离出具有促进3T3细胞转化功能的人蛋白活性的多肽。
9.一种能与权利要求1所述的具有促进3T3细胞转化功能的人蛋白多肽特异性结合的抗体。
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