CN1141651A - 内皮素转化酶(ece) - Google Patents
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Abstract
本发明涉及包括SEQ ID NO:30或SEQ IDNO:36所述多肽序列的内皮素转化酶,或涉及其功能片段,且涉及其编码基因,和涉及其制备的方法及其用途。
Description
本发明涉及内皮素转化酶、该酶的制备方法及其应用。
内皮素水平升高被认为与原发性高血压、心肌梗塞、急性肾功能衰竭、休克或心功能衰竭等多种疾病有关。利用内皮素抗体所能获得的结果也提示了这种相关性。在动物模型中,有可能以此以剂量依赖性方式减少心肌梗塞的面积(Watanabeet al.,Nature 344,114(1990)、对肾功能产生有益的影响(Kon et al.,J.Clin.Invest.83,1762(1989))和降低环孢菌素的肾毒性(Kon et al.,KindeyInt.37,1487(1990))。
内皮素转化酶(ECE-1)可从由38个氨基酸组成的前体分子之大内皮素1释出内皮素1。由内皮素1所引起的不利生物作用可通过例如抑制内皮素转化酶,进而抑制内皮素1的生物合成来加以阻止。已从多种细胞系中分离出由相应的前体分子,即大内皮素释放内皮素或内皮素样分子的酶活性(Takeda et al.,Biochem.Biophys.Res.Comm.176,860(1991),Ohnaka et al.,Biochem.Biophys.Res.Comm.168,1128(1990);Matsumura et al.,FEBS Lett.293,45(1992),Ahn et al.Proc.Natl.Acad.Sci.USA 89,8606(1992),PCTWO 92/13944,Ohnaka et al.Clin.Exp.Hypertension A 14;No.4(1992),Okada et al.Biochem.Biophys.Res.Comm.180,1019(1991),Takada et al.Biochem.Biophys.Res.Comm.182,1383(1992),Opgenorth et al.FASEB 6,2653(1992))
然而,这些酶活性的特征至今还未得以充分地鉴定。该酶以未被浓缩的酶混合物形式存在。但是,这些类型的不纯酶混合物不是非常适用于发现特异性ECE抑制剂的测定法,因为具有非生理相关性的外源蛋白酶极大地干扰了酶测定。
因此,大内皮素也可以由例如丝氨酸蛋白酶胰凝乳蛋白酶[sic]以及木瓜蛋白酶、嗜热菌蛋白酶和NEP 24.11裂解成内皮素和内皮素样产物,而这些酶在测定法中被错误地定为ECE酶。结果,与文献中有关内皮素转化酶的描述产生矛盾。
有关内皮素转化酶的分子量的描述是在65-540KD范围内变动;Km和Vmax值也彼此极不相同(Opgenorth et al.,FASEB 6,2653(1992);Sawamuraet al.Biochem.Biophys.Acta 1161,295(1993))。在是否将内皮素转化酶定为天冬氨酸蛋白酶族或金属蛋白酶族方面也存在着分歧(Sawamura etal.Biochem.Biophys.Acta 1161,295(1993);Biochem.Pharmacol.43,845(1992)。而且,有关金属蛋白酶是否为胞质酶或膜结合酶和哪些底物可被特定的ECE(大Et-1;大Et-3)转化之特征信息的对立意见可见于下列文献:Matsumura et al.(FEBS Lett.293,45(1992):Takahashi et al.(J.Biol.Chem.268;21394(1993));Okada et al.(Biochem.Biophys.Res.Comm.180,1019(1991));Ohnaka et al.(Clin.Exp.Hypretension A14;No.4(1992));Matsumura et al.(FEBS Lett.305;86(1992))。
至今还未见能够使得满意地定义ECE的确定性描述。大概只有通过测定ECE的一级结构其氨基酸序列才可能定义ECE。为此,首先必需制备纯化形式的ECE。
本发明的目的在于制备纯化形式的内皮素转化酶。
我们已经发现,通过分子量为250,000Da的内皮素转化酶和其部分氨基酸序列SEQ ID NO:1可以实现上述目的。
本发明的内皮素转化酶具有下列特征:在非还原条件下经SDS聚丙烯酰胺凝胶电泳测定的分子量约为250,000Da;在还原条件下的SDS聚丙烯酰胺凝胶电泳中显示出大约125,000Da的条带,推测其由同源二聚体组成。
内皮素转化酶是糖基化酶。经酶学方法除去糖基使表观分子量由125,000Da变为大约82,000Da。
内皮素转化酶的胰蛋白酶肽的测序提供了下文的部分氨基酸序列SEQ IDNO:1、2、3、4、5、6。
本发明内皮素转化酶的更进一步特征将在实施中描述。
本发明进一步涉及分子量约为250,000Da且与SEQ ID NO:1显示出[sic]至少80%同源性之部分氨基酸序列的内皮素转化酶。
所述内皮素转化酶可从牛之外的生物体,例如从人细胞中分离。
本发明进一步涉及含有SEQ ID NO:30和SEQ ID NO:36所述多肽序列的内皮素转化酶或其功能性片段。
功能性片段意指仍然具有内皮素转化酶之酶活性、抗原特性或与配体(如结合蛋白质)的亲和力的那些部分序列。
其它的功能性片段包括可从SEQ ID NO:30或NO:36开始经氨基酸或肽的***、缺失或替代能够得到片段,并且仍然基本具有内皮素转化酶的酶活性和/或基本具有与SEQ ID NO:30或NO:36的内皮素转化酶的免疫交叉反应性。
本发明进一步涉及编码内皮素转化酶且选自下组序列的DNA序列:
a)具有SEQ ID NO:29或SEQ ID NO:35所述结构的DNA序列;
b)编码具有SEQ ID NO:30或SEQ ID NO:36所述结构之蛋白质的DNA序列;以及
c)在标准条件下与DNA序列a)或b)杂交的DNA序列;
d)编码能被针对SEQ ID NO:30或36之蛋白质产生的抗体所识别的蛋白质产物或其片段的DNA序列。
标准条件指例如在浓度为0.1-1×SSC(1×SSC:0.15M NaCl,15mM柠檬酸钠,pH 7.2)的水性缓冲液中温度为42-58℃。DNA杂交的实验条件在遗传工程的教科书(如Sambrook et al.,“Molecular Cloning”Cold SpringHarbor Laboratory,1989)中进行了描述。
实施例8描述了所说DNA序列的制备。所说的DNA序列的蛋白质产物可以被已产生的SEQ ID NO:30或36的蛋白质或其片段之抗体识别,从而以已很好描述的方法获得。
在例如Hudson和Hay,F.C.,“Practical Immunology”,Blackwell Sci.Pub.,Oxford,1980书中描述了培育所说蛋白质抗体的方法。
在例如“DNA Clonging Vol.I”,Glover,D.M.,Ed.,IRL PressOxford,1985的书中描述了用抗体发现编码与这些抗体发生交叉反应的蛋白质之cDNA序列的方法。
本发明进一步涉及内皮素转化酶制备的方法,它包括用内皮素转化酶抑制剂刺激哺乳动物细胞,优选内皮细胞,使之过量产生内皮素转化酶,然后用常规蛋白质化学方法从这些细胞中分离内皮素转化酶。
所有的ECE抑制剂均可用于诱导哺乳动物细胞ECE。在下列的文献中描述了用于此目的的ECE抑制剂,例如:JP-146737;JP 05148277-Al;Jpn J.Biochem.64;(8)Abst.2367(1992);Derment NCE-93-0956(1993);Derwent NCE-93-0971(1993);J.Med.Chem.36,173(1993);EP 518299-A2。优选应用Phosphoramidon。通过将这些抑制剂加至细胞培养基中培养6小时至6天,刺激培养的细胞。然而,这种刺激优选进行2-3天。所用的抑制剂的浓度为10-2M-10-7M优选10-3M-10-4M。
然而,根据本发明的内皮素转化酶也可以用遗传学方法制备。基于所描述的部分氨基酸序列制备编码这些序列或互补于编码这些序列之序列的合成寡核苷酸是可能的。将这些寡核苷酸用作杂交探针从基因库如基因bank或cDNA库中分离相应的基因或cDNA分子。专业技术人员可从分子生物学的教科书如Sambrook,Fritsch,Maniatis,Molecular Cloning,Cold Spring Harbor LaboratoryPress,1989,USA中得知这种基因分离的方法。
同样地,从例如cDNA库或第一链cDNA中用适宜的合成寡核苷酸及聚合酶链反应(PCR)(PCR Protocols,Academic Press San Diego;USA;1990)获得部分基因是可能的。从Seq ID No.I至No.6中描述的肽中,通过将氨基酸序列翻译成双链DNA序列(正义及反义)推导出适于此目的的寡核苷酸。在此情况下,由于遗传密码的简并性,有可能从一种肽中推导出简并的寡核苷酸混合物并用作引物。考虑到所谓的“密码子使用”(codon usage),即在一物种中(本发明为牛)优选地使用,推导出氨基酸的确定核苷酸三联密码。然后用两种不同的寡核苷酸作引物进行PCR,在此情况下PCR所需的两个引物可由寡核苷酸的正链(正义链)和互补链(反义链)组成。同样地也可能用cDNA载体的多个部分或对应的mRNA3’部分的寡-dT推导出的寡核苷酸。
借助于基因分离方法获得完整的基因或完整的cDNA,反过来也可应用这些基因部分。
以这种方式能够不仅分离到内皮素转化酶具有与SEQ ID NO:1描述的部分氨基酸序列完全一致序列的基因,也可能分离到变体或其它生物体如人的内皮素转化酶的基因,这些基因在蛋白质水平上显示出与SEQ ID NO:1具有至少80%的同源性。
根据本发明的内皮素转化酶可通过用常规遗传工程方法分离相应基因并在宿主生物体中进行表达的方法得到很好地制备。
根据本发明的ECE或由其衍生的肽可被用于制备抗体或抗体片段如Fv片段,这些抗体或片段可应用于例如鉴定ECE抑制剂的测定方法中或作为ECE抑制剂测定ECE。
这些抗体在内皮素转化酶参与的疾病的诊断中一般均是有价值的辅助手段。
为了达到如测定ECE转录或确定基因型的目的,也可以用根据本发明的DNA序列衍生的PCR引物。
以本领域专业人员熟知的方法,可将所得基因用于修饰动物,使其基因完善。可以是含有一个或多个额外拷贝或可以是关闭掉正常的ECE基因。这些转基因动物特别适于实验目的。
内皮素转化酶本身作为生物活性成分也适于作为药物生产,这些药物用于出现了ECE底物水平升高的疾病,ECE底物如缓激肽、P物质、生长激素、释放抑制因子、神经肽Y。这些疾病的例子为炎症、哮喘、偏头痛、风湿病以及作为生长因子的肽参与的细胞过程及细胞增殖。ECE也可用作药物应用于例如感染性休克、偏头痛、性功能失调等有可以服用ECR消除的血管舒张之疾病。
为将ECE作为药物加以应用必须在某些情况下将ECE的某些部分用突变方式加以改变。这些突变性改变可导致例如一种ECE衍生物的产生,这种衍生物未糖基化或不含膜锚点(可溶性ECE)。不再含有膜锚点的可溶性ECE可由例如表达含胞外催化活性结构域的ECE片段加以制备。该类型的ECE片段可用例如下述的方法获得:将SEQ ID NO:25的氨基酸20-703,优选SEP[sic]ID NO:25的氨基酸27-703的编码序列连接到一信号肽的DNA序列上,该信号肽可在体内消除。在本领域专业人员已知的适宜表达载体中导入真核细胞。适宜的信号肽的一个例子是人组织纤维蛋白溶酶原激活剂自氨基酸-35至-4位的序列(Collen,D.,Nature 301,214至221(1983))。所产生的改变可影响例如特异性、作用的时间或摄取。而且,为产生可药用的新的蛋白质,ECE的功能部分可与其它蛋白质的部分耦联。也可以如PEG、葡聚糖或脂肪酸等非肽分子进行共价修饰而改变ECE。
根据本发明的DNA序列也适用于基因治疗,如用于生产基因治疗的药物。内皮素水平升高的疾病也可用衍生于ECE DNA序列的寡核苷酸治疗。这些寡核苷酸衍生于非编码DNA链(反义链)且可用作药物。基于此的反义治疗优选利用了衍生于寡核苷酸的更稳定的化学衍生物。例如,优选Seq ID[sic]:35的3l-100位定义的区域合成反义寡核苷酸。反义寡核苷酸优选的长度为15-30个残基。疾病的反义治疗或预防与ECE抑制剂具有同样的适应症,如脑缺血、蛛网膜下腔 出血、血管痉挛、冠脉缺血、与细胞增殖过程相关的疾病,例如***增生,如动脉粥样硬化,如再狭窄、哮喘、高血压、肺动脉高压、器官衰竭如心衰和肾衰、肾缺血、Neprotoxizitt、心肌梗塞、冠心病、败血症。
也可能从ECE的DNA序列中衍生出可以产生催化性RNA分子的序列,该种RNA分子称为核酶(ribozymes)。它们通过在服用后裂解或失活ECE RNA或ECE DNA并从而阻止ECE蛋白合成而起作用。
为建立可表达该酶的细胞,可将ECE DNA序列与适宜的表达载体一起使用。这些细胞特别地用作药物。
内皮素转化酶优选用于鉴定ECE抑制剂的测定方法中。这些测定方法通常为酶促反应,其中在潜在的抑制剂存在情况下进行测量ECE-催化的底物裂解。
本发明进一步涉及内皮素转化酶的抑制剂,所说的抑制剂是以用根据本发明的内皮素转化酶进行的酶活测定法鉴定的。
本发明也包括生产能抑制内皮素转化酶之药物的方法,该方法包括应用根据本发明内皮素转化酶的酶法促测定法中所用的已知化合物,和鉴定其是否具有抑制效应,并用常规载剂和辅助物质将以此方式鉴定的化合物配制成药物。
实施例1
用phosphoramidon于FBHE源代牛内皮细胞中刺激ECE-1
将含冷冻FBHE第14代牛内皮细胞(ATCC CRL 1395)的三个试管融化且导入3个各含有40ml DMEM生长培养基(Gibco No.041-01965)的T175细胞培养瓶中。所说培养基加上
+5ml/500ml谷胺酰胺(200mM;Gibco 043-05030)
+1ml/500ml艮他霉素(Gibco 043-05750)
+非必需氨基酸;终浓度:1×(Gibco No.043-01140)
+10%FCS(Gibco No.011-06290;于56℃处理37分钟失活)+
25ng/ml碱性FGF(Intergen 4125-80))。
以标准方法于37℃在7%CO2、100%湿度下温育这些细胞。第二天换培养基。当4天后达到汇合时,以标准方法用胰蛋白酶处理细胞传代,并分至9个T175瓶中,如所描述的那样于37℃温育生长。3天后达到汇合时,再将细胞以标准方法用胰蛋白酶处理后分至20个T瓶中。再过3天可以将细胞分至40个T瓶中。3天后,将其中39个T瓶分至78个T瓶中。由于这78个T瓶用于收获细胞,所以用适宜的细胞培养瓶(Costar;Accell;No.3155)用于此目的。在这些T瓶接种后的第二天向68个瓶中加入phosphoramidon(得自Peptide Institute;No.4082,简称PHAM)使终浓度为10-4M,这68个瓶标为(+PHAM)。这种处理诱导了ECE活性。剩下的10个T瓶含无诱导对照细胞(-PHAM)。在所描述的条件下再温育2天,分别收获PHAM诱导细胞和对照细胞(-PHAM)。过程是:打开瓶子,吸出培养基;对各瓶分别用10ml PBS(Boehringer MannheimNo.210331)于瓶中冲洗细胞(即小心旋转瓶子,使PBS流经细胞层);吸出PBS,再向各瓶中加入5ml PBS。用细胞刮除器(Costar No.3010)将细胞从培养瓶底刮下并以细胞悬液的形式移至150ml离心管(Faleon No.2076)中,在冰中贮存;各T瓶然后用10ml PBS冲洗,洗出的细胞悬液与离心管中的悬液合并;离心(1200rpm;10分钟;Heraeus Christ Minifuge GL No.4400;Rotor2150)使细胞沉淀。弃去不含细胞的上清液,将沉淀物置于其3倍体积的PBS中;再次离心,弃去无细胞上清液,将沉淀物保存在冰中直至使用。
实施例2
非刺激的及phosphoramidon刺激的[sic]FBHE牛内皮细胞的比较性富集;蛋白
质的鉴定
在15ml PBS缓冲液、0.5mM二异丙基氟磷酸中于冰浴里用超声处理实施例1中获得的用及未用phosphoramidon诱导的FBHE细胞(各为湿重500μl)20分钟。离心(1000g,20分钟)后,向上清液中加入5%Pluriol F68,再以10,000g离心使膜沉淀以获得膜。用2ml 100mM Tris缓冲液(pH8.0)消化膜并用1%TritonX-100使之增溶。将增溶物(各为2.5ml)离心,弃去残渣。
Mono-Q层析
在完全相同的条件下于Mono-QHR5/5柱(Phamacia)分别对+/-phosphoramidon诱导的细胞增溶物进行层析分离。层析柱用50ml Tris缓冲液(pH8.0)、0.1%Triton X-100(A缓冲液)平衡。将溶解液上柱并以A缓冲液洗涤15分钟,再用B缓冲液(A缓冲液+1M NaCl)线性梯度洗涤50分钟(流速0.2ml/min)。收集0.5ml的级分20份并用人大Et-1作为底物进行测定。用反相HPLC按所述方法(实施例3)检测并定量所产生的Et-1。在每种情况下将两个最高酶活性的级分合并。
Saperose 12凝胶层析
用Centricon管(Amicon)将得自Mono Q层析的洗脱液浓缩至250μl,并各自在Superose 12 HR 10/36上进行凝胶层析。
条件: 缓冲液: 20mM磷酸钠
250mM NaCl,pH 7.4
0.05 Triton×100
流速: 0.5ml/min
级分: 0.5ml
以实施例3描述的方法测定各级分,并且各自将两个最高酶活性的级分合并。
用Bradford法(Anal.Biochem.72,248(1976))测定所有级分的蛋白质。
结果:
经刺激和未经刺激细胞的纯化在不同的纯化阶段给出了下列结果:
纯化步骤 | 比活[μU/mg]FBHE细胞+phosphoramidon | μg/mgFBHE细胞-phosphoramidon |
膜 | 129 | 44 |
增溶液 | 188 | 22 |
Mono Q层析 | 1470 | 78 |
Superose 12层析 | 4500 | 130 |
将Superose 12层析的两个活性级分上样于8%SDS聚丙烯酰胺凝胶进行比较。phosphoramidon处理的细胞之纯化ECE-1的凝胶图形显示与未处理细胞相比于250,000Dalton处有一另外的条带。当与还原剂二硫苏糖醇(DTT)一起将蛋白质上样时,比较显示在125,000Dalton处有一另外的条带。
实施例3
HPLC的ECE-1活性测定
内皮素转化酶(ECE-1)引起的大内皮素-1向内皮素1的转化
将1μl实施例4中所得的酶溶液与21μl 50mM Tris,50mM咪唑、250mMNaCl(pH7.4)缓冲液及2.5μl的人大内皮素(溶于水中的0.1%乙酸中,2mg/ml)混合。2小时后,用72μl 0.5%三氟乙酸终止反应以分析产生的内皮素。将样本离心,用带UV检测(205nm)的反相高压液相层析分析,按J.Takadaet al.,Biochem.Biophys.Res.Comm.176,860(1991),K.Ohnaka et al.,Biochem.Biophys.Res.Comm.168,1128(1990)描述的方法,且与内皮素标准品比较鉴定上清液并对其进行定量测定。
酶活性可如此计算:
此方法用于在对FBHE细胞的纯化的不同阶段测定ECE活性。与此同时,蛋白质的量由Bradford法(Anal.Biochem.72,248(1976))测定。
实施例4
从FBHE牛内皮细胞中纯化ECE-1
a)获得膜,外源蛋白的胰蛋白酶消化
将6ml FBHE细胞沉淀(得自实施例1)在25ml PBS缓冲液中水消化且冰浴超声破碎15分钟。用10,000g离心20分钟去除细胞碎片。边搅动边向上清液中加入3.6mg胰蛋白酶,室温下搅动混合物1.5小时。以100,000g将样品离心1小时,用25ml 100mM Tris缓冲液(pH8.0)将残余物中的膜悬浮。
b)ECE-1的增溶
将25ml膜悬浮液调至0.5mM二异丙基氟磷酸,然后再调至0.5%Triton X-100,再贮于冰浴中过夜。
c)Nono-Q层析
用50mM Tris缓冲液(pH8.0),0.05%Triton X-100(A缓冲液)平衡Mono-QFPLC柱,HRl6/10(Pharmacia)。
将增溶物上柱,然后以缓冲液A洗30分钟,随后以线性梯度转换成缓冲液B(缓冲液A+1M NaCl)进行洗脱100分钟。收集30份10ml的级分。用Brodford法测定样本的蛋白含量,并按实施例3的方法测定ECE-1活性。
将两个最高特异活性的级分合并。
d)凝胶滤过
通过30KDa Centricon膜(Amicon)将得自4c)将Mono-Q洗脱液合并物浓缩至500μl。
以20mM磷酸钠缓冲液(pH7.4)、250mM NaCl、0.05%Triton X-100平衡Superose 12HR 10/30柱(Pharmacia),将浓缩样品上柱。以流速0.5ml[sic]的平衡缓冲液进行层析,收集0.5ml的各级分。
各级分的比活如4c)中进行测定。有最高比活的级分在e)中示出,在相同的层析条件下用标准蛋白质进行校准,该级分于250,000Dalton处洗脱。
e)结果:
纯化步骤 | 蛋白质[mg] | 比活[μU/mg] |
膜 | 87 | 410 |
溶解物 | 87 | 360 |
Mono Q洗脱液 | 6.6 | 3760 |
Superose 12洗脱液 | 0.35 | 15100 |
实施例5
对自FBHE细胞纯化的ECE-1进行的SDS凝胶分析
在非还原和还原条件(+DTT)下,以Lmmli(Nature 227,680(1979))的方法,8%SDS凝胶上对实施例4中纯化的ECE-1进行分析。结果表明在非还原状态下ECE-1分子量为250,000Dalton,还原后分解成约125,000Dalton的宽带。
实施例6
ECE-1的去糖基化
以10%强SDS溶液将实施例4中纯化的50μl ECE溶液调至SDS含量为0.25%。室温下温育样品20分钟且调至1%Triton X-100。室温下20分钟后,加入5μl 250mM磷酸钠缓冲液(pH4.7)。再加入0.25μl PNG酶和0.5μgEndo F。37℃温育样本8小时。然后再加入等量的酶,于37℃继续温育8小时。浓缩样品至50μl,于4-12%SDS凝胶中分析。
结果:
在还原状态,由PNG酶F/Endo F混合物消除了糖残基,使125,000Dalton的表观分子量改变至约82,000Dalton。
实施例7
牛内皮细胞ECE-1的部分序列
将实施例4中所得的16×25μg ECE-1溶液上样在制备型的4-12%SDS聚丙烯胺梯度凝胶上。以常用的考马斯蓝染色后,在纯水中将凝胶脱色和水合4×40分钟。用刀片切下250KDa处染色的条带并以200μl 100mM NH4HCO3溶液消化。加入0.5μg胰蛋白酶并将混合物温育过夜,去除上清液。室温下将残留物再与200μl 100mM NH4HCO3缓冲液中的0.5μg胰蛋白酶温育5小时。然后在变速真空浓缩器中将样品浓缩干燥。将残留物置入40μl水中并与4μl40mM二硫苏糖醇搅动30分钟。然后于37℃加入4μl 100mM碘乙酰胺溶液。2小时后,将样品浓缩至干燥。
将所得肽3小时内在溶于水之0.5%三氟乙酸线性梯度变化至溶于90%乙腈之0.5%三氟乙酸的1min1×15cm反相HPLC柱上分馏。收集UV活性级分,在气相测序仪上进行初步测序。
结果:
测出下列部分序列:SEQ ID NO: 1:Xaa-Xaa-Pro-Asn-Ala-Leu-Asn-Phe-Gly-Gly-Ile-Gly-Val-Val-Val-Gly-His-Glu-Leu-Thr-His-Ala-Phe...SEQ ID NO: 2:Xaa-Tyr-Xaa-Lys-Xaa-Gly-Asn-Leu-Arg-ProSEQ ID NO: 3:Xaa-Ile-Ala-Xaa-Glu-Thr-Glu-Leu-Glu-Ile...
(Ile)(Ile)SEQ ID NO: 4:Xaa-Pro-Glu-Phe-Leu-Leu-Glu-Gly-Leu-Ile-Thr-Asp-Pro...SEQ ID NO: 5:Xaa-(Gln)-(Ala)-(Glu)-Asn-Val-Ile-Gln-Val-Xaa-Gln...SEQ ID NO: 6:
Val-Glu-Ile-Val-Phe-Pro-Ala-Gly-Ile-Leu-Gln-Ala-Pro-(Phe)-Tyr-Thr
(Thr)
括号内示出的氨基酸没有绝对的确定性。SEQ ID NO:1证明ECE-1是来自金属蛋白酶族的一种新蛋白质,因为与金属内肽酶NEP 24.11和嗜热菌蛋白酶的序列相比,它显示出显著的同源性,分别为72%和40%。
实施例8
编码内皮素转化酶的cDNA的制备
a)分离RNA并制备cDNA库
通过在硫氰酸胍中破坏如实施例1中由phosphoramidon刺激的FBHE细胞或如实施例1中由phosphoramidon刺激的Hela细胞获得完整的RNA。这一过程是按照RNA分离试剂盒(Stratagene,La Jolla,CA,USA(Catalog No.200345))的说明,利用其试剂进行的。
用oligo(dT)亲和分离法从FBHE细胞的上述完整RNA中选出多腺苷信使RNA。此方法是按照PolyATtract mRNA Isolation System(Promega,Madison,WI,USA(Catalog No.25200))的说明,利用其试剂进行的。按照ZAP-cDNA合成试剂盒(Stratagene,La Jolla cA,USA(catalog No.200400))的说明,利用其试剂自多腺苷信使RNA中合成cDNA,并按照Uni-ZAP XR Gigapack II克隆试剂盒(Stratagene,La Jolla,cA,USA(catalogNo.237611))的说明,利用其试剂将cDNA装入lambda噬菌体中。
b)用于RACE PCR的寡核苷酸探针的制备
实施例7中所示的含SEQ ID NO:1和NO:6的肽为通过聚合酶链反应(PCR,参见Molecular Cloning,2nd Edition(1989),Sambrook,J.et al.,CSH Press,page 14.1 et seq.)的cDAN片段克隆之起始点。
基于遗传密码,从SEQ ID NO:1第16-22位推出含核酸序列SEQ IDNO:7:5’-GGSCAYGARYTNACNCAYGC-3’的寡核苷酸混合物是可能的。
同样地从SEQ ID NO:6第2-8位推出含SEQ ID NO:8:5’-GARATYGTSTTYCCYGCYGG-3’的寡核苷酸混合物以及从SEQ ID NO:6第9-16位推出含SEQ ID NO:9:5’-ATYCTSCAGGCYCCYTTYTAYAC-3’的寡核苷酸混合物是可能的。
在这种情况下,SEQ ID NO:7至NO:9相应于编码DNA链的序列。由于已知的遗传密码的简并性,在一些位点已***几个核苷酸。这导致SEQ ID NO:7至9的混合物复杂性增至512倍。合成所说的序列作为寡核苷酸。
下列寡核苷酸另外合成作为RACE PCR中的3’引物:
A-B-T18(SEQ ID NO:10):
5′-CGAGGGGGATGGTCGACGGAAGCGACCTTTTTTTTTTTTTTTTTT-3′;
和
A(from A-B-T18)(SEQ ID NO:11):
5′-CGAGGGGGATGGTCGACGG-3′;
和
B(from A-B-T18)(SEQ ID NO:12):
5′-GATGGTCGACGGAAGCGACC-3′.
用Applied Biosystems Type 360A DNA合成仪进行所说合成。在去除保护性基团后,用丙烯酰胺/尿素凝胶上进行的凝胶电泳纯化寡核苷酸。
c)进行PCR的DNA模板的制备
用1μg寡核苷酸A-B-T18(SEQ ID NO:10)及逆转录酶将a)中所提到的来自phosphoramidon刺激之FBHE细胞的5μg总RNA或1μg RNA制备物的poly(A)+RNA翻译成至单链cDNA(sscDNA)。按照SuperScriptPreamplification System(Gibco BRL Eggenstein Germany(Catalog No.8089SA))的说明,使用其试剂进行此步骤。反应完成后,用GenedeanII试剂盒(BIO101,La Jolla,CA,USA)将合成产物纯化以去除较小的分子和过量的寡核苷酸。
d)PCRs和部分cDNA序列的克隆
用已知的方案(参见Molecular Cloning,.,2nd Edition(1989),Sambrook,J.et al.,CSH Press,Page 14.1 et seq.)进行聚合酶链反应。所用仪器为DNA热循环仪(Perkin Elmer)。也应用了Frohmarm,M.A et al的“internalprimer”原理(Proc.Natl Acad Sci.USA(1988)85,8998-9002)和修改的Fritz,J.D.et al方法(Nucl.Acids.Res.(1991)19,3747)。
特别地在各种情形下以20pmol的寡核苷酸SEQ ID NO:8和A(自A-B-T18)扩增得自c)的sscDNA。条件是:95℃1秒;55℃2秒;72℃3秒;共35个循环。
将PCR产物在1.2%LMP琼脂糖/TBE凝胶上分离。
在整个“拖尾”的长度上从凝胶上切下约10个凝胶盘,并这些凝胶盘熔化成含不断增加分子量的DNA片段的独立级分。
取出部分级分再分别进行第二次PCR,各使用了20pmol寡核苷酸SEQ IDNO:9和B(自A-B-T18)。此过程中琼脂糖含量从未超过PCR混合物体积的1/10。反应条件:95℃1秒;50℃2秒;72℃3秒,共35个循环。
将这些级分的扩增产物进行凝胶电泳结果显示将第一次PCR复杂的产物范围减少至第二次PCR之后的长度为约1000bp的较为限定的条带。
用标准方法洗脱以此方式选出的PCR产物。首先用另一次,也是第三次PCR扩增检验该大小为1000bp的条带与牛内皮素转化酶cDNA的一致性,其中使用了寡核苷酸SEQ ID NO:7和B(自A-B-T18)。该PCR产物为大小约950bp的条带。
在将第二次PCR反应得到的大小为1000bp的条带亚克隆至载体pCRII(TACloning Kit,Invitrogen Corp.,San Diego Cat.No.K2000-01)中并在E.coliDH5alpha中复制质粒后,克隆的序列分析显示-189个氨基酸SEQ ID NO:13、NO:14的开放阅读框架。肽SEQ ID NO:1、NO:2和NO:4的序列也均位于同一阅读框架内,这毫不含糊地确定了cDNA片段的同一性。e)牛内皮素转化酶cDNA的克隆
利用实施例1a)中详述的方案在合成第一链的步骤中用含随机引物混合物产生cDNA文库。合成的引物序列为:
5’-GAGAGAGAGTCGACGGTACCN7;SEQ ID NO:15
与上述cDNA合成方案不同的是,将双链cDNA在2nd Sephacryl S-1000(级:超纯,Pharmacia,Freiburg;Cat No:17-0476-01)柱上进行级分。以常规方法用乙醇沉淀法浓缩含大小至少为1kb cDNA片段的级分,随后按照cDNA合成试剂盒的说明进行处理,以限制性内切酶SalI替代XhoI裂解cDNA制备物并参入lambila载体中。
在转移至尼龙膜上后,用-DNA探针与得自文库的2×106克隆杂交,该探针是用来自部分牛cDNA(SEQ ID NO:13)分别为第136-156位和391-412位的寡核苷酸,5’-CGGCCCTGGTGGAAGAACTCG-3’(SEQ IDNO:16)和5’-TGCGGACGGAACACCAGACCT-3’(SEQ ID NO:17)进行制备的。如实施例1f)所描述,在毛地黄毒苷-dUTP存在条件下于聚合酶链反应中标记DNA探针。在严格条件下进行杂交(参见实施例1f)[sic])在于0.1×SSC,60℃杂交后进行最后一次洗涤步骤,随后用免疫学方法检测结合的DNA探针。所选克隆的测序显示牛cDNA序列C60(SEQ ID NO:18)含有一连续的阅读框架(SEQ ID NO:19)。
然后如上所述,在同样条件下用-DNA探针与来自上述文库的2×106克隆杂交,该探针是用来自cDNA克隆C60(SEQ ID NO:18)分别为第500-522位和14-35位的寡核苷酸5’-GCCAGCGCCGTCTCAAAGTCCAG-3’(SEQ ID NO:20)和5’-TGGGGGACCTTCAGCAACCTCT-3’(SEOID NO:21)制备的。所选克隆的测序显示牛cDAN序列SEQ ID NO:22含有一708氨基酸(SEQ ID NO:23)的连续阅读框架。
将商售的在lambda gtll中之牛肺cDNA文库(Clontech Laboratories,Inc.,4030 Fabian Way;Palo Alto CA 94303-4607,USA;Catalog No.BL10066)5×108噬菌斑形成单位用10%聚乙二醇6000,1M NaCl沉淀浓缩,再悬浮于100μl双蒸水中并于70℃加热5分钟。与引物gtll fwd和5’-GGTGCTTGATGATGGCTTGGTTGT-3’(SEQ ID NO:26)一起将5μl此溶菌产物用于50μl标准PCR反应中(实施例8d)。引物gtll fwd相应于β-半乳糖苷酶基因(基因库:ECLACZ)的第2979-3003位,并且正好位于在用于构建该库的单一EcoRI整合位点之前的lambda gtll克隆载体中(Yonug,R.A.&Dowis R.W.(1985)Genetic Engineeing Ed.by Setlow,J.U.Hollander A.Plenum Press N.Y.29-41)。引物SEQ ID NO:26相应于SEQ ID NO:22的第280-303位。
进行40次循环以上的扩增。在3%低熔点Nu Sieve GTG琼脂糖凝胶(FMCBioProducts,191 Thomaston Street,Rockland,ME04841,USA;CatalogNo:50082)上级分PCR产物,且切下单个凝胶片段分出大小在400-600bp的产物。如实施例8d)中所述熔化凝胶盘,取1.5μl与引物gtll fwd和5’-AGATGGAGCTGGTCACCGAGATGC-3’(SEQ ID NO:27)一起用于50μl新的PCR反应中。引物SEQ ID NO:27相应于SEQ ID NO:22第127-150位。
在于质粒载体pUC18(Yanisch-Perron,D.,Vieira J.&Messing,J.(1985)Gene 33,103-119)中亚克隆后对所产生的PCR片段测序(SEQ IDNO:28)。
该序列以其175-324位核苷酸覆盖了SEQ ID NO:22第1-150位,且以5’方向扩展了174个核苷酸。所得完整序列(SEQ ID NO:29)编码754氨基酸的开放阅读框架(SEQ ID NO:30)。
f)人内皮素转化酶cDNA的克隆
由于已在人胎盘中检出内皮素-1,可以设想内皮素转化酶也在胎盘内表达。因此利用一标记的牛cDNA探针筛选人λgt-1胎盘cDNA文库。为制备标有毛地黄毒苷-dUTP的牛cDNA探针,将1ng(1ng/ml)的部分牛cDNA序列(SEQID NO:16)用作PCR反应中的模板(参见Molecular Cloning,2nd Edition(1989),Sambrook,J.et al.,CSH Press,page 14.1et seq.)正义寡核苷酸(SEQ ID NO:16)包括SEQ ID NO:13第136-156位核苷酸,而反义寡核苷酸(SEQ ID NO:17)包括SEQ ID NO:13第392-412位核苷酸。在DNA热循环仪(Perkin Elmer)中将下列循环进行35次:94℃2分钟,60℃ 2分钟及72℃ 3分钟随后将已标记的276bp牛cDNA探针在琼脂糖凝胶上纯化并煮沸变性。该探针用于在较低严格条件下筛选入胎盘λgt-11cDNA文库(Clontech,HL 10086)的约650,000个克隆。探针滤膜提取物的杂交在5×SSC、2%阻断剂(Bochringer Mamheim;No.1096176)、0.1%N-十二烷肌氨酸、0.02%SDS中于60℃过夜进行。于60℃用2×SSC,0.1%SDS洗涤滤膜2×5分钟,随后于60℃用0.5×SSC、0.1%SDS洗涤20分钟。按照Bochringer Mamheim方案(DIG核酸测定[sic]试剂盒No.1175041)进一步处理滤膜以鉴定结合的DIG探针。
分离含SEQ ID NO:24所示的cDNA序列的克隆。SEQ ID NO:24cDNA之第1-2109位核苷酸编码SEQ ID NO:25所示的氨基酸序列。该氨基酸序列相应于内皮素转化酶一级序列的主要部分,因为从分离的内皮素转化酶胰蛋白酶肽测序中发现SEQ ID NO:1至SEQ ID NO:6的肽在该序列中。也可能如氨基酸序列SEQ ID NO:25第540-544位的HELTH一样鉴定金属蛋白酶的共有序列HEXXH。
如实施例8e)所述用寡核苷酸5’-GAGAGAGAGAGAGAGAGAGAACTAGTCTCGAGCCAAGCAGGCCACCAGTCCTG-3’(SEQ ID NO:31)作为第一链cDNA合成引物,从3μg胎盘polyA(+)RNA中制得cDNA文库。核苷酸32-53相应于SEQ ID NO:24的32-53位。如实施例8a)所述,将cDNA制备物整合入lambda载体Uni-ZAPXR中。扩增所得的4×105[sic]独立的克隆,且如实施例8e)所述,将5×108[sic]噬菌斑形成单位用于100μl PCR反应中。采用引物SEQ ID NO:31和C用于此目的。引物C位于lambda载体的Bluescript SK(-)部分,Pos.881-904,序列文档基因库:ARBLSKM。
在杂交温度为65℃情况下,进行40个循环的PCR反应。将1μl反应产物加至新的50μl PCR反应混合物中,于杂交温度60℃条件下进行40个循环。所用的引物为寡核苷酸D和5’-CCTGCCGCCAGAAGTACCACCAACA-3’(SEQID NO:32)。
引物D位于lambda Uni-ZAP XR载体(Pos.857-880,序列文档基因库:ARBLSKM)的Bluescript SK(-)部分,且引物SEQ ID NO:32相应于SEQID NO:24第11-35位。如上所述,将反应产物在质粒载体pUC18中亚克隆。将选出的克隆测序。
所得序列为人ECE cDNA的新5’部分(SEQ ID NO:33,其3’末端区(Pos.188-222)与SEQ ID NO:24第1-35位一样)。它以5’方向扩展了187个核苷酸,且提供了编码753个氨基酸(SEQ ID NO:36)的完整的人ECE序列SEQ ID NO:35。
实施例9
哺乳动物细胞内重组ECE的制备
1.膜结合人ECE表达载体的构建
为表达完整的人ECE,将核苷酸第29至2720位的cDNA序列(SEQ ID NO:35)以合适的转接体***于表达载体pcDNA3neo(Invitrogen,3985 B SorentoValley Blvd,San Diego,CA 92121,USA;product No.V 790-20)的KphI和XbaI限制位点间。该ECE序列向其自身提供了翻译起始和终止序列以及共有Kozak序列(Kozak,M.(1989)J.Cell Biol.108,229-241)。在该载体中,ECE信使RNA的转录处在强巨细胞病毒启动子调控之下。通过位于质粒中并仅允许G418抗性菌落生长的新霉素抗性基因进行转染细胞的筛选。
2.分泌入ECE的表达***之构建
a)在商售的真核细胞表达载体pcDNA3neo(lnvitrogen 3985 B SorrentoValley Blvd.,San Diego.CA 92121,USA;product No.V790-20)进行克隆。为达到此目的,首先将该载体用限制性酶EcoRI和EcoRV裂解。
b)然后通过用适宜的寡核苷酸引物进行的聚合酶链反应获得人ECE cDNA第241-2396位的核酸片段(SEQ ID NO:35)。由于5’引物序列的选择,第241-245位(SEQ ID NO:35)的核酸序列改变为5’-ACGCG-3’的碱基序列。通过包括246位,该步骤产生限制核酸内切酶MluI的识别序列。在进一步的步骤中,用MluI裂解所产生的ECE片段。
c)从2个合成寡核苷酸(链/互补链)制备已公开序列(D.Collen,Nature 301,214-221(1983))之核苷酸第74位和176位之间的人组织纤维蛋白溶酶原激活物(t-PA)的编码序列作为进一步的片段。除了所说的序列外,与合成的寡核苷酸提供可引起5’EcoRI和3’MluI突出的转接体。
通过共同的MluI限制性裂解位点将b)和c)产生的片段连接,会导致入t-PA基因信号肽及人ECE基因胞外功能域的阅读框架之融合。将该融合产物连接至得自a)并已被EcoRI和EcoRV割切的pcDNA3载体上,会导致分泌人ECE之表达***的产生。
3.哺乳动物细胞内的表达
用Lipofectamine(Gibco,Life Technologies GmbH;Dieselstra β e 5,76334 Eggensteine,Germany;No.530-8324SA)将表达载体的DNA转染至哺乳动物细胞内。细胞用CHO-K1(ATCC CCL 61);BHK-21(ATCCCCL 10);293(ATCC CRL 1573)和C127I(ATCC CRL 1616)。
在每种情况下将2×105细胞导入6孔培养板各孔的3ml生长培养基中。第二天进行转染。为达到这一目的,将细胞以PBS冲洗一次。按照生产者Gibco提供的信息进行用Lipofectamine的转染(Focus(1993),15 No.3,73-78)。各孔中加入1μg DNA和6μl Lipofectamine,将其一起加到1000μl无血清细胞培养基中。37℃温育6小时后,以PBS冲洗细胞一次并与正常生长培养基一起温育过夜。第二天,用胰蛋白酶松解孔中的细胞并分至1、5或10个Petri皿(10cm直径)中。下一天通过以G418(Gibco,Cat No.066-01811Y;商标名:Geneticin)处理来筛选转染的细胞,即以含1.2mg/ml G418的培养基取代原生长培养基。7-10天后,Petri皿中可见G418抗性细胞集落。用克隆滚筒法(cloningcylinder)分离这些集落(DNA Cloning Vol.II;Ed D.M.Glover IRLPress,1985;page 200)。分离的集落各置于24孔皿中的各孔中,每孔中含约1.5ml的生长培养基(包括G418)。当达到汇合时,以胰蛋白酶处理将细胞转至更大的培养器中。
a)膜结合ECE
如实施例1和2所述进行细胞破碎及级分后,检测表达膜结合ECE细胞是否有ECE的存在。(无phosphoramidon刺激进行)。所分析集落的比活高至每毫克基于膜的蛋白质为1550U。集落38的膜进一步处理。结果如下。
纯化步骤 | 比活(μU/mg蛋白质) |
膜 | 1550 |
增溶物 | 2010 |
Nono-Q层析 | 12000 |
b)分泌的ECE
通过检测汇合细胞细胞培养上清液的ECE活性对无膜锚点表达ECE并分泌至细胞培养基中的细胞进行测定确定重组ECE的存在。为达到该目的,培养两天后去除细胞上清液,且用1000g离心去除细胞碎屑。以Centricon 10,000(Amicon,W.R.Grace+CO.,Dauver,Ma.01923,USA product No.4206)并以3220g(约30分钟)离心浓缩6ml上清液。保存流出的含低分子物质的液体(Centricon洗脱液),用1ml PBS+10mM Tris+150mM NaCl(pH 7.5)冲洗Centricon-次。将浓缩液在300μl Centricon洗脱液中调整。
如实施例3中所述,将18μl的浓缩液用于ECE测定。
所分泌的ECE其单位体积的活性可至每毫升10单位,这依赖于所测定重组细胞之集落。
序列表(1)一般信息
(i)申请人:
(A)姓名:BASF Aktiengesellschaft
(B)街道:Carl-Bosch-Strasse 38
(C)城市:Ludwigshafen
(E)国家:Federal Republic of Germany
(F)邮编:D-67056
(G)电话:0621/6048526
(H)传真:0621/6043123
(I)电传:1762175170
(ii)申请名称:内皮素转化酶(ECE)
(iii)序列数:25
(iv)计算机可读形式:
(A)介质类型:软盘
(B)计算机:IBM PC兼容
(C)操作***:PC-DOS/MS-DOS
(D)软件:PatentIn Release#1.0,Version#1.25(EPA)(2)SEQ ID NO:1的信息:
(i)序列特征:
(A)长度:23个氨基酸
(B)类型:氨基酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:肽
(iii)假设序列:不是
(v)片段类型:中间片段
(vi)来源:
(A)生物名:Bos taurus
(F)组织类型:内皮
(H)细胞系:胎牛心内皮细胞系
(xi)序列描述:SEQ ID NO:1:Xaa Xaa Pro Asn Ala Leu Asn Phe Gly Gly Ile Gly Val Val Val Gly1 5 10 15His Glu Leu Thr His Ala Phe
20(2)SEQ ID NO:2的信息:
(i)序列特征:
(A)长度:10个氨基酸
(B)类型:氨基酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:肽
(v)片段类型:中间片段
(vi)来源:
(A)生物名:Bos taurus
(H)细胞系:胎牛心内皮细胞系
(xi)序列描述:SEQ ID NO:2:
Xaa Tyr Xaa Lys Xaa Gly Asn Leu Arg Pro
1 5 10(2)SEQ ID NO:3的信息:
(i)序列特征:
(A)长度:10个氨基酸
(B)类型:氨基酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:肽
(iii)假设序列:不是
(vi)来源:
(A)生物名:Bos taurus
(H)细胞系:胎牛心内皮细胞系
(ix)序列特征:
(A)名称/关键:修饰位点
(B)位置:8
(D)其它信息:/注:“Xaa=Leu or Ile”
(ix)序列特征:
(A)名称/关键:修饰位点
(B)位置:7
(D)其它信息:/注:=“Xaa=Glu or Ile’
(xi)序列描述:SEQ ID NO:3
Xaa Ile Ala Xaa Glu Thr Xaa Xaa Glu Ile
1 5 102)SEQ ID NO:4的信息:
(i)序列特征:
(A)长度:13个氨基酸
(B)类型:氨基酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:肽
(v)片段类型:中间片段
(vi)来源:
(A)生物名:Bos taurus
(H)细胞系:胎牛心内皮细胞系
(xi)序列描述:SEQ ID NO:4.
Xaa Pro Glu Phe Leu Leu Glu Gly Leu Ile Thr Asp Pro
5 10
(2)SEQ ID NO:5的信息:
(i)序列特征:
(A)长度:11个氨基酸
(B)类型:氨基酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:肽
(v)片段类型:中间片段
(vi)来源:
(A)生物名:Bos taurus
(H)细胞系:胎牛心内皮细胞系
(xi)序列描述:SEQ ID NO:5:
Xaa Gln Ala Glu Asn Val Ile Gln Val Xaa Gln
1 5 10(2)SEQ ID NO:6的信息:
(i)序列特征:
(A)长度:16个氨基酸
(B)类型:氨基酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:肽
(v)片段类型:中间片段
(vi)来源:
(A)生物名:Bos taurus
(H)细胞系:胎牛心内皮细胞系
(ix)序列特征:
(A)名称/关键:修饰位点
(B)位置:7
(D)其它信息:/注:“Xaa=Ala or Thr”
(xi)序列描述:SEQ ID NO:6:Val Glu Ile Val Phe Pro Xaa Gly Ile Leu Gln Ala Pro Phe Tyr Thr1 5 10 15(2)SEQ ID NO:7的信息:
(i)序列特征:
(A)长度:20bp
(B)类型:核苷酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(xi)序列描述:SEQ ID NO:7:GGSCAYGARY TNACNCAYGC 20(2)SEQ ID NO:8的信息:
(i)序列特征:
(A)长度:20bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(xi)序列描述:SEQ ID NO:8:GARATYGTST TYCCYGCYGG(2)SEQ ID NO:9的信息:
(i)序列特征:
(A)长度:23bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(xi)序列描述:SEQ ID NO:9:ATYCTSCAGG CYCCYTTYTA YAC 23(2)SEQ ID NO:10的信息:
(i)序列特征:
(A)长度:45bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(xi)序列描述:SEQID NO:10:CGAGGGGGAT GGTCGACGGA AGCGACCTTT TTTTTTTTTT TTTTT 45(2)SEQ ID NO:11的信息:
(i)序列特征:
(A)长度:19bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(xi)序列描述:SEQ ID NO:11:CGAGGGGGAT GGTCGACGG - 19(2)SEQ ID NO:12的信息:
(i)序列特征:
(A)长度:20bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(xi)序列描述:SEQ ID NO:12:GATGGTCGAC GGAAGCGACC 20(2)SEQ ID NO:13的信息:
(i)序列特征:
(A)长度:570bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:mRNA的cDNA(cDNA for mRNA)
(vi)来源:
(A)生物名:Bos taurus
(H)细胞系:胎牛心内皮细胞系
(ix)序列特征:
(A)名称/关键:CDS
(B)位置:1..567
(xi)序列描述:SEQ ID NO:13:ATC CTG CAG GCG CCA TTC TAC ACC CGC TCT TCA CCC AAT GCC TTA AAC 48Ile Leu Gln Ala Pro Phe Tyr Thr Arg Ser Ser Pro Asn Ala Leu Asn1 5 10 15TTC GGC GGC ATC GGC GTC GTC GTG GGC CAC GAG CTG ACT CAT GCT TTT 96Phe Gly Gly Ile Gly Val Val Val Gly His Glu Leu Thr His Ala Phe
20 25 30GAT GAT CAA GGC CGA GAG TAC GAC AAG GAT GGG AAC CTC CGG CCC TGG 144Asp Asp Gln Gly Arg Glu Tyr Asp Lys Asp Gly Asn Leu Arg Pro Trp
35 40 45TGG AAG AAC TCG TCC GTG GAG GCG TTC AAG CAG CAG ACC GCG TGC ATG 192Trp Lys Asn Ser Ser Val Glu Ala Phe Lys Gln Gln Thr Ala Cys Met
50 55 60GTG GAG CAG TAC GGC AAC TAT AGC GTG AAC GGG GAG CCG GTG AAC GGC 240Val Glu Gln Tyr Gly Asn Tyr Ser Val Asn Gly Glu Pro Val Asn Gly65 70 75 80CGG CAC ACC CTC GGC GAA AAC ATC GCC GAC AAC GGG GGC CTC AAG GCG 288Arg His Thr Leu Gly Glu Asn Ile Ala Asp Asn Gly Gly Leu Lys Ala
85 90 95GCC TAT CGG GCC TAC CAG AAC TGG GTC AAG AAG AAT GGG GCT GAG CAG 336Ala Tyr Arg Ala Tyr Gln Asn Trp Val Lys Lys Asn Gly Ala Glu Gln
100 105 110ACA CTG CCC ACC CTG GGT CTC ACC AAC AAC CAG CTC TTC TTC CTG AGT 384Thr Leu Pro Thr Leu Gly Leu Thr Asn Asn Gln Leu Phe Phe Leu Ser
115 120 125TTT GCA CAG GTC TGG TGT TCC GTC CGC ACC CCC GAG AGT TCG CAC GAA 432Phe Ala Gln Val Trp Cys Ser Val Arg Thr Pro Glu Ser Ser His Glu
130 135 140GGT CTC ATC ACC GAT CCC CAC AGC CCC TCC CGC TTC CGG GTC ATC GGC 480Gly Leu Ile Thr Asp Pro His Ser Pro Ser Arg Phe Arg Val Ile Gly145 150 155 160TCC ATC TCC AAC TCC AAG GAG TTC TCG GAA CAC TTC CAC TGC CCG CCC 528Ser Iie Ser Asn Ser Lys Glu Phe Ser Glu His Phe His Cys Pro Pro
165 170 175GGC TCA CCC ATG AAC CCG CAT CAC AAG TGT GAA GTC TGG TGA 570Gly Ser Pro Met Asn Pro His His Lys Cys Glu Val Trp
180 185(2)SEQ ID NO:14的信息:
(i)序列特征:
(A)长度:189个氨基酸
(B)类型:氨基酸
(D)拓扑学:线性
(ii)分子类型:蛋白质:
(xi)序列描述:SEQ ID NO:14:Ile Leu Gln Ala Pro Phe Tyr Thr Arg Ser Ser Pro Asn Ala Leu Asn1 5 10 15Phe Gly Gly Ile Gly Val Val Val Gly His Glu Leu Thr His Ala Phe
20 25 30Asp Asp Gln Gly Arg Glu Tyr Asp Lys Asp Gly Asn Leu Arg Pro Trp
35 40 45Trp Lys Asn Ser Ser Val Glu Ala Phe Lys Gln Gln Thr Ala Cys Met
50 55 60Val Glu Gln Tyr Gly Asn Tyr Ser Val Asn Gly Glu Pro Val Asn Gly65 70 75 80Arg His Thr Leu Gly Glu Asn Ile Ala Asp Asn Gly Gly Leu Lys Ala
85 90 95Ala Tyr Arg Ala Tyr Gln Asn Trp Val Lys Lys Asn Gly Ala Glu Gln
100 105 110Thr Leu Pro Thr Leu Gly Leu Thr Asn Asn Gln Leu Phe Phe Leu Ser
115 120 125Phe Ala Gln Val Trp Cys Ser Val Arg Thr Pro Glu Ser Ser His Glu
130 135 140Gly Leu Ile Thr Asp Pro His Ser Pro Ser Arg Phe Arg Val Ile Gly145 150 155 160Ser Ile Ser Asn Ser Lys Glu Phe Ser Glu His Phe His Cys Pro Pro
165 170 175Gly Ser Pro Met Asn Pro His His Lys Cys Glu Val Trp
180 185(2)SEQ ID NO:15的信息:
(i)序列特征:
(A)长度:27bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(iii)假设序列:是
(xi)序列描述:SEQ ID NO:15:GAGAGAGAGT CGACGGTACC NNNNNNN 27(2)SEQ ID NO:16的信息:
(i)序列特征:
(A)长度:21bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(xi)序列描述:SEQ ID NO:16:CGGCCCTGGT GGAAGAACTC G 21(2)SEQ ID NO:17的信息:
(i)序列特征:
(A)长度:21bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(iii)反义序列:是
(xi)序列描述:SEQ ID NO:17:
TGCGGACGGA ACACCAGACC T 21(2)SEQ ID NO:18的信息:
(i)序列特征:
(A)长度:1703bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:mRNA的cDNA
(vi)来源:
(A)生物名:Bos taurus
(H)细胞系:胎牛心内皮细胞系
(ix)序列特征:
(A)名称/关键:CDS
(B)位置:2..1703
(xi)序列描述:SEQ ID NO:18:T GGC CAC TCG CGC TGG GGG ACC TTC AGC AAC CTC TGG GAA CAC AAC 46Gly His Ser Arg Trp Gly Thr Phe Ser Asn Leu Trp Glu His Asn
1 5 l0 15CAA GCC ATC ATC AAG CAC CTC CTT GAA AAC TCC ACG GCC AGC GTG AGC 94Gln Ala Ile Ile Lys His Leu Leu Glu Asn Ser Thr Ala Ser Val Ser
20 25 30GAG GCA GAG AGG AAG GAC CAG GAG TAC TAC CGA GCC TGC ATG AAC GAA 142Glu Ala Glu Arg Lys Asp Gln Glu Tyr Tyr Arg Ala Cys Met Asn Glu
35 40 45ACC AGG ATT GAG GAG CTC AAG GCC AAA CCC CTG ATG GAG CTC ATT GAG 190Thr Arg Ile Glu Glu Leu Lys Ala Lys Pro Leu Met Glu Leu Ile Glu
50 55 60AAG CTC GGC GGC TGG AAC ATC ACG GGG CCC TGG GAC AAG GAC AAC TTC 238Lys Leu Gly Gly Trp Asn Ile Thr Gly Pro Trp Asp Lys Asp Asn Phe
65 70 75CAG GAC ACC CTG CAG GTG GTC ACA TCC CAC TAC CAC ACC TCC CCC TTC 286Gln Asp Thr Leu Gln Val Val Thr Ser His Tyr His Thr Ser Pro Phe80 85 90 95TTC TCC GTC TAC GTC AGT GCC GAC TCC AAG AAT TCC AAC AGC AAC GTG 334Phe Ser Val Tyr Val Ser Ala Asp Ser Lys Asn Ser Asn Ser Asn Val
l00 105 l10ATC CAA GTG GAC CAG TCT GGC CTG GGC TTA CCC TCA AGA GAT TAT TAC 382Ile Gln Val Asp Gln Ser Gly Leu Gly Leu Pro Ser Arg Asp Tyr Tyr
115 120 125CTG AAC AAA ACC GAG AAT GAG AAG GTG CTG ACG GGA TAC CTG AAC TAC 430Leu Asn Lys Thr Glu Asn Glu Lys Val Leu Thr Gly Tyr Leu Asn Tyr
130 135 140ATG GTC CAG CTG GGG AAG CTG CTG GGA GGA GGG GCC GAG GAC ACC ATC 478Met Val Gln Leu Gly Lys Leu Leu Gly Gly Gly Ala Glu Asp Thr Ile
145 150 155CGG CCC CAG ATG CAG CAG ATC CTG GAC TTT GAG ACG GCG CTG GCC AAC 526Arg Pro Gln Met Gln Gln Ile Leu Asp Phe Glu Thr Ala Leu Ala Asn160 165 170 175ATC ACC ATC CCC CAG GAG AAG CGC CGG GAC GAG GAA CTC ATC TAC CAC 574Ile Thr Ile Pro Gln Glu Lys Arg Arg Asp Glu Glu Leu Ile Tyr His
180 185 190AAA GTG ACG GCG GCT GAG TTG CAG ACC TTG GCG CCC GCC ATC AAC TGG 622Lys Val Thr Ala Ala Glu Leu Gln Thr Leu Ala Pro Ala Ile Asn Trp
195 200 205CTG CCC TTC CTC AAC ACC ATC TTC TAC CCC GTG GAG ATC AAT GAA TCA 670Leu Pro Phe Leu Asn Thr Ile Phe Tyr Pro Val Glu Ile Asn Glu Ser
210 215 220GAG CCT ATT GTC ATC TAC GAC AAA GAA TAC CTG AGC AAG GTC TCC ACC 718Glu Pro Ile Val Ile Tyr Asp Lys Glu Tyr Leu Ser Lys Val Ser Thr
225 230 235CTC ATC AAC AGC ACA GAC AAA TGC CTG CTG AAC AAC TAC ATG ATC TGG 766Leu Ile Asn Ser Thr Asp Lys Cys Leu Leu Asn Asn Tyr Met Ile Trp240 245 250 255AAC CTG GTA CGG AAG ACG AGC TCC TTC CTC GAT CAG CGC TTC CAG GAC 814Asn Leu Val Arg Lys Thr Ser Ser Phe Leu Asp Gln Arg Phe Gln Asp
260 265 270GCC GAC GAG AAG TTC ATG GAA ATC ATG TAT GGG ACC AAG AAG ACG TGT 862Ala Asp Glu Lys Phe Met Glu Val Met Tyr Gly Thr Lys Lys Thr Cys
275 280 285CTT CCC CGC TGG AAG TTT TGT GTG AGT GAT ACA GAG AAC ACC TTG GGC 910Leu Pro Arg Trp Lys Phe Cys Val Ser Asp Thr Glu Asn Thr Leu Gly
290 295 300
TTC GCC CTG GGC CCC ATG TTC GTC AAA GCG ACC TTC GCT GAG GAC AGC 958Phe Ala Leu Gly Pro Met Phe Val Lys Ala Thr Phe Ala Glu Asp Ser
305 310 315AAG AAC ATA GCC AGC GAG ATC ATC CTG GAG ATC AAG AAG GCG TTT GAA 1006Lys Asn Ile Ala Ser Glu Ile Ile Leu Glu Ile Lys Lys Ala Phe Glu320 325 330 335GAG AGC CTG AGC ACC CTG AAG TGG ATG GAT GAA GAT ACT CGG AAA TCG 1054Glu Ser Leu Ser Thr Leu Lys Trp Met Asp Glu Asp Thr Arg Lys Ser
340 345 350GCC AAG GAA AAG GCG GAC GCG ATC TAC AAC ATG ATA GGC TAC CCC AAC 1102Ala Lys Glu Lys Ala Asp Ala Ile Tyr Asn Met Ile Gly Tyr Pro Asn
355 360 365TTT ATC ATG GAC CCC AAG GAG CTG GAC AAA GTG TTC AAT GAC TAC ACC 1150Phe Ile Met Asp Pro Lys Glu Leu Asp Lys Val Phe Asn Asp Tyr Thr
370 375 380GCT GTG CCA GAC CTC TAC TTC GAG AAC GCC ATG CGG TTT TTC AAC TTC 1198Ala Val Pro Asp Leu Tyr Phe Glu Asn Ala Met Arg Phe Phe Asn Phe
385 390 395TCC TGG AGG GTC ACT GCC GAC CAG CTC CGG AAA GCG CCC AAC AGA GAT 1246Ser Trp Arg Val Thr Ala Asp Gln Leu Arg Lys Ala Pro Asn Arg Asp400 405 410 415CAG TGG AGC ATG ACC CCG CCC ATG GTG AAC GCC TAC TAC TCG CCC ACC 1294Gln Trp Ser Met Thr Pro Pro Met Val Asn Ala Tyr Tyr Ser Pro Thr
420 425 430AAG AAC GAG ATC GTG TTT CCG GCC GGA ATC CTG CAG GCG CCA TTC TAC 1342Lys Asn Glu Ile Val Phe Pro Ala Gly Ile Leu Gln Ala Pro Phe Tyr
435 440 445ACC CGC TCT TCA CCC AAT GCC TTA AAC TTC GGC GGC ATC GGC GTC GTC 1390Thr Arg Ser Ser Pro Asn Ala Leu Asn Phe Gly Gly Ile Gly Val Val
450 455 460GTG GGC CAC GAG CTG ACT CAT GCT TTT GAT GAT CAA GGC CGA GAG TAC 1438Val Gly His Glu Leu Thr His Ala Phe Asp Asp Gln Gly Arg Glu Tyr
465 470 475GAC AAG GAT GGG AAC CTC CGG CCC TGG TGG AAG AAC TCG TCC GTG GAG 1486Asp Lys Asp Gly Asn Leu Arg Pro Trp Trp Lys Asn Ser Ser Val Glu480 485 490 495GCG TTC AAG CAG CAG ACC GCG TGC ATG GTG GAG CAG TAC GGC AAC TAT 1534Ala Phe Lys Gln Gln Thr Ala Cys Met Val Glu Gln Tyr Gly Asn Tyr
500 505 510AGC GTG AAC GGG GAG CCG GTG AAC GGC CGG CAC ACC CTC GGC GAA AAC 1582Ser Val Asn Gly Glu Pro Val Asn Gly Arg His Thr Leu Gly Glu Asn
515 520 525ATC GCC GAC AAC GGG GGC CTC AAG GCG GCC TAT CGG GCC TAC CAG AAC 1630Ile Ala Asp Asn Gly Gly Leu Lys Ala Ala Tyr Arg Ala Tyr Gln Asn
530 535 540TGG GTC AAG AAG AAT GGG GCT GAG CAG ACA CTG CCC ACC CTG GGT CTC 1678Trp Val Lys Lys Asn Gly Ala Glu Gln Thr Leu Pro Thr Leu Gly Leu
545 550 555ACC AAC AAC CAG CTC TTC TTC CTG A 1703Thr Asn Asn Gln Leu Phe Phe Leu560 5652)SEQ ID NO:19的信息:
(i)序列特征:
(A)长度:567个氨基酸
(B)类型:氨基酸
(D)拓扑学:线性
(ii)分子类型:蛋白质
(xi)序列描述:SEQ ID NO:19:Gly His Ser Arg Trp Gly Thr Phe Ser Asn Leu Trp Glu His Asn Gln1 5 10 15Ala Ile Ile Lys His Leu Leu Glu Asn Ser Thr Ala Ser Val Ser Glu
20 25 30Ala Glu Arg Lys Asp Gln Glu Tyr Tyr Arg Ala Cys Met Asn Glu Thr
35 40 45Arg Ile Glu Glu Leu Lys Ala Lys Pro Leu Met Glu Leu Ile Glu Lys
50 55 60Leu Gly Gly Trp Asn Ile Thr Gly Pro Trp Asp Lys Asp Asn Phe Gln65 70 75 80Asp Thr Leu Gln Val Val Thr Ser His Tyr His Thr Ser Pro Phe Phe
85 90 95Ser Val Tyr Val Ser Ala Asp Ser Lys Asn Ser Asn Ser Asn Val Ile
100 105 110Gln Val Asp Gln Ser Gly Leu Gly Leu Pro Ser Arg Asp Tyr Tyr Leu
115 120 125Asn Lys Thr Glu Asn Glu Lys Val Leu Thr Gly Tyr Leu Asn Tyr Met
130 135 140Val Gln Leu Gly Lys Leu Leu Gly Gly Gly Ala Glu Asp Thr Ile Arg145 150 155 160Pro Gln Met Gln Gln Ile Leu Asp Phe Glu Thr Ala Leu Ala Asn Ile
165 170 175Thr Ile Pro Gln Glu Lys Arg Arg Asp Glu Glu Leu Ile Tyr His Lys
180 185 190Val Thr Ala Ala Glu Leu Gln Thr Leu Ala Pro Ala Ile Asn Trp Leu
195 200 205Pro Phe Leu Asn Thr Ile Phe Tyr Pro Val Glu Ile Asn Glu Ser Glu
210 215 220Pro Ile Val Ile Tyr Asp Lys Glu Tyr Leu Ser Lys Val Ser Thr Leu225 230 235 240Ile Asn Ser Thr Asp Lys Cys Leu Leu Asn Asn Tyr Met Ile Trp Asn
245 250 255Leu Val Arg Lys Thr Ser Ser Phe Leu Asp Gln Arg Phe Gln Asp Ala
260 265 270Asp Glu Lys Phe Met Glu Val Met Tyr Gly Thr Lys Lys Thr Cys Leu
275 280 285Pro Arg Trp Lys Phe Cys Val Ser Asp Thr Glu Asn Thr Leu Gly Phe
290 295 300Ala Leu Gly Pro Met Phe Val Lys Ala Thr Phe Ala Glu Asp Ser Lys305 310 315 320Asn Ile Ala Ser Glu Ile Ile Leu Glu Ile Lys Lys Ala Phe Glu Glu
325 330 335Ser Leu Ser Thr Leu Lys Trp Met Asp Glu Asp Thr Arg Lys Ser Ala
340 345 350Lys Glu Lys Ala Asp Ala Ile Tyr Asn Met Ile Gly Tyr Pro Asn Phe
355 360 365Ile Met Asp Pro Lys Glu Leu Asp Lys Val Phe Asn Asp Tyr Thr Ala
370 375 380Val Pro Asp Leu Tyr Phe Glu Asn Ala Met Arg Phe Phe Asn Phe Ser385 390 395 400Trp Arg Val Thr Ala Asp Gln Leu Arg Lys Ala Pro Asn Arg Asp Gln
405 410 415Trp Ser Met Thr Pro Pro Met Val Asn Ala Tyr Tyr Ser Pro Thr Lys
420 425 430Asn Glu Ile Val Phe Pro Ala Gly Ile Leu Gln Ala Pro Phe Tyr Thr
435 440 445Arg Ser Ser Pro Asn Ala Leu Asn Phe Gly Gly Ile Gly Val Val Val
450 455 460Gly His Glu Leu Thr His Ala Phe Asp Asp Gln Gly Arg Glu Tyr Asp465 470 475 480Lys Asp Gly Asn Leu Arg Pro Trp Trp Lys Asn Ser Ser Val Glu Ala
485 490 495Phe Lys Gln Gln Thr Ala Cys Met Val Glu Gln Tyr Gly Asn Tyr Ser
500 505 510Val Asn Gly Glu Pro Val Asn Gly Arg His Thr Leu Gly Glu Asn Ile
515 520 525Ala Asp Asn Gly Gly Leu Lys Ala Ala Tyr Arg Ala Tyr Gln Asn Trp
530 535 540Val Lys Lys Asn Gly Ala Glu Gln Thr Leu Pro Thr Leu Gly Leu Thr545 550 555 560Asn Asn Gln Leu Phe Phe Leu
565(2)SEQ ID NO:20的信息:
(i)序列特征:
(A)长度:23bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(iii)反义序列:是
(xi)序列描述:SEQ ID NO:20:GCCAGCGCCG TCTCAAAGTC CAG 23(2)SEQ ID NO:21的信息:
(i)序列特征:
(A)长度:22bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(iii)反义序列:不是
(xi)序列描述:SEQ ID NO:21:
TGGGGGACCT TCAGCAACCT CT 22(2)SEQ ID NO:22的信息:
(i)序列特征:
(A)长度:2129bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:mRNA的cDNA
(vi)来源:
(A)生物名:Bos taurus
(H)细胞系:胎牛心内皮细胞系
(ix)序列特征:
(A)名称/关键:CDS
(B)位置:3..2126
(xi)序列描述:SEQ ID NO:22:GG ACC CCG GTG GAG AAG CGG CTG GTG GTG CTG GTG GCG CTC CTG GCG 47Thr Pro Val Glu Lys Arg Leu Val Val Leu Val Ala Leu Leu Ala
1 5 l0 15GCG GCA TTG GTG GCC TGT TTG GCA GTA CTG GGC ATC CAA TAC CAG ACA 95Ala Ala Leu Val Ala Cys Leu Ala Val Leu Gly Ile Gln Tyr Gln Thr
20 25 30AGA ACG CCC TCG GTG TGC CTA AGT GAG GCC TGC ATC TCG GTG ACC AGC 143Arg Thr Pro Ser Val Cys Leu Ser Glu Ala Cys Ile Ser Val Thr Ser
35 40 45TCC ATC TTG AGT TCC ATG GAC CCC ACG GTG GAC CCC TGC CAG GAC TTC 191Ser Ile Leu Ser Ser Met Asp Pro Thr Val Asp Pro Cys Gln Asp Phe
50 55 60TTC ACC TAT GCC TGT GGC GGC TGG ATC AAA GCC AAC CCC GTG CCG GAT 239Phe Thr Tyr Ala Cys Gly Gly Trp Ile Lys Ala Asn Pro Val Pro Asp
65 70 75GGC CAC TCG CGC TGG GGG ACC TTC AGC AAC CTC TGG GAA CAC AAC CAA 287Gly His Ser Arg Trp Gly Thr Phe Ser Asn Leu Trp Glu His Asn Gln80 85 90 95GCC ATC ATC AAG CAC CTC CTT GAA AAC TCC ACG GCC AGC GTG AGC GAG 335Ala Ile Ile Lys His Leu Leu Glu Asn Ser Thr Ala Ser Val Ser Glu
100 105 110GCA GAG AGG AAG GAC CAG GAG TAC TAC CGA GCC TGC ATG AAC GAA ACC 383Ala Glu Arg Lys Asp Gln Glu Tyr Tyr Arg Ala Cys Met Asn Glu Thr
115 120 125AGG ATT GAG GAG CTC AAG GCC AAA CCC CTG ATG GAG CTC ATT GAG AAG 431Arg Ile Glu Glu Leu Lys Ala Lys Pro Leu Met Glu Leu Ile Glu Lys
130 135 140CTC GGC GGC TGG AAC ATC ACG GGG CCC TGG GAC AAG GAC AAC TTC CAG 479Leu Gly Gly Trp Asn Ile Thr Gly Pro Trp Asp Lys Asp Asn Phe Gln
145 150 155GAC ACC CTG CAG GTG GTC ACA TCC CAC TAC CAC ACC TCC CCC TTC TTC 527Asp Thr Leu Gln Val Val Thr Ser His Tyr His Thr Ser Pro Phe Phe160 165 170 175TCC GTC TAC GTC AGT GCC GAC TCC AAG AAT TCC AAC AGC AAC GTG ATC 575Ser Val Tyr Val Ser Ala Asp Ser Lys Asn Ser Asn Ser Asn Val Ile
180 185 190
CAA GTG GAC CAG TCT GGC CTG GGC TTA CCC TCA AGA GAT TAT TAC CTG 623Gln Val Asp Gln Ser Gly Leu Gly Leu Pro Ser Arg Asp Tyr Tyr Leu
195 200 205AAC AAA ACC GAG AAT GAG AAG GTG CTG ACG GGA TAC CTG AAC TAC ATG 671Asn Lys Thr Glu Asn Glu Lys Val Leu Thr Gly Tyr Leu Asn Tyr Met
210 215 220GTC CAG CTG GGG AAG CTG CTG GGA GGA GGG GCC GAG GAC ACC ATC CGG 719Val Gln Leu Gly Lys Leu Leu Gly Gly Gly Ala Glu Asp Thr Ile Arg
225 230 235
CCC CAG ATG CAG CAG ATC CTG GAC TTT GAG ACG GCG CTG GCC AAC ATC 767Pro Gln Met Gln Gln Ile Leu Asp Phe Glu Thr Ala Leu Ala Asn Ile240 245 250 255ACC ATC CCC CAG GAG AAG CGC CGG GAC GAG GAA CTC ATC TAC CAC AAA 815Thr Ile Pro Gln Glu Lys Arg Arg Asp Glu Glu Leu Ile Tyr His Lys
260 265 270GTG ACG GCG GCT GAG TTG CAG ACC TTG GCG CCC GCC ATC AAC TGG CTG 863Val Thr Ala Ala Glu Leu Gln Thr Leu Ala Pro Ala Ile Asn Trp Leu
275 280 285CCC TTC CTC AAC ACC ATC TTC TAC CCC GTG GAG ATC AAT GAA TCA GAG 911Pro Phe Leu Asn Thr Ile Phe Tyr Pro Val Glu Ile Asn Glu Ser Glu
290 295 300CCT ATT GTC ATC TAC GAC AAA GAA TAC CTG AGC AAG GTC TCC ACC CTC 959Pro Ile Val Ile Tyr Asp Lys Glu Tyr Leu Ser Lys Val Ser Thr Leu
305 310 315
ATC AAC AGC ACA GAC AAA TGC CTG CTG AAC AAC TAC ATG ATC TGG AAC 1007Ile Asn Ser Thr Asp Lys Cys Leu Leu Asn Asn Tyr Met Ile Trp Asn320 325 330 335CTG GTA CGG AAG ACG AGC TCC TTC CTC GAT CAG CGC TTC CAG GAC GCC 1055Leu Val Arg Lys Thr Ser Ser Phe Leu Asp Gln Arg Phe Gln Asp Ala
340 345 350GAC GAG AAG TTC ATG GAA GTC ATG TAT GGG ACC AAG AAG ACG TGT CTT 1103Asp Glu Lys Phe Met Glu Val Met Tyr Gly Thr Lys Lys Thr Cys Leu
355 360 365CCC CGC TGG AAG TTT TGT GTG AGT GAT ACA GAG AAC ACC TTG GGC TTC 1151Pro Arg Trp Lys Phe Cys Val Ser Asp Thr Glu Asn Thr Leu Gly Phe
370 375 380GCC CTG GGC CCC ATG TTC GTC AAA GCG ACC TTC GCT GAG GAC AGC AAG 1199Ala Leu Gly Pro Met Phe Val Lys Ala Thr Phe Ala Glu Asp Ser Lys
385 390 395AAC ATA GCC AGC GAG ATC ATC CTG GAG ATC AAG AAG GCG TTT GAA GAG 1247Asn Ile Ala Ser Glu Ile Ile Leu Glu Ile Lys Lys Ala Phe Glu Glu400 405 410 415AGC CTG AGC ACC CTG AAG TGG ATG GAT GAA GAT ACT CGG AAA TCG GCC 1295Ser Leu Ser Thr Leu Lys Trp Met Asp Glu Asp Thr Arg Lys Ser Ala
420 425 430AAG GAA AAG GCG GAC GCG ATC TAC AAC ATG ATA GGC TAC CCC AAC TTT 1343Lys Glu Lys Ala Asp Ala Ile Tyr Asn Met Ile Gly Tyr Pro Asn Phe
435 440 445ATC ATG GAC CCC AAG GAG CTG GAC AAA GTG TTC AAT GAC TAC ACC GCT 1391Ile Met Asp Pro Lys Glu Leu Asp Lys Val Phe Asn Asp Tyr Thr Ala
450 455 460GTG CCA GAC CTC TAC TTC GAG AAC GCC ATG CGG TTT TTC AAC TTC TCC 1439Val Pro Asp Leu Tyr Phe Glu Asn Ala Met Arg Phe Phe Asn Phe Ser
465 470 475TGG AGG GTC ACT GCC GAC CAG CTC CGG AAA GCG CCC AAC AGA GAT CAG 1487Trp Arg Val Thr Ala Asp Gln Leu Arg Lys Ala Pro Asn Arg Asp Gln480 485 490 495TGG AGC ATG ACC CCG ATG ATG GTG AAC GCC TAC TAC TCG CCC ACC AAG 1535Trp Ser Met Thr Pro Pro Met Val Asn Ala Tyr Tyr Ser Pro Thr Lys
500 505 510AAC GAG ATC GTG TTT CCG GCC GGA ATC CTG CAG GCG CCA TTC TAC ACC 1583Asn Glu Ile Val Phe Pro Ala Gly Ile Leu Gln Ala Pro Phe Tyr Thr
515 520 525CGC TCT TCA CCC AAT GCC TTA AAC TTG GGC GGC ATC GGC GTC GTC GTG 1631Arg Ser Ser Pro Asn Ala Leu Asn Phe Gly Gly Ile Gly Val Val Val
530 535 540GGC CAC GAG CTG ACT CAT GCT TTT GAT GAT CAA GGC CGA GAG TAC GAC 1679Gly His Glu Leu Thr His Ala Phe Asp Asp Gln Gly Arg Glu Tyr Asp
545 550 555AAG GAT GGG AAC CTC CGG CCC TGG TGG AAG AAC TCG TCC GTG GAG GCG 1727Lys Asp Gly Asn Leu Arg Pro Trp Trp Lys Asn Ser Ser Val Glu Ala560 565 570 575TTC AAG CAG CAG ACC GCG TGC ATG GTG GAG CAG TAC GGC AAC TAT AGC 1775Phe Lys Gln Gln Thr Ala Cys Met Val Glu Gln Tyr Gly Asn Tyr Ser
580 585 590GTG AAC GGG GAG CCG GTG AAC GGC CGG CAC ACC CTC GGC GAA AAC ATC 1823Val Asn Gly Glu Pro Val Asn Gly Arg His Thr Leu Gly Glu Asn Ile
595 600 605GCC GAC AAC GGG GGC CTC AAG GCG GCC TAT CGG GCC TAC CAG AAC TGG 1871Ala Asp Asn Gly Gly Leu Lys Ala Ala Tyr Arg Ala Tyr Gln Asn Trp
610 615 620GTC AAG AAG AAT GGG GCT GAG CAG ACA CTG CCC ACC CTG GGT CTC ACC 1919Val Lys Lys Asn Gly Ala Glu Gln Thr Leu Pro Thr Leu Gly Leu Thr
625 630 635AAC AAC CAG CTC TTC TTC CTG AGT TTT GCA CAG GTC TGG TGT TCC GTC 1967Asn Asn Gln Leu Phe Phe Leu Ser Phe Ala Gln Val Trp Cys Ser Val640 645 650 655CGC ACC CCC GAG AGT TCG CAC GAA GGT CTC ATC ACC GAT CCC CAC AGC 2015Arg Thr Pro Glu Ser Ser His Glu Gly Leu Ile Thr Asp Pro His Ser
660 665 670CCC TCC CGC TTC CGG GTC ATC GGC TCC ATC TCC AAC TCC AAG GAG TTC 2063Pro Ser Arg Phe Arg Val Ile Gly Ser Ile Ser Asn Ser Lys Glu Phe
675 680 685TCG GAA CAC TTC CAC TGC CCG CCC GGC TCA CCC ATG AAC CCG CAT CAC 2111Ser Glu His Phe His Cys Pro Pro Gly Ser Pro Met Asn Pro His His
690 695 700AAG TGT GAA GTC TGG TGA 2129Lys Cys Glu Val Trp
705(2)SEQ ID NO:23的信息:
(i)序列特征:
(A)长度:708个氨基酸
(B)类型:氨基酸
(D)拓扑学:线性
(ii)分子类型:蛋白质
(xi)序列描述:SEQ ID NO:23:Thr Pro Val Glu Lys Arg Leu Val Val Leu Val Ala Leu Leu Ala Ala1 5 10 15Ala Leu Val Ala Cys Leu Ala Val Leu Gly Ile Gln Tyr Gln Thr Arg
20 25 30Thr Pro Ser Val Cys Leu Ser Glu Ala Cys Ile Ser Val Thr Ser Ser
35 40 45Ile Leu Ser Ser Met Asp Pro Thr Val Asp Pro Cys Gln Asp Phe Phe50 55 60Thr Tyr Ala Cys Gly Gly Trp Ile Lys Ala Asn Pro Val Pro Asp Gly65 70 75 80His Ser Arg Trp Gly Thr Phe Ser Asn Leu Trp Glu His Asn Gln Ala
85 90 95Ile Ile Lys His Leu Leu Glu Asn Ser Thr Ala Ser Val Ser Glu Ala
100 105 110Glu Arg Lys Asp Gln Glu Tyr Tyr Arg Ala Cys Met Asn Glu Thr Arg
115 120 125Ile Glu Glu Leu Lys Ala Lys Pro Leu Met Glu Leu Ile Glu Lys Leu
130 135 140Gly Gly Trp Asn Ile Thr Gly Pro Trp Asp Lys Asp Asn Phe Gln Asp145 150 155 160Thr Leu Gln Val Val Thr Ser His Tyr His Thr Ser Pro Phe Phe Ser
165 170 175Val Tyr Val Ser Ala Asp Ser Lys Asn Ser Asn Ser Asn Val Ile Gln
180 185 190Val Asp Gln Ser Gly Leu Gly Leu Pro Ser Arg Asp Tyr Tyr Leu Asn
195 200 205Lys Thr Glu Asn Glu Lys Val Leu Thr Gly Tyr Leu Asn Tyr Met Val
210 215 220Gln Leu Gly Lys Leu Leu Gly Gly Gly Ala Glu Asp Thr Ile Arg Pro225 230 235 240Gln Met Gln Gln Ile Leu Asp Phe Glu Thr Ala Leu Ala Asn Ile Thr
245 250 255Ile Pro Gln Glu Lys Arg Arg Asp Glu Glu Leu Ile Tyr His Lys Val
260 265 270Thr Ala Ala Glu Leu Gln Thr Leu Ala Pro Ala Ile Asn Trp Leu Pro
275 280 285Phe Leu Asn Thr Ile Phe Tyr Pro Val Glu Ile Asn Glu Ser Glu Pro
290 295 300Ile Val Ile Tyr Asp Lys Glu Tyr Leu Ser Lys Val Ser Thr Leu Ile305 310 315 320Asn Ser Thr Asp Lys Cys Leu Leu Asn Asn Tyr Met Ile Trp Asn Leu
325 330 335Val Arg Lys Thr Ser Ser Phe Leu Asp Gln Arg Phe Gln Asp Ala Asp
340 345 350Glu Lys Phe Met Glu Val Met Tyr Gly Thr Lys Lys Thr Cys Leu Pro
355 360 365Arg Trp Lys Phe Cys Val Ser Asp Thr Glu Asn Thr Leu Gly Phe Ala
370 375 380Leu Gly Pro Met Phe Val Lys Ala Thr Phe Ala Glu Asp Ser Lys Asn385 390 395 400Ile Ala Ser Glu Ile Ile Leu Glu Ile Lys Lys Ala Phe Glu Glu Ser
405 410 415Leu Ser Thr Leu Lys Trp Met Asp Glu Asp Thr Arg Lys Ser Ala Lys
420 425 430Glu Lys Ala Asp Ala Ile Tyr Asn Met Ile Gly Tyr Pro Asn Phe Ile
435 440 445Met Aso Pro Lys Glu Leu Asp Lys Val Phe Asn Asp Tyr Thr Ala Val
450 455 460Pro Asp Leu Tyr Phe Glu Asn Ala Met Arg Phe Phe Asn Phe Ser Trp465 470 475 480Arg Val Thr Ala Asp Gln Leu Arg Lys Ala Pro Asn Arg Asp Gln Trp
485 490 495Ser Met Thr Pro Pro Met Val Asn Ala Tyr Tyr Ser Pro Thr Lys Asn
500 505 510Glu Ile Val Phe Pro Ala Gly Ile Leu Gln Ala Pro Phe Tyr Thr Arg
515 520 525Ser Ser Pro Asn Ala Leu Asn Phe Gly Gly Ile Gly Val Val Val Gly
530 535 540His Glu Leu Thr His Ala Phe Asp Asp Gln Gly Arg Glu Tyr Asp Lys545 550 555 560Asp Gly Asn Leu Arg Pro Trp Trp Lys Asn Ser Ser Val Glu Ala Phe
565 570 575Lys Gln Gln Thr Ala Cys Met Val Glu Gln Tyr Gly Asn Tyr Ser Val
580 585 590Asn Gly Glu Pro Val Asn Gly Arg His Thr Leu Gly Glu Asn Ile Ala
595 600 605Asp Asn Gly Gly Leu Lys Ala Ala Tyr Arg Ala Tyr Gln Asn Trp Val
610 615 620Lys Lys Asn Gly Ala Glu Gln Thr Leu Pro Thr Leu Gly Leu Thr Asn625 630 635 640Asn Gln Leu Phe Phe Leu Ser Phe Ala Gln Val Trp Cys Ser Val Arg
645 650 655Thr Pro Glu Ser Ser His Glu Gly Leu Ile Thr Asp Pro His Ser Pro
660 665 670Ser Arg Phe Arg Val Ile Gly Ser Ile Ser Asn Ser Lys Glu Phe Ser
675 680 685Glu His Phe His Cys Pro Pro Gly Ser Pro Met Asn Pro His His Lys
690 695 700Cys Glu Val Trp705(2)SEQ ID NO:24的信息:
(i)序列特征:
(A)长度:2533bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:mRNA的cDNA
(vi)来源:
(A)生物名:Homo sapiens
(H)组织类型:胎盘
(ix)序列特征:
(A)名称/关键:CDS
(B)位置:1..2109
(xi)序列描述:SEQ ID NO:24:CGG CTG GTG GTG TTG GTG GTA CTT CTG GCG GCA GGA CTG GTG GCC TGC 48Arg Leu Val Val Leu Val Val Leu Leu Ala Ala Gly Leu Val Ala Cys1 5 10 15TTG GCA GCA CTG GGC ATC CAG TAC CAG ACA AGA TCC CCC TCT GTG TGC 96Leu Ala Ala Leu Gly Ile Gln Tyr Gln Thr Arg Ser Pro Ser Val Cys
20 25 30CTG AGC GAA GCT TGT GTC TCA GTG ACC AGC TCC ATC TTG AGC TCC ATG 144Leu Ser Glu Ala Cys Val Ser Val Thr Ser Ser Ile Leu Ser Ser Met
35 40 45GAC CCC ACA GTG GAC CCC TGC CAT GAC TTC TTC AGC TAC GCC TGT GGG 192Asp Pro Thr Val Asp Pro Cys His Asp Phe Phe Ser Tyr Ala Cys Gly
50 55 60GGC TGG ATC AAG GCC AAC CCA GTC CCT GAT GGC CAC TCA CGC TGG GGG 240Gly Trp Ile Lys Ala Asn Pro Val Pro Asp Gly His Ser Arg Trp Gly65 70 75 80ACC TTC AGC AAC CTC TGG GAA CAC AAC CAA GCA ATC ATC AAG CAC CTC 288Thr Phe Ser Asn Leu Trp Glu His Asn Gln Ala Ile Ile Lys His Leu
85 90 95CTC GAA AAC TCC ACG GCC AGC GTG AGC GAG GCA GAG AGA AAG GCG CAA 336Leu Glu Asn Ser Thr Ala Ser Val Ser Glu Ala Glu Arg Lys Ala Gln
100 105 110GTA TAC TAC CGT GCG TGC ATG AAC GAG ACC AGG ATC GAG GAG CTC AGG 384Val Tyr Tyr Arg Ala Cys Met Asn Glu Thr Arg Ile Glu Glu Leu Arg
115 120 125GCC AAA CCT CTA ATG GAG TTG ATT GAG AGG CTC GGG GGC TGG AAC ATC 432Ala Lys Pro Leu Met Glu Leu Ile Glu Arg Leu Gly Gly Trp Asn Ile
130 135 140ACA GGT CCC TGG GCC AAG GAC AAC TTC CAG GAC ACC CTG CAG GTG GTC 480Thr Gly Pro Trp Ala Lys Asp Asn Phe Gln Asp Thr Leu Gln Val Val145 150 155 160ACC GCC CAC TAC CGC ACC TCA CCC TTC TTC TCT GTC TAT GTC AGT GCC 528Thr Ala His Tyr Arg Thr Ser Pro Phe Phe Ser Val Tyr Val Ser Ala
165 170 175GAT TCC AAG AAC TCC AAC AGC AAC GTG ATC CAG GTG GAC CAG TCT GGC 576Asp Ser Lys Asn Ser Asn Ser Asn Val Ile Gln Val Asp Gln Ser Gly
180 185 190CTG GGC TTG CCC TCG AGA GAC TAT TAC CTG AAC AAA ACT GAA AAC GAG 624Leu Gly Leu Pro Ser Arg Asp Tyr Tyr Leu Asn Lys Thr Glu Asn Glu
195 200 205
AAG GTG CTG ACC GGA TAT CTG AAC TAC ATG GTC CAG CTG GGG AAG CTG 672Lys Val Leu Thr Gly Tyr Leu Asn Tyr Met Val Gln Leu Gly Lys Leu
210 215 220
CTG GGC GGC GGG GAC GAG GAG GCC ATC CGG CCC CAG ATG CAG CAG ATC 720Leu Gly Gly Gly Asp Glu Glu Ala Ile Arg Pro Gln Met Gln Gln Ile225 230 235 240TTG GAC TTT GAG ACG GCA CTG GCC AAC ATC ACC ATC CCA CAG GAG AAG 768Leu Asp Phe Glu Thr Ala Leu Ala Asn Ile Thr Ile Pro Gln Glu Lys
245 250 255CGC CGT GAT GAG GAG CTC ATC TAC CAC AAA GTG ACG GCA GCC GAG CTG 816Arg Arg Asp Glu Glu Leu Ile Tyr His Lys Val Thr Ala Ala Glu Leu
260 265 270CAG ACC TTG GCA CCC GCC ATC AAC TGG TTG CCT TTT CTC AAC ACC ATC 864Gln Thr Leu Ala Pro Ala Ile Asn Trp Leu Pro Phe Leu Asn Thr Ile
275 280 285TTC TAC CCC GTG GAG ATC AAT GAA TCC GAG CCT ATT GTG GTC TAT GAC 912Phe Tyr Pro Val Glu Ile Asn Glu Ser Glu Pro Ile Val Val Tyr Asp
290 295 300AAG GAA TAC CTT GAG CAG ATC TCC ACT CTC ATC AAC ACC ACC GAC AGA 960Lys Glu Tyr Leu Glu Gln Ile Ser Thr Leu Ile Asn Thr Thr Asp Arg305 310 315 320TGC CTG CTC AAC AAC TAC ATG ATC TGG AAC CTG GTG CGG AAA ACA AGC 1008Cys Leu Leu Asn Asn Tyr Met ILe Trp Asn Leu Val Arg Lys Thr Ser
325 330 335TCC TTC CTT GAC CAG CGC TTT CAG GAC GCC GAT GAG AAG TTC ATG GAA 1056Ser Phe Leu Asp Gln Arg Phe Gln Asp Ala Asp Glu Lys Phe Met Glu
340 345 350GTC ATG TAC GGG ACC AAG AAG ACC TGT CTT CCT CGC TGG AAG TTT TGC 1104Val Met Tyr Gly Thr Lys Lys Thr Cys Leu Pro Arg Trp Lys Phe Cys
355 360 365GTG AGT GAC ACA GAA AAC AAC CTG GGC TTT GCG TTG GGC CCC ATG TTT 1152Val Ser Asp Thr Glu Asn Asn Leu Gly Phe Ala Leu Gly Pro Met Phe
370 375 380GTC AAA GCA ACC TTC GCC GAG GAC AGC AAG AGC ATA GCC ACC GAG ATC 1200Val Lys Ala Thr Phe Ala Glu Asp Ser Lys Ser Ile Ala Thr Glu Ile385 390 395 400ATC CTG GAG ATT AAG AAG GCA TTT GAG GAA AGC CTG AGC ACC CTG AAG 1248Ile Leu Glu Ile Lys Lys Ala Phe Glu Glu Ser Leu Ser Thr Leu Lys
405 410 415TGG ATG GAT GAG GAA ACC CGA AAA TCA GCC AAG GAA AAG GCC GAT GCC 1296Trp Met Asp Glu Glu Thr Arg Lys Ser Ala Lys Glu Lys Ala Asp Ala
420 425 430ATC TAC AAC ATG ATA GGA TAC CCC AAC TTC ATC ATG GAT CCC AAG GAG 1344Ile Tyr Asn Met Ile Gly Tyr Pro Asn Phe Ile Met Asp Pro Lys Glu
435 440 445CTG GAC AAA GTG TTT AAT GAC TAC ACT GCA GTT CCA GAC CTC TAC TTT 1392Leu Asp Lys Val Phe Asn Asp Tyr Thr Ala Val Pro Asp Leu Tyr Phe
450 455 460GAA AAT GCC ATG CGG TTT TTC AAC TTC TCA TGG AGG GTC ACT GCC GAT 1440Glu Asn Ala Met Arg Phe Phe Asn Phe Ser Trp Arg Val Thr Ala Asp465 470 475 480CAG CTC AGG AAA GCC CCC AAC AGA GAT CAG TGG AGC ATG ACC CCG CCC 1488Gln Leu Arg Lys Ala Pro Asn Arg Asp Gln Trp Ser Met Thr Pro Pro
485 490 495ATG GTG AAC GCC TAC TAC TCG CCC ACC AAG AAT GAG ATT GTG TTT CCG 1536Met Val Asn Ala Tyr Tyr Ser Pro Thr Lys Asn Glu Ile Val Phe Pro
500 505 510GCC GGG ATC CTG CAG GCA CCA TTC TAC ACA CGC TCC TCA CCC AAG GCC 1584Ala Gly Ile Leu Gln Ala Pro Phe Tyr Thr Arg Ser Ser Pro Lys Ala
515 520 525TTA AAC TTT GGT GGC ATA GGT GTC GTC GTG GGC CAT GAG CTG ACT CAT 1632Leu Asn Phe Gly Gly Ile Gly Val Val Val Gly His Glu Leu Thr His
530 535 540GCT TTT GAT GAT CAA GGA CGG GAG TAT GAC AAG GAC GGG AAC CTC CGG 1680Ala Phe Asp Asp Gln Gly Arg Glu Tyr Asp Lys Asp Gly Asn Leu Arg545 550 555 560CCA TGG TGG AAG AAC TCA TCC GTG GAG GCC TTC AAG CGT CAG ACC GAG 1728Pro Trp Trp Lys Asn Ser Ser Val Glu Ala Phe Lys Arg Gln Thr Glu
565 570 575TGC ATG GTA GAG CAG TAC AGC AAC TAC AGC GTG AAC GGG GAG CCG GTG 1776Cys Met Val Glu Gln Tyr Ser Asn Tyr Ser Val Asn Gly Glu Pro Val
580 585 590AAC GGG CGG CAC ACC CTG GGG GAG AAC ATC GCC GAC AAC GGG GGT CTC 1824Asn Gly Arg His Thr Leu Gly Glu Asn Ile Ala Asp Asn Gly Gly Leu
595 600 605AAG GCG GCC TAT CGG GCT TAC CAG AAC TGG GTG AAG AAG AAC GGG GCT 1872Lys Ala Ala Tyr Arg Ala Tyr Gln Asn Trp VaH Lys Lys Asn Gly Ala
610 615 620
GAG CAC TCG CTC CCC ACC CTG GGC CTC ACC AAT AAC CAG CTC TTC TTC 1920Glu His Ser Leu Pro Thr Leu GHy Leu Thr Asn Asn Gln Leu Phe Phe625 630 635 640CTG GGC TTT GCA CAG GTC TGG TGC TCC GTC CGC ACA CCT GAG AGC TCC 1968Leu Gly Phe Ala Gln Val Trp Cys Ser Val Arg Thr Pro Glu Ser Ser
645 650 655CAC GAA GGC CTC ATC ACC GAT CCC CAC AGC CCC TCT CGC TTC CGG GTC 2016His Glu Gly Leu Ile Thr Asp Pro His Ser Pro Ser Arg Phe Arg Val
660 665 670ATC GGC TCC CTC TCC AAT TCC AAG GAG TTC TCA GAA CAC TTC CGC TGC 2064Ile Gly Ser Leu Ser Asn Ser Lys Glu Phe Ser Glu His Phe Arg Cys
675 680 685CCA CCT GGC TCA CCC ATG AAC CCG CCT CAC AAG TGC GAA GTC TGG 2109Pro Pro Gly Ser Pro Met Asn Pro Pro His Lys Cys Glu Val Trp
690 695 700TAAGGACGAA GCGGAGAGAG CCAAGACGGA GGAGGGGAAG GGGCTGAGGA CGAGACCCCC 2169ATCCAGCCTC CAGGGCATTG CTCAGCCCGC TTGGCCACCC GGGGCCCTGC TTCCTCACAC 2229TGGCGGGTTT TCAGCCGGAA CCGAGCCCAT GGTGTTGGCT CTCAACGTGA CCCGCAGTCT 2289GATCCCCTGT GAAGAGCCGG ACATCCCAGG CACACGTGTG CGCCACCTTC AGCAGGCATT 2349CGGGTGCTGG GCTGGTGGCT CATCAGGCCT GGGCCCCACA CTGACAAGCG CCAGATACGC 2409CACAAATACC ACTGTGTCAA ATGCTTTCAA GATATATTTT TGGGGAAACT ATTTTTTAAA 2469CACTGTGGAA TACACTGGAA ATCTTCAGGG AAAAACACAT TTAAACACTT TTTTTTTTAA 2529GCCC 2533(2)SEQ ID NO:25的信息:
(i)序列特征:
(A)长度:703个氨基酸
(B)类型:氨基酸
(D)拓扑学:线性
(ii)分子类型:蛋白质
(xi)序列描述:SEQ ID NO:25:Arg Leu Val Val Leu Val Val Leu Leu Ala Ala Gly Leu Val Ala Cys1 5 10 15Leu Ala Ala Leu Gly Ile Gln Tyr Gln Thr Arg Ser Pro Ser Val Cys
20 25 30Leu Ser Glu Ala Cys Val Ser Val Thr Ser Ser Ile Leu Ser Ser Met
35 40 45Asp Pro Thr Val Asp Pro Cys His Asp Phe Phe Ser Tyr Ala Cys Gly
50 55 60Gly Trp Ile Lys Ala Asn Pro Val Pro Asp Gly His Ser Arg Trp Gly65 70 75 80Thr Phe Ser Asn Leu Trp Glu His Asn Gln Ala Ile Ile Lys His Leu
85 90 95Leu Glu Asn Ser Thr Ala Ser Val Ser Glu Ala Glu Arg Lys Ala Gln
100 105 110Val Tyr Tyr Arg Ala Cys Met Asn Glu Thr Arg Ile Glu Glu Leu Arg
115 120 125Ala Lys Pro Leu Met Glu Leu Ile Glu Arg Leu Gly Gly Trp Asn Ile
130 135 140Thr Gly Pro Trp Ala Lys Asp Asn Phe Gln Asp Thr Leu Gln Val Val145 150 155 160Thr Ala His Tyr Arg Thr Ser Pro Phe Phe Ser Val Tyr Val Ser Ala
165 170 175Asp Ser Lys Asn Ser Asn Ser Asn Val Ile Gln Val Asp Gln Ser Gly
180 185 190Leu Gly Leu Pro Ser Arg Asp Tyr Tyr Leu Asn Lys Thr Glu Asn Glu
195 200 205Lys Val Leu Thr Gly Tyr Leu Asn Tyr Met Val Gln Leu Gly Lys Leu
210 215 220Leu Gly Gly Gly Asp Glu Glu Ala Ile Arg Pro Gln Met Gln Gln Ile225 230 235 240Leu Asp Phe Glu Thr Ala Leu Ala Asn Ile Thr Ile Pro Gln Glu Lys
245 250 255Arg Arg Asp Glu Glu Leu Ile Tyr His Lys Val Thr Ala Ala Glu Leu
260 265 270Gln Thr Leu Ala Pro Ala Ile Asn Trp Leu Pro Phe Leu Asn Thr Ile
275 280 285Phe Tyr Pro Val Glu Ile Asn Glu Ser Glu Pro Ile Val Val Tyr Asp
290 295 300Lys Glu Tyr Leu Glu Gln Ile Ser Thr Leu Ile Asn Thr Thr Asp Arg305 310 315 320Cys Leu Leu Asn Asn Tyr Met Ile Trp Asn Leu Val Arg Lys Thr Ser
325 330 335Ser Phe Leu Asp Gln Arg Phe Gln Asp Ala Asp Glu Lys Phe Met Glu
340 345 350Val Met Tyr Gly Thr Lys Lys Thr Cys Leu Pro Arg Trp Lys Phe Cys
355 360 365Val Ser Asp Thr Glu Asn Asn Leu Gly Phe Ala Leu Gly Pro Met Phe
370 375 380Val Lys Ala Thr Phe Ala Glu Asp Ser Lys Ser Ile Ala Thr Glu Ile385 390 395 400Ile Leu Glu Ile Lys Lys Ala Phe Glu Glu Ser Leu Ser Thr Leu Lys
405 410 415Trp Met Asp Glu Glu Thr Arg Lys Ser Ala Lys Glu Lys Ala Asp Ala
420 425 430Ile Tyr Asn Met Ile Gly Tyr Pro Asn Phe Ile Met Asp Pro Lys Glu
435 440 445Leu Asp Lys Val Phe Asn Asp Tyr Thr Ala Val Pro Asp Leu Tyr Phe
450 455 460Glu Asn Ala Met Arg Phe Phe Asn Phe Ser Trp Arg Val Thr Ala Asp465 470 475 480Gln Leu Arg Lys Ala Pro Asn Arg Asp Gln Trp Ser Met Thr Pro Pro
485 490 495Met Val Asn Ala Tyr Tyr Ser Pro Thr Lys Asn Glu Ile Val Phe Pro
500 505 510Ala Gly Ile Leu Gln Ala Pro Phe Tyr Thr Arg Ser Ser Pro Lys Ala
515 520 525Leu Asn Phe Gly Gly Ile Gly Val Val Val Gly His Glu Leu Thr His
530 535 540Ala Phe Asp Asp Gln Gly Arg Glu Tyr Asp Lys Asp Gly Asn Leu Arg545 550 555 560Pro Trp Trp Lys Asn Ser Ser Val Glu Ala Phe Lys Arg Gln Thr Glu
565 570 575Cys Met Val Glu Gln Tyr Ser Asn Tyr Ser Val Asn Gly Glu Pro Val
580 585 590Asn Gly Arg His Thr Leu Gly Glu Asn Ile Ala Asp Asn Gly Gly Leu
595 600 605Lys Ala Ala Tyr Arg Ala Tyr Gln Asn Trp Val Lys Lys Asn Gly Ala
610 615 620Glu His Ser Leu Pro Thr Leu Gly Leu Thr Asn Asn Gln Leu Phe Phe625 630 635 640Leu Gly Phe Ala Gln Val Trp Cys Ser Val Arg Thr Pro Glu Ser Ser
645 650 655His Glu Gly Leu Ile Thr Asp Pro His Ser Pro Ser Arg Phe Arg Val
660 665 670Ile Gly Ser Leu Ser Asn Ser Lys Glu Phe Ser Glu His Phe Arg Cys
675 680 685Pro Pro Gly Ser Pro Met Asn Pro Pro His Lys Cys Glu Val Trp
690 695 700(2)SEQ ID NO:26的信息:
(i)序列特征:
(A)长度:24bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(iii)假设序列:不是
(iii)反义序列:是
(xi)序列描述:SEQ ID NO:26:GGTGCTTGAT GATGGCTTGG TTGT 24(2)SEQ ID NO:27的信息:
(i)序列特征:
(A)长度:24bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(iii)假设序列:不是
(iii)反义序列:是
(xi)序列描述:SEQ ID NO:27:AGATGGAGCT GGTCACCGAG ATGC 24(2)SEQ ID NO:28的信息:
(i)序列特征:
(A)长度:324bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型mRNA的cDNA
(iii)假设序列:不是
(vi)来源:
(A)生物名:Bos taurus
(H)组织类型:肺
(xi)序列描述:SEQ ID NO:28:GGGAGCGCGG CGCGGGGCCG GAGCGGAGCG CGCGAGCGAT GATGTCTACC TACAAGCGGC 60CCACGCTGGA CGAGGAGGAC CTGGTGGACT CGCTGTCCGA GAGCGACGTG TACCCCAACC 120ACCTGCAGGT GAACTTCCGA GGCCCCCGGA ACGGCCAGAG ATGCTGGGCC GCCAGGACCC 180CGGTGGAGAA GCGGCTGGTG GTGCTGGTGG CGCTCCTGGC GGCGGCATTG GTGGCCTGTT 240TGGCAGTACT GGGCATCCAA TACCAGACAA GAACGCCCTC GGTGTGCCTA AGTGAGGCCT 300GCATCTCGGT GACCAGCTCC ATCT 324(2)SEQ ID NO:29的信息:
(i)序列特征:
(A)长度:2314bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:mRNA的cDNA
(vi)来源:
(A)生物名:Bos taurus
(H)组织类型:肺
(ix)序列特征:
(A)名称/关键:CDS
(B)位置:39..2301
(xi)序列描述:SEQ ID NO:29:GGGAGCGCGG CGCGGGGCCG GAGCGGAGCG CGCGAGCG ATG ATG TCT ACC TAC 33
Met Met Ser Thr Tyr
1 5AAG CGG CCC ACG CTG GAC GAG GAG GAC CTG GTG GAC TCG CTG TCC GAG 101Lys Arg Pro Thr Leu Asp Glu Glu Asp Leu Val Asp Ser Leu Ser Glu
10 15 20AGC GAC GTG TAC CCC AAC CAC CTG CAG GTG AAC TTC CGA GGC CCC CGG 149Ser Asp Val Tyr Pro Asn His Leu Gln Val Asn Phe Arg Gly Pro Arg
25 30 35AAC GGC CAG AGA TGC TGG GCC GCC AGG ACC CCG GTG GAG AAG CGG CTG 197Asn Gly Gln Arg Cys Trp Ala Ala Arg Thr Pro Val Glu Lys Arg Leu
40 45 50GTG GTG CTG GTG GCG CTC CTG GCG GCG GCA TTG GTG GCC TGT TTG GCA 245Val Val Leu Val Ala Leu Leu Ala Ala Ala Leu Val Ala Cys Leu Ala
55 60 65GTA CTG GGC ATC CAA TAC CAG ACA AGA ACG CCC TCG GTG TGC CTA AGT 293Val Leu Gly Ile Gln Tyr Gln Thr Arg Thr Pro Ser Val Cys Leu Ser70 75 80 85GAG GCC TGC ATC TCG GTG ACC AGC TCC ATC TTG AGT TCC ATG GAC CCC 341Glu Ala Cys Ile Ser Val Thr Ser Ser Ile Leu Ser Ser Met Asp Pro
90 95 100ACG GTG GAC CCC TGC CAG GAC TTC TTC ACC TAT GCC TGT GGC GGC TGG 389Thr Val Asp Pro Cys Gln Asp Phe Phe Thr Tyr Ala Cys Gly Gly Trp
105 110 115ATC AAA GCC AAC CCC GTG CCG GAT GGC CAC TCG CGC TGG GGG ACC TTC 437Ile Lys Ala Asn Pro Val Pro Asp Gly His Ser Arg Trp Gly Thr Phe
120 125 130AGC AAC CTC TGG GAA CAC AAC CAA GCC ATC ATC AAG CAC CTC CTT GAA 485Ser Asn Leu Trp Glu His Asn Gln Ala Ile Ile Lys His Leu Leu Glu
135 140 145AAC TCC ACG GCC AGC GTG AGC GAG GCA GAG AGG AAG GAC CAG GAG TAC 533Asn Ser Thr Ala Ser Val Ser Glu Ala Glu Arg Lys Asp Gln Glu Tyr150 155 160 165TAC CGA GCC TGC ATG AAC GAA ACC AGG ATT GAG GAG CTC AAG GCC AAA 581Tyr Arg Ala Cys Met Asn Glu Thr Arg Ile Glu Glu Leu Lys Ala Lys
170 175 180CCC CTG ATG GAG CTC ATT GAG AAG CTC GGC GGC TGG AAC ATC ACG GGG 629Pro Leu Met Glu Leu Ile Glu Lys Leu Gly Gly Trp Asn Ile Thr Gly
185 190 195CCC TGG GAC AAG GAC AAC TTC CAG GAC ACC CTG CAG GTG GTC ACA TCC 677Pro Trp Asp Lys Asp Asn Phe Gln Asp Thr Leu Gln Val Val Thr Ser
200 205 210CAC TAC CAC ACC TCC CCC TTC TTC TCC GTC TAC GTC AGT GCC GAC TCC 725His Tyr His Thr Ser Pro Phe Phe Ser Val Tyr Val Ser Ala Asp Ser
215 220 225AAG AAT TCC AAC AGC AAC GTG ATC CAA GTG GAC CAG TCT GGC CTG GGC 773Lys Asn Ser Asn Ser Asn Val Ile Gln Val Asp Gln Ser Gly Leu Gly230 235 240 245TTA CCC TCA AGA GAT TAT TAC CTG AAC AAA ACC GAG AAT GAG AAG GTG 821Leu Pro Ser Arg Asp Tyr Tyr Leu Asn Lys Thr Glu Asn Glu Lys Val
250 255 260CTG ACG GGA TAC CTG AAC TAC ATG GTC CAG CTG GGG AAG CTG CTG GGA 869Leu Thr Gly Tyr Leu Asn Tyr Met Val Gln Leu Gly Lys Leu Leu Gly
265 270 275GGA GGG GCC GAG GAC ACC ATC CGG CCC CAG ATG CAG CAG ATC CTG GAC 917Gly Gly Ala Glu Asp Thr Ile Arg Pro Gln Met Gln Gln Ile Leu Asp
280 285 290TTT GAG ACG GCG CTG GCC AAC ATC ACC ATC CCC CAG GAG AAG CGC CGG 965Phe Glu Thr Ala Leu Ala Asn Ile Thr Ile Pro Gln Glu Lys Arg Arg
295 300 305GAC GAG GAA CTC ATC TAC CAC AAA GTG ACG GCG GCT GAG TTG CAG ACC 1013Asp Glu Glu Leu Ile Tyr His Lys Val Thr Ala Ala Glu Leu Gln Thr310 315 320 325TTG GCG CCC GCC ATC AAC TGG CTG CCC TTC CTC AAC ACC ATC TTC TAC 1061Leu Ala Pro Ala Ile Asn Trp Leu Pro Phe Leu Asn Thr Ile Phe Tyr
330 335 340CCC GTG GAG ATC AAT GAA TCA GAG CCT ATT GTC ATC TAC GAC AAA GAA 1109Pro Val Glu Ile Asn Glu Ser Glu Pro Ile Val Ile Tyr Asp Lys Glu
345 350 355TAC CTG AGC AAG GTC TCC ACC CTC ATC AAC AGC ACA GAC AAA TGC CTG 1157Tyr Leu Ser Lys Val Ser Thr Leu Ile Asn Ser Thr Asp Lys Cys Leu
360 365 370CnG AAC AAC TAC ATG ATC TGG AAC CTG GTA CGG AAG ACG AGC TCC TTC 1205Leu Asn Asn Tyr Met Ile Trp Asn Leu Val Arg Lys Thr Ser Ser Phe
375 380 385CTC GAT CAG CGC TTC CAG GAC GCC GAC GAG AAG TTC ATG GAA GTC ATG 1253Leu Asp Gln Arg Phe Gln Asp Ala Asp Glu Lys Phe Met Glu Val Met390 395 400 405TAT GGG ACC AAG AAG ACG TGT CTT CCC CGC TGG AAG TTT TGT GTG AGT 130lTyr Gly Thr Lys Lys Thr Cys Leu Pro Arg Trp Lys Phe Cys Val Ser
410 415 420GAT ACA GAG AAC ACC TTG GGC TTC GCC CTG GGC CCC ATG TTC GTC AAA 1349Asp Thr Glu Asn Thr Leu Gly Phe Ala Leu Gly Pro Met Phe Val Lys
425 430 435GCG ACC TTC GCT GAG GAC AGC AAG AAC ATA GCC AGC GAG ATC ATC CTG 1397Ala Thr Phe Ala Glu Asp Ser Lys Asn Ile Ala Ser Glu Ile Ile Leu
440 445 450GAG ATC AAG AAG GCG TTT GAA GAG AGC CTG AGC ACC CTG AAG TGG ATG 1445Glu Ile Lys Lys Ala Phe Glu Glu Ser Leu Ser Thr Leu Lys Trp Met
455 460 465GAT GAA GAT ACT CGG AAA TCG GCC AAG GAA AAG GCG GAC GCG ATC TAC 1493Asp Glu Asp Thr Arg Lys Ser Ala Lys Glu Lys Ala Asp Ala Ile Tyr470 475 480 485AAC ATG ATA GGC TAC CCC AAC TTT ATC ATG GAC CCC AAG GAG CTG GAC 1541Asn Met Ile Gly Tyr Pro Asn Phe Ile Met Asp Pro Lys Glu Leu Asp
490 495 500AAA GTG TTC AAT GAC TAC ACC GCT GTG CCA GAC CTC TAC TTC GAG AAC 1589Lys Val Phe Asn Asp Tyr Thr Ala Val Pro Asp Leu Tyr Phe Glu Asn
505 510 515GCC ATG CGG TTT TTC AAC TTC TCC TGG AGG GTC ACT GCC GAC CAG CTC 1637Ala Met Arg Phe Phe Asn Phe Ser Trp Arg Val Thr Ala Asp Gln Leu
520 525 530CGG AAA GCG CCC AAC AGA GAT CAG TGG AGC ATG ACC CCG CCC ATG GTG 1685Arg Lys Ala Pro Asn Arg Asp Gln Trp Ser Met Thr Pro Pro Met Val
535 540 545AAC GCC TAC TAC TCG CCC ACC AAG AAC GAG ATC GTG TTT CCG GCC GGA 1733Asn Ala Tyr Tyr Ser Pro Thr Lys Asn Glu Ile Val Phe Pro Ala Gly550 555 560 565ATC CTG CAG GCG CCA TTC TAC ACC CGC TCT TCA CCC AAT GCC TTA AAC 1781Ile Leu Gln Ala Pro Phe Tyr Thr Arg Set Ser Pro Asn Ala Leu Asn
570 575 580TTC GGC GGC ATC GGC GTC GTC GTG GGC CAC GAG CTG ACT CAT GCT TTT 1829Phe Gly Gly Ile Gly Val Val Val Gly His Glu Leu Thr His Ala Phe
585 590 595GAT GAT CAA GGC CGA GAG TAC GAC AAG GAT GGG AAC CTC CGG CCC TGG 1877Asp Asp Gln Gly Arg Glu Tyr Asp Lys Asp Gly Asn Leu Arg Pro Trp
600 605 610TGG AAG AAC TCG TCC GTG GAG GCG TTC AAG CAG CAG ACC GCG TGC ATG 1925Trp Lys Asn Ser Ser Val Glu Ala Phe Lys Gln Gln Thr Ala Cys Met
615 620 625GTG GAG CAG TAC GGC AAC TAT AGC GTG AAC GGG GAG CCG GTG AAC GGC 1973Val Glu Gln Tyr Gly Asn Tyr Ser Val Asn Gly Glu Pro Val Asn Gly630 635 640 645CGG CAC ACC CTC GGC GAA AAC ATC GCC GAC AAC GGG GGC CTC AAG GCG 2021Arg His Thr Leu Gly Glu Asn Ile Ala Asp Asn Gly Gly Leu Lys Ala
650 655 660GCC TAT CGG GCC TAC CAG AAC TGG GTC AAG AAG AAT GGG GCT GAG CAG 2069Ala Tyr Arg Ala Tyr Gln Asn Trp Val Lys Lys Asn Gly Ala Glu Gln
665 670 675ACA CTG CCC ACC CTG GGT CTC ACC AAC AAC CAG CTC TTC TTC CTG AGT 2117Thr Leu Pro Thr Leu Gly Leu Thr Asn Asn Gln Leu Phe Phe Leu Ser
680 685 690TTT GCA CAG GTC TGG TGT TCC GTC CGC ACC CCC GAG AGT TCG CAC GAA 2165Phe Ala Gln Val Trp Cys Ser Val Arg Thr Pro Glu Ser Ser His Glu
695 700 705GGT CTC ATC ACC GAT CCC CAC AGC CCC TCC CGC TTC CGG GTC ATC GGC 2213Gly Leu Ile Thr Asp Pro His Ser Pro Ser Arg Phe Arg Val Ile Gly710 715 720 725TCC ATC TCC AAC TCC AAG GAG TTC TCG GAA CAC TTC CAC TGC CCG CCC 2261Ser Ile Ser Asn Ser Lys Glu Phe Ser Glu His Phe His Cys Pro Pro
730 735 740GGC TCA CCC ATG AAC CCG CAT CAC AAG TGT GAA GTC TGG T GAAGGGCCAG 2311Gly Ser Pro Met Asn Pro His His Lys Cys Glu Val Trp
745 750GCA 2314(2)SEQ ID NO:30的信息:
(i)序列特征:
(A)长度:754个氨基酸
(B)类型:氨基酸
(D)拓扑学:线性
(ii)分子类型:蛋白质
(xi)序列描述:SEQ ID NO:30:Met Met Ser Thr Tyr Lys Arg Pro Thr Leu Asp Glu Glu Asp Leu Val1 5 10 15Asp Ser Leu Ser Glu Ser Asp Val Tyr Pro Asn His Leu Gln Val Asn
20 25 30Phe Arg Gly Pro Arg Asn Gly Gln Arg Cys Trp Ala Ala Arg Thr Pro
35 40 45Val Glu Lys Arg Leu Val Val Leu Val Ala Leu Leu Ala Ala Ala Leu
50 55 60
Val Ala Cys Leu Ala Val Leu Gly Ile Gln Tyr Gln Thr Arg Thr Pro65 70 75 80Ser Val Cys Leu Ser Glu Ala Cys Ile Ser Val Thr Ser Ser Ile Leu
85 90 95Ser Ser Met Asp Pro Thr Val Asp Pro Cys Gln Asp Phe Phe Thr Tyr
100 105 110Ala Cys Gly Gly Trp Ile Lys Ala Asn Pro Val Pro Asp Gly His Ser
115 120 125Arg Trp Gly Thr Phe Ser Asn Leu Trp Glu His Asn Gln Ala Ile Ile
130 135 140Lys His Leu Leu Glu Asn Ser Thr Ala Ser Val Ser Glu Ala Glu Arg145 150 155 160Lys Asp Gln Glu Tyr Tyr Arg Ala Cys Met Asn Glu Thr Arg Ile Glu
165 170 175Glu Leu Lys Ala Lys Pro Leu Met Glu Leu Ile Glu Lys Leu Gly Gly
180 185 190Trp Asn Ile Thr Gly Pro Trp Asp Lys Asp Asn Phe Gln Asp Thr Leu
195 200 205Gln Val Val Thr Ser His Tyr His Thr Ser Pro Phe Phe Ser Val Tyr
210 215 220Val Ser Ala Asp Ser Lys Asn Ser Asn Ser Asn Val Ile Gln Val Asp225 230 235 240Gln Ser Gly Leu Gly Leu Pro Ser Arg Asp Tyr Tyr Leu Asn Lys Thr
245 250 255Glu Asn Glu Lys Val Leu Thr Gly Tyr Leu Asn Tyr Met Val Gln Leu
260 265 270Gly Lys Leu Leu Gly Gly Gly Ala Glu Asp Thr Ile Arg Pro Gln Met
275 280 285Gln Gln Ile Leu AsP Phe Glu Thr Ala Leu Ala Asn Ile Thr Ile Pro
290 295 300Gln Glu Lys Arg Arg Asp Glu Glu Leu Ile Tyr His Lys Val Thr Ala305 310 315 320Ala Glu Leu Gln Thr Leu Ala Pro Ala Ile Asn Trp Leu Pro Phe Leu
325 330 335Asn Thr Ile Phe Tyr Pro Val Glu Ile Asn Glu Ser Glu Pro Ile Val
340 345 350Ile Tyr Asp Lys Glu Tyr Leu Ser Lys Val Ser Thr Leu Ile Asn Ser
355 360 365Thr Asp Lys Cys Leu Leu Asn Asn Tyr Met Ile Trp Asn Leu Val Arg
370 375 380
Lys Thr Ser Ser Phe Leu Asp Gln Arg Phe Gln Asp Ala Asp Glu Lys385 390 395 400
~Phe Met Glu Val Met Tyr Gly Thr Lys Lys Thr Cys Leu Pro Arg Trp
405 410 415Lys Phe Cys Val Ser Asp Thr Glu Asn Thr Leu Gly Phe Ala Leu Gly
420 425 430Pro Met Phe Val Lys Ala Thr Phe Ala Glu Asp Ser Lys Asn Ile Ala
435 440 445Ser Glu Ile Ile Leu Glu Ile Lys Lys Ala Phe Glu Glu Ser Leu Ser
450 455 460Thr Leu Lys Trp Met Asp Glu Asp Thr Arg Lys Ser Ala Lys Glu Lys465 470 475 480Ala Asp Ala Ile Tyr Asn Met Ile Gly Tyr Pro Asn Phe Ile Met Asp
485 490 495Pro Lys Glu Leu Asp Lys Val Phe Asn Asp Tyr Thr Ala Val Pro Asp
500 505 510Leu Tyr Phe Glu Asn Ala Met Arg Phe Phe Asn Phe Ser Trp Arg Val
515 520 525Thr Ala Asp Gln Leu Arg Lys Ala Pro Asn Arg Asp Gln Trp Ser Met
530 535 540Thr Pro Pro Met Val Asn Ala Tyr Tyr Ser Pro Thr Lys Asn Glu Ile545 550 555 560Val Phe Pro Ala Gly Ile Leu Gln Ala Pro Phe Tyr Thr Arg Ser Ser
565 570 575Pro Asn Ala Leu Asn Phe Gly Gly Ile Gly Val Val Val Gly His Glu
580 585 590Leu Thr His Ala Phe Asp Asp Gln Gly Arg Glu Tyr Asp Lys Asp Gly
595 600 605Asn Leu Arg Pro Trp Trp Lys Asn Ser Ser Val Glu Ala Phe Lys Gln
610 615 620Gln Thr Ala Cys Met Val Glu Gln Tyr Gly Asn Tyr Ser Val Asn Gly625 630 635 640Glu Pro Val Asn Gly Arg His Thr Leu Gly Glu Asn Ile Ala Asp Asn
645 650 655Gly Gly Leu Lys Ala Ala Tyr Arg Ala Tyr Gln Asn Trp Val Lys Lys
660 665 670Asn Gly Ala Glu Gln Thr Leu Pro Thr Leu Gly Leu Thr Asn Asn Gln
675 680 685Leu Phe Phe Leu Ser Phe Ala Gln Val Trp Cys Ser Val Arg Thr Pro
690 695 700Glu Ser Ser His Glu Gly Leu Ile Thr Asp Pro His Ser Pro Ser Arg705 710 715 720Phe Arg Val Ile Gly Ser Ile Ser Asn Ser Lys Glu Phe Ser Glu His
725 730 735Phe His Cys Pro Pro Gly Ser Pro Met Asn Pro His His Lys Cys Glu
740 745 750Val Trp(2)SEQ ID NO:31的信息:
(i)序列特征:
(A)长度:53bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(iii)假设序列:否
(iii)反义序列:是
(xi)序列描述:SEQ ID NO 31:GAGAGAGAGA GAGAGAGAGA ACTAGTCTCG AGCCAAGCAG GCCACCAGTC CTG 53(2)SEQ ID NO:32的信息:
(i)序列特征:
(A)长度:25bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:DNA(基因组)
(iii)假设序列:否
(iii)反义序列:是
(xi)序列描述:SEQ ID NO 32:CCTGCCGCCA GAAGTACCAC CAACA 25(2)SEQ ID NO:33的信息:
(i)序列特征:
(A)长度:222bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:mRNA的cDNA
(iii)假设序列:否
(vi)来源:
(A)生物名:Homo sapiens
(H)组织类型:胎盘
(ix)序列特征:
(A)名称/关键:CDS
(B)位置:38..222
(xi)序列描述:SEQ ID NO:33:CGCCCCCCCG GTGTCCGCCC TGCTGTCGGC GCTGGGG ATG TCG ACG TAC AAG CGG 55
Met Ser Thr Tyr Lys Arg
1 5GCC ACG CTG GAC GAG GAG GAC CTG GTG GAC TCG CTC TCC GAG GGC GAC 103Ala Thr Leu Asp Glu Glu Asp Leu Val Asp Ser Leu Ser Glu Gly Asp
10 15 20GCA TAC CCC AAC GGC CTG CAG GTG AAC TTC CAC AGC CCC CGG AGT GGC 151Ala Tyr Pro Asn Gly Leu Gln Val Asn Phe His Ser Pro Arg Ser Gly
25 30 35CAG AGG TGC TGG GCT GCA CGG ACC CAG GTG GAG AAG CGG CTG GTG GTG 199Gln Arg Cys Trp Ala Ala Arg Thr Gln Val Glu Lys Arg Leu Val Val
40 45 50TTG GTG GTA CTT CTG GCG GCA GG 222Leu Val Val Leu Leu Ala Ala55 60(2)SEQ ID NO:34的信息:
(i)序列特征:
(A)长度:61个氨基酸
(B)类型:氨基酸
(D)拓扑学:线性
(ii)分子类型:蛋白质
(xi)序列描述:SEQ ID NO:34:Met Ser Thr Tyr Lys Arg Ala Thr Leu Asp Glu Glu Asp Leu Val Asp1 5 10 15Ser Leu Ser Glu Gly Asp Ala Tyr Pro Asn Gly Leu Gln Val Asn Phe
20 25 30His Ser Pro Arg Ser Gly Gln Arg Cys Trp Ala Ala Arg Thr Gln Val
35 40 45Glu Lys Arg Leu Val Val Leu Val Val Leu Leu Ala Ala
50 55 60(2)SEQ ID NO:35的信息:
(i)序列特征:
(A)长度:2720bp
(B)类型:核酸
(C)链型:单链
(D)拓扑学:线性
(ii)分子类型:mRNA的cDNA
(iii)假设序列:否
(vi)来源:
(A)生物名:Homo sapiens
(H)组织类型:胎盘
(ix)序列特征:
(A)名称/关键:CDS
(B)位置:38..2297
(xi)序列描述:SEQ ID NO:35:CGCCCCCCCG GTGTCCGCCC TGCTGTCGGC GCTGGGG ATG TCG ACG TAC AAG CGG 55
Met Ser Thr Tyr Lys Arg
1 5GCC ACG CTG GAC GAG GAG GAC CTG GTG GAC TCG CTC TCC GAG GGC GAC 103Ala Thr Leu Asp Glu Glu Asp Leu Val Asp Ser Leu Ser Glu Gly Asp
10 15 20GCA TAC CCC AAC GGC CTG CAG GTG AAC TTC CAC AGC CCC CGG AGT GGC 151Ala Tyr Pro Asn Gly Leu Gln Val Asn Phe His Ser Pro Arg Ser Gly
25 30 35CAG AGG TGC TGG GCT GCA CGG ACC CAG GTG GAG AAG CGG CTG GTG GTG 199Gln Arg Cys Trp Ala Ala Arg Thr Gln Val Glu Lys Arg Leu Val Val
40 45 50TTG GTG GTA CTT CTG GCG GCA GGA CTG GTG GCC TGC TTG GCA GCA CTG 247Leu Val Val Leu Leu Ala Ala Gly Leu Val Ala Cys Leu Ala Ala Leu55 60 65 70GGC ATC CAG TAC CAG ACA AGA TCC CCC TCT GTG TGC CTG AGC GAA GCT 295
Gly Ile Gln Tyr Gln Thr Arg Ser Pro Ser Val Cys Leu Ser Glu Ala
75 80 85TGT GTC TCA GTG ACC AGC TCC ATC TTG AGC TCC ATG GAC CCC ACA GTG 343Cys Val Ser Val Thr Ser Ser Ile Leu Ser Ser Met Asp Pro Thr Val
90 95 100GAC CCC TGC CAT GAC TTC TTC AGC TAC GCC TGT GGG GGC TGG ATC AAG 391Asp Pro Cys His Asp Phe Phe Ser Tyr Ala Cys Gly Gly Trp Ile Lys
105 110 115GCC AAC CCA GTC CCT GAT GGC CAC TCA CGC TGG GGG ACC TTC AGC AAC 439Ala Asn Pro Val Pro Asp Gly His Ser Arg Trp Gly Thr Phe Ser Asn
120 125 130CTC TGG GAA CAC AAC CAA GCA ATC ATC AAG CAC CTC CTC GAA AAC TCC 487Leu Trp Glu His Asn Gln Ala Ile Ile Lys His Leu Leu Glu Asn Ser135 140 145 150ACG GCC AGC GTG AGC GAG GCA GAG AGA AAG GCG CAA GTA TAC TAC CGT 535Thr Ala Ser Val Ser Glu Ala Glu Arg Lys Ala Gln Val Tyr Tyr Arg
155 160 165GCG TGC ATG AAC GAG ACC AGG ATC GAG GAG CTC AGG GCC AAA CCT CTA 583Ala Cys Met Asn Glu Thr Arg Ile Glu Glu Leu Arg Ala Lys Pro Leu
170 175 180ATG GAG TTG ATT GAG AGG CTC GGG GGC TGG AAC ATC ACA GGT CCC TGG 631Met Glu Leu Ile Glu Arg Leu Gly Gly Trp Asn Ile Thr Gly Pro Trp
185 190 195GCC AAG GAC AAC TTC CAG GAC ACC CTG CAG GTG GTC ACC GCC CAC TAC 679Ala Lys Asp Asn Phe Gln Asp Thr Leu Gln Val Val Thr Ala His Tyr
200 205 210CGC ACC TCA CCC TTC TTC TCT GTC TAT GTC AGT GCC GAT TCC AAG AAC 727Arg Thr Ser Pro Phe Phe Ser Val Tyr Val Ser Ala Asp Ser Lys Asn215 220 225 230TCC AAC AGC AAC GTG ATC CAG GTG GAC CAG TCT GGC CTG GGC TTG CCC 775Ser Asn Ser Asn Val Ile Gln Val Asp Gln Ser Gly Leu Gly Leu Pro
235 240 245TCG AGA GAC TAT TAC CTG AAC AAA ACT GAA AAC GAG AAG GTG CTG ACC 823Ser Arg Asp Tyr Tyr Leu Asn Lys Thr Glu Asn Glu Lys Val Leu Thr
250 255 260
GGA TAT CTG AAC TAC ATG GTC CAG CTG GGG AAG CTG CTG GGC GGC GGG 871Gly Tyr Leu Asn Tyr Met Val Gln Leu Gly Lys Leu Leu Gly Gly Gly
265 270 275GAC GAG GAG GCC ATC CGG CCC CAG ATG CAG CAG ATC TTG GAC TTT GAG 919Asp Glu Glu Ala Ile Arg Pro Gln Met Gln Gln Ile Leu Asp Phe Glu
280 285 290ACG GCA CTG GCC AAC ATC ACC ATC CCA CAG GAG AAG CGC CGT GAT GAG 967Thr Ala Leu Ala Asn Ile Thr Ile Pro Gln Glu Lys Arg Arg Asp Glu295 300 305 310GAG CTC ATC TAC CAC AAA GTG ACG GCA GCC GAG CTG CAG ACC TTG GCA
1015Glu Leu Ile Tyr His Lys Val Thr Ala Ala Glu Leu Gln Thr Leu Ala
315 320 325CCC GCC ATC AAC TGG TTG CCT TTT CTC AAC ACC ATC TTC TAC CCC GTG 1063Pro Ala Ile Asn Trp Leu Pro Phe Leu Asn Thr Ile Phe Tyr Pro Val
330 335 340GAG ATC AAT GAA TCC GAG CCT ATT GTG GTC TAT GAC AAG GAA TAC CTT 1111Glu Ile Asn Glu Ser Glu Pro Ile Val Val Tyr Asp Lys Glu Tyr Leu
345 350 355GAG CAG ATC TCC ACT CTC ATC AAC ACC ACC GAC AGA TGC CTG CTC AAC 1159Glu Gln Ile Ser Thr Leu Ile Asn Thr Thr Asp Arg Cys Leu Leu Asn
360 365 370AAC TAC ATG ATC TGG AAC CTG GTG CGG AAA ACA AGC TCC TTC CTT GAC 1207Asn Tyr Met Ile Trp Asn Leu Val Arg Lys Thr Ser Ser Phe Leu Asp375 380 385 390CAG CGC TTT CAG GAC GCC GAT GAG AAG TTC ATG GAA GTC ATG TAC GGG 1255Gln Arg Phe Gln Asp Ala Asp Glu Lys Phe Met Glu Val Met Tyr Gly
395 400 405ACC AAG AAG ACC TGT CTT CCT CGC TGG AAG TTT TGC GTG AGT GAC ACA 1303Thr Lys Lys Thr Cys Leu Pro Arg Trp Lys Phe Cys Val Ser Asp Thr
410 415 420GAA AAC AAC CTG GGC TTT GCG TTG GGC CCC ATG TTT GTC AAA GCA ACC 1351Glu Asn Asn Leu Gly Phe Ala Leu Gly Pro Met Phe Val Lys Ala Thr
425 430 435TTC GCC GAG GAC AGC AAG AGC ATA GCC ACC GAG ATC ATC CTG GAG ATT 1399Phe Ala Glu Asp Ser Lys Ser Ile Ala Thr Glu Ile Ile Leu Glu Ile
440 445 450AAG AAG GCA TTT GAG GAA AGC CTG AGC ACC CTG AAG TGG ATG GAT GAG 1447Lys Lys Ala Phe Glu Glu Ser Leu Ser Thr Leu Lys Trp Met Asp Glu455 460 465 470
GAA ACC CGA AAA TCA GCC AAG GAA AAG GCC GAT GCC ATC TAC AAC ATG 1495Glu Thr Arg Lys Ser Ala Lys Glu Lys Ala Asp Ala Ile Tyr Asn Met
475 480 485ATA GGA TAC CCC AAC TTC ATC ATG GAT CCC AAG GAG CTG GAC AAA GTG 1543Ile Gly Tyr Pro Asn Phe Ile Met Asp Pro Lys Glu Leu Asp Lys Val
490 495 500TTT AAT GAC TAC ACT GCA GTT CCA GAC CTC TAC TTT GAA AAT GCC ATG 1591Phe Asn Asp Tyr Thr Ala Val Pro Asp Leu Tyr Phe Glu Asn Ala Met
505 510 515CGG TTT TTC AAC TTC TCA TGG AGG GTC ACT GCC GAT CAG CTC AGG AAA 1639Arg Phe Phe Asn Phe Ser Trp Arg Val Thr Ala Asp Gln Leu Arg Lys
520 525 530GCC CCC AAC AGA GAT CAG TGG AGC ATG ACC CCG CCC ATG GTG AAC GCC 1687Ala Pro Asn Arg Asp Gln Trp Ser Met Thr Pro Pro Met Val Asn Ala535 540 545 550TAC TAC TCG CCC ACC AAG AAT GAG ATT GTG TTT CCG GCC GGG ATC CTG 1735Tyr Tyr Ser Pro Thr Lys Asn Glu Ile Val Phe Pro Ala Gly Ile Leu
555 560 565CAG GCA CCA TTC TAC ACA CGC TCC TCA CCC AAG GCC TTA AAC TTT GGT 1783Gln Ala Pro Phe Tyr Thr Arg Ser Ser Pro Lys Ala Leu Asn Phe Gly
570 575 580GGC ATA GGT GTC GTC GTG GGC CAT GAG CTG ACT CAT GCT TTT GAT GAT 1831Gly Ile Gly Val Val Val Gly His Glu Leu Thr His Ala Phe Asp Asp
585 590 595CAA GGA CGG GAG TAT GAC AAG GAC GGG AAC CTC CGG CCA TGG TGG AAG 1879Gln Gly Arg Glu Tyr Asp Lys Asp Gly Asn Leu Arg Pro Trp Trp Lys
600 605 610AAC TCA TCC GTG GAG GCC TTC AAG CGT CAG ACC GAG TGC ATG GTA GAG 1927Asn Ser Ser Val Glu Ala Phe Lys Arg Gln Thr Glu Cys Met Val Glu615 620 625 630CAG TAC AGC AAC TAC AGC GTG AAC GGG GAG CCG GTG AAC GGG CGG CAC 1975Gln Tyr Ser Asn Tyr Ser Val Asn Gly Glu Pro Val Asn Gly Arg His
635 640 645ACC CTG GGG GAG AAC ATC GCC GAC AAC GGG GGT CTC AAG GCG GCC TAT 2023Thr Leu Gly Glu Asn Ile Ala Asp Asn Gly Gly Leu Lys Ala Ala Tyr
650 655 660CGG GCT TAC CAG AAC TGG GTG AAG AAG AAC GGG GCT GAG CAC TCG CTC 2071Arg Ala Tyr Gln Asn Trp Val Lys Lys Asn Gly Ala Glu His Ser Leu
665 670 675CCC ACC CTG GGC CTC ACC AAT AAC CAG CTC TTC TTC CTG GGC TTT GCA 2119Pro Thr Leu Gly Leu Thr Asn Asn Gln Leu Phe Phe Leu Gly Phe Ala
680 685 690CAG GTC TGG TGC TCC GTC CGC ACA CCT GAG AGC TCC CAC GAA GGC CTC 2167Gln Val Trp Cys Ser Val Arg Thr Pro Glu Ser Ser His Glu Gly Leu695 700 705 710ATC ACC GAT CCC CAC AGC CCC TCT CGC TTC CGG GTC ATC GGC TCC CTC 2215Ile Thr Asp Pro His Ser Pro Ser Arg Phe Arg Val Ile Gly Ser Leu
715 720 725TCC AAT TCC AAG GAG TTC TCA GAA CAC TTC CGC TGC CCA CCT GGC TCA 2263Ser Asn Ser Lys Glu Phe Ser Glu His Phe Arg Cys Pro Pro Gly Ser
730 735 740CCC ATG AAC CCG CCT CAC AAG TGC GAA GTC TGG T AAGGACGAAG 2307Pro Met Asn Pro Pro His Lys Cys Glu Val Trp
745 750CGGAGAGAGC CAAGACGGAG GAGGGGAAGG GGCTGAGGAC GAGACCCCCA TCCAGCCTCC 2367AGGGCATTGC TCAGCCCGCT TGGCCACCCG GGGCCCTGCT TCCTCACACT GGCGGGTTTT 2427CAGCCGGAAC CGAGCCCATG GTGTTGGCTC TCAACGTGAC CCGCAGTCTG ATCCCCTGTG 2487AAGAGCCGGA CATCCCAGGC ACACGTGTGC GCCACCTTCA GCAGGCATTC GGGTGCTGGG 2547CTGGTGGCTC ATCAGGCCTG GGCCCCACAC TGACAAGCGC CAGATACGCC ACAAATACCA 2607CTGTGTCAAA TGCTTTCAAG ATATATTTTT GGGGAAACTA TTTTTTAAAC ACTGTGGAAT 2667ACACTGGAAA TCTTCAGGGA AAAACACATT TAAACACTTT TTTTTTTAAG CCC 2720(2)SEQ ID NO:36的信息:
(i)序列特征:
(A)长度:735个氨基酸
(B)类型:氨基酸
(D)拓扑学:线性
(ii)分子类型:蛋白质
(xi)序列描述:SEQ ID NO:36:Met Ser Thr Tyr Lys Arg Ala Thr Leu Asp Glu Glu Asp Leu Val Asp1 5 10 15Ser Leu Ser Glu Gly Asp Ala Tyr Pro Asn Gly Leu Gln Val Asn Phe
20 25 30His Ser Pro Arg Ser Gly Gln Arg Cys Trp Ala Ala Arg Thr Gln Val
35 40 45Glu Lys Arg Leu Val Val Leu Val Val Leu Leu Ala Ala Gly Leu Val
50 55 60Ala Cys Leu Ala Ala Leu Gly Ile Gln Tyr Gln Thr Arg Ser Pro Ser65 70 75 80Val Cys Leu Ser Glu Ala Cys Val Ser Val Thr Ser Ser Ile Leu Ser
85 90 95Ser Met Asp Pro Thr Val Asp Pro Cys His Asp Phe Phe Ser Tyr Ala
100 105 110Cys Gly Gly Trp Ile Lys Ala Asn Pro Val Pro Asp Gly His Ser Arg
115 120 125Trp Gly Thr Phe Ser Asn Leu Trp Glu His Asn Gln Ala Ile Ile Lys
130 135 140His Leu Leu Glu Asn Ser Thr Ala Ser Val Ser Glu Ala Glu Arg Lys145 150 155 160Ala Gln Val Tyr Tyr Arg Ala Cys Met Asn Glu Thr Arg Ile Glu Glu
165 170 175Leu Arg Ala Lys Pro Leu Met Glu Leu Ile Glu Arg Leu Gly Gly Trp
180 185 190Asn Ile Thr Gly Pro Trp Ala Lys Asp Asn Phe Gln Asp Thr Leu Gln
195 200 205Val Val Thr Ala His Tyr Arg Thr Ser Pro Phe Phe Ser Val Tyr Val
210 215 220Ser Ala Asp Ser Lys Asn Ser Asn Ser Asn Val Ile Gln Val Asp Gln225 230 235 240Ser Gly Leu Gly Leu Pro Ser Arg Asp Tyr Tyr Leu Asn Lys Thr Glu
245 250 255Asn Glu Lys Val Leu Thr Gly Tyr Leu Asn Tyr Met Val Gln Leu Gly
260 265 270Lys Leu Leu Gly Gly Gly Asp Glu Glu Ala Ile Arg Pro Gln Met Gln
275 280 285Gln Ile Leu Asp Phe Glu Thr Ala Leu Ala Asn Ile Thr Ile Pro Gln
290 295 300Glu Lys Arg Arg Asp Glu Glu Leu Ile Tyr His Lys Val Thr Ala Ala305 310 315 320Glu Leu Gln Thr Leu Ala Pro Ala Ile Asn Trp Leu Pro Phe Leu Ash
325 330 335Thr Ile Phe Tyr Pro Val Glu Ile Asn Glu Ser Glu Pro Ile Val Val
340 345 350Tyr Asp Lys Glu Tyr Leu Glu Gln Ile Ser Thr Leu Ile Asn Thr Thr
355 360 365Asp Arg Cys Leu Leu Asn Asn Tyr Met Ile Trp Asn Leu Val Arg Lys
370 375 380Thr Ser Ser Phe Leu Asp Gln Arg Phe Gln Asp Ala Asp Glu Lys Phe385 390 395 400Met Glu Val Met Tyr Gly Thr Lys Lys Thr Cys Leu Pro Arg Trp Lys
405 410 415Phe Cys Val Ser Asp Thr Glu Asn Asn Leu Gly Phe Ala Leu Gly Pro
420 425 430Met Phe Val Lys Ala Thr Phe Ala Glu Asp Ser Lys Ser Ile Ala Thr
435 440 445Glu Ile Ile Leu Glu Ile Lys Lys Ala Phe Glu Glu Ser Leu Ser Thr
450 455 460Leu Lys Trp Met Asp Glu Glu Thr Arg Lys Ser Ala Lys Glu Lys Ala465 470 475 480Asp Ala Ile Tyr Asn Met Ile Gly Tyr Pro Asn Phe Ile Met Asp Pro
485 490 495Lys Glu Leu Asp Lys Val Phe Asn Asp Tyr Thr Ala Val Pro Asp Leu
500 505 510Tyr Phe Glu Asn Ala Met Arg Phe Phe Asn Phe Ser Trp Arg Val Thr
515 520 525Ala Asp Gln Leu Arg Lys Ala Pro Asn Arg Asp Gln Trp Ser Met Thr
530 535 540Pro Pro Met Val Asn Ala Tyr Tyr Ser Pro Thr Lys Asn Glu Ile Val545 550 555 560Phe Pro Ala Gly Ile Leu Gln Ala Pro Phe Tyr Thr Arg Ser Ser Pro
565 570 575Lys Ala Leu Asn Phe Gly Gly Ile Gly Val Val Val Gly His Glu Leu
580 585 590Thr His Ala Phe Asp Asp Gln Gly Arg Glu Tyr Asp Lys Asp Gly Asn
595 600 605Leu Arg Pro Trp Trp Lys Asn Ser Ser Val Glu Ala Phe Lys Arg Gln
610 615 620Thr Glu Cys Met Val Glu Gln Tyr Ser Asn Tyr Ser Val Asn Gly Glu625 630 635 640Pro Val Asn Gly Arg His Thr Leu Gly Glu Asn Ile Ala Asp Asn Gly
645 650 655Gly Leu Lys Ala Ala Tyr Arg Ala Tyr Gln Asn Trp Val Lys Lys Asn
660 665 670Gly Ala Glu His Ser Leu Pro Thr Leu Gly Leu Thr Asn Asn Gln Leu
675 680 685Phe Phe Leu Gly Phe Ala Gln Val Trp Cys Ser Val Arg Thr Pro Glu
690 695 700Ser Ser His Glu Gly Leu Ile Thr Asp Pro His Ser Pro Ser Arg Phe705 710 715 720Arg Val Ile Gly Ser Leu Ser Asn Ser Lys Glu Phe Ser Glu His Phe
725 730 735Arg Cys Pro Pro Gly Ser Pro Met Asn Pro Pro His Lys Cys Glu Val
740 745 750Trp
Claims (16)
1.一种内皮素转化酶,包括SEQ ID NO:30或SEQ ID NO:36所示的多肽序列或其功能性片段。
2.编码如权利要求1所要求的内皮素转化酶的DNA序列,选自下列的一组序列:
a)具有SEQ ID NO:29或SEQ ID NO:35所述结构的DNA序列;
b)编码具有SEQ ID NO:30或SEQ ID NO:36所述结构之蛋白质的DNA序列;以及
c)在标准条件下与DNA序列a)或b)杂交的DNA序列;
d)编码能被针对SEQ ID NO:30或36之蛋白质或其片段所产生的抗体识别的蛋白质产物之DNA序列。
3.利用如权利要求2所要求的DNA序列,通过基因操作以制备内皮素转化酶的方法。
4.制备如权利要求1所要求之内皮素转化酶的方法,包括用内皮素转化酶抑制剂刺激哺乳动物细胞以过量产生内皮素转化酶,和用常规的蛋白质化学方法从所述细胞分离内皮素转化酶。
5.如权利要求4的方法,其中的phosphoramidon用作抑制剂。
6.制备权利要求1之内皮素转化酶的方法,包括使用编码SEQ ID NO:36部分序列的寡核苷酸或序列互补性寡核苷酸作为杂交探针,以从基因库中分离出内皮素转化酶的基因,以及利用常规的遗传工程方法在宿主生物体中表达所述基因。
7.权利要求1要求的内皮素转化酶,在鉴定内皮素转化酶抑制剂中的应用。
8.内皮素转化酶抑制剂,它是通过应用权利要求1之内皮素转化酶的酶测定法鉴定的。
9.权利要求8之抑制剂作为药物的应用。
10.生产能够抑制内皮素转化酶之药物的方法,包括在应用了权利要求1之内皮素转化酶的酶测定法中使用已知的化合物,鉴别具有抑制作用的化合物,和用常规载体和辅助剂与如此鉴别的化合物配制成药物。
11.权利要求1之内皮素转化酶在抗体生产中的应用。
12.权利要求11所述之抗体在疾病诊断中的应用。
13.权利要求2所要求的DNA序列在制备转基因动物中的应用,所述转基因动物包含所述DNA序列的一个或多个额外拷贝,或者在该转基因动物中的正常EXE基因是关闭的。
14.权利要求1之内皮素转化酶在药物生产中的应用。
15.SEQ ID NO:35的部分序列在制备反义寡核苷酸中的应用。
16.权利要求15所述的部分序列在药物生产中的应用。
Applications Claiming Priority (7)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
DEP4339100.1 | 1993-11-16 | ||
DE19934339100 DE4339100A1 (de) | 1993-11-16 | 1993-11-16 | Endothelinkonversionsenzym-1 (ECE-1) |
DE19944403665 DE4403665A1 (de) | 1994-02-07 | 1994-02-07 | Endothelinkonversionsenzym (ECE) |
DEP4403665.5 | 1994-02-07 | ||
DE19944412372 DE4412372A1 (de) | 1994-04-12 | 1994-04-12 | Endothelinkonversionsenzym (ECE) |
DEP4412372.8 | 1994-04-12 | ||
PCT/EP1994/003706 WO1995014095A1 (de) | 1993-11-16 | 1994-11-10 | Endothelinkonversionsenzym (ece) |
Publications (1)
Publication Number | Publication Date |
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CN1141651A true CN1141651A (zh) | 1997-01-29 |
Family
ID=27205762
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN94194828A Pending CN1141651A (zh) | 1993-11-16 | 1994-11-10 | 内皮素转化酶(ece) |
Country Status (18)
Country | Link |
---|---|
US (1) | US6066502A (zh) |
EP (1) | EP0728209B1 (zh) |
JP (1) | JPH09505733A (zh) |
KR (1) | KR960705934A (zh) |
CN (1) | CN1141651A (zh) |
AT (1) | ATE315656T1 (zh) |
AU (1) | AU8107094A (zh) |
BR (1) | BR9408077A (zh) |
CA (1) | CA2176507A1 (zh) |
CZ (1) | CZ139496A3 (zh) |
DE (1) | DE59410427D1 (zh) |
FI (1) | FI962047A (zh) |
HU (1) | HUT75554A (zh) |
NO (1) | NO962023L (zh) |
NZ (1) | NZ275817A (zh) |
PL (1) | PL314480A1 (zh) |
SK (1) | SK59896A3 (zh) |
WO (1) | WO1995014095A1 (zh) |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN1918473B (zh) * | 2004-02-13 | 2011-08-10 | B.R.A.H.M.S有限公司 | 出于医学诊断目的而测定内皮素形成的方法,以及用于实施所述方法的抗体和试剂盒 |
Families Citing this family (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6255468B1 (en) * | 1998-05-07 | 2001-07-03 | Smithkline Beecham Plc | MPROT12 polynucleotides and methods thereof |
EP1409690A2 (en) | 1999-09-24 | 2004-04-21 | Lexicon Genetics Incorporated | Human endothelin converting enzyme-like proteins and polynucleotides encoding the same |
WO2009018209A1 (en) | 2007-07-27 | 2009-02-05 | The Regents Of The University Of Colorado, A Body Corporate | ENDOTHELIN SINGLE NUCLEOTIDE POLYMORPHISMS AND METHODS OF PREDICTING β-ADRENERGIC RECEPTOR TARGETING AGENT EFFICACY |
Family Cites Families (2)
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JP2966939B2 (ja) * | 1990-12-07 | 1999-10-25 | 日清製粉株式会社 | ヒト細胞由来のエンドセリン変換酵素 |
CA2100355A1 (en) * | 1991-02-04 | 1992-08-05 | Lynne H. Parker Botehlo | Endothelin converting enzyme |
-
1994
- 1994-11-10 US US08/646,273 patent/US6066502A/en not_active Expired - Lifetime
- 1994-11-10 AT AT95900130T patent/ATE315656T1/de not_active IP Right Cessation
- 1994-11-10 BR BR9408077A patent/BR9408077A/pt not_active Application Discontinuation
- 1994-11-10 WO PCT/EP1994/003706 patent/WO1995014095A1/de active IP Right Grant
- 1994-11-10 AU AU81070/94A patent/AU8107094A/en not_active Abandoned
- 1994-11-10 HU HU9601298A patent/HUT75554A/hu unknown
- 1994-11-10 SK SK598-96A patent/SK59896A3/sk unknown
- 1994-11-10 PL PL94314480A patent/PL314480A1/xx unknown
- 1994-11-10 CN CN94194828A patent/CN1141651A/zh active Pending
- 1994-11-10 CZ CZ961394A patent/CZ139496A3/cs unknown
- 1994-11-10 EP EP95900130A patent/EP0728209B1/de not_active Expired - Lifetime
- 1994-11-10 CA CA002176507A patent/CA2176507A1/en not_active Abandoned
- 1994-11-10 DE DE59410427T patent/DE59410427D1/de not_active Expired - Fee Related
- 1994-11-10 NZ NZ275817A patent/NZ275817A/en not_active IP Right Cessation
- 1994-11-10 KR KR1019960702556A patent/KR960705934A/ko not_active Application Discontinuation
- 1994-11-10 JP JP7514199A patent/JPH09505733A/ja not_active Ceased
-
1996
- 1996-05-14 FI FI962047A patent/FI962047A/fi unknown
- 1996-05-15 NO NO962023A patent/NO962023L/no not_active Application Discontinuation
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN1918473B (zh) * | 2004-02-13 | 2011-08-10 | B.R.A.H.M.S有限公司 | 出于医学诊断目的而测定内皮素形成的方法,以及用于实施所述方法的抗体和试剂盒 |
Also Published As
Publication number | Publication date |
---|---|
EP0728209B1 (de) | 2006-01-11 |
HUT75554A (en) | 1997-05-28 |
CZ139496A3 (en) | 1997-10-15 |
EP0728209A1 (de) | 1996-08-28 |
FI962047A0 (fi) | 1996-05-14 |
ATE315656T1 (de) | 2006-02-15 |
NZ275817A (en) | 1997-02-24 |
BR9408077A (pt) | 1997-08-12 |
AU8107094A (en) | 1995-06-06 |
KR960705934A (ko) | 1996-11-08 |
WO1995014095A1 (de) | 1995-05-26 |
HU9601298D0 (en) | 1996-07-29 |
NO962023D0 (no) | 1996-05-15 |
PL314480A1 (en) | 1996-09-16 |
US6066502A (en) | 2000-05-23 |
FI962047A (fi) | 1996-07-12 |
CA2176507A1 (en) | 1995-05-26 |
NO962023L (no) | 1996-07-16 |
DE59410427D1 (en) | 2006-04-06 |
SK59896A3 (en) | 1997-07-09 |
JPH09505733A (ja) | 1997-06-10 |
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