CHARACTERIZATION OF A CDNA CODING FOR HUMAN PROTEIN-C
CHARACTERIZATION OF A CDNA CODING FOR HUMAN PROTEIN-C
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DOI：
10.1073/pnas.81.15.4766
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发表时间：
1984-01-01
期刊：
影响因子：
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通讯作者：
DAVIE, EW
中科院分区：
文献类型：
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作者：
FOSTER, D;DAVIE, EW
蛋白 C 是存在于哺乳动物血浆中的丝氨酸蛋白酶的前体。在其激活形式下，它很容易使因子 Va 和因子 VIIIa 失活，这两种蛋白质在凝血级联中作为辅助因子参与。用抗人蛋白C的抗体筛选含有从人肝脏mRNA制备的cDNA[互补DNA]插入片段的λgt11文库。从2×中分离出7个阳性克隆。 106噬菌体并进行噬菌斑纯化。对其中 2 个噬菌体的 cDNA 插入片段进行了测序，结果显示其编码人蛋白 C。每个 cDNA 插入片段编码分子轻链的一部分、连接区、重链、终止密码子、3''-非编码区和聚腺苷酸尾。 2 个克隆中 3'' 末端非编码序列的长度不同，但每个克隆均包含加工信号或多聚腺苷酸化信号，后跟聚腺苷酸尾。从cDNA确定的氨基酸序列表明，蛋白C被合成为含有通过Lys-Arg二肽连接的轻链和重链的单链多肽。然后通过切割 2 个或更多个内部肽键，将单链分子转化为轻链和重链。在血浆中，蛋白 C 的重链和轻链通过二硫键连接在一起。人C蛋白的氨基酸序列与牛分子的氨基酸序列具有高度的同源性。编码人 C 蛋白活性位点丝氨酸附近催化区域的 DNA 序列也显示出与凝血酶原、因子 IX 和因子 X（血浆中存在的其他 3 种维生素 K 依赖性丝氨酸蛋白酶）具有高度的 DNA 和氨基酸序列同一性。
Protein C is a precursor to a serine protease that is present in mammalian plasma. In its activated form, it readily inactivates factor Va and factor VIIIa, 2 proteins that participate as cofactors in the blood coagulation cascade. A .lambda.gt11 library containing cDNA [complementary DNA] inserts prepared from human liver mRNA was screened with an antibody to human protein C. Seven positive clones were isolated from 2 .times. 106 phage and were plaque-purified. The cDNA inserts of 2 of these phage were sequenced and shown to code for human protein C. Each cDNA insert coded for a portion of the light chain of the molecule, a connecting region, the heavy chain, a stop codon, a 3''-noncoding region and a poly(A) tail. The length of the noncoding sequence on the 3'' end differed in the 2 clones, but each contained a processing or polyadenylylation signal followed by a poly(A) tail. The amino acid sequence as determined from the cDNA indicates that protein C is synthesized as a single-chain polypeptide containing the light chain and the heavy chain connected by a dipeptide of Lys-Arg. The single-chain molecule is then converted to the light and heavy chains by cleavage of 2 or more internal peptide bonds. In plasma, the heavy and light chains of protein C are linked together by a disulfide bond. The amino acid seqeunce of human protein C shows a high degree of homology with that of the bovine molecule. The DNA sequence coding for the catalytic region near the active site serine in human protein C also showed a high degree of DNA and amino acid sequence identity with prothrombin, factor IX and factor X, 3 of the other vitamin K-dependent serine proteases that are present in plasma.