眼镜蛇血清解毒蛋白基因的分子克隆和序列分析

既往研究中作者发现，眼镜蛇血清解毒蛋白（ＣＳＡＰ）经胰蛋白酶水解后纯化所得的几个肽段中，至少在一３９肽显示其序列与大鼠血清白蛋白相关。为进一步弄清ＣＳＡＰ的毒素结合活性与其相应的分子结构关系，以眼镜蛇肝细胞建立了ｃＤＮＡ文库，借已知的胰蛋白酶水解ＣＳＡＰ所得的肽段氨基酸序列设计了一组ＰＣＲ引物。采用ＰＣＲ扩增方法从文库中筛选得ＣＳＡＰ特异性克隆，测定了ＣＳＡＰ全ｃＤＮＡ序列，然后推导出ＣＳＡＰ氨基酸序列。并得知ＣＳＡＰ为一由６１４个氨基酸残基组成的单一多肽链，分子量６９８００。根据其全序列的结构特征，尤其是其信号肽与已知的几种哺乳动物白蛋白的信号肽序列非常一致，其中半胱氨酸残基的分布规律也与其他血清白蛋白非常相似，可确认ＣＳＡＰ即为眼镜蛇血清白蛋白（ＣＳＡ）。

The Molecular Cloning and Sequencing of the Cobra Serum Antitoxic Protein Gene

In the previous study the authors found at least one peptide of 39 amino acid residues of several purified tryptic-fragments of the cobra serum antitoxic protein(CSAP),whose sequence had shown certain correlation with the rat serum albumin.For further clarification of the toxin-binding activity of CSAP in relation with its corresponding structure,the cDNA gene library of cobra liver cells was established and DNA primers were designed according to the known amino acid sequences of the tryptic peptides of CSAP. The specific probes were prepared and the specific clone was screened out from the library by PCR,Finally,the CSAP cDNA was sequenced from its sub-clones. Then the complete amino acid sequence of CSAP was elucidated.It is a polypeptide chain of 6l4 amino acid residues with a molecular size of 69,8 KDa,In comparing the whole sequence of CSAP,especially its signal peptide,with mammalian serum albumins,the authors have come to realize that CSAP is just the cobra serum albumin(CSA).