蝮蛇类凝血酶基因的分析及表达研究  

Cloning and Expression of cDNA for Thrombin like Enzyme from Agkistrodon halys Pallas Snake Venom

作　　者：潘华[1] 周元聪[1] 杨冠珍[1] 吴祥甫[1] 

机构地区：[1]中国科学院上海生物化学研究所

出　　处：《生物化学与生物物理学报》1999年第1期79-82,共4页

摘　　要：从蝮蛇（ＡｇｋｉｓｔｒｏｄｏｎｈａｌｙｓＰａｌａｓ）毒腺中抽提总ＲＮＡ，经ＲＴＰＣＲ扩增该基因，克隆后经全序列测定，蝮蛇类凝血酶ｐａｌａｓｅ的ｃＤＮＡ长７０８个核苷酸，即编码２３６个氨基酸；根据同源性，推测该类凝血酶ｐａｌａｓｅ的活性中心为Ｈｉｓ４１、Ａｓｐ８６和Ｓｅｒ１８２；二硫键为Ｃｙｓ７Ｃｙｓ１３９、Ｃｙｓ２６Ｃｙｓ４２、Ｃｙｓ７４Ｃｙｓ２３４、Ｃｙｓ１１８Ｃｙｓ１８８、Ｃｙｓ１５０Ｃｙｓ１６７和Ｃｙｓ１７８Ｃｙｓ２０３；该蝮蛇毒类凝血酶ｃＤＮＡ序列及推导的氨基酸序列均为首次报道。构建Ｔ７启动子控制下的ｐａｌａｓｅ的大肠杆菌表达质粒，ＩＰＴＧ诱导ｐａｌａｓｅ获得表达。Total RNAs were extracted from the venom gland of the Agkistrodon halys Pallas snake. The thrombin like enzyme gene was amplified by RT PCR, cloned, and its nucleotide sequences have been determined. The enzyme called pallase cDNA encodes 708 nucleotides, namely 236 amino acids. Based on the homology, the catalytic residues and disulfide bridges of pallase were deduced as follows: catalytic residues, His 41 , Asp 86 和Ser 182 ; and disulfide bridges, Cys 7 Cys 139 , Cys 26 Cys 42 , Cys 74 Cys 234 , Cys 118 Cys 188 , Cys 150 Cys 167 和Cys 178 Cys 203 . The pallase expression plasmids under the control of the T7 promoter was constructed and the pallase was expressed in E.coli .

关 键 词：蝮蛇 类凝血酶基因 克隆 序列分析 表达 

分 类 号：Q753[生物学—分子生物学] Q959.62