五步蛇毒血小板聚集抑制因子cDNA的克隆及表达

采用一步法抽提五步蛇毒腺总ＲＮＡ，通过ＲＴ－ＰＣＲ的扩增出低分子量金属蛋白酶酶原的ｃＤＮＡ，克隆并测定了全序列。根据推导的氨基酸序列，发现其中一个ｃＤＮＡ除编码一个低分子量金属蛋白酶外，羧基端还包括一个血小板聚集抑制因子，这一结果证实了蛇毒金属蛋白酶和血小板聚集抑制因子起源于蛇毒金属蛋白酶酶原的前体。

Molecular Cloning and Expression of the cDNA for Disintegrin from Agkistrodon acutus

In order to clone novel and valuable genes from natural resources and develop genetic engineering drugs, total RNA was extracted from the venom gland of Agkistrodon acutus by one step method. The cDNAs for low molecular weight metalloproteinase were amplified by RT PCR and cloned into the pGEMT vector. Their sequence were determined. One of the deduced amino acid sequences included a metalloproteinase and a disintegrin (acugrin) at the carboxyl terminus. This result further confirms that metalloproteinase and disintegrin are derived from the proenzyme of snake venom metalloproeinase. Acugrin exhibits a certain amino acid homology with other disintegrins from snake venoms. Acugrin has four disulfide bonds and a consensus motif: RGD. The cDNA for the acugrin was cloned into the plasmid pT7ZZa and expressed in E.coli. ZZ acugrin fusion protein could strongly inhibit aggregation of platelet induced by ADP and the IC 50 was about 4 nmol/L.