Minh Le Thi Nguyet, Reza Md Abu, Swarup Sanjay, Kini R Manjunatha
Department of Biological Sciences, Faculty of Science, National University of Singapore, Singapore, Republic of Singapore.
Thromb Haemost. 2005 Mar;93(3):420-9. doi: 10.1160/TH04-11-0707.
The origin and evolution of venom toxins is a mystery that has evoked much interest. We have recently shown that pseutarin C, a prothrombin activator from Pseudonaja textilis venom, is structurally and functionally similar to mammalian coagulation factor Xa-factor Va complex. Its catalytic subunit is homologous to factor Xa while the nonenzymatic subunit is homologous to factor Va. P. textilis therefore has two parallel prothrombin activator systems: one expressed in its venom gland as a toxin and the other expressed in its liver and released into its plasma as a haemostatic factor. Here we report the complete amino acid sequence of factor V (FV) from its liver determined by cDNA cloning and sequencing. The liver FV shows 96% identity to pseutarin C nonenzymatic subunit. Most of the functional sites involved in its interaction with factor Xa and prothrombin are conserved. However, many potential sites of post-translational modifications and one critical cleavage site for activated protein C are different. The absence of the latter cleavage site makes pseutarin C nonenzymatic subunit resistant to inactivation and enhances its potential as an excellent toxin. By PCR and real-time quantitative analysis, we show that pseutarin C nonenzymatic subunit gene is expressed specifically in the venom gland at approximately 280 fold higher than that of FV gene in liver. These two are thus encoded by two separate genes that express in a highly tissue-specific manner. Our results imply that the gene encoding pseutarin C nonenzymatic subunit was derived by the duplication of plasma FV gene and they have evolved to perform distinct functions.
毒液毒素的起源与进化是一个引发诸多关注的谜团。我们最近发现，来自东部拟眼镜蛇毒液的凝血酶原激活剂——拟眼镜蛇毒素C，在结构和功能上与哺乳动物凝血因子Xa - 因子Va复合物相似。其催化亚基与因子Xa同源，而非酶亚基与因子Va同源。因此，东部拟眼镜蛇有两个平行的凝血酶原激活剂系统：一个在其毒腺中作为毒素表达，另一个在其肝脏中表达并作为止血因子释放到血浆中。在此，我们报告通过cDNA克隆和测序确定的来自其肝脏的因子V（FV）的完整氨基酸序列。肝脏FV与拟眼镜蛇毒素C非酶亚基的序列一致性为96%。其与因子Xa和凝血酶原相互作用所涉及的大多数功能位点是保守的。然而，许多潜在的翻译后修饰位点以及一个对活化蛋白C至关重要的切割位点有所不同。后者切割位点的缺失使得拟眼镜蛇毒素C非酶亚基对失活具有抗性，并增强了其作为优良毒素的潜力。通过PCR和实时定量分析，我们发现拟眼镜蛇毒素C非酶亚基基因在毒腺中特异性表达，其表达水平比肝脏中的FV基因高约280倍。因此，这两者由两个单独的基因编码，并以高度组织特异性的方式表达。我们的结果表明，编码拟眼镜蛇毒素C非酶亚基的基因源自血浆FV基因的复制，并且它们已经进化以执行不同的功能。
