HCV NS5B蛋白对不同HCV基因型来源RNA模板复制活性的比较

The Comparison of the RdRp Activities of HCV NS5B Protein Between RNA Templates from Different HCV Genotypes

目的分析HCV NS5B蛋白以不同HCV型来源的负链3’末端RNA为模板的复制活性,为优化以NS5B为靶点的抗HCV药物模型提供理论依据。方法利用原核表达系统表达HCV NS5B蛋白,Ni-NTA亲和层析柱纯化,分别以体外转录的1a(-)3′T RNA、1b(-)3′T RNA和2a(-)3′T RNA为模板进行体外RdRP反应,Northen blot检测RdRp产物。结果当以2a(-)3′T RNA为模板时,RdRp产物量最高,约为以1a(-)3′T RNA为模板时的RdRp产物量的220%,当以1b(-)3′T RNA为模板时产物量最低,约为1a(-)3′T RNA为模板时的RdRp产物量的60%。结论HCV NS5B对不同基因型来源的RNA模板具有选择性。

Objective To compare the RdRp activities of HCV NS5B protein to three RNA templates from different HCV genotypes in vitro and offer some useful data to optimize the model for screening of anti - HCV drugs targeting at NS5B. Methods HCV NS5B was prokaryon - expressed and purified by the Ni - NTA affinity chromatography, its activities to three RNA templates transcripted via in vitro was detected through RdRp assay in vitro, Northem blot was employed to detect the RdRp products. Results The RdRp production from 2a（ - ）3＇T RNA is 220 percent of the production from la（ - ）3＇T RNA and the production from lb（ - ）3＇T RNA is 60 percent of it. Conclusion HCV NS5B showed distinct activities to RNA templates from different HCV genotyoes.