HIV-1 Tat-TAR结合抑制剂筛选模型的构建  

Model Establishment for Screening HIV-1 Tat-TAR Interaction Inhibitors

作　　者：徐皓亮[1] 黄永平[1] 韩伟[1] 刘建文[1] 肖蒙蒙[1] 

机构地区：[1]华东理工大学药学院,上海200237

出　　处：《华东理工大学学报（自然科学版）》2011年第2期175-178,共4页Journal of East China University of Science and Technology

基　　金：上海市浦江人才计划项目(06PJ14026)

摘　　要：将HIV-1 Tat(1-86aa)编码序列插入原核表达质粒pET28a中,转化E.coliBL21(DE3),IPTG诱导Tat蛋白表达,并用Ni+金属螯合层析介质纯化;把TAR DNA序列插入到含有T7启动子的载体pBluescript Ⅱ SK(+)中,用T7 RNA多聚酶体外转录合成TAR RNA。用毛细管电泳观察Tat与TAR的结合活性,并用转录抑制活性化合物AP064检验对Tat-TAR结合的阻断作用。结果显示,制备的Tat和TAR能相互结合,并被AP064有效阻断,表明此模型可用于筛选抑制Tat-TAR结合的化合物。HIV-1 Tat（1-86aa） coding sequence was inserted into the prokaryotic expression plasmid pET28a and transformed E.coli BL21（DE3）.Tat protein expression was induced by IPTG and purified with Ni-NTA column affinity chromatography.Then,the TAR sequence was inserted into pBluescript II SK（＋）,which contain T7 promoter.The TAR RNA was synthesized by T7 RNA polymerase transcription in vitro.The binding activity of Tat with TAR was observed by capillary electrophoresis,positive compound AP064 was used to test the blocking effect on Tat-TAR binding.The results show that Tat can combine TAR effectively,and AP064 can block the binding.So the model can be used as screening compounds which can inhibit the binding of Tat and TAR.

关 键 词：TAT蛋白 TAR RNA 毛细管电泳 

分 类 号：Q812[生物学—生物工程]