HIV-1核心蛋白的表达与纯化  

Expression and Isolation of HIV-1 gag p24 Protein in E. coli

作　　者：金宁一[1] 李体远[1] 刘晓明[1] 王玉红[1] 王宏伟[1] 郭志儒[1] 安汝国[2] 殷震[1] 

机构地区：[1]解放军农牧大学研究所病毒室,长春130062 [2]白求恩医科大学,长春130021

出　　处：《高技术通讯》2000年第1期28-31,共4页Chinese High Technology Letters

基　　金：国家自然科学基金!( 3 9770 6 6 1);杰出青年基金!( 3 982 5 119)资助项目

摘　　要：将编码人I型免疫缺陷病毒(HIV1)核心蛋白p24gag的基因序列克隆到原核表达载体pET28(b)中,高效表达了N,C端融合His·Tagp24蛋白,所表达的重组p24蛋白占菌体总蛋白的46%。在变性条件下,使用NiNTA亲和层析法纯化了p24蛋白,纯度为94%。菌体中及纯化、复性后的目的蛋白均能与抗HIV1p24单克隆抗体发生特异性反应。用纯化的p24蛋白免疫小鼠,4周时小鼠血清抗p24抗体效价达1∶400。实验结果表明:大肠杆菌表达的HIV1p24蛋白纯化后可用作HIV1检测试剂的原料。The p24gag gene fragment encoding the capsid protein of HIV 1 was cloned to the pET 28(b) plasmid and overexpressed at a level of approximately 46% of the total cellular protein in E. coli BL21(DE3)plysS. A procedure, which allows the rapid purification of the protein fused to both amino and carboxy terminal tag of six histidines, is described. Extracts from the induced E. coli were loaded onto Ni 2+ nitrilotriacetic acid (Ni 2+ NTA) agarose and histidine tagged proteins were selectively eluted to >94% purity under denaturation condition. The purified p24 gag protein could be recognized by anti p24 antibody in western blot, and was capable of priming mice for the humoral response to HIV 1 gag protein expressed in the recombinant vaccinia viruses. The part of experiment promises to be useful for the preparation of p24 gag specific antibody and diagnostic material.

关 键 词：大肠杆菌 纯化 基因表达 HZV-1 核心蛋白 

分 类 号：Q939.91[生物学—微生物学]