重组蛋白sHLA-G1的原核表达与纯化  

Prokaryotic expression and purification of recombinant protein soluble HLA-G1

作　　者：张劲[1] 郭霭光[1] 丰美福[2] 

机构地区：[1]西北农林科技大学生命科学学院,陕西杨凌712100 [2]中国科学院动物研究所生物膜与膜生物工程国家重点实验室,北京100080

出　　处：《西北农林科技大学学报（自然科学版）》2007年第8期180-184,共5页Journal of Northwest A&F University(Natural Science Edition)

基　　金：国家自然科学基金项目(30370383)

摘　　要：为了进一步在蛋白质水平对人白细胞抗原HLA-G进行分析,利用已经构建好的克隆质粒pGEM-T-HLA-G1,进一步构建了重组原核表达质粒pET28a-sHLA-G1。SDS-PAGE检测表明,重组蛋白sHLA-G1在大肠杆菌(E.coli)中得到稳定表达,原核表达的重组蛋白sHLA-G1主要以包涵体形式存在。包涵体蛋白经充分洗涤、尿素溶解后用钴离子树脂纯化,并用SDS-PAGE和Western印迹进行了分析,鉴定结果表明,该蛋白纯度达到90%以上,并能与HLA-G特异性抗体87G反应。该研究结果为进一步的复性工作奠定了基础。In order to study human leukocyte antigen （HLA-G） protein,on the basis of a recombinant cloning plasmid pGEM-T-HLA-G1, prokaryotic expression plasmid pET28a-sHLA-G1 was constructed, which could express soluble recombinant protein HLA-G1 in Escherichia coli （E. coli）. Most of His- sHLA-G1 existed in the inclusion body. The inclusion body was washed extensively and solubilized with u- rea,and then purified with Co^2＋ resin. Finally,the purified protein was analyzed by using SDS-PAGE and Western blot. The results indicated that the purity of His-sHLA-G1 could reach 90%,and the recombinant protein could react with monoclonal antigen 87G. This research provided the basis for the further refolding study on HI.A-G molecule.

关 键 词：HLA-G 原核表达 重组蛋白 包涵体 

分 类 号：Q78[生物学—分子生物学]