人S100A6-GST融合蛋白的原核表达和纯化  

Expression and purification of human S100A6-GST fusion protein

作　　者：苗静琨[1] 赖天霞[1] 左国伟[1] 李星星[1] 王嫣[1] 蒋薇[2] 何通川[2] 周兰[1] 

机构地区：[1]重庆医科大学生物医学工程系,重庆400016 [2]美国芝加哥大学医学中心分子肿瘤研究室,美国chicagoil60637

出　　处：《重庆医科大学学报》2007年第10期1009-1012,共4页Journal of Chongqing Medical University

基　　金：国家自然科学基金(30340039);2003年教育部"春晖计划"科研启动经费资助项目;2004年重庆市留学回国人员启动基金。

摘　　要：目的:制备人S100A6-GST融合蛋白,为研究人S100A6((hS100A6)蛋白的功能奠定基础。方法:从pHAHA-hS100A6中双酶切获取hS100A6基因,亚克隆至原核表达载体pGST-moluc,构建pGST-moluc-hS100A6,转化BL21后,IPTG诱导重组菌表达hS100A6蛋白,SDS-PAGE及WesternBlot鉴定表达产物,Glutathion-Sepharose4B球珠分离纯化。结果:重组菌株表达出与理论预测值相符的一个约36ku的hS100A6融合蛋白,纯化后的融合蛋白的产量为3mg/L菌液,纯度约92%。结论:成功构建了hS100A6-GST原核表达质粒,在大肠杆菌中表达纯化后得到较高浓度的hS100A6-GST融合蛋白,为hS100A6蛋白功能的研究打下了基础。Objective：To obtain human S100A6-GST fusion protein as material of the study of function of hS100A6. Methods：The hS100A6 gene from pHAHA-hS100A6 was subcloned into the prokaryotic GST fusion protein expression plasmid, pGST-moluc,to form pGST-moluc-hS100A6, The recombinant plasmid was transformed to E. coli BL21 and the expression of hS100A6-GST fusion protein was induced with IPTG. The protein was detected by SDS-PAGE and purified with Glutathion-Sepharose 4B beads,then identified by western blot assays. Results：A 36 000 Dalton protein,as expected, was obtained evidently. The concentration of the purified fusion protein was 3mg/L bacterial culture, and the purity was about 92%, Conclusion：A prokaryotic system expressing hSI00A6-GST fusion protein was successfully constructed, hS100A6-GST fusion protein with high purity could be obtained after it was efficiently expressed in E. coli BL21 and purified with Glutathion-Sepharose 4B beads. This will facilitate our study of the function of hS100A6.

关 键 词：人S100A6基因 融合蛋白 原核表达 蛋白纯化 

分 类 号：Q786[生物学—分子生物学]