谷胱甘肽S-转移酶的诱导表达及提纯  

Expression and Purification of Glutathione S-Transferase

作　　者：杨丹[1] 叶佩莹[2] 万津[1] 刘永平[1] 马辉文[1] 

机构地区：[1]武汉大学药学院,湖北武汉430072 [2]武汉大学生命科学学院,湖北武汉430072

出　　处：《氨基酸和生物资源》2004年第3期33-37,共5页Amino Acids & Biotic Resources

摘　　要：含有日本血吸虫谷胱甘肽S -转移酶基因的质粒 pGEX - 5X - 1在大肠杆菌BL2 1 (DE3)菌株中得到表达。 37℃[1 ]下用 1 %乳糖诱导谷胱甘肽S -转移酶 (GST)的最适表达时间为 3h。盐析粗提酶液并以Habig法[2 ] 监测GST的活性 ,用pH6 .0 ,饱和度为 30 %硫酸铵去除杂蛋白 ,并调 pH7.5 ,饱和度 70 %硫酸铵使GST大量析出并保持活性。用透析法脱盐 ,并用Sephadex -G5 0凝胶对GST进行了初步纯化。Glutathione S-transferase (GST)gene of Schistosoma japonicum harboring in the plasmid pGEX-5X-1 was expressed in Escherichia coli BL21(DE3) induced with lactose. The optimal inducing time was 3 hours at the temperature of 37℃. The expressed GST was salted out by 70% saturated ammonium sulphate, dialysed and partially purified with Sephadex-G50 gel filtration-chromatography. The enzymatic activity of GST in the cell lysate was determined with spectrophotometericrely described by Habig et al. and the expressing level of GST evaluated with SDS-PAGE was used as indexes to optimize the process of induction and purification of GST.

关 键 词：谷胱甘肽S-转移酶 基因 表达 纯化 质粒 大肠杆菌 

分 类 号：Q814[生物学—生物工程] Q78