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