摘要
从草鱼肝胰脏中分离纯化出一种胰蛋白酶.纯化过程包括:硫酸铵分级沉淀,DEAE-Sepharose离子交换,Sephacryl S-200凝胶过滤和Q-Sepharose离子交换层析.SDS-PAGE显示在分子量约为28.5 ku处有单一电泳带,表明该酶已得到高度纯化.以Boc-Leu-Arg-Arg-MCA为底物测得该酶的最适温度为40℃,最适pH值为9.0.丝氨酸蛋白酶抑制剂(Pefab loc SC,PMSF,Benzam id ine)对该酶的活力有明显抑制作用.底物特异性实验表明该酶特异性分解精氨酸和赖氨酸残基的C末端,进一步证明纯化蛋白为草鱼胰蛋白酶,其酶学性质与鲤鱼胰蛋白酶相似.
An anionic trypsin was purified from the hepatopancreas of Grass carp. The purification procedures consisted of ammonium sulfate fractionation, and chromatographies on DEAE-Sepharose fast flow, Sephacryl S-200 and Q-Sepharose. Grass carp trypsin was purified to high homogeneity with the molecular mass of approximately 28.5 ku. The optimum temperature of the trypsin was 40℃ using Boc-Leu-Arg-Arg-MCA as substrate and the optimum pH was 9.0. Substrate specificity analysis revealed that the trypsin specifically cleaved at the carboxyl side of the arginine and lysine residues. Inhibitor susceptibility analysis showed that the trypsin was inhibited effectively by serine protease inhibitors such as Pefabloc SC, PMSF and benzamidine. Enzymatic properties of the enzyme were similar to common carp trypsins.
出处
《集美大学学报（自然科学版）》
CAS
2005年第4期300-304,共5页
Journal of Jimei University：Natural Science
基金
福建省自然科学基金资助项目(C0410036)
集美大学科研启动基金资助项目(F03003)
