酵母蔗糖酶的提取方法_李楠.pdfVIP

第26 卷第4 期 食 品 与生 物 技 术学 报 Vol.26　No.4 　 　　　　　　　 2007 年7 月 Journal of Food Science and Biotechnology 　　　　　　 Jul.　2007 :1673-1689(2007)04-0083-05 　 酵母蔗糖酶的提取方法 李 楠, 　庄苏星, 　 丁益 (南京大学生命科学学院, 医药生物技术国家重点实验室, 江苏 南京210093) 　:采用甲苯自溶法、冻融法、SDS 抽提法3 种方法从酵 中提取蔗糖酶, 经质量分数50%的 乙醇分级沉淀﹑Mono Q 阴离子交换柱层析纯化后, 制得高纯度的酵 蔗糖酶。比较了上述 3 种 提取方法并对该酶的部分性质进行了研究。纯化的酶经等电聚焦测定, 等电点为5.6, SDS- AGE 鉴定其相对分子质量为60 kD。以蔗糖为底物测得酵 蔗糖酶的表观米氏常数K m 为0.013 mol/ L。结果显示3 种提取方法各有优劣, 冻融法和SDS 抽提法的提取效率远高于传统的甲苯自溶法。 其中SDS 抽提法的效率最高, 加之其操作简便, 更适合于酵 蔗糖酶大规模的制备提取。 :酵母;蔗糖 ;提取;纯化 :Q 556.2;TS 201.25 :A Study on Purification of Invertase from Yeast LI Nan, 　ZHUANG Su-xing, 　DING Yi (School of Life Sciences, State Key Laboratory of harmaceutical Biotechnology, Nanjing University, Nanjing 210093, China) Abstract:T his paper used three methods to extract invertase from yeast, which included in this manuscript, three different extraction method breaking cells by adding methylbenzene, frost grinding, and adding SDS for extracting invertase from yeast were investigated.Then the purified invertase was obtained by precipitatation with 50% ethyl alcohol and Mono Q chromatography .The molecular weight of the enzyme was 60kD by SDS- AGE, and the pI was 5.6 by isoelectric focusing.The invertase followed typical Michaelis-Menten Kinetics with apparent K m of 0.013 mol/L.The results showed all three methods had both advantages and disadvantages.The invertase extracted by adding SDS and frost grinding had much more total activity than that of extracted by adding methylbenzene.A highest total invertase activity was found in the SDS extraction process, and it was a convenient and economical method for commerical producti