低温脂肪酶的分离纯化及酶学性质

Purification and Enzymatic Properties of Cryogenic Lipase

投稿时间：2012-10-30

DOI：

中文关键词: 脂肪酶 分离纯化 酶学性质

英文关键词: lipase purification enzymatic properties

基金项目:国家863计划项目(2007AA021306)

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中文摘要:

苏云金芽胞杆菌CZW001脂肪酶发酵上清液经硫酸铵沉淀、透析、超滤离心、DEAE-纤维素-52离子交换层析和Sephadex G-100凝胶过滤层析得到电泳纯的脂肪酶,纯化倍数为75.5倍,活性回收率为37.6%,12%SDS-PAGE电泳估计其相对分子质量约40 000。对纯化后的脂肪酶进行酶性质研究表明:酶的最适作用温度为25℃,对热敏感,60℃处理30 min仅残留30%酶活性;酶的适宜作用pH范围在7～9,最适pH为8;该脂肪酶的水解活性对Ca2+表现明显的依赖性,Al3+、Zn2+和Fe2+对脂肪酶有显著的抑制作用;脂肪酶对醇类有机溶剂的耐受性随碳链增加而减小;脂肪酶对豆油表现出显著的特异性,而蓖麻油抑制脂肪酶水解活力。

英文摘要:

A lipase from Bacillus thuringiensis CZW001 was purified to homogeneity using ammonium sulfate precipitation,dialysis,DEAE-cellulose-52 anion exchange chromatography and Sephadex G-75 gel filtration chromatography.This purification protocol resulted in a 75.5 fold purification of lipase with 37.6% final yield,and the relative molecular weight of the enzyme was determined to be approximately 40kD using SDS-PAGE.Preliminary stuies have shown that the enzymatic properties of bacteria lipase,the enzyme’s optimum temperature is 25 ℃ and the enzyme was thermal lability,only 30% of its activity was remained after 30 min incubation at 60 ℃.The enzyme showed high lipolytic from ph 7.0 to 9.0 and its optimal ph for activity was 8.0.Ca2+ions stimulated lipolytic activity,whereas Al3+、Zn2+and Fe2+ions caused inhibition.Lipase tolerance of organic solvents of alcohols with carbon chain decreases.Lipase on soybean oil showed a significant the hydrolysis of the lipase.

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