文献类型：期刊文献

中文题名：Serratia marcescens H30脂肪酶的重组可溶表达、固定化及其酶学性质

英文题名：Recombinant soluble expression,immobilization and characterization of lipase from Serratia marcescens H30

作者：徐晶晶[1];苏二正[2];吴向萍[1];马昱澍[1];魏东芝[1]

机构：[1]华东理工大学鲁华生物技术研究所生物反应器工程国家重点实验室,上海200237;[2]南京林业大学轻工科学与工程学院酶与发酵工程实验室,南京210037

年份：2014

卷号：12

期号：6

起止页码：33

中文期刊名：生物加工过程

外文期刊名：Chinese Journal of Bioprocess Engineering

收录：CSTPCD;;CSCD:【CSCD_E2013_2014】；

基金：国家重点基础研究发展计划(973计划)(2012CB721103;2012CB721003);国家高技术研究发展计划(863计划)(2012AA022206B;2012AA020403);国家自然科学基金(31201296/C200101);中央高校基本科研业务费专项资金;生物反应器工程国家重点实验室开放课题

语种：中文

中文关键词：黏质沙雷氏菌;脂肪酶;可溶表达;固定化;手性拆分

外文关键词：Serratia marcescens;lipase;soluble expression;immobilization;characterization;resolution

摘要：为了提高Serratia marcescens H30脂肪酶的可溶表达水平,分别将目的基因与p GEX-4T-1、p ET28a和p ET32a构建重组表达载体,转入大肠杆菌BL21(DE3),通过优化诱导过程,发现可溶性酶的最高活性可达25 000 U/L。再经Ni2+亲和柱纯化、LH-EP固定化后,固定化酶的比酶活为214 U/g(以1 g湿质量计),酶活回收率为51%。固定化后重组脂肪酶的最适温度由30℃提高到35℃,最适p H从7.0偏移至8.0左右,并且稳定性也有所增加。该固定化重组脂肪酶同样能够拆分消旋体反式-4-甲氧苯基缩水甘油酸甲酯,光学选择性没有改变。反应14 h,转化率为48.5%,底物的e.e.值为99.2%,表明该固定化脂肪酶能有效拆分消旋体反式-4-甲氧苯基缩水甘油酸甲酯,为工业生物催化制备地尔硫卓提供了可能。
To enhance the recombinant soluble expression of lipase from Serratia marcescens H30 in E-coli, different expression vectors such as pGEX-4T-1, pET28a and pET32a were studied-After optimization of inducing conditions, the maximum lipase activity reached to 25 000 U/L in shake flasks-The recombinant lipase was purified by Ni-NTA superflow column-Two carriers were used to immobilize the recombinant lipase, and LH-EP was the best one-Optimal temperature and pH of the immobilized lipase were 35 ℃ and 8-0, respectively-The thermal and pH stability of the lipase was improved after being immobilized onto LH-EP-The kinetic resolution of （ ＆#177; ）-MPGM by immobilized lipase was studied, and a conversion of 48-5% was achieved along with an e-e. value of 98-2%-Our study reveals the good potential of immobilized Serratia marcescens H30 lipase for application in industrial＆amp;nbsp;production of diltiazem.