文献类型：期刊文献

英文题名：Biocatalytic Properties of a Recombinant Fusarium proliferatum Lactonase with Significantly Enhanced Production by Optimal Expression in Escherichia coli

作者：Chen, Bing[1];Fan, Li-Qiang[1];Xu, Jian-He[1];Zhao, Jian[1];Zhang, Xian[1];Ouyang, Li-Ming[1]

机构：[1]E China Univ Chem Technol, State Key Lab Bioreactor Engn, Lab Biocatalysis & Bioproc, Shanghai 200237, Peoples R China

年份：2010

卷号：162

期号：3

起止页码：744

外文期刊名：APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY

收录：;EI(收录号：20103013097532);WOS:【SCI-EXPANDED(收录号:WOS:000278178500012)】；

基金：This research was financially supported by National Natural Science Foundation of China (grant No. 20506037 & 20773038), Ministry of Science and Technology (grant No. 2007AA02Z225 & 2009CB724706), and China National Special Fund for State Key Laboratory of Bioreactor Engineering (grant No. 2060204).

语种：英文

外文关键词：Levo-lactonase; DL-pantoyl lactone; Recombinant protein; Immobilized metal affinity chromatography; Biocatalytic property; Substrate specificity

摘要：The levo-lactonase gene of Fusarium proliferatum ECU2002 (EC3.1.1.25) was cloned and expressed in Escherichia coli JM109 (DE3) for biocatalytic resolution of industrially important chiral lactones, including DL-pantoyl lactone which was a key precursor to calcium d-pantothenate. By increasing the biomass concentration and lowering the inducer (isopropyl-beta-d-thiogalactoside) concentration and induction temperature, the lactonase production was significantly enhanced up to 20 kU/L, which was 20 times higher than that of wild-type strain F. proliferatum ECU2002. The recombinant Fusarium lactonase was purified using immobilized metal affinity chromatography, and its SDS-PAGE revealed a molecular mass of 50 kDa for the recombinant protein, suggesting that the enzyme was a simplex protein. Furthermore, biocatalytic properties of the recombinant lactonase were investigated, including kinetic parameters, additive's effect, and substrate specificity. The results reported in this paper provide a feasible method to make the whole cells of E. coli JM109 (DE3) expressing lactonase gene to be a highly efficient and easy-to-make biocatalyst for asymmetric synthesis of chiral compounds.