Hydrolysis mechanism of carbendazim hydrolase from the strain Microbacterium sp.djl-6F  

作　　者：Ji Lei Shaopeng Wei Lijun Ren Shibin Hu Peng Chen 

机构地区：[1]Key Laboratory of Disaster Survey and Mechanism Simulation of Shaanxi Province,College of Chemistry and Chemical Engineering,Baoji University of Arts and Sciences [2]College of Resources and Environment,College of Life Sciences and College of Plant Protection,Northwest A&F University

出　　处：《Journal of Environmental Sciences》2017年第4期171-177,共7页环境科学学报（英文版）

基　　金：supported by the Hi-Tech Research and Development Program(863) of China(No.2012AA101404);the National Natural Science Foundation of China(No.31301700);the Science and Technology Program of BaoJi(No.2013R6-3);the Key Subject Project of Baoji University of Arts and Sciences(No.ZK0919)

摘　　要：The carbendazim（MBC） hydrolyzing enzyme gene was cloned and heterologously expressed in Escherichia coli BL21（DE3） from a newly isolated MBC-degrading bacterium strain Microbacterium sp. strain djl-6F. High performance liquid chromatography-mass spectrometry（HPLC-MS）analysis revealed that purified MheI-6F protein catalyzes direct hydrolysis of MBC into2-aminobenzimidazole（2-AB） with a high turnover rate and moderate affinity（Kmof6.69 μmol/L and kcatof 160.88/min） without the need for any cofactors. The optimal catalytic condition of MheI-6F was identified as 45°C, pH 7.0. The enzymatic activity of MheI-6F was found to be diminished by metal ions, and strongly inhibited by sodium dodecyl sulfate（SDS）.Through generating amino acid mutations in MheI-6F, Cys16 and Cys222 were identified as the catalytic groups that are essential for the hydrolysis of MBC. This is the first report on the biodegradation of MBC at the enzymatice level.The carbendazim（MBC） hydrolyzing enzyme gene was cloned and heterologously expressed in Escherichia coli BL21（DE3） from a newly isolated MBC-degrading bacterium strain Microbacterium sp. strain djl-6F. High performance liquid chromatography-mass spectrometry（HPLC-MS）analysis revealed that purified MheI-6F protein catalyzes direct hydrolysis of MBC into2-aminobenzimidazole（2-AB） with a high turnover rate and moderate affinity（Kmof6.69 μmol/L and kcatof 160.88/min） without the need for any cofactors. The optimal catalytic condition of MheI-6F was identified as 45°C, pH 7.0. The enzymatic activity of MheI-6F was found to be diminished by metal ions, and strongly inhibited by sodium dodecyl sulfate（SDS）.Through generating amino acid mutations in MheI-6F, Cys16 and Cys222 were identified as the catalytic groups that are essential for the hydrolysis of MBC. This is the first report on the biodegradation of MBC at the enzymatice level.

关 键 词：Carbendazim Microbacterium sp.djl-6F Hydrolase MheI-6F 

分 类 号：X172[环境科学与工程—环境科学]