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.
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)
