title: Characterization of a Novel Butachlor Biodegradation Pathway and Cloning of the Debutoxylase (Dbo) Gene Responsible for Debutoxylation of Butachlor in Bacillus sp. hys-1.
abstract: Bacillus sp. strain hys-1, which was isolated from active sludge, could degrade >90% butachlor at a concentration of 100 mg/L within 7 days. The present work revealed that strain hys-1 could mineralize butachlor via the following pathway: butachlor was initially metabolized to 2-chloro-N-(2,6-diethylphenyl)-N-methylacetamide by debutoxylation and then transformed to form 2-chloro-N-(2,6-diethylphenyl)acetamide by N-demethylation. Subsequently, it was converted to 2,6-diethylaniline and further mineralized into CO2 and H2O. In addition, the catalytic efficiency of crude cell extracts descended as follows: alachlor > acetochlor > butachlor. Furthermore, a novel 744 bp gene responsible for transforming butachlor into 2-chloro-N-(2,6-diethylphenyl)-N-methylacetamide was cloned from strain hys-1 and the encoding debutoxylase was designated Dbo. Then Dbo was expressed in Escherichia coli BL21 (DE3) and purified using Ni-nitrilotriacetic acid affinity chromatography. Dbo displayed the highest activity against butachlor at pH 6.5 and 30 °C. Metal ions played an important role in Dbo activity. To the best of the authors' knowledge, this is the first report that strain hys-1 can mineralize butachlor by a novel metabolic mechanism and the first identification of a gene encoding butachlor debutoxylase.
authors.affiliation: Zhejiang Ocean University , Zhoushan 316000, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Gao
authors.affiliation: Zhejiang Ocean University , Zhoushan 316000, China.|Marine Fishery Research Institute of Zhejiang Province , Zhoushan 316021, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Jin
authors.affiliation: Marine Fishery Research Institute of Zhejiang Province , Zhoushan 316021, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Shi
authors.affiliation: Zhejiang Ocean University , Zhoushan 316000, China.
authors.forename: Zhangjie
authors.initials: Z
authors.lastname: Chu
mesh_terms: D000083:Acetanilides / Q000737:chemistry / Q000378:metabolism*; D001407:Bacillus / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D001673:Biodegradation, Environmental; D003001:Cloning, Molecular; D004795:Enzyme Stability; D006540:Herbicides / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010802:Phylogeny; D013379:Substrate Specificity
chemical_list: D000083:Acetanilides; D001426:Bacterial Proteins; D006540:Herbicides; C054409:butachlor
keywords: Bacillus sp. hys-1; Dbo gene; butachlor; debutoxylation; enzyme purification; mineralization