Metabolic Pathway Involved in 6-Chloro-2-Benzoxazolinone Degradation by Pigmentiphaga sp Strain DL-8 and Identification of the Novel Metal-Dependent Hydrolase CbaA
Metabolic Pathway Involved in 6-Chloro-2-Benzoxazolinone Degradation by Pigmentiphaga sp Strain DL-8 and Identification of the Novel Metal-Dependent Hydrolase CbaA
复制标题
色素噬菌体 DL-8 菌株降解 6-氯-2-苯并恶唑啉酮的代谢途径及新型金属依赖性水解酶 CbaA 的鉴定
DOI：
10.1128/aem.00532-16
复制
发表时间：
2016
影响因子：
4.4
通讯作者：
Cui Zhongli
中科院分区：
文献类型：
--
作者：
Dong Weiliang;Wang Fei;Huang Fei;Wang Yicheng;Zhou Jie;Ye Xianfeng;Li Zhoukun;Hou Ying;Huang Yan;Ma Jiangfeng;Jiang Min;Cui Zhongli
6-Chloro-2-benzoxazolinone (CDHB) is a precursor of herbicide, insecticide, and fungicide synthesis and has a broad spectrum of biological activity. Pigmentiphaga sp. strain DL-8 can transform CDHB into 2-amino-5-chlorophenol (2A5CP), which it then utilizes as a carbon source for growth. The CDHB hydrolase (CbaA) was purified from strain DL-8, which can also hydrolyze 2-benzoxazolinone (BOA), 5-chloro-2-BOA, and benzamide. The specific activity of purified CbaA was 5,900 U · mg protein−1for CDHB, withKmandkcatvalues of 0.29 mM and 8,500 s−1, respectively. The optimal pH for purified CbaA was 9.0, the highest activity was observed at 55°C, and the inactive metal-free enzyme could be reactivated by Mg2+, Ni2+, Ca2+, or Zn2+. Based on the results obtained for the CbaA peptide mass fingerprinting and draft genome sequence of strain DL-8,cbaA(encoding 339 amino acids) was cloned and expressed in Escherichia coli BL21(DE3). CbaA shared 18 to 21% identity with some metal-dependent hydrolases of the PF01499 family and contained the signature metal-binding motif Q127XXXQ131XD133XXXH137. The conserved amino acid residues His288 and Glu301 served as the proton donor and acceptor. E. coli BL21(DE3-pET-cbaA) resting cells could transform 0.2 mM CDHB into 2A5CP. The mutant strain DL-8ΔcbaAlost the ability to degrade CDHB but retained the ability to degrade 2A5CP, consistent with strain DL-8. These results indicated thatcbaAwas the key gene responsible for CDHB degradation by strain DL-8.IMPORTANCE2-Benzoxazolinone (BOA) derivatives are widely used as synthetic intermediates and are also an important group of allelochemicals acting in response to tissue damage or pathogen attack in gramineous plants. However, the degradation mechanism of BOA derivatives by microorganisms is not clear. In the present study, we reported the identification of CbaA and metabolic pathway responsible for the degradation of CDHB in Pigmentiphaga sp. DL-8. This will provide microorganism and gene resources for the bioremediation of the environmental pollution caused by BOA derivatives.
