Identification and characterization of a meta-cleavage product hydrolase involved in biphenyl degradation from Arthrobacter sp. YC-RL1

Autor:	Yanchun Yan, Jiayi Wang, Ruth Nahurira, Chao Ren, Ibatsam Khokhar, Junhuan Wang, Yang Jia, Shuanghu Fan
Rok vydání:	2019
Předmět:	Hydrolases Stereochemistry DNA Mutational Analysis Gene Expression medicine.disease_cause Applied Microbiology and Biotechnology 03 medical and health sciences chemistry.chemical_compound Catalytic Domain Enzyme Stability Catalytic triad Hydrolase Escherichia coli medicine Enzyme kinetics Arthrobacter Cloning Molecular Enzyme Inhibitors Biotransformation 030304 developmental biology Biphenyl chemistry.chemical_classification 0303 health sciences 030306 microbiology Chemistry Biphenyl Compounds Temperature General Medicine Hydrogen-Ion Concentration Biodegradation Fungicides Industrial Kinetics Enzyme Mutagenesis Site-Directed bacteria PMSF Biotechnology
Zdroj:	Applied Microbiology and Biotechnology. 103:6825-6836
ISSN:	1432-0614 0175-7598
DOI:	10.1007/s00253-019-09956-z
Popis:	Polychlorinated biphenyls (PCBs) are a group of persistent organic pollutants (POPs) widely existing in the environment. Arthrobacter sp. YC-RL1 is a biphenyl-degrading bacterium that shows metabolic versatility towards aromatic compounds. A 2-hydroxy-6-oxo-6-phenylhexa-2, 4-dienoate (HOPDA) hydrolase (BphD) gene involved in the biodegradation of biphenyl was cloned from strain YC-RL1 and heterologously expressed in Escherichia coli BL21 (DE3). The recombinant BphDYC-RL1 was purified and characterized. BphDYC-RL1 showed the highest activity at 45 °C and pH 7. It was stable under a wide range of temperature (20–50 °C). The enzyme had a Km value of 0.14 mM, Kcat of 11.61 s−1, and Vmax of 0.027 U/mg. Temperature dependence catalysis exhibited a biphasic Arrhenius Plot with a transition at 20 °C. BphDYC-RL1 was inactivated by SDS, Tween 20, Tween 80, Trition X-100, DTT, CHAPS, NBS, PMSF, and DEPC, but insensitive to EDTA. Site-directed mutagenesis of the active-site residues revealed that the catalytic triad residues (Ser115, His275, and Asp247) of BphDYC-RL1 were necessary for its activity. The investigation of BphDYC-RL1 not only provides new potential enzyme resource for the biodegradation of biphenyl but also helps deepen our understanding on the catalytic process and mechanism.
Databáze:	OpenAIRE
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