title: Cloning and characterisation of a novel 2,4-dichlorophenol hydroxylase from a metagenomic library derived from polychlorinated biphenyl-contaminated soil.
abstract: A novel 2,4-dichlorophenol hydroxylase (TfdB, EC 1.14.13.20) gene, designated as tfdB-JLU, was identified from a metagenome constructed from polychlorinated biphenyl-contaminated soil by functional screening and heterologously expressed in Escherichia coli. The deduced amino acid sequence of tfdB-JLU exhibited less than 48% homology with other known TfdBs. The enzyme exhibited a wider substrate spectrum than the previously reported TfdBs and higher relative activity towards ortho-substituted dichlorophenols, 2-chlorophenol, and 3-chlorophenol than towards 2,4-dichlorophenol, the preferred substrate of other known TfdBs. The enzyme had a K ( m ) of 5 μM for 2,4-dichlorophenol and 6 μM for NADPH. The optimal temperature and pH of the enzyme were 25°C and 7.5, respectively. Activity of the purified TfdB-JLU was slightly enhanced by Ca(2+), Mn(2+), Co(2+), and Fe(2+), and completely inhibited by Cu(2+), Hg(2+), and Zn(2+). This study is the first report to identify a novel TfdB from a metagenome.
authors.affiliation: Key Laboratory for Molecular Enzymology and Engineering of the Ministry of Education, Jilin University, 2699 Qianjin Street, Changchun, 130012, People's Republic of China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Lu
authors.forename: Ying
authors.initials: Y
authors.lastname: Yu
authors.forename: Rui
authors.initials: R
authors.lastname: Zhou
authors.forename: Wei
authors.initials: W
authors.lastname: Sun
authors.forename: Chunyan
authors.initials: C
authors.lastname: Dai
authors.forename: Pei
authors.initials: P
authors.lastname: Wan
authors.forename: Lanying
authors.initials: L
authors.lastname: Zhang
authors.forename: Dongyun
authors.initials: D
authors.lastname: Hao
authors.forename: Hejun
authors.initials: H
authors.lastname: Ren
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001673:Biodegradation, Environmental; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D006540:Herbicides / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D054892:Metagenome; D056186:Metagenomics; D006899:Mixed Function Oxygenases / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D011078:Polychlorinated Biphenyls / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012988:Soil Microbiology; D012989:Soil Pollutants / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D017931:DNA Primers; D006540:Herbicides; D011994:Recombinant Proteins; D012989:Soil Pollutants; D011078:Polychlorinated Biphenyls; D006899:Mixed Function Oxygenases; C034354:2,4-dichlorophenol hydroxylase