Molecular Cloning and Characterization of a New Cold-active Extradiol Dioxygenase from a Metagenomic Library Derived from Polychlorinated Biphenyl-contaminated Soil  

作　　者：REN He-jun LU Yang ZHOU Rui DAI Chun-yan WANG Yah ZHANG Lan-ying 

机构地区：[1]Key Laboratory of Groudwater Resources and Environment,Ministry of Education,College of Environment and Resources,Jilin University,Changehun 130021,P.R.China [2]Jilin Academy of Agricultural Sciences,Changehun 130033,P.R.China [3]Institute of Virology and AIDS Research,the First Hospital of Jilin University,Changchun 130061,P.R.China

出　　处：《Chemical Research in Chinese Universities》2012年第4期666-671,共6页高等学校化学研究（英文版）

基　　金：Supported by the National Natural Science Foundation of China(Nos.41101226,50879029);the Technology Development Project of Jilin Province,China(Nos.201101020,20090415)

摘　　要：To find new extradiol dioxygenases（EDOs,EC 1.13.11.2）,a metagenomics library was constructed from polychlorinated biphenyl-contaminated soil and was screened for some dioxygenase with aromatic ring cleavage activity.A novel EDO,designated as BphC_A,was identified and heterologously expressed in Escherichia coli.The deduced amino acid sequence of BphC_A exhibited a homology of less than 60% with other known EDOs.Phylogenetic analysis of BphC_A suggests that the protein is a novel member of the EDO family.The enzyme exhibits higher substrate affinity and catalytic efficiency toward 3-methylcatechol than toward 2,3-dihydroxybiphenyl or catechol,the preferred substrate of other known EDOs.The optimum activity of purified BphC_A occurred at pH=8.5 and 35 °C,and BphC_A showed more than 40% of its initial activity at 5 °C.The activity of purified BphC_A was significantly induced by Mn^2＋ and slightly reduced by Al^3＋,Cu^2＋ and Zn^2＋.To find new extradiol dioxygenases（EDOs,EC 1.13.11.2）,a metagenomics library was constructed from polychlorinated biphenyl-contaminated soil and was screened for some dioxygenase with aromatic ring cleavage activity.A novel EDO,designated as BphC_A,was identified and heterologously expressed in Escherichia coli.The deduced amino acid sequence of BphC_A exhibited a homology of less than 60% with other known EDOs.Phylogenetic analysis of BphC_A suggests that the protein is a novel member of the EDO family.The enzyme exhibits higher substrate affinity and catalytic efficiency toward 3-methylcatechol than toward 2,3-dihydroxybiphenyl or catechol,the preferred substrate of other known EDOs.The optimum activity of purified BphC_A occurred at pH=8.5 and 35 °C,and BphC_A showed more than 40% of its initial activity at 5 °C.The activity of purified BphC_A was significantly induced by Mn^2＋ and slightly reduced by Al^3＋,Cu^2＋ and Zn^2＋.

关 键 词：Extradiol dioxygenase METAGENOME Cold-active enzyme Gene cloning Functional characterization 

分 类 号：Q933[生物学—微生物学] X172[环境科学与工程—环境科学]