Molecular cloning and functional characterization of the genes encoding benzoate and p-hydroxybenzoate degradation by the halophilic Chromohalobacter sp. strain HS-2.
Chromohalobacter sp. strain HS-2 was isolated from salted fermented clams and analyzed for the ability to grow on benzoate and p-hydroxybenzoate as the sole carbon and energy source. HS-2 was characterized as moderately halophilic, with an optimal NaCl concentration of 10%. The genes encoding the benzoate metabolism were cloned into a cosmid vector, sequenced, and then analyzed to reveal the benzoate (benABCD) and catechol (catBCA) catabolic genes, both of which are flanked on either side by LysR-type transcriptional regulator (catR) and membrane transport protein for benzoate (benE) in the gene order catRBCAbenABCDE. Near the large cat-ben cluster, a p-hydroxybenzoate hydroxylase gene (pobA) and two putative regulatory genes (pcaQ and pobR) were additionally detected. The HS-2 genes involved in benzoate and p-hydroxybenzoate degradation are tightly clustered within a c. 19 kb region, and show quite a different genetic organization from those of other benzoate catabolic genes. Reverse transcriptase-PCR experiments show that benzoate induces the expression of benzoate 1,2-dioxygenase, catechol 1,2-dioxygenase, and protocatechuate 3,4-dioxygenase while p-hydroxybenzoate only induced the expression of p-hydroxybenzoate hydroxylase. When expressed in Escherichia coli, benzoate 1,2-dioxygenase (BenABC) and p-hydroxybenzoate hydroxylase (PobA) transformed benzoate and p-hydroxybenzoate into cis-benzoate dihydrodiol and protocatechuate, respectively.
Kim
Dockyu
D
Department of Environmental Engineering, BK21 Team for Biohydrogen Production, Chosun University, Gwangju, Korea.
Kim
Si Wouk
SW
Choi
Ki Young
KY
Lee
Jong Suk
JS
Kim
Eungbin
E
D001565:Benzoates / Q000378:metabolism*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000254:growth & development; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005809:Genes, Regulator; D044129:Halomonadaceae / Q000201:enzymology / Q000235:genetics*; D010226:Parabens / Q000378:metabolism*
D001565:Benzoates; D010226:Parabens; C038193:4-hydroxybenzoic acid