Open AccessMolecular cloning and functional characterization of the genes encoding benzoate and p ‐hydroxybenzoate degradation by the halophilic Chromohalobacter sp. strain HS‐2
Author(s) -
Kim Dockyu,
Kim Si Wouk,
Choi Ki Young,
Lee Jong Suk,
Kim Eungbin
Publication year - 2008
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2008.01067.x
Subject(s) - hydroxybenzoate , biology , gene , biochemistry , dioxygenase , escherichia coli , catabolism , microbiology and biotechnology , enzyme
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.