bph genes of the thermophilic PCB degrader, Bacillus sp. JF8: characterization of the divergent ring-hydroxylating dioxygenase and hydrolase genes upstream of the Mn-dependent BphC.
Bacillus sp. JF8 is a thermophilic polychlorinated biphenyl (PCB) degrader, which utilizes biphenyl and naphthalene. A thermostable, Mn-dependent 2,3-dihydroxybiphenyl 1,2-dioxygenase, BphC_JF8, has been characterized previously. Upstream of bphC are five ORFs exhibiting low homology with, and a different gene order from, previously characterized bph genes. From the 5' to 3' direction the genes are: a putative regulatory gene (bphR), a hydrolase (bphD), the large and small subunits of a ring-hydroxylating dioxygenase (bphA1A2), and a cis-diol dehydrogenase (bphB). Hybridization studies indicate that the genes are located on a plasmid. Ring-hydroxylating activity of recombinant BphA1A2_JF8 towards biphenyl, PCB, naphthalene and benzene was observed in Escherichia coli cells, with complementation of non-specific ferredoxin and ferredoxin reductase by host cell proteins. PCB degradation by recombinant BphA1A2_JF8 showed that the congener specificity of the recombinant enzyme was similar to Bacillus sp. JF8. BphD_JF8, with an optimum temperature of 85 degrees C, exhibited a narrow substrate preference for 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid. The Arrhenius plot of BphD_JF8 was biphasic, with two characteristic energies of activation and a break point at 47 degrees C.
Mukerjee-Dhar
Gouri
G
Environmental Biotechnology Laboratory, Railway Technical Research Institute, Kokubunji, Tokyo 185-8540, Japan.
Shimura
Minoru
M
Environmental Biotechnology Laboratory, Railway Technical Research Institute, Kokubunji, Tokyo 185-8540, Japan.
Miyazawa
Daisuke
D
Department of Built Environment, Tokyo Institute of Technology, Yokohama 226-8502, Japan.
Kimbara
Kazuhide
K
Department of Built Environment, Tokyo Institute of Technology, Yokohama 226-8502, Japan.|Environmental Biotechnology Laboratory, Railway Technical Research Institute, Kokubunji, Tokyo 185-8540, Japan.
Hatta
Takashi
T
Research Institute of Technology, Okayama University of Science, Okayama 703-8232, Japan.
D001407:Bacillus / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D001673:Biodegradation, Environmental; D049308:Dioxygenases / Q000235:genetics* / Q000378:metabolism; D006867:Hydrolases / Q000235:genetics* / Q000378:metabolism; D007506:Iron-Sulfur Proteins / Q000235:genetics / Q000378:metabolism; D008345:Manganese / Q000378:metabolism*; D008969:Molecular Sequence Data; D010105:Oxygenases / Q000235:genetics / Q000378:metabolism; D011078:Polychlorinated Biphenyls / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity; D013696:Temperature
D007506:Iron-Sulfur Proteins; D008345:Manganese; D011078:Polychlorinated Biphenyls; D010105:Oxygenases; D049308:Dioxygenases; C074272:2,3-dihydroxybiphenyl oxygenase; C074161:biphenyl-2,3-dioxygenase; D006867:Hydrolases; C062907:2,6-dioxo-6-phenylhexa-3-enoate hydrolase