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A Novel p-Nitrophenol Degradation Gene Cluster from a Gram-Positive Bacterium, Rhodococcus opacus SAO101. Wataru Kitagawa, 2004.p-Nitrophenol (4-NP) is recognized as an environmental contaminant; it is used primarily for manufacturing medicines and pesticides . To date, several 4-NP-degrading bacteria have been isolated; however, the genetic information remains very limited . In this study, a novel 4-NP degradation gene cluster from a gram-positive bacterium, Rhodococcus opacus SAO101, was identified and characterized . The deduced amino acid sequences of npcB, npcA, and npcC showed identity with phenol 2-hydroxylase component B (reductase, PheA2) of Geobacillus thermoglucosidasius A7 (32%), with 2,4,6-trichlorophenol monooxygenase (TcpA) of Ralstonia eutropha JMP134 (44%), and with hydroxyquinol 1,2-dioxygenase (ORF2) of Arthrobacter sp . strain BA-5-17 (76%), respectively . The npcB, npcA, and npcC genes were cloned into pET-17b to construct the respective expression vectors pETnpcB, pETnpcA, and pETnpcC . Conversion of 4-NP was observed when a mixture of crude cell extracts of Escherichia coli containing pETnpcB and pETnpcA was used in the experiment . The mixture converted 4-NP to hydroxyquinol and also converted 4-nitrocatechol (4-NCA) to hydroxyquinol . Furthermore, the crude cell extract of E . coli containing pETnpcC converted hydroxyquinol to maleylacetate . These results suggested that npcB and npcA encode the two-component 4-NP/4-NCA monooxygenase and that npcC encodes hydroxyquinol 1,2-dioxygenase . The npcA and npcC mutant strains, SDA1 and SDC1, completely lost the ability to grow on 4-NP as the sole carbon source . These results clearly indicated that the cloned npc genes play an essential role in 4-NP mineralization in R . opacus SAO101 . Microbiology of the Phyllosphere. Steven E. Lindow, 2003.
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