Identification of the Monooxygenase Gene Clusters Responsible for the Regioselective Oxidation of Phenol to Hydroquinone in Mycobacteria

• Published in: Applied and Environmental Microbiology Vol. 77; no. 4; pp. 1214 - 1220
• Main Authors: Furuya, Toshiki, Hirose, Satomi, Osanai, Hisashi, Semba, Hisashi, Kino, Kuniki
• Format: Journal Article
• Language: English
• Published: Washington, DC American Society for Microbiology 01.02.2011; American Society for Microbiology (ASM)
• Subjects: Acetone; Acetone - metabolism; Actinomycetes; Amino acids; Bacteria; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bacteriology; Base Sequence; Biological and medical sciences; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Bacterial; Genes; Genes, Bacterial; Genetics and Molecular Biology; Hydroquinones - chemistry; Hydroquinones - metabolism; Metabolism; Microbiology; Mixed Function Oxygenases - genetics; Mixed Function Oxygenases - metabolism; Molecular Sequence Data; Multigene Family; Mycobacterium - enzymology; Mycobacterium - genetics; Mycobacterium - metabolism; Mycobacterium goodii; Mycobacterium smegmatis; Oxidation; Oxidation-Reduction; Phenol - chemistry; Phenol - metabolism; Polymerase Chain Reaction; RNA, Ribosomal, 16S - genetics; Sequence Analysis, DNA; Sequence Deletion; Hydroquinone; Gene cluster; Phenol; Regioselectivity; Phenols; Identification; Oxidation
• ISSN: 0099-2240; 1098-5336; 1098-6596
• DOI: 10.1128/aem.02316-10

Mycobacterium goodii strain 12523 is an actinomycete that is able to oxidize phenol regioselectively at the para position to produce hydroquinone. In this study, we investigated the genes responsible for this unique regioselective oxidation. On the basis of the fact that the oxidation activity of M. goodii strain 12523 toward phenol is induced in the presence of acetone, we first identified acetone-induced proteins in this microorganism by two-dimensional electrophoretic analysis. The N-terminal amino acid sequence of one of these acetone-induced proteins shares 100% identity with that of the protein encoded by the open reading frame Msmeg_1971 in Mycobacterium smegmatis strain mc²155, whose genome sequence has been determined. Since Msmeg_1971, Msmeg_1972, Msmeg_1973, and Msmeg_1974 constitute a putative binuclear iron monooxygenase gene cluster, we cloned this gene cluster of M. smegmatis strain mc²155 and its homologous gene cluster found in M. goodii strain 12523. Sequence analysis of these binuclear iron monooxygenase gene clusters revealed the presence of four genes designated mimABCD, which encode an oxygenase large subunit, a reductase, an oxygenase small subunit, and a coupling protein, respectively. When the mimA gene (Msmeg_1971) of M. smegmatis strain mc²155, which was also found to be able to oxidize phenol to hydroquinone, was deleted, this mutant lost the oxidation ability. This ability was restored by introduction of the mimA gene of M. smegmatis strain mc²155 or of M. goodii strain 12523 into this mutant. Interestingly, we found that these gene clusters also play essential roles in propane and acetone metabolism in these mycobacteria.