Comamonas testosteroni TA441 degrades phenol by a meta-cleavage pathway after the occurrence of a spontaneous mutation that derepresses the aphKLMNOPQB operon encoding phenol hydroxylase and catechol 2,3-dioxygenase, the enzymes for the initial two steps of the degradation pathway. A gene cluster, aphCEFGHJI, encoding the meta-pathway enzymes for degradation of 2-hydroxymuconic semialdehyde (HMS) to TCA cycle intermediates was found downstream of the aphK operon. The upstream operon and the downstream gene cluster were found to be separated by two open reading frames of unknown function and an oppositely oriented aphT gene, which is similar to regulatory genes for ortho-cleavage of catechol or chlorinated catechols. A promoter assay using an aphC::lacZ transcriptional fusion plasmid revealed that the aphC promoter activity is induced by both phenol and HMS. The phenol-dependent induction was mediated by AphR and the HMS-dependent induction was mediated by AphT. The aphC promoter in strain TA441 was not silenced, unlike the cases of the aphK and aphR promoters, and was highly induced by HMS.
AraiH., AkahiraS., OhishiT., MaedaM., KudoT.1998; Adaptation of Comamonas testosteroni TA441 to utilize phenol: organization and regulation of the genes involved in phenol degradation. Microbiology 144:2895–2903[CrossRef]
AraiH., AkahiraS., OhishiT., KudoT.1999a; Adaptation of Comamonas testosteroni TA441 to utilization of phenol by spontaneous mutation of the gene for a trans-acting factor. Mol Microbiol 33:1132–1140
BradfordM. M.1976; A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein–dye binding. Anal Biochem 72:248–254[CrossRef]
CocoW. M., RothmelR. K., HenikoffS., ChakrabartyA. M.1993; Nucleotide sequence and initial functional characterization of the clcR gene encoding a LysR family activator of the clcABD chlorocatechol operon in Pseudomonas putida. J Bacteriol 175:417–427
DanielR., StuertzK., GottschalkG.1995; Biochemical and molecular characterization of the oxidative branch of glycerol utilization by Citrobacter freundii. J Bacteriol 177:4392–4401
EricksonB. D., MondelloF. J.1992; Nucleotide sequencing and transcriptional mapping of genes encoding biphenyl dioxygenase, a multicomponent PCB-degrading enzyme in Pseudomonas strain LB400. J Bacteriol 174:2903–2912
FrantzB., ChakrabartyA. M.1987; Organization and nucleotide sequence determination of a gene cluster involved in 3-chlorocatechol degradation. Proc Natl Acad Sci USA 84:4460–4464[CrossRef]
GrimmA. C., HarwoodC. S.1999; NahY, a catabolic plasmid-encoding receptor required for chemotaxis of Pseudomonas putida to the aromatic hydrocarbon naphthalene. J Bacteriol 181:3310–3316
HarayamaS., RekikM., NgaiK.-L., OrnstonL. N.1989; Physically associated enzymes produce and metabolize 2-hydroxypent-2,4-dienoate, a chemically unstable intermediate formed in catechol metabolism via meta cleavage in Pseudomonas putida. J Bacteriol 171:6251–6258
LauP. C., BergeronH., LabbeD., WangY., BrousseauR., GibsonD. T.1994; Sequence and expression of the todGIH genes involved in the last three steps of toluene degradation by Pseudomonas putida F1. Gene 146:7–13[CrossRef]
LodgeJ., WilliamsR., BellA., ChanB., BusbyS.1990; Comparison of promoter activities in Escherichia coli and Pseudomonas aeruginosa: use of a new broad-host-range promoter-probe plasmid. FEMS Microbiol Lett 67:221–226[CrossRef]
McFallS. M., ParsekM. R., ChakrabartyA. M.1997; 2-Chloromuconate and ClcR-mediated activation of the clcABC operon: in vitro transcriptional and DNase I footprint analyses. J Bacteriol 179:3655–3663
MarsA. E., KingmaJ., KaschabekS. R., ReinekeW., JanssenD. B.1999; Conversion of 3-chlorocatechol by various catechol 2,3-dioxygenases and sequence analysis of the chlorocatechol dioxygenase region of Pseudomonas putida GJ31. J Bacteriol 181:1309–1318
van der MeerJ. R., EggenR. I., ZehnderA. J., de VosW. M.1991a; Sequence analysis of the Pseudomonas sp. strain P51 tcb gene cluster, which encodes metabolism of chlorinated catechols: evidence for specialization of catechol 1,2-dioxygenases for chlorinated substrates. J Bacteriol 173:2425–2434
van der MeerJ. R., FrijtersA. C., LeveauJ. H., EggenR. I., ZehnderA. J., de VosW. M.1991b; Characterization of the Pseudomonas sp. strain P51 gene tcbR, a LysR-type transcriptional activator of the tcbCDEF chlorocatechol oxidative operon, and analysis of the regulatory region. J Bacteriol 173:3700–3708
MillerJ. H.1992A Short Course in Bacterial Genetics: a Laboratory Manual and Handbook for Escherichia coli and Related Bacteria Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
ParalesR. E., OntlT. A., GibsonD. T.1997; Cloning and sequence analysis of a catechol 2,3-dioxygenase gene from the nitrobenzene-degrading strain Comamonas sp. JS765. J Ind Microbiol Biotechnol 19:385–391[CrossRef]
ParsekM. E., ShinabargerD. L., RothmelR. K., ChakrabartyA. M.1992; Roles of CatR and cis,cis-muconate in activation of the catBC operon, which is involved in benzoate degradation in Pseudomonas putida. J Bacteriol 174:7798–7806
RothmelR. K., AldrichT. L., HoughtonJ. E., CocoW. M., OrnstonL. N., ChakrabartyA. M.1990; Nucleotide sequencing and characterization of Pseudomonas putidacatR: a positive regulator of the catBC operon is a member of the lysR family. J Bacteriol 172:922–931
SalomoneJ.-Y., CrouzetP., De RuffrayP., OttenL.1996; Characterization and distribution of tartrate utilization genes in the grapevine pathogen Agrobacterium vitis. Mol Plant–Microbe Interact 9:401–408[CrossRef]
ShinglerV.1996; Metabolic and regulatory check points in phenol degradation by Pseudomonas sp. CF600. In Molecular Biology of Pseudomonas pp. 153–154Edited byNakazawaT., FurukawaK., HaasD., SilverS. Washington, DC: American Society for Microbiology;
ShinglerV., BartilsonM., MooreT.1993; Cloning and nucleotide sequence of the gene encoding the positive regulator (DmpR) of the phenol catabolic pathway encoded by pVI150 and identification of DmpR as a member of the NtrC family of transcriptional activators. J Bacteriol 175:1596–1604
SimonR.1984; High frequency mobilization of gram-negative bacterial replicons by the in vitro constructed Tn5-Mob transposon. Mol Gen Genet 196:413–420[CrossRef]
ValentinH. E., ZwingmannG., SchonebaumA., SteinbüchelA.1995; Metabolic pathway for biosynthesis of poly(3-hydroxybutyrate-co-4-hydroxybutyrate) from 4-hydroxybutyrate by Alcaligenes eutrophus. Eur J Biochem 227:43–60[CrossRef]
WigmoreH. J., BaylyR. C., BerardinoD.1974; Pseudomonas putida mutants defective in metabolism of the products of meta-fission of catechol and its methyl analogues. J Bacteriol 120:31–37
Arrangement and regulation of the genes for meta-pathway enzymes required for degradation of phenol in Comamonas testosteroni TA441The DDBJ/EMBL/GenBank accession number for the sequence reported in this paper is AB029044.