Abstract
The TOL catabolic genes in Pseudomonas putida (pWW0) are clustered in the upper operon, encoding enzymes for the conversion of toluene and xylenes to benzoate and toluates, and the meta-cleavage operon, encoding enzymes for the conversion of the benzoate and toluates to tricarboxylic acid cycle intermediates. In this study, it was shown that cells growing in a chemostat under succinate growth-limiting conditions express both the upper and meta-cleavage pathways in response to o-xylene, a nonmetabolizable effector of the XylR regulatory protein. The dilution rate maintained in the succinate-limited chemostat cultures influenced the synthesis levels of TOL pathway enzymes, their steady-state levels, and their turnover rates. Cells growing in the presence of nonlimiting concentrations of succinate in continuous culture did not express pathway enzymes in response to the addition of o-xylene, which was due to a blockage at the transcriptional level. Expression of the meta-cleavage pathway in response to 2,3-dimethylbenzoate, a nonmetabolizable effector of the XylS regulatory protein, was 93% lower in cultures exposed to succinate at nonlimiting concentrations than in the succinate-limited chemostats. The mRNA level of xylS during nonlimited growth on succinate was very low compared with that in succinate-limited cultures, suggesting that suppression of expression of the meta-cleavage pathway is regulated mainly by the level of the XylS regulator.
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- Abril M. A., Buck M., Ramos J. L. Activation of the Pseudomonas TOL plasmid upper pathway operon. Identification of binding sites for the positive regulator XylR and for integration host factor protein. J Biol Chem. 1991 Aug 25;266(24):15832–15838. [PubMed] [Google Scholar]
- Abril M. A., Michan C., Timmis K. N., Ramos J. L. Regulator and enzyme specificities of the TOL plasmid-encoded upper pathway for degradation of aromatic hydrocarbons and expansion of the substrate range of the pathway. J Bacteriol. 1989 Dec;171(12):6782–6790. doi: 10.1128/jb.171.12.6782-6790.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Assinder S. J., Williams P. A. The TOL plasmids: determinants of the catabolism of toluene and the xylenes. Adv Microb Physiol. 1990;31:1–69. doi: 10.1016/s0065-2911(08)60119-8. [DOI] [PubMed] [Google Scholar]
- Duetz W. A., Winson M. K., van Andel J. G., Williams P. A. Mathematical analysis of catabolic function loss in a population of Pseudomonas putida mt-2 during non-limited growth on benzoate. J Gen Microbiol. 1991 Jun;137(6):1363–1368. doi: 10.1099/00221287-137-6-1363. [DOI] [PubMed] [Google Scholar]
- Duetz W. A., van Andel J. G. Stability of TOL plasmid pWW0 in Pseudomonas putida mt-2 under non-selective conditions in continuous culture. J Gen Microbiol. 1991 Jun;137(6):1369–1374. doi: 10.1099/00221287-137-6-1369. [DOI] [PubMed] [Google Scholar]
- Harder W., Dijkhuizen L. Strategies of mixed substrate utilization in microorganisms. Philos Trans R Soc Lond B Biol Sci. 1982 Jun 11;297(1088):459–480. doi: 10.1098/rstb.1982.0055. [DOI] [PubMed] [Google Scholar]
- Hugouvieux-Cotte-Pattat N., Köhler T., Rekik M., Harayama S. Growth-phase-dependent expression of the Pseudomonas putida TOL plasmid pWW0 catabolic genes. J Bacteriol. 1990 Dec;172(12):6651–6660. doi: 10.1128/jb.172.12.6651-6660.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Inouye S., Nakazawa A., Nakazawa T. Determination of the transcription initiation site and identification of the protein product of the regulatory gene xylR for xyl operons on the TOL plasmid. J Bacteriol. 1985 Sep;163(3):863–869. doi: 10.1128/jb.163.3.863-869.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Marqués S., Ramos J. L., Timmis K. N. Analysis of the mRNA structure of the Pseudomonas putida TOL meta fission pathway operon around the transcription initiation point, the xylTE and the xylFJ regions. Biochim Biophys Acta. 1993 Nov 16;1216(2):227–236. doi: 10.1016/0167-4781(93)90149-8. [DOI] [PubMed] [Google Scholar]
- Marqués S., Ramos J. L. Transcriptional control of the Pseudomonas putida TOL plasmid catabolic pathways. Mol Microbiol. 1993 Sep;9(5):923–929. doi: 10.1111/j.1365-2958.1993.tb01222.x. [DOI] [PubMed] [Google Scholar]
- Peterson G. L. A simplification of the protein assay method of Lowry et al. which is more generally applicable. Anal Biochem. 1977 Dec;83(2):346–356. doi: 10.1016/0003-2697(77)90043-4. [DOI] [PubMed] [Google Scholar]
- Ramos J. L., Mermod N., Timmis K. N. Regulatory circuits controlling transcription of TOL plasmid operon encoding meta-cleavage pathway for degradation of alkylbenzoates by Pseudomonas. Mol Microbiol. 1987 Nov;1(3):293–300. doi: 10.1111/j.1365-2958.1987.tb01935.x. [DOI] [PubMed] [Google Scholar]
- Ramos J. L., Stolz A., Reineke W., Timmis K. N. Altered effector specificities in regulators of gene expression: TOL plasmid xylS mutants and their use to engineer expansion of the range of aromatics degraded by bacteria. Proc Natl Acad Sci U S A. 1986 Nov;83(22):8467–8471. doi: 10.1073/pnas.83.22.8467. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sala-Trepat J. M., Evans W. C. The meta cleavage of catechol by Azotobacter species. 4-Oxalocrotonate pathway. Eur J Biochem. 1971 Jun 11;20(3):400–413. doi: 10.1111/j.1432-1033.1971.tb01406.x. [DOI] [PubMed] [Google Scholar]
- Shields M. S., Montgomery S. O., Cuskey S. M., Chapman P. J., Pritchard P. H. Mutants of Pseudomonas cepacia G4 defective in catabolism of aromatic compounds and trichloroethylene. Appl Environ Microbiol. 1991 Jul;57(7):1935–1941. doi: 10.1128/aem.57.7.1935-1941.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Topp E., Akhtar M. H. Identification and characterization of a pseudomonas strain capable of metabolizing phenoxybenzoates. Appl Environ Microbiol. 1991 May;57(5):1294–1300. doi: 10.1128/aem.57.5.1294-1300.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Worsey M. J., Williams P. A. Metabolism of toluene and xylenes by Pseudomonas (putida (arvilla) mt-2: evidence for a new function of the TOL plasmid. J Bacteriol. 1975 Oct;124(1):7–13. doi: 10.1128/jb.124.1.7-13.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zylstra G. J., Olsen R. H., Ballou D. P. Cloning, expression, and regulation of the Pseudomonas cepacia protocatechuate 3,4-dioxygenase genes. J Bacteriol. 1989 Nov;171(11):5907–5914. doi: 10.1128/jb.171.11.5907-5914.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]