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. 1996 Aug;62(8):2716–2722. doi: 10.1128/aem.62.8.2716-2722.1996

Chloroform mineralization by toluene-oxidizing bacteria.

K McClay 1, B G Fox 1, R J Steffan 1
PMCID: PMC168056  PMID: 8702263

Abstract

Seven toluene-oxidizing bacterial strains (Pseudomonas mendocina KR1, Burkholderia cepacia G4, Pseudomonas putida F1, Pseudomonas pickettii PKO1, and Pseudomonas sp. strains ENVPC5, ENVBF1, and ENV113) were tested for their ability to degrade chloroform (CF). The greatest rate of CF oxidation was achieved with strain ENVBF1 (1.9 nmol/min/mg of cell protein). CF also was oxidized by P. mendocina KR1 (0.48 nmol/min/mg of cell protein), strain ENVPC5 (0.49 nmol/min/mg of cell protein), and Escherichia coli DH510B(pRS202), which contained cloned toluene 4-monooxygenase genes from P. mendocina KR1 (0.16 nmol/min/mg of cell protein). Degradation of [14C]CF and ion analysis of culture extracts revealed that CF was mineralized to CO2 (approximately 30 to 57% of the total products), soluble metabolites (approximately 15%), a total carbon fraction irreversibly bound to particulate cellular constituents (approximately 30%), and chloride ions (approximately 75% of the expected yield). CF oxidation by each strain was inhibited in the presence of trichloroethylene, and acetylene significantly inhibited trichloroethylene oxidation by P. mendocina KR1. Differences in the abilities of the CF-oxidizing strains to degrade other halogenated compounds were also identified. CF was not degraded by B. cepacia G4, P. putida F1, P. pickettii PKO1, Pseudomonas sp. strain ENV113, or P. mendocina KRMT, which contains a tmo mutation.

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Selected References

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  1. Alvarez-Cohen L., McCarty P. L., Boulygina E., Hanson R. S., Brusseau G. A., Tsien H. C. Characterization of a methane-utilizing bacterium from a bacterial consortium that rapidly degrades trichloroethylene and chloroform. Appl Environ Microbiol. 1992 Jun;58(6):1886–1893. doi: 10.1128/aem.58.6.1886-1893.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Alvarez-Cohen L., McCarty P. L. Effects of toxicity, aeration, and reductant supply on trichloroethylene transformation by a mixed methanotrophic culture. Appl Environ Microbiol. 1991 Jan;57(1):228–235. doi: 10.1128/aem.57.1.228-235.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Brusseau G. A., Tsien H. C., Hanson R. S., Wackett L. P. Optimization of trichloroethylene oxidation by methanotrophs and the use of a colorimetric assay to detect soluble methane monooxygenase activity. Biodegradation. 1990;1(1):19–29. doi: 10.1007/BF00117048. [DOI] [PubMed] [Google Scholar]
  4. Byrne A. M., Kukor J. J., Olsen R. H. Sequence analysis of the gene cluster encoding toluene-3-monooxygenase from Pseudomonas pickettii PKO1. Gene. 1995 Feb 27;154(1):65–70. doi: 10.1016/0378-1119(94)00844-i. [DOI] [PubMed] [Google Scholar]
  5. Ensley B. D., Ratzkin B. J., Osslund T. D., Simon M. J., Wackett L. P., Gibson D. T. Expression of naphthalene oxidation genes in Escherichia coli results in the biosynthesis of indigo. Science. 1983 Oct 14;222(4620):167–169. doi: 10.1126/science.6353574. [DOI] [PubMed] [Google Scholar]
  6. Fox B. G., Borneman J. G., Wackett L. P., Lipscomb J. D. Haloalkene oxidation by the soluble methane monooxygenase from Methylosinus trichosporium OB3b: mechanistic and environmental implications. Biochemistry. 1990 Jul 10;29(27):6419–6427. doi: 10.1021/bi00479a013. [DOI] [PubMed] [Google Scholar]
  7. Fox B. G., Shanklin J., Ai J., Loehr T. M., Sanders-Loehr J. Resonance Raman evidence for an Fe-O-Fe center in stearoyl-ACP desaturase. Primary sequence identity with other diiron-oxo proteins. Biochemistry. 1994 Nov 1;33(43):12776–12786. doi: 10.1021/bi00209a008. [DOI] [PubMed] [Google Scholar]
  8. Haigler B. E., Pettigrew C. A., Spain J. C. Biodegradation of mixtures of substituted benzenes by Pseudomonas sp. strain JS150. Appl Environ Microbiol. 1992 Jul;58(7):2237–2244. doi: 10.1128/aem.58.7.2237-2244.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hanahan D. Studies on transformation of Escherichia coli with plasmids. J Mol Biol. 1983 Jun 5;166(4):557–580. doi: 10.1016/s0022-2836(83)80284-8. [DOI] [PubMed] [Google Scholar]
  10. Hareland W. A., Crawford R. L., Chapman P. J., Dagley S. Metabolic function and properties of 4-hydroxyphenylacetic acid 1-hydroxylase from Pseudomonas acidovorans. J Bacteriol. 1975 Jan;121(1):272–285. doi: 10.1128/jb.121.1.272-285.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Herrero M., de Lorenzo V., Timmis K. N. Transposon vectors containing non-antibiotic resistance selection markers for cloning and stable chromosomal insertion of foreign genes in gram-negative bacteria. J Bacteriol. 1990 Nov;172(11):6557–6567. doi: 10.1128/jb.172.11.6557-6567.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Jahng D., Wood T. K. Trichloroethylene and chloroform degradation by a recombinant pseudomonad expressing soluble methane monooxygenase from Methylosinus trichosporium OB3b. Appl Environ Microbiol. 1994 Jul;60(7):2473–2482. doi: 10.1128/aem.60.7.2473-2482.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Johnson G. R., Olsen R. H. Nucleotide sequence analysis of genes encoding a toluene/benzene-2-monooxygenase from Pseudomonas sp. strain JS150. Appl Environ Microbiol. 1995 Sep;61(9):3336–3346. doi: 10.1128/aem.61.9.3336-3346.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lewis T. A., Crawford R. L. Transformation of carbon tetrachloride via sulfur and oxygen substitution by Pseudomonas sp. strain KC. J Bacteriol. 1995 Apr;177(8):2204–2208. doi: 10.1128/jb.177.8.2204-2208.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Little C. D., Palumbo A. V., Herbes S. E., Lidstrom M. E., Tyndall R. L., Gilmer P. J. Trichloroethylene biodegradation by a methane-oxidizing bacterium. Appl Environ Microbiol. 1988 Apr;54(4):951–956. doi: 10.1128/aem.54.4.951-956.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. McClay K., Streger S. H., Steffan R. J. Induction of toluene oxidation activity in Pseudomonas mendocina KR1 and Pseudomonas sp. strain ENVPC5 by chlorinated solvents and alkanes. Appl Environ Microbiol. 1995 Sep;61(9):3479–3481. doi: 10.1128/aem.61.9.3479-3481.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Miller V. L., Mekalanos J. J. A novel suicide vector and its use in construction of insertion mutations: osmoregulation of outer membrane proteins and virulence determinants in Vibrio cholerae requires toxR. J Bacteriol. 1988 Jun;170(6):2575–2583. doi: 10.1128/jb.170.6.2575-2583.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Nelson M. J., Montgomery S. O., Mahaffey W. R., Pritchard P. H. Biodegradation of trichloroethylene and involvement of an aromatic biodegradative pathway. Appl Environ Microbiol. 1987 May;53(5):949–954. doi: 10.1128/aem.53.5.949-954.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Nelson M. J., Montgomery S. O., Pritchard P. H. Trichloroethylene metabolism by microorganisms that degrade aromatic compounds. Appl Environ Microbiol. 1988 Feb;54(2):604–606. doi: 10.1128/aem.54.2.604-606.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Oldenhuis R., Vink R. L., Janssen D. B., Witholt B. Degradation of chlorinated aliphatic hydrocarbons by Methylosinus trichosporium OB3b expressing soluble methane monooxygenase. Appl Environ Microbiol. 1989 Nov;55(11):2819–2826. doi: 10.1128/aem.55.11.2819-2826.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Olsen R. H., Kukor J. J., Kaphammer B. A novel toluene-3-monooxygenase pathway cloned from Pseudomonas pickettii PKO1. J Bacteriol. 1994 Jun;176(12):3749–3756. doi: 10.1128/jb.176.12.3749-3756.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Rosenzweig A. C., Nordlund P., Takahara P. M., Frederick C. A., Lippard S. J. Geometry of the soluble methane monooxygenase catalytic diiron center in two oxidation states. Chem Biol. 1995 Sep;2(9):409–418. [PubMed] [Google Scholar]
  23. Shields M. S., Montgomery S. O., Chapman P. J., Cuskey S. M., Pritchard P. H. Novel pathway of toluene catabolism in the trichloroethylene-degrading bacterium g4. Appl Environ Microbiol. 1989 Jun;55(6):1624–1629. doi: 10.1128/aem.55.6.1624-1629.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Shields M. S., Reagin M. J., Gerger R. R., Campbell R., Somerville C. TOM, a new aromatic degradative plasmid from Burkholderia (Pseudomonas) cepacia G4. Appl Environ Microbiol. 1995 Apr;61(4):1352–1356. doi: 10.1128/aem.61.4.1352-1356.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Tsien H. C., Brusseau G. A., Hanson R. S., Waclett L. P. Biodegradation of trichloroethylene by Methylosinus trichosporium OB3b. Appl Environ Microbiol. 1989 Dec;55(12):3155–3161. doi: 10.1128/aem.55.12.3155-3161.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Wackett L. P., Gibson D. T. Degradation of trichloroethylene by toluene dioxygenase in whole-cell studies with Pseudomonas putida F1. Appl Environ Microbiol. 1988 Jul;54(7):1703–1708. doi: 10.1128/aem.54.7.1703-1708.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Whited G. M., Gibson D. T. Separation and partial characterization of the enzymes of the toluene-4-monooxygenase catabolic pathway in Pseudomonas mendocina KR1. J Bacteriol. 1991 May;173(9):3017–3020. doi: 10.1128/jb.173.9.3017-3020.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]
  29. Yen K. M., Karl M. R., Blatt L. M., Simon M. J., Winter R. B., Fausset P. R., Lu H. S., Harcourt A. A., Chen K. K. Cloning and characterization of a Pseudomonas mendocina KR1 gene cluster encoding toluene-4-monooxygenase. J Bacteriol. 1991 Sep;173(17):5315–5327. doi: 10.1128/jb.173.17.5315-5327.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Yen K. M., Karl M. R. Identification of a new gene, tmoF, in the Pseudomonas mendocina KR1 gene cluster encoding toluene-4-monooxygenase. J Bacteriol. 1992 Nov;174(22):7253–7261. doi: 10.1128/jb.174.22.7253-7261.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. de Lorenzo V., Eltis L., Kessler B., Timmis K. N. Analysis of Pseudomonas gene products using lacIq/Ptrp-lac plasmids and transposons that confer conditional phenotypes. Gene. 1993 Jan 15;123(1):17–24. doi: 10.1016/0378-1119(93)90533-9. [DOI] [PubMed] [Google Scholar]

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