1887

Microbiology Society logo

Volume 149, Issue 7

, and , genes involved in regulating the expression of soluble methane monooxygenase in OB3b Free

Abstract

The methanotrophic bacterium OB3b converts methane to methanol using two distinct forms of methane monooxygenase (MMO) enzyme: a cytoplasmic soluble form (sMMO) and a membrane-bound form (pMMO). The transcription of these two operons is known to proceed in a reciprocal fashion with sMMO expressed at low copper-to-biomass ratios and pMMO at high copper-to-biomass ratios. Transcription of the operon is initiated from a promoter 5′ of . In this study the genes encoding () and a typical -dependent transcriptional activator () were cloned and sequenced. , a regulatory gene, and , a gene encoding a GroEL homologue, lie 5′ of the structural genes for the sMMO enzyme. Subsequent mutation of and by marker-exchange mutagenesis resulted in strains Gm1 and JS1, which were unable to express functional sMMO or initiate transcription of . An mutant was also unable to fix nitrogen or use nitrate as sole nitrogen source, indicating that plays a role in both nitrogen and carbon metabolism in OB3b. The data also indicate that is transcribed in a - and MmoR-independent manner. Marker-exchange mutagenesis of revealed that MmoG is necessary for gene transcription and activity and may be an MmoR-specific chaperone required for functional assembly of transcriptionally competent MmoR . The data presented allow the proposal of a more complete model for copper-mediated regulation of gene expression.

  • Received:
  • Accepted:
  • Revised:
  • Published Online:
Keyword(s): (p)(s)MMO, (particulate) (soluble) methane monooxygenase , Ap, ampicillin , EBP, enhancer-binding protein , Gm, gentamicin , GOGAT, glutamate synthase , GS, glutamine synthetase , Km, kanamycin , NMS, nitrate mineral salts and UAS, upstream activator sequence
SGM
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.26060-0
2003-07-01
2024-04-18
Loading full text...

Full text loading...

/deliver/fulltext/micro/149/7/mic1491771.html?itemId=/content/journal/micro/10.1099/mic.0.26060-0&mimeType=html&fmt=ahah

References

  1. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. 1990; Basic Local Alignment Search Tool. J Mol Biol 215:403–410
     [Google Scholar]
  2. Arenghi F. L. G., Pinti M., Galli E., Barbieri P. 1999; Identification of the Pseudomonas stutzeri OX1 toluene- o -xylene monooxygenase regulatory gene ( touR ) and its cognate promoter. Appl Environ Microbiol 65:4057–4063
     [Google Scholar]
  3. Barrios H., Valderrama B., Morett E. 1999; Compilation and analysis of σ 54-dependent promoter sequences. Nucleic Acids Res 27:4305–4313
     [Google Scholar]
  4. Brusseau G. A., Tsien H.-C., Hanson R. S., Wackett L. P. 1990; Optimization of trichloroethylene oxidation by methanotrophs and the use of a colorimetric assay to detect soluble methane monooxygenase activity. Biodegradation 1:19–29
     [Google Scholar]
  5. Buck M., Gallegos M.-T., Studholme D. J., Guo Y., Gralla J. D. 2000; The bacterial enhancer-dependent σ 54 ( σ N) transcription factor. J Bacteriol 182:4129–4136
     [Google Scholar]
  6. Cardy D. L. N., Laidler V., Salmond G. P. C., Murrell J. C. 1991; Molecular analysis of the methane monooxygenase (MMO) gene cluster of Methylosinus trichosporium OB3b. Mol Microbiol 5:335–342
     [Google Scholar]
  7. Csáki R., Bodrossy L., Klem J., Murrell J. C., Kovács K. 2003; Genes involved in the copper-dependent regulation of soluble methane monooxygenase of Methylococcus capsulatus Bath: cloning, sequencing and mutational analysis. Microbiology 149:1785–1795
     [Google Scholar]
  8. Dennis J. J., Zylstra G. J. 1998; Plasposons: modular self-cloning minitransposon derivatives for rapid genetic analysis of gram-negative bacterial genomes. Appl Environ Microbiol 64:2710–2715
     [Google Scholar]
  9. Feinberg A. P., Vogelstein B. 1984; A technique for radiolabelling DNA restriction endonuclease fragments to high specific activity. Anal Biochem 137:266–267
     [Google Scholar]
  10. Felsenstein J. 1993 phylip (Phylogeny Interface Package), version 3.5c. Seattle: Department of Genetics, University of Washington;
     [Google Scholar]
  11. Fischer H. M., Babst M., Kaspar T., Acuna G., Arigoni F., Hennecke H. 1993; One member of a groESL -like chaperonin multigene family of Bradyrhizobium japonicum is co-regulated with the symbiotic nitrogen fixation genes. EMBO J 12:2901–2912
     [Google Scholar]
  12. Garmendia J., Devos D., Valencia A., de Lorenzo V. 2001; A la carte transcriptional regulators: unlocking responses of the prokaryotic enhancer-binding protein XylR to non-natural effectors. Mol Microbiol 42:47–59
     [Google Scholar]
  13. Gilbert B., McDonald I. R., Finch R., Stafford G. P., Nielsen A. K., Murrell J. C. 2000; Molecular analysis of the pmo (particulate methane monooxygenase) operons from the two type II methanotrophs. Appl Environ Microbiol 66:966–975
     [Google Scholar]
  14. Huala E., Stigter J., Ausubel F. M. 1992; The central domain of Rhizobium leguminosarum DctD functions independently to activate transcription. J Bacteriol 174:1428–1431
     [Google Scholar]
  15. Kaneko T., Nakamura Y., Sato S. 21 other authors 2000; Complete genome sequence of the nitrogen-fixing symbiotic bacterium Mesorhizobium loti . DNA Res 7:331–338
     [Google Scholar]
  16. Koch K. A., Pena M. M. O., Thiele D. J. 1997; Copper-binding motifs in catalysis, transport, detoxification and signaling. Chem Biol 4:549–560
     [Google Scholar]
  17. Krüger N., Steinbüchel A. 1992; Identification of acoR , a regulatory gene for the expression of genes essential for acetoin catabolism in Alcaligenes eutrophus HI6. J Bacteriol 174:4391–4400
     [Google Scholar]
  18. Kullik I., Fritsche S., Knobel H., Sanjuan J., Hennecke H., Fischer H. M. 1991; Bradyrhizobium japonicum has two differentially regulated, functional homologs of the sigma 54 gene ( rpoN ). J Bacteriol 173:1125–1138
     [Google Scholar]
  19. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685
     [Google Scholar]
  20. Lee W. T., Terlesky K. C., Tabita F. R. 1997; Cloning and characterisation of two groESL operons of Rhodobacter sphaeroides : transcriptional regulation of the heat-induced groESL operon. J Bacteriol 179:487–495
     [Google Scholar]
  21. Lemos J. A. C., Chen Y. M., Burne R. A. 2001; Genetic and physiological analysis of the groE operon and role of the HrcA repressor in stress regulation and acid tolerance in Streptococcus mutans . J Bacteriol 183:6074–6084
     [Google Scholar]
  22. Lipscomb J. D. 1994; Biochemistry of soluble methane monooxygenase. Annu Rev Microbiol 48:371–399
     [Google Scholar]
  23. Lund P. 2001; Microbial molecular chaperones. Adv Microbial Physiology 44:93–140
     [Google Scholar]
  24. Martin H., Murrell J. C. 1995; Methane monooxygenase mutants of Methylosinus trichosporium constructed by marker-exchange mutagenesis. FEMS Microbiol Lett 127:243–248
     [Google Scholar]
  25. McDonald I. R., Uchiyama H., Kambe S., Yagi O., Murrell J. C. 1997; The soluble methane monooxygenase gene cluster of the trichloroethylene-degrading methanotroph Methylocystis sp. strain M. Appl Environ Microbiol 63:1898–1904
     [Google Scholar]
  26. Meijer W., Tabita F. 1992; Isolation and characterisation of the nifUSVW - rpoN gene cluster from Rhodobacter sphaeroides . J Bacteriol 174:3855–3566
     [Google Scholar]
  27. Merkx M., Lippard S. J. 2002; Why OrfY? Characterization of mmoD , a long overlooked component of the soluble methane monooxygenase from Methylococcus capsulatus (Bath). J Biol Chem 277:5858–5865
     [Google Scholar]
  28. Merrick M. J. 1993; In a class of its own – the RNA polymerase sigma factor σ 54 ( σ N). Mol Microbiol 10:903–909
     [Google Scholar]
  29. Merrick M. J., Edwards R. A. 1995; Nitrogen control in bacteria. Microbiol Rev 59:604–622
     [Google Scholar]
  30. Michiels J., Van Soom T., D'Hooghe I., Dombrecht B., Benhassine T., De Wilde P., Vanderleyden J. 1998; The Rhizobium etli rpoN locus: sequence analysis and phenotypical characterisation of rpoN , ptsN , and ptsA mutants. J Bacteriol 180:1729–1740
     [Google Scholar]
  31. Morrett E., Segovia L. 1993; The σ 54 bacterial enhancer-binding protein family: mechanism of action and phylogenetic relationship of their functional domains. J Bacteriol 175:6067–6074
     [Google Scholar]
  32. Murrell J. C., Dalton H. 1983a; Nitrogen fixation in obligate methanotrophs. J Gen Microbiol 129:3481–3496
     [Google Scholar]
  33. Murrell J. C., Dalton H. 1983b; Ammonia assimilation in Methylococcus capsulatus (Bath) and other obligate methanotrophs. J Gen Microbiol 129:1197–1206
     [Google Scholar]
  34. Nguyen H.-H. T., Shiemke A. K., Jacobs S. J., Hales B. J., Lidstrom M. E., Chan S. I. 1994; The nature of the copper ions in the membranes containing the particulate methane monooxygenase from Methylococcus capsulatus (Bath). J Biol Chem 269:14995–15005
     [Google Scholar]
  35. Nielsen A. K., Gerdes K., Degn H., Murrell J. C. 1996; Regulation of bacterial methane oxidation: transcription of the soluble methane monooxygenase operon of Methylococcus capsulatus (Bath) is repressed by copper ions. Microbiology 142:1289–1296
     [Google Scholar]
  36. Nielsen A. K., Gerdes K., Murrell J. C. 1997; Copper-dependent reciprocal transcriptional regulation of methane monooxygenase genes in Methylococcus capsulatus and Methylosinus trichosporium . Mol Microbiol 25:399–409
     [Google Scholar]
  37. Oakley C. J., Murrell J. C. 1988; nifH genes in obligate methane oxidizing bacteria. FEMS Microbiol Lett 49:53–57
     [Google Scholar]
  38. Parkhill J., Achtman M., James K. D. 25 other authors 2000; Complete DNA sequence of a serogroup A strain of Neisseria meningitidis Z2491. Nature 404:502–506
     [Google Scholar]
  39. Powell B. S., Court D. L., Inada T. 7 other authors 1995; Novel proteins of the phosphotransferase system encoded within the rpoN operon of Escherichia coli . J Biol Chem 270:4822–4839
     [Google Scholar]
  40. Reitzer L., Schneider B. L. 2001; Metabolic context and possible physiological themes of σ 54 dependent genes in Escherichia coli . Microbiol Mol Biol Rev 65:422–444
     [Google Scholar]
  41. Ronson C. W., Nixon B. T., Albright L. M., Ausubel F. M. 1987; Rhizobium meliloti ntrA ( rpoN ) gene is required for diverse metabolic functions. J Bacteriol 169:2424–2431
     [Google Scholar]
  42. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning: a Laboratory Manual , 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
     [Google Scholar]
  43. Schäfer A., Tauch A., Jäger W., Lalinowski J., Thierbach G., Pühler A. 1994; Small mobilisable multi-purpose cloning vector derived from the Escherichia coli plasmids pK18 and pK19: selection of defined deletions in the chromosome of Corynebacterium glutamicum . Gene 145:69–73
     [Google Scholar]
  44. Segal G., Ron E. Z. 1996; Regulation and organization of the groE and dnaK operons in eubacteria. FEMS Microbiol Lett 138:1–10
     [Google Scholar]
  45. Semrau J. D., Chistoserdov A., Lebron J. 7 other authors 1995; Particulate methane monooxygenase genes in methanotrophs. J Bacteriol 177:3071–3079
     [Google Scholar]
  46. Shigematsu T., Handa S., Eguchi M., Kamagata Y., Kanagawa T., Kurane R. 1999; Soluble methane monooxygenase gene clusters from trichloroethylene-degrading Methylomonas sp. strains and detection of methanotrophs during in situ bioremediation. Appl Environ Microbiol 65:5198–5206
     [Google Scholar]
  47. Simon R., Priefer U., Pühler A. 1983; A broad host range mobilisation system for in vivo genetic engineering: transposon mutagenesis on Gram-negative bacteria. Biotechnol Lett 1:784–791
     [Google Scholar]
  48. Skärfstad E., O'Neill E., Garmendia J., Shingler V. 2000; Identification of an effector specificity subregion within the aromatic-responsive regulators DmpR and XylR by DNA shuffling. J Bacteriol 182:3008–3016
     [Google Scholar]
  49. Stainthorpe A. C., Lees V., Salmond G. P. C., Dalton H., Murrell J. C. 1990; The methane monooxygenase gene cluster of Methylococcus capsulatus (Bath). Gene 91:27–34
     [Google Scholar]
  50. Stanley S. H., Prior S. D., Leak D. J., Dalton H. 1983; Copper stress underlies the fundamental change in intracellular location of methane monooxygenase in methane-oxidizing organisms: studies in batch and continuous cultures. Biotechnol Lett 5:487–492
     [Google Scholar]
  51. Stolyar S., Costello A. M., Peeples T. L., Lidstrom M. E. 1999; Role of multiple gene copies in particulate methane monooxygenase activity in the methane-oxidizing bacterium Methylococcus capsulatus Bath. Microbiology 145:1235–1244
     [Google Scholar]
  52. Stolyar S., Franke M., Lidstrom M. E. 2001; Expression of individual copies of Methylococcus capsulatus Bath particulate methane monooxygenase genes. J Bacteriol 183:1810–1812
     [Google Scholar]
  53. Strausak D., La Fontaine S., Hill J., Firth S. D., Lockhart P. J., Mercer J. F. B. 1999; The role of GMXCXXC metal binding sites in the copper-induced redistribution of the Menkes protein. J Biol Chem 274:11170–11177
     [Google Scholar]
  54. Studholme D. J., Buck M. 2000; The biology of enhancer-dependent transcriptional regulation in bacteria: insights from genome sequences. FEMS Microbiol Lett 186:1–9
     [Google Scholar]
  55. Takeguchi M., Miyakawa K., Okura I. 1999; The role of copper in particulate methane monooxygenase from Methylosinus trichosporium OB3b. J Mol Catal A Chem 137:161–168
     [Google Scholar]
  56. Tanaka N., Hiyama T., Nakamoto H. 1997; Cloning, characterization and functional analysis of groESL operon from thermophilic cyanobacterium Synechococcus vulcanus . Biochim Biophys Acta 1343:335–348
     [Google Scholar]
  57. Warrelmann J., Eitinger M., Schwartz E., Rommermann D., Friedrich B. 1992; Nucleotide sequence of the rpoN ( hno ) gene region of Alcaligenes eutrophus : evidence for a conserved gene cluster. Arch Microbiol 158:107–114
     [Google Scholar]
  58. Whittenbury R., Phillips K. C., Wilkinson J. F. 1970; Enrichment, isolation and some properties of methane-utilizing bacteria. J Gen Microbiol 61:205–218
     [Google Scholar]
  59. Zahn J. A., Dispirito A. A. 1996; Membrane-associated methane monooxygenase from Methylococcus capsulatus (Bath). J Bacteriol 178:1018–1029
     [Google Scholar]
  60. Zharkikh A., Li W.-H. 1992; Statistical properties of bootstrap estimation of phylogenetic variability from nucleotide sequences. I. Four taxa with a molecular clock. FEMS Microbiol Lett 55:181–186
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.26060-0
Loading
rpoN, mmoR and mmoG, genes involved in regulating the expression of soluble methane monooxygenase in Methylosinus trichosporium OB3b
Microbiology 149, 1771 (2003); https://doi.org/10.1099/mic.0.26060-0
/content/journal/micro/10.1099/mic.0.26060-0
/content/journal/micro/10.1099/mic.0.26060-0
Loading

Data & Media loading...

Most cited Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error