Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1987 May;169(5):1853–1860. doi: 10.1128/jb.169.5.1853-1860.1987

Molybdenum-sensitive transcriptional regulation of the chlD locus of Escherichia coli.

J B Miller, D J Scott, N K Amy
PMCID: PMC212035  PMID: 3106322

Abstract

The chlD gene in Escherichia coli is required for the incorporation and utilization of molybdenum when the cells are grown with low concentrations of molybdate. We constructed chlD-lac operon fusions and measured expression of the fusion, Mo cofactor, and nitrate reductase activities under a variety of growth conditions. The chlD-lac fusion was highly expressed when cells were grown with less than 10 nm molybdate. Increasing concentrations of molybdate caused loss of activity, with less than 5% of the activity remaining at 500 nM molybdate; when tungstate replaced molybdate, it had an identical affect on chlD expression. Expression of chlD-lac was increased in cells grown with nitrate. Strains with chlD-lac plus an additional mutation in a chl or nar gene were constructed to test whether the regulation of chlD-lac required the concerted action of gene products involved with Mo cofactor or nitrate reductase synthesis. Mutations in narL prevented the increase in activity in response to nitrate; mutations in chlB, narC, or narI resulted in partial constitutive expression of the chlD-lac fusion: the fusion was regulated by molybdate, but it no longer required the presence of nitrate for maximal activity. Mutations in chlA, chlE, or chlG which affect Mo cofactor metabolism, did not affect the expression of chlD-lac.

Full text

PDF
1853

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Amy N. K. Identification of the molybdenum cofactor in chlorate-resistant mutants of Escherichia coli. J Bacteriol. 1981 Oct;148(1):274–282. doi: 10.1128/jb.148.1.274-282.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Amy N. K., Rajagopalan K. V. Characterization of molybdenum cofactor from Escherichia coli. J Bacteriol. 1979 Oct;140(1):114–124. doi: 10.1128/jb.140.1.114-124.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Casadaban M. J., Cohen S. N. Lactose genes fused to exogenous promoters in one step using a Mu-lac bacteriophage: in vivo probe for transcriptional control sequences. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4530–4533. doi: 10.1073/pnas.76.9.4530. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chippaux M., Bonnefoy-Orth V., Ratouchniak J., Pascal M. C. Operon fusions in the nitrate reductase operon and study of the control gene nir R in Escherichia coli. Mol Gen Genet. 1981;182(3):477–479. doi: 10.1007/BF00293938. [DOI] [PubMed] [Google Scholar]
  5. Dixon R., Eady R. R., Espin G., Hill S., Iaccarino M., Kahn D., Merrick M. Analysis of regulation of Klebsiella pneumoniae nitrogen fixation (nif) gene cluster with gene fusions. Nature. 1980 Jul 10;286(5769):128–132. doi: 10.1038/286128a0. [DOI] [PubMed] [Google Scholar]
  6. Edwards E. S., Rondeau S. S., DeMoss J. A. chlC (nar) operon of Escherichia coli includes structural genes for alpha and beta subunits of nitrate reductase. J Bacteriol. 1983 Mar;153(3):1513–1520. doi: 10.1128/jb.153.3.1513-1520.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Giordano G., Saracino L., Grillet L. Identification in various chlorate-resistant mutants of a protein involved in the activation of nitrate reductase in the soluble fraction of a chlA mutant of Escherichia coli K-12. Biochim Biophys Acta. 1985 Apr 17;839(2):181–190. doi: 10.1016/0304-4165(85)90035-2. [DOI] [PubMed] [Google Scholar]
  8. Giordano G., Violet M., Medani C. L., Pommier J. A common pathway for the activation of several molybdoenzymes in Escherichia coli K12. Biochim Biophys Acta. 1984 Apr 10;798(2):216–225. doi: 10.1016/0304-4165(84)90307-6. [DOI] [PubMed] [Google Scholar]
  9. Glaser J. H., DeMoss J. A. Phenotypic restoration by molybdate of nitrate reductase activity in chlD mutants of Escherichia coli. J Bacteriol. 1971 Nov;108(2):854–860. doi: 10.1128/jb.108.2.854-860.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Imperial J., Ugalde R. A., Shah V. K., Brill W. J. Mol- mutants of Klebsiella pneumoniae requiring high levels of molybdate for nitrogenase activity. J Bacteriol. 1985 Sep;163(3):1285–1287. doi: 10.1128/jb.163.3.1285-1287.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Imperial J., Ugalde R. A., Shah V. K., Brill W. J. Role of the nifQ gene product in the incorporation of molybdenum into nitrogenase in Klebsiella pneumoniae. J Bacteriol. 1984 Apr;158(1):187–194. doi: 10.1128/jb.158.1.187-194.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Johnson J. L., Hainline B. E., Rajagopalan K. V. Characterization of the molybdenum cofactor of sulfite oxidase, xanthine, oxidase, and nitrate reductase. Identification of a pteridine as a structural component. J Biol Chem. 1980 Mar 10;255(5):1783–1786. [PubMed] [Google Scholar]
  13. Kennedy C., Postgate J. R. Expression of Klebsiella pneumoniae nitrogen fixation genes in nitrate reductase mutants of Escherichia coli. J Gen Microbiol. 1977 Feb;98(2):551–557. doi: 10.1099/00221287-98-2-551. [DOI] [PubMed] [Google Scholar]
  14. Komeda Y., Iino T. Regulation of expression of the flagellin gene (hag) in Escherichia coli K-12: analysis of hag-lac gene fusions. J Bacteriol. 1979 Sep;139(3):721–729. doi: 10.1128/jb.139.3.721-729.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Li S., Rabi T., DeMoss J. A. Delineation of two distinct regulatory domains in the 5' region of the nar operon of Escherichia coli. J Bacteriol. 1985 Oct;164(1):25–32. doi: 10.1128/jb.164.1.25-32.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. MacGregor C. H., Schnaitman C. A., Normansell D. E. Purification and properties of nitrate reductase from Escherichia coli K12. J Biol Chem. 1974 Aug 25;249(16):5321–5327. [PubMed] [Google Scholar]
  17. Miller J. B., Amy N. K. Molybdenum cofactor in chlorate-resistant and nitrate reductase-deficient insertion mutants of Escherichia coli. J Bacteriol. 1983 Aug;155(2):793–801. doi: 10.1128/jb.155.2.793-801.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Pascal M. C., Burini J. F., Ratouchniak J., Chippaux M. Regulation of the nitrate reductase operon: effect of mutations in chlA, B, D and E genes. Mol Gen Genet. 1982;188(1):103–106. doi: 10.1007/BF00333001. [DOI] [PubMed] [Google Scholar]
  19. Pascal M. C., Chippaux M. Involvement of a gene of the chl E locus in the regulation of the nitrate reductase operon. Mol Gen Genet. 1982;185(2):334–338. doi: 10.1007/BF00330808. [DOI] [PubMed] [Google Scholar]
  20. Riviere C., Giordano G., Pommier J., Azoulay E. Membrane reconstitution in chl-r mutants of Escherichia coli K 12. VIII. Purification and properties of the FA factor, the product of the chl B gene. Biochim Biophys Acta. 1975 May 6;389(2):219–235. doi: 10.1016/0005-2736(75)90317-x. [DOI] [PubMed] [Google Scholar]
  21. Showe M. K., DeMoss J. A. Localization and regulation of synthesis of nitrate reductase in Escherichia coli. J Bacteriol. 1968 Apr;95(4):1305–1313. doi: 10.1128/jb.95.4.1305-1313.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Skotnicki M. L., Rolfe B. G. Interaction between the nitrate respiratory system of Escherichia coli K12 and the nitrogen fixation genes of Klebsiella pneumoniae. Biochem Biophys Res Commun. 1977 Sep 23;78(2):726–733. doi: 10.1016/0006-291x(77)90239-x. [DOI] [PubMed] [Google Scholar]
  23. Sperl G. T., DeMoss J. A. chlD gene function in molybdate activation of nitrate reductase. J Bacteriol. 1975 Jun;122(3):1230–1238. doi: 10.1128/jb.122.3.1230-1238.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Stewart V., MacGregor C. H. Nitrate reductase in Escherichia coli K-12: involvement of chlC, chlE, and chlG loci. J Bacteriol. 1982 Aug;151(2):788–799. doi: 10.1128/jb.151.2.788-799.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Stewart V. Requirement of Fnr and NarL functions for nitrate reductase expression in Escherichia coli K-12. J Bacteriol. 1982 Sep;151(3):1320–1325. doi: 10.1128/jb.151.3.1320-1325.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Ugalde R. A., Imperial J., Shah V. K., Brill W. J. Biosynthesis of the iron-molybdenum cofactor and the molybdenum cofactor in Klebsiella pneumoniae: effect of sulfur source. J Bacteriol. 1985 Dec;164(3):1081–1087. doi: 10.1128/jb.164.3.1081-1087.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. del Campillo-Campbell A., Campbell A. Molybdenum cofactor requirement for biotin sulfoxide reduction in Escherichia coli. J Bacteriol. 1982 Feb;149(2):469–478. doi: 10.1128/jb.149.2.469-478.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES