Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1996 Jul;178(13):3949–3952. doi: 10.1128/jb.178.13.3949-3952.1996

Cloning, characterization, and regulation of nifF from Rhodobacter capsulatus.

G Gennaro 1, P Hübner 1, U Sandmeier 1, A F Yakunin 1, P C Hallenbeck 1
PMCID: PMC232658  PMID: 8682802

Abstract

The Rhodobacter capsulatus nifF gene and upstream sequence were cloned by using a probe based on the N-terminal sequence of NifF. nifF was found to not be contained in the previously described nif regions I, II, and III. Comparison of the deduced amino acid sequence showed that it is highly similar to NifF from Azotobacter vinelandii and NifF from Klebsiella pneumoniae. Analysis of translational fusions demonstrated that the regulation of transcription was the same as previously reported at the protein level. Insertional mutagen esis showed that NifF contributes significantly to nitrogenase activity under normal nitrogen-fixing conditions and that it is absolutely required for nitrogen fixation under iron limitation.

Full Text

The Full Text of this article is available as a PDF (314.8 KB).

Selected References

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

  1. Bennett L. T., Jacobson M. R., Dean D. R. Isolation, sequencing, and mutagenesis of the nifF gene encoding flavodoxin from Azotobacter vinelandii. J Biol Chem. 1988 Jan 25;263(3):1364–1369. [PubMed] [Google Scholar]
  2. Deistung J., Cannon F. C., Cannon M. C., Hill S., Thorneley R. N. Electron transfer to nitrogenase in Klebsiella pneumoniae. nifF gene cloned and the gene product, a flavodoxin, purified. Biochem J. 1985 Nov 1;231(3):743–753. doi: 10.1042/bj2310743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Drummond M. H. The base sequence of the nifF gene of Klebsiella pneumoniae and homology of the predicted amino acid sequence of its protein product to other flavodoxins. Biochem J. 1985 Dec 15;232(3):891–896. doi: 10.1042/bj2320891. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Fonstein M., Zheng S., Haselkorn R. Physical map of the genome of Rhodobacter capsulatus SB 1003. J Bacteriol. 1992 Jun;174(12):4070–4077. doi: 10.1128/jb.174.12.4070-4077.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Grabau C., Schatt E., Jouanneau Y., Vignais P. M. A new [2Fe-2S] ferredoxin from Rhodobacter capsulatus. Coexpression with a 2[4Fe-4S] ferredoxin in Escherichia coli. J Biol Chem. 1991 Feb 15;266(5):3294–3299. [PubMed] [Google Scholar]
  6. Hill S., Kavanagh E. P. Roles of nifF and nifJ gene products in electron transport to nitrogenase in Klebsiella pneumoniae. J Bacteriol. 1980 Feb;141(2):470–475. doi: 10.1128/jb.141.2.470-475.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hillmer P., Gest H. H2 metabolism in the photosynthetic bacterium Rhodopseudomonas capsulata: H2 production by growing cultures. J Bacteriol. 1977 Feb;129(2):724–731. doi: 10.1128/jb.129.2.724-731.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Hübner P., Masepohl B., Klipp W., Bickle T. A. nif gene expression studies in Rhodobacter capsulatus: ntrC-independent repression by high ammonium concentrations. Mol Microbiol. 1993 Oct;10(1):123–132. doi: 10.1111/j.1365-2958.1993.tb00909.x. [DOI] [PubMed] [Google Scholar]
  9. Hübner P., Willison J. C., Vignais P. M., Bickle T. A. Expression of regulatory nif genes in Rhodobacter capsulatus. J Bacteriol. 1991 May;173(9):2993–2999. doi: 10.1128/jb.173.9.2993-2999.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Jacobson M. R., Brigle K. E., Bennett L. T., Setterquist R. A., Wilson M. S., Cash V. L., Beynon J., Newton W. E., Dean D. R. Physical and genetic map of the major nif gene cluster from Azotobacter vinelandii. J Bacteriol. 1989 Feb;171(2):1017–1027. doi: 10.1128/jb.171.2.1017-1027.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Jouanneau Y., Meyer C., Naud I., Klipp W. Characterization of an fdxN mutant of Rhodobacter capsulatus indicates that ferredoxin I serves as electron donor to nitrogenase. Biochim Biophys Acta. 1995 Nov 21;1232(1-2):33–42. doi: 10.1016/0005-2728(95)00106-x. [DOI] [PubMed] [Google Scholar]
  12. Klipp W., Masepohl B., Pühler A. Identification and mapping of nitrogen fixation genes of Rhodobacter capsulatus: duplication of a nifA-nifB region. J Bacteriol. 1988 Feb;170(2):693–699. doi: 10.1128/jb.170.2.693-699.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kranz R. G., Foster-Hartnett D. Transcriptional regulatory cascade of nitrogen-fixation genes in anoxygenic photosynthetic bacteria: oxygen- and nitrogen-responsive factors. Mol Microbiol. 1990 Nov;4(11):1793–1800. doi: 10.1111/j.1365-2958.1990.tb02027.x. [DOI] [PubMed] [Google Scholar]
  14. Nieva-Gómez D., Roberts G. P., Klevickis S., Brill W. J. Electron transport to nitrogenase in Klebsiella pneumoniae. Proc Natl Acad Sci U S A. 1980 May;77(5):2555–2558. doi: 10.1073/pnas.77.5.2555. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Roberts G. P., MacNeil T., MacNeil D., Brill W. J. Regulation and characterization of protein products coded by the nif (nitrogen fixation) genes of Klebsiella pneumoniae. J Bacteriol. 1978 Oct;136(1):267–279. doi: 10.1128/jb.136.1.267-279.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Saeki K., Suetsugu Y., Tokuda K., Miyatake Y., Young D. A., Marrs B. L., Matsubara H. Genetic analysis of functional differences among distinct ferredoxins in Rhodobacter capsulatus. J Biol Chem. 1991 Jul 15;266(20):12889–12895. [PubMed] [Google Scholar]
  17. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Schatt E., Jouanneau Y., Vignais P. M. Molecular cloning and sequence analysis of the structural gene of ferredoxin I from the photosynthetic bacterium Rhodobacter capsulatus. J Bacteriol. 1989 Nov;171(11):6218–6226. doi: 10.1128/jb.171.11.6218-6226.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Schneider K., Müller A., Johannes K. U., Diemann E., Kottmann J. Selective removal of molybdenum traces from growth media of N2-fixing bacteria. Anal Biochem. 1991 Mar 2;193(2):292–298. doi: 10.1016/0003-2697(91)90024-n. [DOI] [PubMed] [Google Scholar]
  20. Schneider K., Müller A., Schramm U., Klipp W. Demonstration of a molybdenum- and vanadium-independent nitrogenase in a nifHDK-deletion mutant of Rhodobacter capsulatus. Eur J Biochem. 1991 Feb 14;195(3):653–661. doi: 10.1111/j.1432-1033.1991.tb15750.x. [DOI] [PubMed] [Google Scholar]
  21. Schüddekopf K., Hennecke S., Liese U., Kutsche M., Klipp W. Characterization of anf genes specific for the alternative nitrogenase and identification of nif genes required for both nitrogenases in Rhodobacter capsulatus. Mol Microbiol. 1993 May;8(4):673–684. doi: 10.1111/j.1365-2958.1993.tb01611.x. [DOI] [PubMed] [Google Scholar]
  22. Shah V. K., Stacey G., Brill W. J. Electron transport to nitrogenase. Purification and characterization of pyruvate:flavodoxin oxidoreductase. The nifJ gene product. J Biol Chem. 1983 Oct 10;258(19):12064–12068. [PubMed] [Google Scholar]
  23. St John R. T., Johnston H. M., Seidman C., Garfinkel D., Gordon J. K., Shah V. K., Brill W. J. Biochemistry and genetics of Klebsiella pneumoniae mutant strains unable to fix N2. J Bacteriol. 1975 Mar;121(3):759–765. doi: 10.1128/jb.121.3.759-765.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Wall J. D., Braddock K. Mapping of Rhodopseudomonas capsulata nif genes. J Bacteriol. 1984 May;158(2):404–410. doi: 10.1128/jb.158.2.404-410.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Weaver P. F., Wall J. D., Gest H. Characterization of Rhodopseudomonas capsulata. Arch Microbiol. 1975 Nov 7;105(3):207–216. doi: 10.1007/BF00447139. [DOI] [PubMed] [Google Scholar]
  26. Yakunin A. F., Gennaro G., Hallenbeck P. C. Purification and properties of a nif-specific flavodoxin from the photosynthetic bacterium Rhodobacter capsulatus. J Bacteriol. 1993 Nov;175(21):6775–6780. doi: 10.1128/jb.175.21.6775-6780.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES