Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1991 Oct;173(19):6159–6161. doi: 10.1128/jb.173.19.6159-6161.1991

Glycine 100 in the dinitrogenase reductase of Rhodospirillum rubrum is required for nitrogen fixation but not for ADP-ribosylation.

L J Lehman 1, G P Roberts 1
PMCID: PMC208365  PMID: 1917849

Abstract

Dinitrogenase reductase (Rr2) is required for reduction of the molybdenum dinitrogenase in the nitrogen fixation reaction and is the target of posttranslational regulation in Rhodospirillum rubrum. This posttranslational regulation involves the ADP-ribosylation of Rr2. To study the structural requirements for these two functions of Rr2, i.e., activity and regulation, two site-directed mutations in nifH, the gene encoding Rr2, were constructed and analyzed. The mutations both affected a region of the protein known to be highly conserved in evolution and to be relevant to both of the above properties. These mutants were both Nif-, but one of the altered Rr2s was a substrate for ADP-ribosylation. This demonstrates that the ability of Rr2 to participate in nitrogen fixation can be separated from its ability to act as a substrate for ADP-ribosylation.

Full text

PDF
6159

Images in this article

6161Image
on p.6161

Selected References

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

  1. Bonam D., Lehman L., Roberts G. P., Ludden P. W. Regulation of carbon monoxide dehydrogenase and hydrogenase in Rhodospirillum rubrum: effects of CO and oxygen on synthesis and activity. J Bacteriol. 1989 Jun;171(6):3102–3107. doi: 10.1128/jb.171.6.3102-3107.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Chang C. L., Davis L. C., Rider M., Takemoto D. J. Characterization of nifH mutations of Klebsiella pneumoniae. J Bacteriol. 1988 Sep;170(9):4015–4022. doi: 10.1128/jb.170.9.4015-4022.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. 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]
  4. Hausinger R. P., Howard J. B. Thiol reactivity of the nitrogenase Fe-protein from Azotobacter vinelandii. J Biol Chem. 1983 Nov 25;258(22):13486–13492. [PubMed] [Google Scholar]
  5. Kanemoto R. H., Ludden P. W. Effect of ammonia, darkness, and phenazine methosulfate on whole-cell nitrogenase activity and Fe protein modification in Rhodospirillum rubrum. J Bacteriol. 1984 May;158(2):713–720. doi: 10.1128/jb.158.2.713-720.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Lehman L. J., Fitzmaurice W. P., Roberts G. P. The cloning and functional characterization of the nifH gene of Rhodospirillum rubrum. Gene. 1990 Oct 30;95(1):143–147. doi: 10.1016/0378-1119(90)90426-r. [DOI] [PubMed] [Google Scholar]
  7. Lehman L. J., Roberts G. P. Identification of an alternative nitrogenase system in Rhodospirillum rubrum. J Bacteriol. 1991 Sep;173(18):5705–5711. doi: 10.1128/jb.173.18.5705-5711.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Lowery R. G., Chang C. L., Davis L. C., McKenna M. C., Stephens P. J., Ludden P. W. Substitution of histidine for arginine-101 of dinitrogenase reductase disrupts electron transfer to dinitrogenase. Biochemistry. 1989 Feb 7;28(3):1206–1212. doi: 10.1021/bi00429a038. [DOI] [PubMed] [Google Scholar]
  9. Lowery R. G., Ludden P. W. Purification and properties of dinitrogenase reductase ADP-ribosyltransferase from the photosynthetic bacterium Rhodospirillum rubrum. J Biol Chem. 1988 Nov 15;263(32):16714–16719. [PubMed] [Google Scholar]
  10. Lowery R. G., Saari L. L., Ludden P. W. Reversible regulation of the nitrogenase iron protein from Rhodospirillum rubrum by ADP-ribosylation in vitro. J Bacteriol. 1986 May;166(2):513–518. doi: 10.1128/jb.166.2.513-518.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ludden P. W., Preston G. G., Dowling T. E. Comparison of active and inactive forms of iron protein from Rhodospirillum rubrum. Biochem J. 1982 Jun 1;203(3):663–668. doi: 10.1042/bj2030663. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. MacNeil T., MacNeil D., Roberts G. P., Supiano M. A., Brill W. J. Fine-structure mapping and complementation analysis of nif (nitrogen fixation) genes in Klebsiella pneumoniae. J Bacteriol. 1978 Oct;136(1):253–266. doi: 10.1128/jb.136.1.253-266.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Morrissey J. H. Silver stain for proteins in polyacrylamide gels: a modified procedure with enhanced uniform sensitivity. Anal Biochem. 1981 Nov 1;117(2):307–310. doi: 10.1016/0003-2697(81)90783-1. [DOI] [PubMed] [Google Scholar]
  14. Pope M. R., Murrell S. A., Ludden P. W. Covalent modification of the iron protein of nitrogenase from Rhodospirillum rubrum by adenosine diphosphoribosylation of a specific arginine residue. Proc Natl Acad Sci U S A. 1985 May;82(10):3173–3177. doi: 10.1073/pnas.82.10.3173. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Pretorius I. M., Rawlings D. E., O'Neill E. G., Jones W. A., Kirby R., Woods D. R. Nucleotide sequence of the gene encoding the nitrogenase iron protein of Thiobacillus ferrooxidans. J Bacteriol. 1987 Jan;169(1):367–370. doi: 10.1128/jb.169.1.367-370.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. 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]
  17. Russel M., Kidd S., Kelley M. R. An improved filamentous helper phage for generating single-stranded plasmid DNA. Gene. 1986;45(3):333–338. doi: 10.1016/0378-1119(86)90032-6. [DOI] [PubMed] [Google Scholar]
  18. Saari L. L., Triplett E. W., Ludden P. W. Purification and properties of the activating enzyme for iron protein of nitrogenase from the photosynthetic bacterium Rhodospirillum rubrum. J Biol Chem. 1984 Dec 25;259(24):15502–15508. [PubMed] [Google Scholar]
  19. Vandeyar M. A., Weiner M. P., Hutton C. J., Batt C. A. A simple and rapid method for the selection of oligodeoxynucleotide-directed mutants. Gene. 1988 May 15;65(1):129–133. doi: 10.1016/0378-1119(88)90425-8. [DOI] [PubMed] [Google Scholar]
  20. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]

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

RESOURCES