Advertisement

Archives of Microbiology

, Volume 156, Issue 6, pp 463–470 | Cite as

Reversible interconversion of the functional state of the gene regulator FNR from Escherichia coli in vivo by O2 and iron availability

  • P. Engel
  • M. Trageser
  • G. Unden
Original Papers
  • 28 Downloads

Abstract

FNR, the gene regulator of anaerobic respiratory genes of Escherichia coli is converted in vivo by O2 and by chelating agents to an inactive state. The interconversion process was studied in vivo in a strain with temperature controlled synthesis of FNR by measuring the expression of the frd (fumarate reductase) operon and the reactivity of FNR with the alkylating agent iodoacetic acid. FNR from aerobic bacteria is, after arresting FNR synthesis and shifting to anaerobic conditions, able to activate frd expression and behaves in the alkylation assay like anaerobic FNR. After shift from anaerobic to aerobic conditions, FNR no longer activates the expression of frd and reacts similar to aerobic FNR in the alkylation assay. The conversion of aerobic (inactive) to anaerobic (active) FNR occurs in the presence of chloramphenicol, an inhibitor of protein synthesis. Anaerobic FNR can also be converted post-translationally to inactive, metal-depleted FNR by growing the bacteria in the presence of chelating agents. The reverse is also possible by incubating metal-depleted bacteria with Fe2+. From the experiments it is concluded that the aerobic and the metal-depleted form of FNR can be transferred post-translationally and reversibly to the anaerobic (active) form. The response of FNR to changes in O2 supply therefore occurs at the FNR protein level in a reversible mode.

Key words

FNR protein Anaerobic respiration Fumarate reductase Oxygen regulation Iron(II) Gene regulation 

Abbreviation

BVred =

reduced benzyl viologen

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Bode C, Goebell H, Stähler E (1968) Zur Eliminierung von Trübungsfehlern bei der Eiweißbestimmung mit der Biuretmethode. Z Klin Chem Biochem 6:419–422Google Scholar
  2. Cole ST (1982) Nucleotide sequence coding for the flavoprotein subunit of the fumarate reductase of Escherichia coli. Eur J Biochem 122:479–484CrossRefGoogle Scholar
  3. Ditta G, Virts E, Palomares A, Kim CH (1987) The nifA gene of Rhizobium meliloti is oxygen regulated. J Bacteriol 169:3217–3223CrossRefGoogle Scholar
  4. Fischer HM, Bruderer T, Hennecke H (1988) Essential and nonessential domains in the Bradyrhizobium japonicum NifA protein: identification of indispensable cysteine residues potentially involved in redox reactivity and/or metal binding. Nucleic Acids Res 16:2207–2224CrossRefGoogle Scholar
  5. Fischer H-M, Hennecke H (1987) Direct response of Bradyrhizobium japonicum, nifA-mediated nif gene regulation to cellular oxygen status. Mol Gen Genet 209:621–626CrossRefGoogle Scholar
  6. Green J, Trageser M, Six S, Unden G, Guest JR (1991) Characterization of the FNR protein of E. coli, an iron-binding transcriptional regulator. Proc R Soc Lond Biol 244:137–144CrossRefGoogle Scholar
  7. Hanson RS, Phillips JA (1981) Chemical composition. In: Gerhardt D (ed) Methodology for general bacteriology. American Society for Microbiology, Washington, DC, pp 349–350Google Scholar
  8. Jones HM, Gunsalus RP (1987) Regulation of Escherichia coli fumarate reductase (frdABCD) operon expression by respiratory electron acceptors and the fnr gene product. J Bacteriol 169:3340–3349CrossRefGoogle Scholar
  9. Kullik I, Hennecke H, Fischer H-M (1989) Inhibition of Bradyrhizobium japonicum nifA-dependent nif gene activation by oxygen occurs at the NifA protein level and is irreversible. Arch Microbiol 151:191–197CrossRefGoogle Scholar
  10. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685CrossRefGoogle Scholar
  11. Lambden PR, Guest JR (1976) Mutants of Escherichia coli unable to use fumarate as an electron acceptor. J Gen Microbiol 97:145–160CrossRefGoogle Scholar
  12. Maniatis T, Fritsch EF, Sambrock J (1989) Molecular cloning. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NYGoogle Scholar
  13. Melville SB, Gunsalus RP (1990) Mutations in fnr that alter anerobic regulation of electron transport-associated genes in Escherichia coli. J Biol Chem 256:18733–18736Google Scholar
  14. Miller JH (1972) Experiments in molecular genetics. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New YorkGoogle Scholar
  15. Pascal M-C, Bonnefoy V, Fons M, Chippaux M (1986) Use of gene fusions to study the expression of fnr, the regulatory gene of anaerobic electron transfer in Escherichia coli. FEMS Microbiol Lett 36:35–39CrossRefGoogle Scholar
  16. Schneider E, Blundell M, Kennell D (1978) Translation and mRNA decay. Mol Gen Genet 160:121–129CrossRefGoogle Scholar
  17. Sharrocks AD, Green J, Guest JR (1990) In vivo and in vitro mutants of FNR, the anaerobic transcriptional regulator of E. coll. FEBS Lett 270:119–122CrossRefGoogle Scholar
  18. Spiro S, Guest JR (1987) Regulation and over-expression of the fnr gene of Escherichia coli. J Gen Microbiol 133:3279–3288PubMedGoogle Scholar
  19. Spiro S, Roberts RE, Guest JR (1989) FNR-dependent repression of the ndh gene of Escherichia coli and metal ion requirement for FNR-regulated gene expression. Mol Microbiol 3:601–608CrossRefGoogle Scholar
  20. Spiro S, Guest JR (1990) FNR and its role in oxygen-regulated gene expression in Escherichia coli. FEMS Microbiol Rev 75:399–428Google Scholar
  21. Thöny B, Fischer HM, Anthamatten D, Bruderer R, Hennecke H (1987) The symbiotic nitrogen fixation regulatory operon (fixRnifA) of Bradyrhizobium japonicum is expressed aerobically and is subject to a novel, nifA-independent type of activation. Nucleic Acids Res 15:8479–8499CrossRefGoogle Scholar
  22. Trageser M, Unden G (1989) Role of cysteine residues and metal ions in the regulatory functioning of FNR, the transcriptional regulator of anaerobic respiration in E. coli. Mol Microbiol 3:593–599CrossRefGoogle Scholar
  23. Trageser M, Spiro S, Duchêne A, Kojro E, Fahrenholtz F, Guest JR, Unden G (1990) Isolation of intact FNR protein (Mr 30000) of Escherichia coli. Mol Microbiol 4:21–27CrossRefGoogle Scholar
  24. Unden G, Guest JR (1985) Isolation and characterization of the FNR protein, the transcriptional regulator of anaerobic electron transport in E. coli. Eur J Biochem 146:193–199CrossRefGoogle Scholar
  25. Unden G, Duchêne A (1987) On the role of cyclic AMP and the FNR protein in E. coli growing anaerobically. Arch Microbiol 150:499–503CrossRefGoogle Scholar
  26. Unden G, Trageser M, Duchêne A (1990) Effect of positive redox potentials (>+400 mV) on the expression of anaerobic respiratory enzymes in E. coli. Mol Microbiol 4:315–319CrossRefGoogle Scholar
  27. Unden G, Trageser M (1991) Oxygen regulated gene expression in Escherichia coli: control of anaerobic respiration by the FNR protein. Antonie van Leeuwenhoek 59:65–76CrossRefGoogle Scholar
  28. Vieira J, Messing J (1982) The pUC plasmid, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene 19259–268CrossRefGoogle Scholar
  29. Wallace BJ, Young IG (1977) Role of quinones in electron transport to oxygen and nitrate in Escherichia coli. Biochim Biophys Acta 461:258–263Google Scholar
  30. Williams RJP (1982) Free manganese(II) and iron(II) cations can act as intracellular cell controls. FEBS Lett 140:3–10CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 1991

Authors and Affiliations

  • P. Engel
    • 1
  • M. Trageser
    • 1
  • G. Unden
    • 1
  1. 1.Institut für MikrobiologieJ. W. Goethe-UniversitätFrankfurt/MainGermany

Personalised recommendations