Cell Biochemistry and Biophysics

, Volume 41, Issue 3, pp 357–365

Hydrogenase 3 but not hydrogenase 4 is major in hydrogen gas production by Escherichia coli formate hydrogenlyase at acidic pH and in the presence of external formate

  • Nelli Mnatsakanyan
  • Karine Bagramyan
  • Armen Trchounian
Original Article


Fermenting Escherichia coli is able to produce formate and molecular hydrogen (H2) when grown on glucose. H2 formation is possessed by two hydrogenases, 3 (Hyd-3) and 4 (Hyd-4), those, in conjunction with formate dehydrogenase H (Fdh-H), constitute distinct membrane-associated formate hydrogenylases. At slightly alkaline pH (pH 7.5), the production of H2 was found to be dependent on Hyd-4 and the F0F1-adenosine triphosphate (ATPase), whereas external formate increased the activity of Hyd-3. In this study with cells grown without and with external formate H2 production dependent on pH was investigated. In both types of cells, H2 production was increased after lowering of pH. At acidic pH (pH 5.5), this production became insensitive either to N,N′-dicyclohexylcarbodiimide or to osmotic shock and it became largely dependent on Fdh-H and Hyd-3 but not Hyd-4 and the F0F1-ATPase. The results indicate that Hyd-3 has a major role in H2 production at acidic pH independently on the F0F1-ATPase.

Index Entries

Formate hydrogenlyase pH Escherichia coli 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Böck, A. and Sawers, G. (1996) Fermentation, in Escherichia coli and Salmonella. Cellular and Molecular Biology (Neidhardt, F. C., Curtiss, J. R. II, Ingraham, J. L., Lin, E. C. C., Low, K. B., Magasanik, B., et al., ed.), 2nd ed. ASM Press, Washington, D.C., pp. 262–282.Google Scholar
  2. 2.
    Kobayashi, H. (2000) Why do bacteria have multiple transport systems for each kind of ions?. Recent Res. Dev. Bioenerg. 1, 33–39.Google Scholar
  3. 3.
    Zakharyan, E. and Trchounian, A. (2001) K+ influx by Kup in Escherichia coli is accompanied by a decrease in H+ efflux. FEMS Microbiol. Lett. 204, 61–64.PubMedCrossRefGoogle Scholar
  4. 4.
    Suppmann, B. and Sawers, G. (1994) Isolation and characterization of hypophosphate-resistant mutants of Escherichia coli: identification of the FocA protein, encoded by the pfl operon, as a putative formate transporter. Mol. Microbiol. 11, 965–982.PubMedCrossRefGoogle Scholar
  5. 5.
    Rossmann, R., Mailer, T., Lottspeich, F., and Böck, A. (1995) Characterization of a protease from Escherichia coli involved in hydrogenase maturation. Eur. J. Biochem. 227, 545–550.PubMedCrossRefGoogle Scholar
  6. 6.
    Self, W. T., Hasona, A., and Shanmugam, K. T. (2001) N-terminal truncations in the FhlA protein result in formate- and MoeA-independent expression of the hyc (formate hydrogenlyase) operon of Escherichia coli. Microbiology 147, 3093–3104.PubMedGoogle Scholar
  7. 7.
    Andrews, S. C., Berks, B. C., McClay, J., Ambler, A., Quail, M. A., Golby, P., et al. (1997) A 12-cistron Escherichia coli operon (hyf) encoding a putative proton-translocating formate hydrogenlyase system. Microbiology 143, 3633–3647.PubMedGoogle Scholar
  8. 8.
    Bagramyan, K., Mnatsakanyan, N., Poladian, A., Vassilian, A., and Trchounian, A. (2002) The roles of hydrogenases 3 and 4, and the F0F1-ATPase in H2 production by Escherichia coli at alkaline and acidic pH. FEBS Lett. 516, 172–178.PubMedCrossRefGoogle Scholar
  9. 9.
    Mnatsakanyan, N., Vassilian, A., Navasardian, L., Bagramyan, K., and Trchounian, A. (2002) Regulation of Escherichia coli formate hydrogenlyase activity by formate at alkaline pH. Curr. Microbiol. 45, 281–286.PubMedCrossRefGoogle Scholar
  10. 10.
    Axley, M. J., Grahame, D. A., and Stadtman, T. C. (1990) Escherichia coli formate hydrogenlyase. Purification and properties of the selenium-dependent formate dehydrogenase component. J. Biol. Chem. 265, 18213–18218.PubMedGoogle Scholar
  11. 11.
    Sawers, G. R., Ballantine, S. P., and Boxer, D. H. (1985) Differential expression of hydrogenase isoenzyme in Escherichia coli K-12: evidence for a third isoenzyme. J. Bacteriol. 164, 1324–1331.PubMedGoogle Scholar
  12. 12.
    Abaibou, H., Giordano, G., and Mandrand-Berthelot, M. A. (1997) Suppression of Escherichia coli formate hydrogenlyase activity by trimethylamine N-oxide is due to drainage of the inducer formate. Microbiology 143, 2657–64.PubMedCrossRefGoogle Scholar
  13. 13.
    Self, R. M. and Shanmugam, K. T. (2000) Isolation and characterization of mutated FhlA proteins, which activate transcription of the hyc operon (formate hydrogenlyase) of Escherichia coli in the absence of molybdate. FEMS Microbiol. Lett. 184, 47–52.PubMedGoogle Scholar
  14. 14.
    Casalot, L. and Rousset, M. (2001) Maturation of the [NiFe] hydrogenases. Trends Mcrobiol. 9, 228–237.CrossRefGoogle Scholar
  15. 15.
    Skibinski, D. A. G., Golby, P., Chang, Y-S., Sargent, F., Hoffman, R., Harper, R., et al. (2002) Regulation of the hydrogenase-4 operon of Escherichia coli by the σ54-dependent transcriptional activators FhlA and HyfR. J. Bacteriol. 184, 6642–6653.PubMedCrossRefGoogle Scholar
  16. 16.
    Dosch, D. C., Helmer, G. L., Sutton, S. H., Salvacion, F. F., and Epstein, W. (1991) Genetic analysis of potassium transport loci in Escherichia coli: evidence for three constitutive systems mediating uptake of potassium. J. Bacteriol. 173, 687–696.PubMedGoogle Scholar
  17. 17.
    Trchounian, A. and Vassilian, A. (1994) Relationship between the F0F1-ATPase and the K+-transport system within the membrane of anaerobically grown Escherichia coli. N.N′-dicyclohexylcarbodiimide-sensitive. ATPase activity in trk mutants. J. Bioenerg. Biomembr. 26, 563–571.PubMedCrossRefGoogle Scholar
  18. 18.
    Trchounian, A., Ohandjanyan, Y., Bagramyan, K., Vardanian, V., Zakharyan, E., Vassilian, A., et al. (1998) Relationship of the Escherichia coli TrkA system of potassium ion uptake with the F0F1-ATPase under growth conditions without aerobic and anaerobic respiration. Biosci. Rep. 18, 143–154.PubMedCrossRefGoogle Scholar
  19. 19.
    Bagramyan, K. A. and Martirosov, S. M. (1989) Formation of an ion transport supercomplex in Escherichia coli. An experimental model of direct transduction of energy. FEBS Lett. 246, 149–152PubMedCrossRefGoogle Scholar
  20. 20.
    Rossmann, R., Sawers, G., and Bock, A. (1991) Mechanisms of regulation of the formate hydrogen lyase pathway by oxygen, nitrate, and pH: determination of the formate regulon. Mol. Microbiol. 5, 2807–2814.PubMedCrossRefGoogle Scholar
  21. 21.
    Roe, A. J., McLaggan, D., Davidson, I., O'Byrne, C., and Booth, I. R. (1998) Perturbation of anion balance during inhibition growth of Escherichia coli by weak acids. J. Bacteriol. 180, 767–772.PubMedGoogle Scholar
  22. 22.
    Russell, J. B. and Diez-Gonsalez, F. (1998) The effect of fermentation acids on bacterial growth. Adv. Microb. Physiol. 39, 205–234.PubMedCrossRefGoogle Scholar
  23. 23.
    Brown, M. H. and Booth, I. R. (1991) Acidulants and pH, in Food Preservatives (Russel, N. J. and Gould, G. W., eds.), Blackie, Glasgow, UK, pp. 22–43.Google Scholar
  24. 24.
    Solioz, M. (1984) Dicyclohexylcarbodiimide as a probe for proton translocating enzymes. Trends Biochem. Sci. 9, 309–312.CrossRefGoogle Scholar
  25. 25.
    Trchounian, A., Bagramyan, K., and Poladian, A. (1997) Formate hydrogenlyase is needed for proton-potassium exchange through the F0F1 ATPase and the TrkA system in anaerobically grown and glycolysing Escherichia coli. Curr. Microbiol. 35, 201–206.PubMedCrossRefGoogle Scholar
  26. 26.
    Sasahara, K. C., Heinzinger, N. K., and Barrett, E. L. (1997) Hydrogen sulfide production and fermentative gas production by Salmonella typhimurium require F0F1 ATPase activity. J. Bacteriol. 179, 6736–6740.PubMedGoogle Scholar
  27. 27.
    Valiyaveetil, F., Hermolin, J., and Fillingame, H. (2002) pH dependent inactivation of solubilized F1F0 ATP synthase by dicyclohexylcarbodiimide: pK a of a detergent unmasked aspartyl-61 in Escherichia coli subunit c. Biochim. Biophys. Acta. 1553, 296–301.PubMedCrossRefGoogle Scholar
  28. 28.
    Gouesbet, G., Abaibou, H., Wu, L. F., Mandrand-Berthelot, M. A., and Blanco, C. (1993) Osmotic repression of anaerobic metabolic systems in Escherichia coli. J. Bacteriol. 175, 214–221.PubMedGoogle Scholar
  29. 29.
    Trchounian, A. and Kobayashi, H. (1999) Kup is the major K+ uptake system of Escherichia coli upon hyper-osmotic stress at a low pH. FEBS Lett. 447, 144–148.PubMedCrossRefGoogle Scholar
  30. 30.
    Künkel, A., Vorholt, J. A., Thauer, R. K., and Hedderich, R. (1998) An Escherichia coli hydrogenase-3-type hydrogenase in methanogenic archaea. Eur. J. Biochem. 252, 467–476.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc 2004

Authors and Affiliations

  • Nelli Mnatsakanyan
    • 1
  • Karine Bagramyan
    • 1
  • Armen Trchounian
    • 1
  1. 1.Department of Biophysics of the Biological FacultyYerevan State UniversityYerevanArmenia

Personalised recommendations