Biochemistry (Moscow)

, Volume 73, Issue 2, pp 123–129 | Cite as

Site-directed mutagenesis of conserved cysteine residues in NqrD and NqrE subunits of Na+-translocating NADH:quinone oxidoreductase

  • M. S. Fadeeva
  • Y. V. Bertsova
  • M. I. Verkhovsky
  • A. V. BogachevEmail author
Accelerated Publication


Each of two hydrophobic subunits of Na+-translocating NADH:quinone oxidoreductase (NQR), NqrD and NqrE, contain a pair of strictly conserved cysteine residues within their transmembrane α-helices. Site-directed mutagenesis showed that substitutions of these residues in NQR of Vibrio harveyi blocked the Na+-dependent and 2-n-heptyl-4-hydroxyquinoline N-oxide-sensitive quinone reductase activity of the enzyme. However, these mutations did not affect the interaction of NQR with NADH and menadione. It was demonstrated that these conserved cysteine residues are necessary for the correct folding and/or the stability of the NQR complex. Mass and EPR spectroscopy showed that NQR from V. harveyi bears only a 2Fe-2S cluster as a metal-containing prosthetic group.

Key words

Na+-translocating NADH:quinone oxidoreductase sodium potential Vibrio respiratory chain 





reduced nicotinamide hypoxanthine dinucleotide


midpoint redox potential


electronic paramagnetic resonance


2-n-heptyl-4-hydroxyquinoline N-oxide




Na+-translocating NADH:quinone oxidoreductase




sub-bacterial particles




Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Hayashi, M., Nakayama, Y., and Unemoto, T. (2001) Biochim. Biophys. Acta, 1505, 37–44.PubMedCrossRefGoogle Scholar
  2. 2.
    Bogachev, A. V., and Verkhovsky, M. I. (2005) Biochemistry (Moscow), 70, 143–149.CrossRefGoogle Scholar
  3. 3.
    Hayashi, M., Hirai, K., and Unemoto, T. (1995) FEBS Lett., 363, 75–77.PubMedCrossRefGoogle Scholar
  4. 4.
    Rich, P. R., Meinier, B., and Ward, B. (1995) FEBS Lett., 375, 5–10.PubMedCrossRefGoogle Scholar
  5. 5.
    Nakayama, Y., Hayashi, M., and Unemoto, T. (1998) FEBS Lett., 422, 240–242.PubMedCrossRefGoogle Scholar
  6. 6.
    Zhou, W., Bertsova, Y. V., Feng, B., Tsatsos, P., Verkhovskaya, M. L., Gennis, R. B., Bogachev, A. V., and Barquera, B. (1999) Biochemistry, 38, 16246–16252.PubMedCrossRefGoogle Scholar
  7. 7.
    Nakayama, Y., Yasui, M., Sugahara, K., Hayashi, M., and Unemoto, T. (2000) FEBS Lett., 474, 165–168.PubMedCrossRefGoogle Scholar
  8. 8.
    Hayashi, M., Nakayama, Y., Yasui, M., Maeda, M., Furuishi, K., and Unemoto, T. (2001) FEBS Lett., 488, 5–8.PubMedCrossRefGoogle Scholar
  9. 9.
    Bogachev, A. V., Bertsova, Y. V., Barquera, B., and Verkhovsky, M. I. (2001) Biochemistry, 40, 7318–7323.PubMedCrossRefGoogle Scholar
  10. 10.
    Turk, K., Puhar, A., Neese, F., Bill, E., Fritz, G., and Steuber, J. (2004) J. Biol. Chem., 279, 21349–21355.PubMedCrossRefGoogle Scholar
  11. 11.
    Barquera, B., Nilges, M. J., Morgan, J. E., Ramirez-Silva, L., Zhou, W., and Gennis, R. B. (2004) Biochemistry, 43, 12322–12330.PubMedCrossRefGoogle Scholar
  12. 12.
    Bogachev, A. V., Bertsova, Y. V., Ruuge, E. K., Wikstrom, M., and Verkhovsky, M. I. (2002) Biochim. Biophys. Acta, 1556, 113–120.PubMedCrossRefGoogle Scholar
  13. 13.
    Barquera, B., Morgan, J. E., Lukoyanov, D., Scholes, C. P., Gennis, R. B., and Nilges, M. J. (2003) J. Am. Chem. Soc., 125, 265–275.PubMedCrossRefGoogle Scholar
  14. 14.
    Bogachev, A. V., Murtazina, R. A., and Skulachev, V. P. (1997) FEBS Lett., 409, 475–477.PubMedCrossRefGoogle Scholar
  15. 15.
    Bogachev, A. V., Bertsova, Y. V., Bloch, D. A., and Verkhovsky, M. I. (2006) Biochemistry, 45, 3421–3428.PubMedCrossRefGoogle Scholar
  16. 16.
    Bogachev, A. V., Bertsova, Y. V., Aitio, O., Permi, P., and Verkhovsky, M. I. (2007) Biochemistry, 46, 10186–10191.PubMedCrossRefGoogle Scholar
  17. 17.
    Fadeeva, M. S., Yakovtseva, E. A., Belevich, G. A., Bertsova, Y. V., and Bogachev, A. V. (2007) Arch. Microbiol., 188, 341–348.PubMedCrossRefGoogle Scholar
  18. 18.
    Smith, P. K., Krohn, R. I., Hermanson, G. T., Mallia, A. K., Gartner, F. H., Provenzano, M. D., Fujimoto, E. K., Goeke, N. M., Olson, B. J., and Klenk, D. C. (1985) Analyt. Biochem., 150, 76–85.PubMedCrossRefGoogle Scholar
  19. 19.
    Li, Q., Li, L., Rejtar, T., Karger, B. L., and Ferry, J. G. (2005) J. Proteome Res., 4, 112–128.PubMedCrossRefGoogle Scholar
  20. 20.
    Saaf, A., Johansson, M., Wallin, E., and Heijne, G. (1999) Proc. Natl. Acad. Sci. USA, 96, 8540–8544.PubMedCrossRefGoogle Scholar
  21. 21.
    Duffy, E. B., and Barquera, B. (2006) J. Bacteriol., 188, 8343–8351.PubMedCrossRefGoogle Scholar
  22. 22.
    Peisach, J., Blumberg, W. E., Lode, E. T., and Coon, M. J. (1971) J. Biol. Chem., 246, 5877–5881.PubMedGoogle Scholar
  23. 23.
    Hayashi, M., Miyoshi, T., Sato, M., and Unemoto, T. (1992) Biochim. Biophys. Acta, 1099, 145–151.PubMedCrossRefGoogle Scholar
  24. 24.
    Fadeeva, M. S., Nunez, C., Bertsova, Y. V., Espin, G., and Bogachev, A. V. (2008) FEMS Microbiol. Lett., 279, 116–123.PubMedCrossRefGoogle Scholar
  25. 25.
    Bertsova, Y. V., and Bogachev, A. V. (2004) FEBS Lett., 563, 207–212.PubMedCrossRefGoogle Scholar
  26. 26.
    Barquera, B., Hellwig, P., Zhou, W., Morgan, J. E., Hase, C. C., Gosink, K. K., Nilges, M., Bruesehoff, P. J., Roth, A., Lancaster, C. R., and Gennis, R. B. (2002) Biochemistry, 41, 3781–3789.PubMedCrossRefGoogle Scholar

Copyright information

© Pleiades Publishing, Ltd. 2008

Authors and Affiliations

  • M. S. Fadeeva
    • 1
  • Y. V. Bertsova
    • 2
  • M. I. Verkhovsky
    • 3
  • A. V. Bogachev
    • 2
    Email author
  1. 1.Department of Bioinformatics and BioengineeringLomonosov Moscow State UniversityMoscowRussia
  2. 2.Department of Molecular Energetics of Microorganisms, Belozersky Institute of Physico-Chemical BiologyLomonosov Moscow State UniversityMoscowRussia
  3. 3.Helsinki Bioenergetics Group, Institute of BiotechnologyUniversity of HelsinkiHelsinkiFinland

Personalised recommendations