Advertisement

A New Intermediate in TPNH-Linked Flavoproteins

  • Vincent Massey
  • Rowena G. Matthews
  • Gordon P. Foust
  • Larry G. Howell
  • Charles H. WilliamsJr.
  • Giuliana Zanetti
  • Severino Ronchi

Abstract

The early studies of Michaelis et. al. (1) clearly indicated that partial reduction of the flavin coenzymes led to the production of colored radical intermediates. Except for the catio-nic radical, which is produced in high yield at very acid pH values (pH<l) the ascription of discrete spectra to the flavin semiquinone was very much hampered by the low radical yield in half reduced flavin-mixtures (2). Further complications arose from the recognition of the existence of charge transfer complexes between reduced and oxidized flavin (2,3,4) with considerable absorption at wavelengths greater than 500 mμ. In fact systematic studies were needed with flavoproteins (5) as well as chemical models (6), (where practically quantitative reduction to the semiquinone can be achieved) in order to show that the neutral flavin semiquinone had a characteristic blue color due to high absorption at 600 my whereas the anionic semiquinone had a characteristic red color, due to the absorption maximum in the region 470–490 my. Figure 1 shows the two semiquinoid forms of glucose oxidase. Except for lipoyl dehydrogenase and glutathione reductase which are complicated by interaction of the flavin with the active center disulfide-dithiol (7,8) all simple flavoproteins so far tested yield either the neutral or anion radical with spectra similar to the two shown in Figure 1. In only two flavoprotein enzymes hitherto studied has the true semiquinoid form been shown unequivocally to be concerned in catalysis (9,10). Instead, with a number of enzymes, the catalytic intermediate appears to be a complex of the flavin and substrate in a partial reduction state, as in the case of D- and L-amino acid oxidases (11,12) or of the two redox active groups of the enzyme (flavin and disulfide) as in the case of lipoyl dehydrogenase (13) and glutathione reductase (8).

Keywords

Charge Transfer Complex Turnover Number Full Reduction Catalytic Intermediate Anaerobic Reduction 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

Bibliography

  1. 1.
    MICHAELIS, L., SCHUBERT, M. P., AND SMYTHE, C. V., J. Biol. Chem., 116, 587 (1963).Google Scholar
  2. 2.
    GIBSON, Q. H., MASSEY, V., AND ATHERTON, N. M., Biochem. J., 85, 369 (1962).PubMedGoogle Scholar
  3. 3.
    EHRENBERG, A., Arkiv Kemi, 17, 97 (1962).Google Scholar
  4. 4.
    MASSEY, V., AND PALMER, G., J. Biol. Chem., 237, 2347 (1962).PubMedGoogle Scholar
  5. 5.
    MASSEY, V., AND PALMER, G., Biochemistry, 5, 3181 (1966).PubMedCrossRefGoogle Scholar
  6. 6.
    EHRENBERG, A., MULLER, F., AND HEMMERICH, P., European J. Biochem., 2, 286 (1967).CrossRefGoogle Scholar
  7. 6a.
    MÜLLER, F., HEMMERICH, P., EHRENBERG, A., PALMER, G., AND MASSEY, V., European J. Biochem. submitted.Google Scholar
  8. 7.
    MASSEY, V., AND VEEGER, C., Biochim. Biophys. Acta, 40, 184 (1960).PubMedCrossRefGoogle Scholar
  9. 8.
    MASSEY, V., AND WILLIAMS, C. H., J. Biol. Chem., 240, 4470 (1965).PubMedGoogle Scholar
  10. 9.
    STRITTMATTER, P., Fed. Proc., 24, 1156 (1956).Google Scholar
  11. 10.
    KAMIN, H., MASTERS, B. S. S., GIBSON, Q. H., AND WILLIAMS, C. H., Fed. Proc., 24, 1164 (1965).PubMedGoogle Scholar
  12. 11.
    MASSEY, V., AND GIBSON, Q. H., Fed. Proc., 23, 18 (1964).PubMedGoogle Scholar
  13. 12.
    MASSEY, V., AND CURTI, B., J. Biol. Chem., 242, 1259 (1967).PubMedGoogle Scholar
  14. 13.
    MASSEY, V., GIBSON, Q. H., AND VEEGER, C., Biochem. J., 77, 341 (1960).PubMedGoogle Scholar
  15. 14.
    MATTHEWS, R. G., AND MASSEY, V., J. Biol. Chem., 244, 1779 (1969).PubMedGoogle Scholar
  16. 15.
    THEORELL, H., AND ÅKESON, A., Arch. Biochem. Biophys., 65, 439 (1956).PubMedCrossRefGoogle Scholar
  17. 16.
    HAAS, E., Biochem. Z., 290, 291 (1937).Google Scholar
  18. 17.
    BEINERT, H., AND SANDS, R. H., in Free Radicals in Biological Systems, Edited by M. S. Blois, W. H. Brown, R. M. Lemmon, R. O. Lindblom and M. Weissbluth. Academic Press, New York, 1961, p. 17.Google Scholar
  19. 18.
    NAKAMURA, T., YOSHIMURA, J., AND OGURA, Y., J. Biochem., 57, 554 (1965).PubMedGoogle Scholar
  20. 19.
    EHRENBERG, A., AND LUDWIG, G. D., Science, 127, 1177 (1958).PubMedCrossRefGoogle Scholar
  21. 20.
    BEINERT, H., J. Biol. Chem., 225, 465 (1957).PubMedGoogle Scholar
  22. 21.
    GIBSON, Q. H., AND MILNES, L., Biochem. J., 91, 161 (1964).PubMedGoogle Scholar
  23. 22.
    BENESI, H. A., AND HILDEBRAND, T. A., J. Am. Chem. Soc., 71, 2703 (1949).CrossRefGoogle Scholar
  24. 23.
    SHIN, M., TAGAWA, K., AND ARNON, D. I., Biochem. Z., 338, 84 (1963).PubMedGoogle Scholar
  25. 24.
    FOUST, G. P., AND MASSEY, V., in K. Yagi, Ed., Flavins and Flavoproteins, U. of Tokyo Press (1968), p. 7.Google Scholar
  26. 25.
    FOUST, G. P., MAYHEW, S. G., AND MASSEY, V., J. Biol. Chem., 244, 964 (1969).PubMedGoogle Scholar
  27. 26.
    DALZIEL, K., Biochem. J., 84, 244 (1962).PubMedGoogle Scholar
  28. 27.
    PALMER, G., AND MASSEY, V., in T. P. Singer, Ed., Biological Oxidations, Interscience, New York, 1968, p. 263.Google Scholar
  29. 28.
    HOSOKAWA, K., AND STANIER, R. Y., J. Biol. Chem., 241, 2453 (1966).PubMedGoogle Scholar
  30. 29.
    YANO, K., HIGASHI, N., AND ARIMA, K., Biochem. Biophys. Res. Comm., 34, 1 (1969).PubMedCrossRefGoogle Scholar
  31. 30.
    YANO, K., HIGASHI, N., NAKAMURA, S., AND ARIMA, K., Biochem. Biophys. Res. Comm., 34, 277 (1969).PubMedCrossRefGoogle Scholar
  32. 31.
    KATAGIRI, M., MAENO, H., YAMAMOTO, S., HAYAISHI, O., KITAO, T., AND OAE, S., J. Biol. Chem., 240, 3414 (1965).PubMedGoogle Scholar
  33. 32.
    MAKI, Y., YAMAMOTO, S., NOZAKI, M., AND HAYAISHI, O., J. Biol. Chem., 244, 2942 (1969).PubMedGoogle Scholar
  34. 33.
    WILLIAMS, C. H., ZANETTI, G., ARSCOTT, L. D., AND MCALLISTER, J. K., J. Biol. Chem., 242, 5226 (1967).PubMedGoogle Scholar
  35. 34.
    THELANDER, L., J. Biol. Chem., 242, 852 (1967).PubMedGoogle Scholar
  36. 35.
    ZANETTI, G., AND WILLIAMS, C. H., J. Biol. Chem., 242, 5232 (1967).PubMedGoogle Scholar
  37. 36.
    ZANETTI, G., WILLIAMS, C. H., AND MASSEY, V., J. Biol. Chem., 243, 4013 (1968).PubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin · Heidelberg 1970

Authors and Affiliations

  • Vincent Massey
    • 1
    • 2
  • Rowena G. Matthews
    • 1
    • 2
  • Gordon P. Foust
    • 1
    • 2
  • Larry G. Howell
    • 1
    • 2
  • Charles H. WilliamsJr.
    • 1
    • 2
  • Giuliana Zanetti
    • 1
    • 2
  • Severino Ronchi
    • 1
    • 2
  1. 1.Department of Biological ChemistryThe University of MichiganAnn ArborUSA
  2. 2.Biophysics Research DivisionVeterans Administration HospitalAnn ArborUSA

Personalised recommendations