Abstract
The effects of Tinopals (cationic benzoxazoles) AMS-GX and 5BM-GX on NADH-oxidase, NADH:ferricyanide reductase, and NADH → APAD+ transhydrogenase reactions and energy-linked NAD+ reduction by succinate, catalyzed by NADH:ubiquinone oxidoreductase (Complex I) in submitochondrial particles (SMP), were investigated. AMS-GX competes with NADH in NADH-oxidase and NADH:ferricyanide reductase reactions (K i = 1 μM). 5BM-GX inhibits those reactions with mixed type with respect to NADH (K i = 5 μM) mechanism. Neither compound affects reverse electron transfer from succinate to NAD+. The type of the Tinopals' effect on the NADH → APAD+ transhydrogenase reaction, occurring with formation of a ternary complex, suggests the ordered binding of nucleotides by the enzyme during the reaction: AMS-GX and 5BM-GX inhibit this reaction uncompetitively just with respect to one of the substrates (APAD+ and NADH, correspondingly). The competition between 5BM-GX and APAD+ confirms that NADH is the first substrate bound by the enzyme. Direct and reverse electron transfer reactions demonstrate different specificity for NADH and NAD+ analogs: the nicotinamide part of the molecule is significant for reduced nucleotide binding. The data confirm the model suggesting that during NADH → APAD+ reaction, occurring with ternary complex formation, reduced nucleotide interacts with the center participating in NADH oxidation, whereas oxidized nucleotide reacts with the center binding NAD+ in the reverse electron transfer reaction.
Similar content being viewed by others
REFERENCES
Brandt, U. (ed.) (1998) Biochim. Biophys. Acta, 1364 (special issue).
Fearnley, I. M., and Walker, J. F. (1992) Biochim. Biophys.Acta, 1140, 105-135.
Walker, J. E. (1992) Q. Rev. Biophys., 25, 253-324.
Rao, N. A., Felton, S. P., Huennekens, F. M., and Mackler, B. (1963) J. Biol. Chem., 238, 449-455.
Ohnishi, T. (1998) Biochim. Biophys. Acta, 1364, 186-206.
Vinogradov, A. D., Sled, V. D., Burbaev, D. Sh., Grivennikova, V. G., Moroz, I. A., and Ohnishi, T. (1995) FEBS Lett., 370, 83-87.
Kang, D., Narabayashi, H., Sata, T., and Takeshige, K. (1983) J. Biochem., 94, 1301-1306.
Krishnamoorthy, G., and Hinkle, P. C. (1988) J. Biol. Chem., 263, 17566-17575.
Takeshige, K., and Minakami, S. (1979) Biochem. J., 180, 129-135.
Turrens, J. F., and Boveris, A. (1980) Biochem. J., 191, 421-427.
Dooijewaard, G., and Slater, E. C. (1976) Biochim. Biophys. Acta, 440, 1-15.
Dooijewaard, G., and Slater, E. C. (1976) Biochim. Biophys. Acta, 440, 16-35.
Gavricova, E. V., Grivennicova, V. G., Sled, V. D., Ohnishi, T., and Vinogradov, A. D. (1995) Biochim. Biophys. Acta, 1230, 23-30.
Sled, V. D., and Vinogradov, A. D. (1992) Biochim. Biophys. Acta, 1141, 262-268.
Ragan, C. I. (1976) Biochim. Biophys. Acta, 456, 249-290.
Degli Esposti, M. D., Ngo, A., McMullen, G. L., Chelli, A., Sparla, F., Benelli, B., Patta, M., and Linnane, A. W. (1996) Biochem. J., 313, 327-334.
Lenaz, G. (1998) Biochim. Biophys. Acta, 1364, 207-221.
Hatefi, Y., and Hanstein, W. G. (1973) Biochemistry, 12, 3515-3522.
Hatefi, Y., and Galante, Y. M. (1977) Proc. Natl. Acad. Sci. USA, 74, 846-850.
Minakami, S., Cremona, T., Ringler, R. L., and Singer, T. P. (1963) J. Biol. Chem., 238, 1528-1537.
Chance, B., and Hollunger, G. (1961) J. Biol. Chem., 236, 1555-1561.
Hommes, F. A. (1963) Biochim. Biophys. Acta, 77, 173-182.
Vallin, I., and Low, H. (1964) Biochim. Biophys. Acta, 92, 446-457.
Kotlyar, A. B., and Vinogradov, A. D. (1990) Biochim. Biophys. Acta, 1019, 151-158.
Galante, Y., and Hatefi, Y. (1978) Meth. Enzymol., 53, 15-21.
Chen, S., and Guillory, R. J. (1981) J. Biol. Chem., 256, 8318-8323.
Yamaguchi, M., Belogrudov, G. I., Matsuno-Yagi, A., and Hatefi, Y. (2000) Eur. J. Biochem., 267, 329-336.
Zharova, T. V., and Vinogradov, A. D. (1997) Biochim. Biophys. Acta, 1320, 256-264.
Frenkin, M. V., and Kotlyar, A. B. (1999) Biochim. Biophys. Acta, 1413, 139-146.
Avraam, R., and Kotlyar, A. B. (1991) Biokhimiya, 56, 2253-2260 (Russ.).
Vinogradov, A. D. (1993) J. Bioenerg. Biomembr., 25, 367-375.
Zakharova, N. V., Zharova, T. V., and Vinogradov, A. D. (1999) FEBS Lett., 444, 211-216.
Zakharova, N. V. (2002) Biochemistry (Moscow), 67, 651-661.
Anderson, W. M., and Delinck-Gordon, D. L. (1988) in Integration of Mitochondrial Function (Lemasters, J. J., Hackenbrock, Ch. R., and Westerhoff, H. V., eds.) Plenum Publishing Corporation, pp. 63-70.
Anderson, W. M., and Delinck, D. L. (1989) Biophys. J., 55, 568a.
Chambers, B. B., Wood, J. M., Delinck, D. L., and Anderson, W. M. (1991) FASEB J., 5, A1193.
Hatefi, Y. (1978) Meth. Enzymol., 53, 3-14.
Stein, A. M., Kaplan, N. O., and Ciotti, M. M. (1959) J. Biol. Chem., 234, 979-986.
Siegel, J. M., and Montgomery, G. A. (1959) Arch. Biochem. Biophys., 82, 288-299.
Hatefi, Y., and Stempel, K. E. (1969) J. Biol. Chem., 244, 2350-2357.
Belogrudov, G., and Hatefi, Y. (1994) Biochemistry, 33, 4571-4576.
Vinogradov, A. D., Gavrikova, E. V., Grivennikova, V. G., Zharova, T. V., and Zakharova, N. V. (1999) Biochemistry (Moscow), 64, 136-152.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Zakharova, N.V., Zharova, T.V. Kinetic Mechanism of Mitochondrial NADH:Ubiquinone Oxidoreductase Interaction with Nucleotide Substrates of the Transhydrogenase Reaction. Biochemistry (Moscow) 67, 1395–1404 (2002). https://doi.org/10.1023/A:1021818312040
Issue Date:
DOI: https://doi.org/10.1023/A:1021818312040