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Role of Cytochrome b5 in the NADH Synergism of NADPH-Dependent Reactions of the Cytochrome P-450 Monooxygenase System of Hepatic Microsomes

  • G. J. Mannering
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 58)

Abstract

Two systems are known to transfer electrons in hepatic microsomes: 1) the NADPH-dependent cytochrome P-450 monooxygenase system, and 2) the NADH dependent cytochrome b5 system. Their coexistence in the same organelle suggested to Estabrook and associates (Estabrook et al., 1971; Hildebrandt and Estabrook, 1971) that the systems might interact during the oxidation of drug substrates in much the same way electron transfer systems interact in mitochrondria. As evidence for the interaction of the two systems, they showed that the rate of NADH oxidation by liver microsomes was enhanced in the presence of NADPH and drug substrate, and that the rate of oxidation of NADH was related to the rate of oxidation of the substrate. They further implicated cytochrome b5 in cytochrome P-450 monooxygenase reactions by showing that the steady state of reduced cytochrome b5 in the presence of NADPH and NADH was decreased by the addition of drug substrate. Other experiments eliminated some alternative possibilities that might explain these observations, e.g., the possibility that NADH was converted to NADPH, or that NADH was sparing NADPH used in competing reactions occurring simultaneously in microsomes.

Keywords

Hepatic Microsome Mixed Function Oxidase Monooxygenase System Reconstituted System Electron Transfer System 
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.

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References

  1. Björkhem, I. and Danielsson, H. 1973. Heterogeneity of hepatic mixed function oxidases. Biochem. Biophys. Res. Comm. 51: 766–774.PubMedCrossRefGoogle Scholar
  2. Buening, M. and Franklin, M. 1974. Limitations in the use of the 340 nm absorbance maximum of NADPH for the determination of oxidation rates and stoichiometry during rat hepatic microsomal metabolism. Mol. Pharmacol, in press.Google Scholar
  3. Chaplin, M. D. and Mannering, G. J. 1970. Role of phospholipids in the hepatic microsomal drug metabolizing system. Mol. Pharmacol. 6: 631–640.PubMedGoogle Scholar
  4. Cinti, D. L. and Ozols, J. 1975. The role of cytochrome b5 in mixed function oxidations: effect of microsomal binding of the hemoprotein on hepatic N-demethylations. This publication.Google Scholar
  5. Cinti, D. L., Moldeus, P. and Schenkman, J. B. 1972. The role of the mitochondria in rat liver mixed function oxidation reactions. Biochem. Biophys. Res. Comm. 47: 1028–1035.PubMedCrossRefGoogle Scholar
  6. Cohen, B. S. and Estabrook, R. W. 1971. Microsomal electron transport reactions. II. The use of reduced triphospho-pyridine nucleotide for the oxidative N-demethylation of aminopyrine and other drug substrates. Arch. Biochem. Biophys. 143: 46–53.PubMedCrossRefGoogle Scholar
  7. Cohen, G. M. and Mannering, G. J. 1974. Sex-dependent differences in drug metabolism in the rat. III. Temporal changes in Type I binding and NADPH-cytochrome P-450 reductase during sexual maturation. Drug Metab. Disp. 2: 285–292.Google Scholar
  8. Conney, A. H. 1967. Pharmacological implications of microsomal enzyme induction. Pharmacol. Rev. 19: 317–366.PubMedGoogle Scholar
  9. Conney, A. H., Brown, R. R., Miller, J. A. and Miller, E. C. 1957. The metabolism of methylated aminoazo dyes. VI. Intracellular distribution and properties of the demethylase system. Cancer Res. 17: 628–633.PubMedGoogle Scholar
  10. Coon, M. J., van der Hoeven, T. A., Haugen, D. A., Guengerich, F. P., Vermilion, J. L. and Bailou, D. P. 1975. Biochemical characterization of highly purified cytochrome P-450 and other components of the mixed function oxidase system of liver microsomal membranes. This volume, p. 26.Google Scholar
  11. Correia, M. A. and Mannering, G. J. 1973a. DPNH synergism of TPNH-dependent mixed function oxidase reactions. Drug Metab. Disp. 1: 139–149.Google Scholar
  12. Correia, M. A. and Mannering, G. J. 1973b. Reduced diphospho-pyridine nucleotide synergism of the reduced triphospho-pyridine. I. Effects of activation and inhibition of the fatty acyl coenzyme A desaturation. Mol. Pharmacol. 9: 455–469.PubMedGoogle Scholar
  13. Correia, M. A. and Mannering, G. J. 1973c. Reduced diphospho-pyridine nucleotide synergism of the reduced triphospho-pyridine. II. Role of Type I drug-binding site of cytochrome P-450. Mol. Pharmacol. 9: 470–485.PubMedGoogle Scholar
  14. Enomoto, K. and Sato, R. 1973. Incorporation in vitro of purified cytochrome b2 into liver microsomal membranes. Biochem. Biophys. Res. Comm. 51: 1–7.PubMedCrossRefGoogle Scholar
  15. Estabrook, R. W., Shigematsu, A. and Schenkman, J. B. 1970. The contribution of the microsomal electron transport pathway to the oxidative metabolism of liver. Adv. Enzyme Reg. 8: 121–130.CrossRefGoogle Scholar
  16. Estabrook, R. W., Franklin, M., Baron, J., Shigematsu, A. and Hildebrandt, A. 1971. Properties of the membrane-bound electron transfer complex of the hepatic endoplasmic reticulum associated with drug metabolism. IN E. Mihich, Ed., Drugs and Cell Regulation, 227–257.Google Scholar
  17. Gigon, P. L., Gram, T. E. and Gillette, J. R. 1968. Effect of drug substrates on the reduction of hepatic microsomal cytochrome P-450 by NADPH. Biochem. Biophys. Res. Comm. 31: 558–562.PubMedCrossRefGoogle Scholar
  18. Gigon, P. L., Gram, T. E. and Gillette, J. R. 1969. Studies on the rate of reduction of hepatic microsomal cytochrome P-450 by reduced nicotinamide adenine dinucleotide phosphate: effect of drug substrates. Mol. Pharmacol. 5: 109–122.PubMedGoogle Scholar
  19. Gillette, J. R. 1966. Biochemistry of drug oxidation and reduction by enzymes in hepatic endoplasmic reticulum. Advan. Pharmacol. 4: 219–261.CrossRefGoogle Scholar
  20. Gillette, J. R. 1971. Factors affecting drug metabolism. Ann. N. Y. Acad. Sci. 179: 43–66.PubMedCrossRefGoogle Scholar
  21. Hildebrandt, A. and Estabrook, R. W. 1971. Evidence for the participation of cytochrome b5 in hepatic microsomal mixed function oxidation reactions. Arch. Biochem. Biophys. 143: 66–79.PubMedCrossRefGoogle Scholar
  22. Holtzman, J. L. 1970. Effect of 4,4-dideuteration of reduced nicotinamide adenine dinucleotide phosphate on the mixed function oxidases of hepatic microsomes. Biochemistry 9: 995–1001.PubMedCrossRefGoogle Scholar
  23. Holtzman, J. L. and Carr, M. L. 1970. Inhibition of hepatic microsomal mixed function oxidases by D2O. Life Sci. 9: 1033–1038.CrossRefGoogle Scholar
  24. Holtzman, J. L. and Rumack, B. H. 1971. The kinetics of cytochrome P-450 reductase stimulation by ethylmorphine. Life Sci. 10: 669–677.CrossRefGoogle Scholar
  25. Hrycay, E. G. and Estabrook, R. W. 1974. The effect of extra bound cytochrome b5 on cytochrome P-450-dependent enzyme activities in liver microsomes. Biochem. Biophys. Res. Comm. 60: 771–778.PubMedCrossRefGoogle Scholar
  26. Imai, Y. and Omura, T. 1974. Unpublished results cited by Mannering, Kuwahara and Omura (Biochem. Biophys. Res. Comm. 57:476–481).CrossRefGoogle Scholar
  27. Imai, Y. and Sato, R. 1966. Substrate interaction with hydroxylase system in liver microsomes. Biochem. Biophys. Res. Comm. 22: 620–626.PubMedCrossRefGoogle Scholar
  28. Jansson, I. and Schenkman, J. B. 1973. Evidence against participation of cytochrome b5 in the hepatic microsomal mixed-function oxidase reaction. Mol. Pharmacol. 9: 840–845.PubMedGoogle Scholar
  29. Jeffery, E. and Mannering, G. J. 1974a. Discrepancy in the measurement of TPNH oxidized during N-demethylation due to the presence of nucleotide pyrophosphatase. Mol. Pharmacol, in press.Google Scholar
  30. Jeffery, E. and Mannering, G. J. 1974b. Unpublished results.Google Scholar
  31. Lichtenberger, F. 1975. Discussion in this publication.Google Scholar
  32. Lu, A. Y. H. 1974. Private communication.Google Scholar
  33. Lu, A. Y. H., West, S. B., Vore, M., Ryan, D. and Levin, W. 1974. Role of cytochrome b5 in hydroxylation by a reconstituted cytochrome P-450-containing system. J. Biol. Chem. 249, in press.Google Scholar
  34. Lu, A. Y. H., Levin, W., West, S. B., Vore, M., Ryan, D. Kuntzman, and Conney, A. H. 1975. Role of cytochrome b5 in NADPH-and NADH-dependent hydroxylation by the reconstituted cytochrome P-450- or P-448-containing system. This publication.Google Scholar
  35. Mannering, G. J. 1968. Significance of stimulation and inhibition of drug metabolism in pharmacological testing. In A. Burger, Ed. Selected Pharmacological Testing Methods, 51–119.Google Scholar
  36. Mannering, G. J. 1971a. Role of substrate binding to P-450 hemoprotein in drug metabolism. IN E. Mihich, Ed., Drugs and Cell Regulation, 197–225.Google Scholar
  37. Mannering, G. J. 1971b. Properties of cytochrome P-450 as affected by environmental factors: qualitative changes due to administration of polycyclic hydrocarbons. Metabolism 20: 228–245.PubMedCrossRefGoogle Scholar
  38. Mannering, G. J. 1971c. Microsomal enzyme systems which catalyze drug metabolism. IN B. N. LaDu, H. G. Mandel and E. L. Way, Eds. Fundamentals of Drug Metabolism and Drug Disposition, 206–252.Google Scholar
  39. Mannering, G. J., Kuwahara, S. and Omura, T. 1974. Immunochemical evidence for the participation of cytochrome b5 in the NADH synergism of the NADH-dependent mono-oxidase system of hepatic microsomes. Biochem. Biophys. Res. Comm. 57: 476–481.PubMedCrossRefGoogle Scholar
  40. Modurzadeh, J. and Kamin, H. 1965. Reduction of microsomal cytochromes by pyridine nucleotides. Biochem. Biophys. Acta. 99: 205–226.Google Scholar
  41. Netter, K. J. and Illing, H. P. A. 1974. Kinetic experiments on the synergistic effect of NADH on microsomal drug oxidation. Xenobiotica 4: 549–561.PubMedCrossRefGoogle Scholar
  42. Nelson, D. and Mannering, G. J. 1974. Unpublished results.Google Scholar
  43. Oshino, N. 1972. Dynamic behavior during dietary induction of the terminal enzyme (cyanide-sensitive factor) of the stearyl CoA desaturation system of rat liver microsomes. Arch. Biochem. Biophys. 149: 378–387.PubMedCrossRefGoogle Scholar
  44. Oshino, N., Imai, Y. and Sato, R. 1971. A function of cytochrome b5 in fatty acid desaturation by rat liver microsomes. J. Biochem. 69: 155–168.PubMedGoogle Scholar
  45. Oshino, N. and Sato, R. 1971. Stimulation by phenols of the re-oxidation of microsomal bound cytochrome b5 and its implication to fatty acid desaturation. J. Biochem. 69: 169–180.PubMedGoogle Scholar
  46. Oshino, N. and Sato, R. 1972. Dietary control of the microsomal stearyl CoA desaturation enzyme system in rat liver. Arch.Biochem. Biophys. 149: 369–377.PubMedCrossRefGoogle Scholar
  47. Sasame, H. A., Mitchell, J. R., Thorgeirsson, S. and Gillette, J. R. 1973. Relationship between NADH and NADPH oxidation during drug metabolism. Drug Metab. Disp. 1: 150–155.Google Scholar
  48. Sasame, H. A., Thorgeirsson, S. S., Mitchell, J. R. and Gillette, J. R. 1974a. The possible involvement of cytochrome b5 in the oxidation of lauric acid by microsomes from kidney cortex and liver of rats. Life Sci. 14: 35–46.PubMedCrossRefGoogle Scholar
  49. Sasame, H. A., Thorgeirsson, S. S., Menard, R. H., Hinson, J. A., Mitchell, J. R., and Gillette, J. R. 1974b. A role of cytochrome b5 in both NADH and NADPH-mediated cytochrome P-450 enzymatic reaction in mammalian tissues. Fed. Proc. 33: 1437.Google Scholar
  50. Sasame, H. A., Thorgeirsson, S. S., Mitchell, J. R. and Gillette, J. R. 1975. The role of cytochrome b5 in cytochrome P-450 enzymes. This publication.Google Scholar
  51. Schenkman, J. B. 1968. Effect of substrates on hepatic microsomal cytochrome P-450. Hoppe-Seyler’s Z. Physiol. Chem. 349: 1624–1628.PubMedGoogle Scholar
  52. Schenkman, J. B. and Jansson, I. 1975. Interaction between microsomal electron transfer pathways. This publication.Google Scholar
  53. Schenkman, J. B., Frey, I., Remmer, H. and Estabrook, R. W. 1967. Sex differences in drug metabolism by rat liver microsomes. Mol. Pharmacol. 3: 516–525.PubMedGoogle Scholar
  54. Shoeman, D. W., Chaplin, M. D. and Mannering, G. J. 1969. Induction of drug metabolism. III. Further evidence for the formation of a new P-450 hemoprotein after treatment of rats with 3-methylcholanthrene. Mol. Pharmacol. 5: 412–419.Google Scholar
  55. Sitar, D. S. and Mannering, G. J. 1973. Determination of apparent kinetic constants of the microsomal hydroxylation of amo-barbital, hexobarbital and pentobarbital. Drug Metab. Disp. 1: 663–668.Google Scholar
  56. Strittmatter, P., Rogers, M. J. and Spatz, L. 1972. The binding of cytochrome b5 to liver microsomes. J. Biol. Chem. 247: 7188–7194.PubMedGoogle Scholar
  57. Staudt, H., Lichtenberger, F. and Ullrich, V. 1974. The role of NADH in uncoupled microsomal monoxygenations. Eur. J. Biochem. 46: 99–106.PubMedCrossRefGoogle Scholar
  58. van der Hoeven, T. A., Haugen, D. A., and Coon, M. J. 1974. Cytochrome P-450 purified to apparent homogeneity from phenobar-bital-induced rabbit liver microsomes: catalytic activity and other properties. Biochem. Biophys. Res. Comm. 60: 569–575.Google Scholar
  59. West, S. B., Levin, W., Ryan, D., Vore, M. and Lu, A. Y. H. 1974. Liver microsomal electron transport systems. II. The involvement of cytochrome b5 in the NADH-dependent hydroxylation of 3,4-benzpyrene by a reconstituted cytochrome P-448-containing system. Biochem. Biophys. Res. Comm. 58: 516–522.Google Scholar

Copyright information

© Plenum Press, New York 1975

Authors and Affiliations

  • G. J. Mannering
    • 1
  1. 1.Department of Pharmacology, Medical SchoolUniversity of MinnesotaMinneapolisUSA

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