The Topology of the Mammalian Cytochrome P-450 Active Site

  • Gerald T. Miwa
  • Anthony Y. H. Lu

Abstract

Currently, the only direct method for visualizing the active site of an enzyme is through high-resolution X-ray crystallography. Indeed, for P450cam, a soluble enzyme crystallized from Pseudomonas putida, this has yielded the most complete three-dimensional information to date on a P450 active site (Chapter 13). Similar data are not available for any of the mammalian P-450 isozymes, however, since no crystals have yet been successfully produced for these hydrophobic proteins. Consequently, investigations have relied on much more indirect means to obtain information about the topology of the active site of these enzymes.

Keywords

Pyrrole Ring Complete Amino Acid Sequence Porphyrin Iron Substrate Binding Domain Polycyclic Aromatic Hydro 
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. 1.
    Lu, A. Y. H., and West, S. B., 1980, Multiplicity of mammalian microsomal cytochromes P-450, Pharmacol. Rev. 31: 277–291.Google Scholar
  2. 2.
    Johnson, E. F., 1979, Multiple forms of cytochrome P-450: Criteria and significance, in: Reviews in Biochemical Toxicology ( E. Hodgson, J. R. Bend, and R. M. Philpot, eds.) Elsevier/North Holland, Amsterdam, pp. 1–26.Google Scholar
  3. 3.
    Ryan, D. E., Thomas, P. E. Reik, L. M., and Levin, W., 1982, Purification, characterization and regulation of five rat hepatic microsomal cytochrome P-450 isozymes, Xenobiotica 12: 727–744.Google Scholar
  4. 4.
    Guengerich, F. P., Wang, P., and Davidson, N. K., 1982, Estimation of isozymes of microsomal cytochrome P-450 in rats, rabbits, and humans using immunochemical staining coupled with sodium dodecyl sulfate—polyacrylamide gel electrophoresis, Biochemistry 21: 1698–1706.PubMedCrossRefGoogle Scholar
  5. 5.
    Pickett, C. B., Jeter, R. L., Morin, J., and Lu, A. Y. H., 1981, Electroimmunochemical quantitation of cytochrome P-450, cytochrome P-448, and epoxide hydrolase in rat liver microsomes, J. Biol. Chem. 256: 8815–8820.PubMedGoogle Scholar
  6. 6.
    Koop, D. R., Persson, A. V., and Coon, M. J., 1981, Properties of electrophoretically homogeneous constitutive forms of liver microsomal cytochrome P-450, J. Biol. Chem. 256: 10704–10711.PubMedGoogle Scholar
  7. 7.
    Cheng, K.-C., and Schenkman, J. B., 1982, Purification and characterization of two constitutive forms of rat liver microsomal cytochrome P-450, 1. Biol. Chenu. 257: 2378 2385.Google Scholar
  8. 8.
    Canady, W. J., Robinson, D. A., and Colby, H. D., 1974, A partition model for hepatic cytochrome P-450—hydrocarbon complex formation, Biochem. Pharmacol. 23: 30753078.Google Scholar
  9. 9.
    White, R. E., Oprian, D. D., and Coon, M. J., 1980, Resolution of multiple equilibria in binding of small molecules to cytochrome P-450LM, in: Microsomes, Drug Oxidations, and Chemical Carcinogenesis, Volume I (M. J. Coon, A. H. Conney, R. W. Estabrook, H. V. Gelboin, J. R. Gillette, and P. J. O’Brien, eds.), Academic Press, New York, pp. 243–251.Google Scholar
  10. 10.
    Jefcoate, C. R., Gaylor, J. L., and Calabrese, R. L., 1969, Ligand interactions with cytochrome P-450. I. Binding of primary amines, Biochemistry 8: 3455–3463.PubMedCrossRefGoogle Scholar
  11. 11.
    Jansson, I. Orrenius, S., Ernster, L. and Schenkman, J. B., 1972, A study of the interactions of a series of substituted barbituric acids with the hepatic microsomal monooxygenase, Arch. Biochem. Biophys. 151:391–400.Google Scholar
  12. 12.
    McMahon, R. E., 1961, Demethylation studies. I. The effect of chemical structure and lipid solubility, J. Med. Pharm. Chem. 4: 67–78.CrossRefGoogle Scholar
  13. 13.
    Martin, Y. C., and Hansch, C., 1971, Influence of hydrophobic character on the relative rate of oxidation of drugs by rat liver microsomes, J. Med. Chem 14: 777–779.PubMedCrossRefGoogle Scholar
  14. 14.
    Cohen, G. M., and Mannering, G. J., 1973, Involvement of a hydrophobic site in the inhibition of the microsomal p-hydroxylation of aniline by alcohols, Mol. Pharmacol. 9: 383–397.PubMedGoogle Scholar
  15. 15.
    Cho, A. K., and Miwa, G. T., 1973, The role of ionization in the N-demethylation of some N,N-dimethylamines, Drug Metab. Dispos. 2: 477–483.Google Scholar
  16. 16.
    Backes, W. L., and Canady, W. J., 1981, The interaction of hepatic cytochrome P-450 with organic solvents; The effect of organic solvents on apparent spectral binding constants for hydrocarbon substrates, J. Biol. Chem. 256: 7213–7227.PubMedGoogle Scholar
  17. 17.
    Kaminsky, L. S., Fasco, M. J., and Guengerich, F. P., 1980, Comparison of different forms of purified cytochrome P-450 from rat liver by immunological inhibition of regio-and stereoselective metabolism of warfarin, J. Biol. Chem. 255: 85–91.PubMedGoogle Scholar
  18. 18.
    Cho, A. K., and Wright, J., 1978, Minireview: Pathways of metabolism of amphetamine and related compounds, Life Sci. 22: 363–372.PubMedCrossRefGoogle Scholar
  19. 19.
    Jerina, D. M., Michaud, D. P., Feldmann, R. J., Armstrong, R. N., Vyas, K. P., Thakker, D. R., Yagi, H., Thomas, P. E., Ryan, D. E., and Levin, W., 1982, Stereochemical modeling of the catalytic site of cytochrome P-450c, in: Microsomes, Drug Oxidations and Drug Toxicity (R. Sato and R. Kato, eds.), Japan Scientific Societies Press, Tokyo, pp. 195–201.Google Scholar
  20. 20.
    Ortiz de Montellano, P. R., Mangold, B. L. K., Wheeler, C., Kunze, K. L., and Reich, N.O., 1983, Stereochemistry of cytochrome P-450-catalyzed epoxidation and prosthetic heure alkylation, J. Biol. Chem. 258: 4208–4213.PubMedGoogle Scholar
  21. 21.
    Waxman, D. J., Ko, A., and Walsh, C., 1983, Regioselectivity and stereoselectivity of androgen hydroxylations catalyzed by cytochrome P-450 isozymes purified from phenobarbital-induced rat liver, J. Biol. Chem. 258: 11937–11947.PubMedGoogle Scholar
  22. 22.
    Waxman, D. J., Light, D. R., and Walsh, C., 1982, Chiral sulfoxidations catalyzed by rat liver cytochrome P-450, Biochemistry 21: 2499–2507.PubMedCrossRefGoogle Scholar
  23. 23.
    Tullman, R. H., Walsh, J. S., and Miwa, G. T., 1984, The stereochemistry of P-450 and P-448 catalyzed 0-dealkylation of 7-ethoxycoumarin, Fed. Proc. 43: 346.Google Scholar
  24. 24.
    van der Hoeven, T. A., Haugen, D. A., and Coon, M. J., 1974, Cytochrome P-450 purified to apparent homogeneity from phenobarbital-induced rabbit liver microsomes: Catalytic activity and other properties, Biochem. Biophys. Res. Commun. 60: 569–575.PubMedCrossRefGoogle Scholar
  25. 25.
    Imai, Y., and Sato, R., 1974, A gel-electrophoretically homogeneous preparation of cytochrome P-450 from liver microsomes of phenobarbital-pretreated rabbits, Biochem. Biophys. Res. Commun. 60: 8–14.PubMedCrossRefGoogle Scholar
  26. 26.
    Heinemann, F. S., and Ozols, J., 1983, The complete amino acid sequence of rabbit phenobarbital-induced liver microsomal cytochrome P-450, J. Biol. Chem. 258: 41954201.Google Scholar
  27. 27.
    Tarr, G. E., Black, S. D., Fujita, V. S., and Coon, M. J., 1983, Complete amino acid sequence and predicted membrane topology of phenobarbital-induced cytochrome P450 (isozyme 2) from rabbit liver microsomes, Proc. Natl. Acad. Sci. USA 80: 65526556.Google Scholar
  28. 28.
    Novak, R. F., and Vatsis, K. P., 1982, ‘H Fourier transform nuclear magnetic resonance relaxation rate studies on the interaction of acetanilide with purified isozymes of rabbit liver microsomal cytochrome P-450 and with cytochrome b5, Mol. Pharmacol. 21: 70 1709.Google Scholar
  29. 29.
    Novak, R. F., Kapetanovic, 1. M., and Mieyal, J. J., 1977, Nuclear magnetic resonance studies of substrate—hemeprotein complexes in solution, Mol. Pharmacol. 13: 15–30.Google Scholar
  30. 30.
    Coon, M. J., and Vatsis, K. P., 1978, Biochemical studies on chemical carcinogenesis: Role of multiple forms of liver microsomal cytochrome P-450 in the metabolism of benzo[a]pyrene and other foreign compounds, in: Polycyclic Hydrocarbons and Cancer: Environment, Chemistry and Metabolism, Volume I, ( H. V. Gelboin and P.O.P. Ts’o, eds.), Academic Press, New York, pp. 335–360.Google Scholar
  31. 31.
    White, R. E., McCarthy, M.-B., Egeberg, K. D., and Sligar, S. G., 1984, Regioselectivity in the cytochromes P-450: Control by protein constraints and by chemical reactivities, Arch. Biochem. Biophys. 228: 493–502.PubMedCrossRefGoogle Scholar
  32. 32.
    Imai, Y., 1982, Interaction of polycyclic hydrocarbons with cytochrome P-450. III. Effects of hydrocarbon binding on the interaction of some ligands with P-448, heure, J. Biochem. 92: 77–88.PubMedGoogle Scholar
  33. 33.
    Thakker, D. R., Yagi, H., Akagi, H., Koreeda, M., Lu, A. Y. H., Levin, W., Wood, A. W., Conney, A. H., and Jerina, D. M., 1977, Metabolism of benzo[a]pyrene. VI. Stereoselective metabolism of benzo[a]pyrene and benzo[a]pyrene 7,8-dihydrodiol to diol epoxides, Chem. Biol. Interact. 16: 281–300.PubMedCrossRefGoogle Scholar
  34. 34. Armstrong, R. N., Levin, W., Ryan, D. E., Thomas, P. E., Mah, H. D., and Jerina, D. M., 1981, Stereoselectivity of rat liver cytochrome P-450c in formation of benzo[a]pyrene 4,5-oxide, Biochem. Biophys. Res. Commun.100:1077–1084.Google Scholar
  35. 35.
    Boyd, D. R., Gadaginamath, G. S., Kher, A., Malone, J. F., Yagi, H., and Jerina. D. M., 1980, (+)- and (—)-benzo[a]pyrene 7,8-oxide: Synthesis, absolute stereochemistry, and stereochemical correlation with other mammalian metabolites of benzo[a]pyrene, J. Chem. Soc. Perkin Trans. 1 1980: 2112–2116.CrossRefGoogle Scholar
  36. 36.
    Levin, W., Buening, M. K., Wood, A. W., Chang, R. L., Kezierski, B., Thakker, D. R., Boyd, D. R., Gadaginamath, G. S., Armstrong, R. N., Yagi, H., Karle, J. M., Slaga, T. J., Jerina, D. M., and Conney, A. H., 1980, An enantiomeric interaction in the metabolism and tumorigenicity of (+)- and (—)-benzo[a]pyrene 7,8-oxide, J. Biol. Chem. 255: 9067–9074.PubMedGoogle Scholar
  37. 37.
    Thakker, D. R., Levin, W., Yagi, H., Turujman, S., Kapadia, D., Conney, A. H., and Jerina, D. M., 1979, Absolute stereochemistry of the trans-dihydrodiols formed from benzo[a]anthracene by liver microsomes, Chem. Biol. Interact. 27: 145–161.PubMedCrossRefGoogle Scholar
  38. 38.
    van Bladeren, P. J., Armstrong, R. N., Cobb, D., Thakker, D. R., Ryan, D. E., Thomas, P. E., Sharma, N. D., Boyd, D. R., Levin, W., and Jerina, D. M., 1982, Stereoselective formation of benz[a]anthracene (+)-(5S,6R)-oxide and (+)-(8R,9S)-oxide by a highly purified and reconstituted system containing cytochrome P-450e, Biochem. Biophys. Res. Commun. 106: 602–609.PubMedCrossRefGoogle Scholar
  39. 39.
    Nordqvist, M., Thakker, D. R., Vyas, K. P., Yagi, H., Levin, W., Ryan, D. E., Thomas, P. E., Conney, A. H., and Jerina, D. M., 1981, Metabolism of chrysene and phenanthrene to bay-region diol epoxides by rat liver enzymes, Mol. Pharmacy!. 19: 168–178.Google Scholar
  40. 40.
    Jerina, D. M., Michaud, D. P., Feldmann, R. J., Armstrong, R. N., Vyas, K. P., Thakker, D. R., Yagi, H., Thomas, P. E., Ryan, D. E., and Levin, W., 1982, Stereochemical modeling of the catalytic site of cytochrome P-450e, in: Microsomes, Drug Oxidations and Drug Toxicity (R. Sato and R. Kato, eds.), Japan Scientific Societies Press, Tokyo, pp. 195–201.Google Scholar
  41. 41.
    Thakker, D. R., Levin, W., Yagi, H., Conney, A. H., and Jerina, D. M., 1982, Regio-and stereoselectivity of hepatic cytochrome P-450 toward polycyclic aromatic hydrocarbon substrates, in: Biological Reactive Intermediates, Volume IlA (R. Snyder, D. V. Parke, D. J. Jollow, C. G. Gibson, and C. M. Witmer, eds.), Plenum Press, New York, pp. 525–539.Google Scholar
  42. 42.
    Miwa, G. T., Walsh, J. S., and Lu, A. Y. H., 1984, Kinetic isotope effects on cytochrome P-450-catalyzed oxidation reactions: The oxidative 0-dealkylation of 7-ethoxycoumarin, J. Biol. Chem. 259: 3000–3004.PubMedGoogle Scholar
  43. 43.
    Harada, N., Miwa, G. T., Walsh, J. S., and Lu, A. Y. H., 1984, Kinetic isotope effects on cytochrome P-450-catalyzed oxidation reactions: Evidence for the irreversible formation of activated oxygen intermediate of cytochrome P-450, J. Biol. Chem. 259: 30053010.Google Scholar
  44. 44.
    Walsh, J. S., and Miwa, G. T., 1984, The mechanism of the cytochrome P-448 mediated 6-hydroxylation of 7-ethoxycoumarin, Biochem. Biophys. Res. Commun. 121: 960–965.PubMedCrossRefGoogle Scholar
  45. 45.
    Fujii-Kuriyama, Y., Mizukami, Y., Kawajiri, K., Sogawa, K., and Muramatsu, M., 1982, Primary structure of a cytochrome P-450: Coding nucleotide sequence of phenobarbital-inducible cytochrome P-450 cDNA from rat liver, Proc. Natl. Acad. Sci. USA 79: 2793–2797.PubMedCrossRefGoogle Scholar
  46. 46.
    Kunze, K. L., Mangold, B. L. K., Wheeler, C., Beilan, H. S., and Ortiz de Montellano, P. R., 1983, The cytochrome P-450 Active Site: Regiospecificity of prosthetic heure alkylation by olefins and acetylenes. J. Biol. Chem. 258: 4202–4207.PubMedGoogle Scholar
  47. 47.
    Ortiz de Montellano, P. R., Kunze, K. L., and Beilan, H. S., 1983, Chiral orientation of prosthetic heure in the cytochrome P-450 active site, J. Biol. Chem. 258: 45–47.PubMedGoogle Scholar
  48. 48.
    Ortiz de Montellano, P. R., and Kunze, K. L., 1981, Cytochrome P-450 inactivation: Structure of the prosthetic heure adduct with propyne, Biochemistry 20: 7266–7271.PubMedCrossRefGoogle Scholar
  49. 49.
    Yagi, H., and Jerina, D. M., 1982, Absolute configuration of the trans-9,10-dihydrodiol metabolite of the carcinogen benzo[a]pyrene, J. Am. Chem. Soc. 104: 4026–4027.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 1986

Authors and Affiliations

  • Gerald T. Miwa
    • 1
  • Anthony Y. H. Lu
    • 1
  1. 1.Department of Animal Drug MetabolismMerck Sharp and Dohme Research LaboratoriesRahwayUSA

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