Catalytic activity of cytochrome P-450 isozyme 3a isolated from liver microsomes of ethanol-treated rabbits. Oxidation of alcohols

E. T. Morgan, Dennis Koop, M. J. Coon

Research output: Contribution to journalArticle

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Abstract

Cytochrome P-450 isozyme 3a, isolated from hepatic microsomes of rabbits treated chronically with ethanol, was found to have a unique substrate specificity when compared with isozymes 2, 3b, 3c, and 4. Form 3a has unusually high activity in the p-hydroxylation of aniline and in the oxidation of alcohols to aldehydes. These properties are reflected in the increased activities of these substrates in microsomes from ethanol-treated rabbits as compared to microsomes from untreated animals or those administered phenobarbital or 5,6-benzoflavone. The ethanol-oxidizing activity of isozyme 3a, which requires the presence of NADPH and NADPH cytochrome P-450 reductase and is stimulated by the presence of phospholipid, was shown not to be due to contaminating catalase or an NAD- or NADP-dependent alcohol dehydrogenase. Isozyme 3a catalyzes the oxidation of methanol, 1-propanol, and 1-butanol as well as ethanol; the relationships between the apparent K(m) values for these alcohols and their octanol-water partition coefficients is in accord with the known hydrophobic nature of the P-450 binding site. Whereas typical substrates of isozyme 2 are known to be metabolized with the stoichiometry predicted of a monooxygenase reaction, with isozyme 3a the sum of acetaldehyde formed from ethanol and of hydrogen peroxide generated is inadequate to account for the NADPH and oxygen consumed. Free hydroxyl radicals appear to mediate the slow oxidation of ethanol in the presence of the reductase alone but not the faster rate catalyzed by P-450 isozyme 3a. The results obtained, however, do not rule out the involvement of hydroxyl radical equivalent generated and bound at the active site of the cytochrome.

Original languageEnglish (US)
Pages (from-to)13951-13957
Number of pages7
JournalJournal of Biological Chemistry
Volume257
Issue number23
StatePublished - 1982
Externally publishedYes

Fingerprint

Liver Microsomes
Liver
Cytochrome P-450 Enzyme System
Isoenzymes
Catalyst activity
Ethanol
Alcohols
Rabbits
Oxidation
Microsomes
NADP
Hydroxyl Radical
Substrates
beta-Naphthoflavone
Octanols
1-Propanol
NADPH-Ferrihemoprotein Reductase
Hydroxylation
1-Butanol
Acetaldehyde

ASJC Scopus subject areas

  • Biochemistry

Cite this

Catalytic activity of cytochrome P-450 isozyme 3a isolated from liver microsomes of ethanol-treated rabbits. Oxidation of alcohols. / Morgan, E. T.; Koop, Dennis; Coon, M. J.

In: Journal of Biological Chemistry, Vol. 257, No. 23, 1982, p. 13951-13957.

Research output: Contribution to journalArticle

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N2 - Cytochrome P-450 isozyme 3a, isolated from hepatic microsomes of rabbits treated chronically with ethanol, was found to have a unique substrate specificity when compared with isozymes 2, 3b, 3c, and 4. Form 3a has unusually high activity in the p-hydroxylation of aniline and in the oxidation of alcohols to aldehydes. These properties are reflected in the increased activities of these substrates in microsomes from ethanol-treated rabbits as compared to microsomes from untreated animals or those administered phenobarbital or 5,6-benzoflavone. The ethanol-oxidizing activity of isozyme 3a, which requires the presence of NADPH and NADPH cytochrome P-450 reductase and is stimulated by the presence of phospholipid, was shown not to be due to contaminating catalase or an NAD- or NADP-dependent alcohol dehydrogenase. Isozyme 3a catalyzes the oxidation of methanol, 1-propanol, and 1-butanol as well as ethanol; the relationships between the apparent K(m) values for these alcohols and their octanol-water partition coefficients is in accord with the known hydrophobic nature of the P-450 binding site. Whereas typical substrates of isozyme 2 are known to be metabolized with the stoichiometry predicted of a monooxygenase reaction, with isozyme 3a the sum of acetaldehyde formed from ethanol and of hydrogen peroxide generated is inadequate to account for the NADPH and oxygen consumed. Free hydroxyl radicals appear to mediate the slow oxidation of ethanol in the presence of the reductase alone but not the faster rate catalyzed by P-450 isozyme 3a. The results obtained, however, do not rule out the involvement of hydroxyl radical equivalent generated and bound at the active site of the cytochrome.

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