Human liver alcohol dehydrogenase: purification and kinetic characterization of the beta 2 beta 2, beta 2 beta 1, alpha beta 2, and beta 2 gamma 1 "Oriental" isoenzymes.
Four alcohol dehydrogenase isoenzymes with "atypical" pH optima for ethanol oxidation at 8.8 were isolated from Japanese livers with the homozygous ADH2 2-2 and the heterozygous ADH2 2-1 phenotypes. Agarose gel isoelectric focusing patterns after dissociation--recombination of three isoenzymes purified from the homozygous livers indicate that they are beta 2 beta 2, alpha beta 2, and beta 2 gamma 1. A fourth isoenzyme, purified from livers with the heterozygous phenotype by agarose-hexane--AMP affinity chromatography, was identified as beta 2 beta 1 by dissociation-recombination studies. The kinetic properties of the three heterodimers, beta 2 beta 1, alpha beta 2, and beta 2 gamma 1, are intermediate between those of the respective homodimers, suggesting that the two subunits act independently. Product inhibition studies indicate that beta 2 beta 2 obeys an ordered sequential mechanism, as do the alpha alpha, beta 1 beta 1, gamma 1 gamma 1, and gamma 2 gamma 2 homodimers which have the "typical" pH optimum for ethanol oxidation at pH 10.0-10.5. The kinetic constants of beta 2 beta 2 differ substantially from those of the other homodimers. At pH 7.5, the Vmax for ethanol oxidation of beta 2 beta 2 is 5-40 times higher than that of alpha alpha, beta 1 beta 1, gamma 1 gamma 1, and gamma 2 gamma 2. The Km and Ki values of beta 2 beta 2 for NAD+ and NADH are also considerably higher than those of the other homodimers.(ABSTRACT TRUNCATED AT 250 WORDS)
D000426:Alcohol Dehydrogenase; D000429:Alcohol Oxidoreductases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D000818:Animals; D002845:Chromatography; D006224:Cricetinae; D000431:Ethanol; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D007564:Japan; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D046911:Macromolecular Substances; D010084:Oxidation-Reduction; D010641:Phenotype
D007527:Isoenzymes; D046911:Macromolecular Substances; D000431:Ethanol; D000429:Alcohol Oxidoreductases; D000426:Alcohol Dehydrogenase