Human liver alcohol dehydrogenase: purification, composition, and catalytic features.
Alcohol dehydrogenase has been purified from human liver by affinity chromatography. Ultracentrifugation, Sephadex G-200 chromatography, and amino acid analyses of multiple preparations demonstrate homogeneity of molecular weight. Sodium dodecyl sulfate disc gel electrophoresis reveals a single species of molecular weight 42 000. Based on a molecular weight of 85 000 for the dimer obtained from the amino acid composition and a molar absorptivity of A280nm0.1% = 0.58, the enzyme contains 3.6-4.2 g-atoms of zinc, as determined by emission spectrography, microwave-induced emission, and atomic absorption spectrometry. Inhibition by o-phenanthroline, (ethylenedinitrilo)tetraacetic acid, and alpha,alpha'-bipyridine demonstrates that zinc is essential to enzymatic function. Detailed kinetic analyses using primary alcohols of the homologous series CH3(CH2)nOH, n = 0-5, and the corresponding aldehydes as substrates show that KM values become smaller as n increases. This suggest that hydrophobic interactions play a role in substrate binding. The availability of well-defined preparations of human liver alcohol dehydrogenase now allows definitive genetic and functional studies of this enzyme to elucidate human ethanol metabolism.
D000429:Alcohol Oxidoreductases / Q000302:isolation & purification / Q000378:metabolism; D000596:Amino Acids / Q000032:analysis; D002614:Chelating Agents / Q000494:pharmacology; D002846:Chromatography, Affinity; D002942:Circular Dichroism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007501:Iron / Q000032:analysis; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008274:Magnesium / Q000032:analysis; D010618:Phenanthrolines / Q000494:pharmacology; D011487:Protein Conformation; D015032:Zinc / Q000032:analysis
D000596:Amino Acids; D002614:Chelating Agents; D010618:Phenanthrolines; D007501:Iron; D000429:Alcohol Oxidoreductases; D008274:Magnesium; D015032:Zinc