title: Kinetic and mechanistic studies of oxidation of vitamin A alcohol to vitamin A aldehyde by horse liver alcohol dehydrogenase. The inhibition by ethanol and pyrazole.
abstract: The present investigation shows that all-trans retinol (Vitamin A alcohol), an alcohol of great physiological importance, is efficiently oxidized to all-trans retinaldehyde by the enzyme horse liver alcohol dehydrogenase. We observe a Km retinol value of 145 microM and a turnover number of 0.45s-1 for the oxidation of all-trans retinol in the presence of Triton X-100, a surfactant used as a solubilizer. Over the concentration range of surfactant used (up to 0.1% Triton X-100) our studies on the oxidation of ethanol and all-trans retinol show that turnover numbers for both reactions remain constant as does the value for Km ethanol. On increasing the concentration of Triton X-100 from 0.025% to 0.10%, however, the Km retinol value increases by a factor of two. This behavior for retinol oxidation can be attributed to the partitioning of retinol between enzyme and surfactant. Pyrazole, a known inhibitor of alcohol oxidation by horse liver alcohol dehydrogenase, is a competitive inhibitor of both all-trans retinol and ethanol, with observed Ki values of 3.3 X 10(-7) M and 3.9 x 10(-7) M, respectively. We also find that ethanol inhibits all-trans retinol oxidation in a complex fashion, an observation which may have important consequences in view of the physiological role of retinol and its oxidation products. Our present studies indicate that all-trans retinol binds in the same region of the enzyme as does ethanol and is oxidized with an efficiency approaching that of ethanol itself.
mesh_terms: D000429:Alcohol Oxidoreductases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D000431:Ethanol / Q000494:pharmacology; D006736:Horses; D066298:In Vitro Techniques; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D010084:Oxidation-Reduction; D011720:Pyrazoles / Q000494:pharmacology; D012172:Retinaldehyde / Q000378:metabolism*; D014801:Vitamin A / Q000031:analogs & derivatives* / Q000378:metabolism*
chemical_list: D011720:Pyrazoles; D014801:Vitamin A; D000431:Ethanol; D000429:Alcohol Oxidoreductases; D012172:Retinaldehyde