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KMID : 0043319970200020115
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Archives of Pharmacal Research
1997 Volume.20 No. 2 p.115 ~ p.121
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1269S mutation in horse liver alcohol dehydrogenase S isoenzyme and its reactivity for steroids and retinoids
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Ryu Ji-Won
Lee Kang-Man
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Abstract
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Ile-269 in horse liver alcohol dehydrogenase isoenzyme S(HLADH-S) was mutated to serine by phosphorothioate-based site-directed mutagenesis in order to study the role of the residue in coenzyme binding. The specific activity of the mutant(1269S) enzyme to ethanol was increased 49-fold. All turnover numbers of 1269S enzyme toward 9 primary alcohols were increased. The mutant enzyme showed 3.6, 4.6, 11.6-fold higher catalytic efficiency for -androstane-3, 17-dione, -cholanic acid-3-one and retinal than wild-type, respectively. The reaction mechanism of 1269S enzyme was ordered bi bi as wild-type’s. These results indicate that the hydrophobic interaction of Ile-269 residue with coenzyme plays an important role in dissociation of coenzyme from enzyme-coenzyme complex, which has been known as the rate limiting step of ADH reaction.
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KEYWORD
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1269S, Horse liver alcohol dehydrogenase S isoenzyme, Site-directed mutagenesis, Coenzyme binding site, Steroids, Retinoids
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