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KMID : 0617319940040010078
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Journal of Pharmacetical Sceiences Ewha Womans University
1994 Volume.4 No. 1 p.78 ~ p.83
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Substitution of Gly-224 Residue to Ile in Yeast Alcohol Dehydrogenase and Enzyme Reaction Mechanism
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Lee Kang-Man
Ryu Ji-Won
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Abstract
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Gly-224 residue of yeast alcohol dehydrogenase was mutated by site-directed mutagenesis to isoleucine, which is the corresponding amino acid residue of horse liver alcohol dehydrogenase. The mutated gene on M13 vector was subcloned in YEp13 and used to transform Saccharomyces cerevisiae 302-21#2 strain, and the expressed protein was purified. The turnover numbers of mutant enzyme for ethanol and acetaldehyde were decreased compared to wild-type enzyme. The results of product inhibition studies indicated that the reaction mechanism was changed to Iso Theorell-Chance from Ordered Bi Bi. We supposed that Gly-224 was related to the enzyme reaction mechanism.
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