J. Biochem, 1992, Vol. 112, No. 6 840-844
© 1992 Japanese Biochemical Society
research-article |
Inhibition of Dimeric Dihydrodiol Dehydrogenase by 4-Hydroxyphenylketone Derivatives: Aspects of Inhibitor Structure and Binding Specificity1
*Gihoku General Hospital, Takatomi, Gifu 501-21
**Laboratory of Biochemistry, Department of Manufacturing Pharmacy, Gifu Pharmaceutical University, Mitahora-higashi, Gifu, Gifu 502
2To whom correspondence should b addressed
Dimeric dihydrodiol dehydrogenases from pig liver, monkey kidney, and rabbit lens were inhibited more potently by 4-hydroxyphenylketones such as 4-hydroxybenzaldehyde, 4-hydroxyphenylglyoxal, and 4-hydroxyacetophenone than by isoascorbate and ascorbate, known inhibitors of the enzymes. No significant inhibition was observed with 2- or 3-hydroxyphenylketones, phenylketones with a functional group other than a hydroxy group at the 4-position, and 4-hydroxyphenyl derivatives without a carbonyl group. The steady-state kinetic analyses of the inhibition of the pig liver enzyme indicated that the 4-hydroxyphenylketones, similarly to ascorbate and its epimer, bound to an enzyme-NADP+ binary complex as competitive inhibitors with respect to dihydrodiol substrate. The inhibition by the 4-hydroxyphenylketones was uncomoetiti ve with respect to isoascorbate, and the addition of one of the 4-hydroxyphenylketones or isoascorbate with NADP+ afforded a great protective effect against inactivation of the enzyme by diethylpyrocar-bonate or by heat treatment, which indicates that 4-hydroxyphenylketones and isoascorbate bind at the same site in or near the active center of the enzyme. The structural comparison of 4-hydroxybenzaldehyde and ascorbate suggests that the hydroxy group at C-5, carbonyl group at C-1 and lactone ring of ascorbate are important for the binding to the enzyme.
1This study was supported in part by a grant for Cooperative Research from the Primate Research Institute, Kyoto University.