Characterization of a major form of human isatin reductase and the reduced metabolite.
Isatin, an endogenous indole, has been shown to inhibit monoamine oxidase, and exhibit various pharmacological actions. However, the metabolism of isatin in humans remains unknown. We have found high isatin reductase activity in the 105,000 g supernatants of human liver and kidney homogenates, and have purified and characterized a major form of the enzyme in the two tissues. The hepatic and renal enzymes showed the same properties, including an M(r) of 31 kDa, substrate specificity for carbonyl compounds and inhibitor sensitivity, which were also identical to those of recombinant human carbonyl reductase. The identity of the isatin reductase with carbonyl reductase was immunologically demonstrated with an antibody against the recombinant carbonyl reductase. About 90% of the soluble isatin reductase activity in the liver and kidney was immunoprecipitated by the antibody. The Km (10 microm) and k(cat)/K(m) (1.7 s(-1) x microm(-1)) values for isatin at pH 7.0 were comparable to those for phenanthrenequinone, the best xenobiotic substrate of carbonyl reductase. The reduced product of isatin was chemically identified with 3-hydroxy-2-oxoindole, which is also excreted in human urine. The inhibitory potency of the reduced product for monoamine oxidase A and B was significantly lower than that of isatin. The results indicate that the novel metabolic pathway of isatin in humans is mediated mainly by carbonyl reductase, which may play a critical role in controlling the biological activity of isatin.
Usami
N
N
Biochemistry Laboratory, Gifu Pharmaceutical University, Gifu, Japan.
Kitahara
K
K
Ishikura
S
S
Nagano
M
M
Sakai
S
S
Hara
A
A
D000429:Alcohol Oxidoreductases / Q000037:antagonists & inhibitors / Q000652:urine; D000449:Aldehyde Reductase; D000074408:Aldo-Keto Reductases; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007211:Indoles / Q000494:pharmacology; D007510:Isatin / Q000378:metabolism*; D013379:Substrate Specificity
D004791:Enzyme Inhibitors; D007211:Indoles; C442334:3-hydroxy-2-oxoindole; D007510:Isatin; D000429:Alcohol Oxidoreductases; D000074408:Aldo-Keto Reductases; D000449:Aldehyde Reductase