Maser E, Gebel T, Netter K J
Department of Pharmacology and Toxicology, School of Medicine, University of Marburg, Lahn, Federal Republic of Germany.
Biochem Pharmacol. 1991 Dec 11;42 Suppl:S93-8. doi: 10.1016/0006-2952(91)90397-n.
Carbonyl reduction was investigated in cytosolic and microsomal fractions of human liver using the ketone metyrapone as a substrate. The cytosolic enzyme has a stronger preference for NADPH over NADH than the microsomal enzyme: the former shows only 14% of the NADPH-supported activity while the latter exhibits 36% activity with NADH. Barbitone and quercitrin, the classic inhibitors of carbonyl reductases, do not affect metyrapone reduction in either fraction. Dicumarol and indomethacin, the specific inhibitors of NAD(P)H: quinone-oxidoreductase and dihydrodiol dehydrogenase, respectively, only slightly decreased metyrapol formation. In contrast, 5 alpha-dihydrotestosterone, the active form of the androgen steroid testosterone, inhibited metyrapone reduction very strongly in the microsomal fractions and is postulated to be the physiological substrate of the enzyme. This resembles the situation in mouse liver [E. Maser and K. J. Netter, Biochem Pharmacol 38: 3049-3054, 1989] where microsomal metyrapone reductase was inhibited by steroids and the purified enzyme was demonstrated to mediate androsterone oxidation. Immunoblot analysis revealed antigenic cross-reaction of antibodies against the 34 kDa metyrapone reductase from mouse liver microsomes with the homologous protein in human liver microsomes pointing to structural homologies between the respective enzymes of the two species. These results--together with previous findings, which have shown that there exist functional as well as structural relationships between microsomal mouse liver metyrapone reductase and 3 alpha-hydroxysteroid dehydrogenase from Pseudomonas testosteroni [E. Maser, U. Oppermann and K. J. Netter, Eur J Pharmacol 183:1366, 1990]--suggest that metyrapone reduction in human liver microsomes might be catalysed by a microsomal hydroxysteroid dehydrogenase.
以酮替米哌酮为底物，研究了人肝脏胞质和微粒体部分中的羰基还原反应。与微粒体酶相比，胞质酶对NADPH的偏好性强于NADH：前者在NADPH支持下的活性仅为14%，而后者在NADH存在时的活性为36%。羰基还原酶的经典抑制剂巴比妥和槲皮苷对两部分中的替米哌酮还原反应均无影响。NAD(P)H：醌氧化还原酶和二氢二醇脱氢酶的特异性抑制剂双香豆素和吲哚美辛仅略微降低了替米哌醇的生成。相反，雄激素类固醇睾酮的活性形式5α-二氢睾酮在微粒体部分中强烈抑制替米哌酮还原反应，据推测它是该酶的生理底物。这与小鼠肝脏中的情况相似[E. 马泽尔和K. J. 内特尔，《生物化学与药物学》38: 3049 - 3054, 1989]，在小鼠肝脏中，微粒体替米哌酮还原酶被类固醇抑制，且纯化后的酶被证明可介导雄甾酮氧化。免疫印迹分析显示，抗小鼠肝脏微粒体中34 kDa替米哌酮还原酶的抗体与人肝脏微粒体中的同源蛋白发生抗原交叉反应，表明这两个物种各自的酶之间存在结构同源性。这些结果——连同先前的研究发现，即已表明小鼠肝脏微粒体替米哌酮还原酶与睾丸酮假单胞菌的3α-羟基类固醇脱氢酶之间存在功能及结构关系[E. 马泽尔、U. 奥佩曼和K. J. 内特尔，《欧洲药理学杂志》183:1366, 1990]——表明人肝脏微粒体中的替米哌酮还原反应可能由微粒体羟基类固醇脱氢酶催化。
