The Purification and Properties of NADPH-Dependent Carbonyl Reductases from Rat Ovary

Abstract

Two carbonyl reductases have been highly purified from rat ovary to apparent homogeneity. Though they have similarities in terms of molecular weight (33,000), substrate specificities, inhibitor sensitivities, amino acid composition, and immunological properties, they differed in pI values (6.0 and 5.9). Both enzymes reduced aromatic aldehydes, ketones, and quinones at higher rates, compared to prostaglandins and 3-ketosteroids, whereas they showed higher affinity for prostaglandins and 3-ketosteroids. The enzymes also catalyzed oxidation of the 9-hydroxy group of prostaglandin F2 alpha. Moreover, they showed the remarkable characteristic of catalyzing the reduction of not only the 9-keto group of prostaglandin E2 but also the 15-keto group of 13,14-dihydro-15-keto-prostaglandin F2 alpha. Both enzymes were inhibited by SH-reagents, quercitrin, indomethacin, furosemide, and disulfiram. The results of immunoinhibition, using antibody against the purified enzymes, indicated that the enzymes were solely responsible for the overall catalytic activities of prostaglandin E series reduction, as well as 13,14-dihydro-15-keto-prostaglandin F2 alpha reduction and prostaglandin F2 alpha oxidation in rat ovarian cytosol. Western-blot analysis revealed that immunoreactive proteins were present in adrenal gland and various reproductive tissues except uterus of rats.

Carbonyl reductases from rat testis and vas deferens

“…Multiple forms of the enzyme differing in size and charge have been isolated from various sources and even from the same tissue [l -31. These carbonyl reductases have been classified into two groups according to substrate specificity. The enzymes in the first group have higher affinity for steroids and have been isolated mostly from hepatic tissues [4 -71, while the enzymes in the second group display higher affinity for prostaglandins (PGs) and have been purified from extrahepatic tissues, such as brain [8], lung [9], kidney [lo] and ovary [l 11. These enzymes are considered to play an important role in the metabolism of both endogenous and xenobiotic carbonyl compounds.…”

“…Recently, we reported that two enzyme forms (CR1 and CR2) gf carbonyl reductase from rat ovary can catalyze the interconversion of prostaglandin Ez (PGEJ to prostaglandin F2, (PGF2a) as well as the reduction of 13,14-dihydro-l~-oxo-PGF2, [11]. CR1 and CR2 are charge isomers with similar molecular mass, substrate specificity, inhibitor susceptibility and immunochemical cross-reactivity.…”

“…Sweden). 13,14-Dihydro-I 5-0x0-PGF,, was obtained from Upjohn Pharmaceuticals Ltd (Kalamazoo, MI, USA) and [5,6,8,9,11 The reaction was initiated by addition of cofactor to the assay mixture. Blanks without substrate or enzyme were routinely included.…”

Effects of Antiestrogens on Ovarian Aldo-Keto Reductase in Relation to Ovulation in Rats.

“…1 and 2. The reduc tion of 15KD-PGF2a, , which is not only a spe cific substrate for ovarian CR1 and CR2 but also an endogenous substrate in rat ovary (13), was significantly decreased to 39.2, 49.2, 62.9, 38.8 and 39.8% of the control levels by tamoxifen, clomiphene, nafoxidine, U-23469 and MPA, respectively (Fig. 1).…”

“…The car bonyl reductase (CR1, CR2) catalyzing con version of 15KD-PGF2a to 13,14H2-PGF2a in rat ovary is a member of the aldo-keto reduct ase family (13) and is localized in the ovarian cytosolic fraction (14). The activities of the ovarian CRs were significantly increased after the endogenous LH surge during the estrous cycle of rats (15) and were markedly stimu lated by co-administration of estradiol and hCG in immature rats (16).…”