Prostaglandin H synthase catalyzes regiospecific release of tritium from labeled estradiol

• Published in: Steroids Vol. 49; no. 6; pp. 561 - 580
• Main Authors: Degen, Gisela H., Jellinck, Peter H., Hershcopf, Richard J.
• Format: Journal Article
• Language: English
• Published: New York, NY Elsevier Inc 01.06.1987; Elsevier Science
• Subjects: Animals; Binding Sites; Biological and medical sciences; Estradiol - metabolism; Fundamental and applied biological sciences. Psychology; General aspects. Hormone interactions. Hormone actions on several organ systems. Adaptive reactions; In Vitro Techniques; Male; Prostaglandin-Endoperoxide Synthases - metabolism; Seminal Vesicles - enzymology; Sheep; Substrate Specificity; Tritium - metabolism; Vertebrates: endocrinology; Biodegradation; Hydroxylation; Radiolabelling; Seminal vesicle; Enzyme; Arachidonic acid derivatives; Tritium; Metabolism; Male genital system; Estradiol; Prostaglandin synthase; Vertebrata; Mammalia; Metabolic interrelation; Sheep; Artiodactyla; Sex steroid hormone; Ungulata
• ISSN: 0039-128X; 1878-5867
• DOI: 10.1016/0039-128X(87)90096-1

Prostaglandin H synthase (PHS) from ram seminal vesicle microsomes was found to catalyze the release of tritium ( 3H) from estradiol (E 2) regiospecifically labeled in position C-2 or C-4 of ring A but not from positions C-17α, C-16α, or C-6, 7. Formation of 3H 2O from ring A of E 2 is dependent upon native enzyme supplemented with either arachidonic acid, eicosapentaenoic acid, or hydrogen peroxide and proceeds very rapidly as do other cooxidation reactions catalyzed by PHS-peroxidase. The 3H-loss from ring A of E 2 reflecting oxidative displacement of this isotope by PHS increases linearly up to 100 μM under our conditions (8–45 nmol/mg × 5 min). Loss of tritium in various blanks is negligible by comparison. Indomethacin (0.07 and 0.2 mM) inhibited the PHS-dependent release of 3H 2O from estradiol but less efficiently than it inhibited DES-cooxidation measured in parallel incubations under similar conditions. Addition of EDTA (0.5 mM) had no effect on the regiospecific transfer of 3H from E 2 or on DES-oxidation; ascorbic acid (0.5 mM) or NADH (0.33 mM) clearly inhibited both reactions and to a similar extent. These data suggest that estradiol- 2 4 -hydroxylation can be catalyzed by PHS in vitro probably via its peroxidase activity and point to PHS as an enzyme that could contribute to catechol estrogen formation in vitro by tissue preparations in the presence of unsaturated fatty acids or peroxides.