Purification and regulatory properties of chicken heart prostaglandin E 9-ketoreductase.
Prostaglandin E 9-ketoreductase was purified from chicken heart by ammonium sulfate fractionation, and DEAE-Sephadex, hydroxylapatite and phosphocellulose chromatography. Two peaks of activity were resolved during the phosphocellulose chromatographic step. Both peaks were stimulated by a substance that was not bound to the phosphocellulose column. This stimulatory substance was destroyed by treatment with phosphodiesterase and 0.1 M NaOH. It was heat-stable (100 degrees, 2 min), nondialyzable, and resistant to treatment with pronase, ribonuclease, and deoxyribonuclease; but it was dialyzable after heating or digestion with pronase. Sodium pyrophosphate also enhanced the activities of the prostaglandin E 9-ketoreductases as did angiotensin I; but not angiotensin II. In the presence of 3':5'-cyclic AMP, AMP, or several other ribonucleotides, the enhancing effects of the natural stimulatory substance, sodium pyrophosphate or angiotensin I were blocked, but these ribonucleotides themselves had little effect on the enzymes activity. The substrate specificities of the two prostaglandin E 9-ketoreductases were also studied. Both the 9-keto group and the 15-keto group of 15-ketoprostaglandin F2 alpha could be converted to the corresponding hydroxyl group; the 15-keto group was reduced faster than the 9-keto group. Prostaglandin D2, a prostaglandin with a 9-hydroxyl and an 11-keto group, could not be converted to prostaglandin F2 alpha nor could cyclohexanone be converted to cyclohexanol by the prostaglandin E 9-ketoreductase.
D000429:Alcohol Oxidoreductases / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D000818:Animals; D002645:Chickens; D002845:Chromatography; D002852:Chromatography, Ion Exchange; D003851:Deoxyribonucleases; D004355:Drug Stability; D006041:Goats / Q000276:immunology; D006358:Hot Temperature; D006882:Hydroxyapatites; D007700:Kinetics; D009206:Myocardium / Q000201:enzymology*; D010756:Phosphoric Acids / Q000494:pharmacology; D010727:Phosphoric Diester Hydrolases; D011402:Pronase; D011458:Prostaglandins E / Q000378:metabolism*; D011863:Radioimmunoassay; D012260:Ribonucleases; D013997:Time Factors
D006882:Hydroxyapatites; D010756:Phosphoric Acids; D011458:Prostaglandins E; D000429:Alcohol Oxidoreductases; D003851:Deoxyribonucleases; D012260:Ribonucleases; D010727:Phosphoric Diester Hydrolases; D011402:Pronase