Isolation and characterization of 15-hydroxyprostaglandin dehydrogenase from human placenta
A method for the purification of a homogeneous 15-hydroxyprostaglandin dehydrogenase from human placenta has been developed which allows 7,700-fold purification with 26% yield, and gives the final specific activity of 15,800 nmol/min/mg (PGE2 as substrate, pH 7.4, 37°C). The active enzyme was found to be a dimer by comparing its molecular weight (about 50,000) with that determined for the subunit (about 28,000), using SDS-polyacrylamide gel electrophoresis. The amino acid composition of the enzyme was calculated for subunit of 28,000. The enzyme was similar to those of the two 'short' dehydrogenases, Drosophila melanogaster alcohol dehydrogenase and Klebsiella aerogenes ribitol dehydrogenase, but not the 'long' alcohol dehydrogenase of horse liver. Inactivation by some modification reagents indicated that histidine, cysteine, lysine and arginine may be functionally or structurally important for the enzyme activity. The anti-inflammatory and analgesic drugs, aspirin and indomethacin, and p-aminosalicylic acid, an antitubercular drug, were found to be inhibitors of the enzyme. Some diuretic drugs, bendrofluazine, bumetanide, frusemide and triamterene were found to cause inhibition of the enzyme. Two anti-depressant drugs, imipramine and amitriptyline, were found to activate the enzyme, and the possible presence of a regulatory site is discussed. Heptan-4-ol, a small organic molecule which is structurally different from prostaglandins, was shown to be a poor substrate of the enzyme, and the first substrate discovered that is not a prostaglandin or closely related compound.