15-Ketoprostaglandin delta 13-reductase activity in bovine ocular tissues.

Abstract

The present study provides the first evidence for delta 13-reduction of 15-ketoprostaglandins (15-keto-PGs) by bovine ocular tissues. The 9,000xg supernatants of cornea, iris, ciliary body, retina, and RPE-choroid except lens exhibited delta 13-reductase activity toward 15-keto-PG E2 and F2 alpha in the presence of NADPH or NADH as an electron donor. Among the tissues tested, the highest activity was observed in ciliary body and iris, followed by RPE-choroid, retina, and cornea. The NADPH- and NADH-linked double bond reductase activities were inhibited by dicumarol, quercitrin, indomethacin and disulfiram, but not by potassium cyanide. NADPH-linked 15-keto-PG F2 alpha delta 13-reductase was purified from bovine iris-ciliary body cytosol by fractionation with ammonium sulfate and high performance liquid chromatography (HPLC) with TSK gel DEAE-5PW, and TSK gel Blue-5PW. The purified enzyme was homogenous by the criterion of sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Its molecular weight was estimated to be about 57,000 by electrophoresis, and about 55,000 by gel filtration HPLC with Superose 12.