Glutathione-prostaglandin A1 conjugate as substrate in the purification of prostaglandin 9-ketoreductase from rabbit kidney

Using GSH-PGA₁ as substrate for determination of enzyme activity a pI 4.8 form of rabbit kidney prostaglandin 9-keto-reductase has been purified 95 times to a specific activity of 1755 nmol/min per mg protein. The purification procedures involve ion-exchange chromatography, gel-filtration and affinity chromatography. The latter procedure comprises Blue Sepharose affinity chromatography, and GSH-PGA₁-Sepharose affinity chromatography.The purified enzyme preparation also showed a weak NADP⁺-dependent 15-hydroxyprostaglandin dehydrogenase activity, 20 nmol/min per mg protein with PGE₁ as substrate. K_m(PGE₁) for the dehydrogenase is 142.6 ± 45.1 μM (S.E., n=7).