A. Finsterbusch, P. Lindemann, R. Grimm
Oct 1, 1999
Citations
3
Influential Citations
59
Citations
Quality indicators
Journal
Planta
Abstract
Abstract.Δ5-3β-Ηydroxysteroid dehydrogenase (Δ5-3β-HSD; EC 1.1.1.145), an enzyme converting pregn-5-ene-3β-ol-20-one (pregnenolone) to pregn-5-ene-3,20-dione (isoprogesterone), was isolated from the soluble fraction of suspension-cultured cells of Digitalis lanata L. strain VIII. Starting with acetone dry powder the enzyme was purified in three steps using column chromatography on Fractogel-TSK DEAE, hydroxyapatite and Sephacryl G-200. Fractions with highest Δ5-3β-HSD activity were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. After in-situ digestion the resulting bands were sequenced N-terminally. The 29-kDa band yielded three fragments with high sequence homology to members of the superfamily of short-chain dehydrogenases/reductases. High similarity was found to microbial hydroxysteroid dehydrogenases. The band may therefore represent the Δ5-3β-HSD. The purified enzyme was characterized with respect to kinetic parameters, substrate specificity and localization. The function of the enzyme in steroid metabolism is discussed.