Comparison of the 3α- and 20β-hydroxysteroid dehydrogenase activities of the cortisone reductase of Streptomyces hydrogenans

Abstract

The 3alpha- and 20beta-hydroxysteroid dehydrogenase (HSD) activities of cortisone reductase in Streptomyces hydrogenans have been examined to determine whether both activities are due to one enzyme. This question was raised when changes in the commercial preparations of the enzyme reduced the 3alpha-HSD activity to 5% of its original level while retaining full 20beta-HSD activity. In our experiments, the enzyme was purified to crystallinity and partially characterized. The 3alpha- and 20beta-HSD activities were both coinduced and copurified. The 3alpha- and 20beta-HSD activities were compared using the crystalline enzyme for studies of substrate competition, thermal inactivation at 52 degrees C, loss of activity with three haloacetoxysteroids, and the effects of Me2SO and temperature on the reaction rate. These studies support the conclusion that the 3alpha- and 20beta-HSD activities are due to the same enzyme molecule. In addition, it appears that the binding sites for the two activities do not act independently.

Cloning and expression of chicken 20-hydroxysteroid dehydrogenase

“…Similar to other steroid dehydrogenases, ch20HSD displays polyfunctional enzyme activity, 3-and 20HSD activity, but in comparison to some other enzymes, it shows an unusual absence of stereochemical specificity as both 3a/3b-and 20a/20b-epimers were identified. In contrast, 20HSD in pigs operates as 3a/3b,20bHSD (Ohno et al 1991) and in Streptomyces hydrogenans only as 3a,20bHSD (Edwards & Orr 1978), whereas cyprinid fishes coexpress both 20a-and 20bHSD activities although it is not known whether these activities reflect one single or two different enzymes (Thibaut & Porte 2004). However, the results in the present study point to a possibility that ch20HSD has similar widespread distribution as 20bHSD in fishes and neonatal pigs (Kobayashi et al 1996, Guan et al 1999.…”

Prokaryotic 20β‐hydroxysteroid dehydrogenase is an enzyme of the ‘short‐chain, non‐metalloenzyme’ alcohol dehydrogenase type

“…multiple, structurally closely related enzymes to explain differences, should also not be completely excluded, especially not since the enzyme is reported as bifunctional [12,13], and preparations have varied in both activity (cf. [12]) and composition (cf. Table I and Table in [12]).…”

The 20α‐hydroxysteroid dehydrogenase of Streptomyces hydrogenans

“…Mueller-Hinton broth was obtained from Oxoid Ltd (Basingstoke, England); after dilution with water as directed it contained meat infusion (6 g/l), casein hydrolyzate (17.5 g/l) and starch (1.5 g/l) at pH 7. 4. Oatmeal agar and constituents of other media were purchased from Difco Labs (Detroit, Michigan).…”

“…Ammonium sulfate precipitation in the presence of glycerol was performed as previously described [4].…”