Steroid transforming enzymes from microorganisms—III: Properties of 4-ene-3-oxosteroid-5α-reductase from Mycobacterium smegmatis

Abstract

Sonicated cell-free extract of M. smegmatis is capable of reducing 4-ene-3-oxosteroids to the corresponding 5α-dihydro-products. In the presence of sodium dithionite and with 4-androstenedione as substrate 5α-reduetion takes place as the predominant transformation reaction. The enzyme remains in the 105,000 g supernatant and can be precipitated by ammonium sulfate at 30–45% saturation. The enzyme can be stored at — 20° for at least 3 months without inactivation. At room temperature it loses about 50% activity in 48 h. In phosphate buffer the pH optimum is at 6.3 and the temperature optimum at 40°. Molecular weight is about 40,000. Thiol reagents, heavy metal ions and o-phenanthroline are strong inhibitors of the 5α-reductase, The inhibition by acriflavin and the increasing effect of sodium dithionite indicates the participation of flavin-nucleotides. Progesterone, testosterone and derivatives with additional double bonds and substituents in positions 4 and 17α, as well as both d- and l-19-nortestosterone are transformed to 5α-dihydrocompounds. The apparent k m for androstenedione is 7.5 × 10 −6 M. Cyproterone and Chlormadinonacetate are powerful steroid inhibitors of the enzyme.