Subcellular Localization and Properties of Mouse Adrenal C , , - Steroid 5 P - Reductase

The localization and some characteristics of mouse adrenal C19-steroid 5fi-reductase were determined by the incubation of subcellular fractions of mouse adrenal tissue with [7a-3H]androst-4-ene-3,17-dione. This enzyme was present only in the soluble fraction and was NADPH-dependent, although a small activity in the presence ofNADH was also detected. The soluble fraction also contained 3 a-, 3,6and a small amount of 17f,-hydroxy steroid dehydrogenase. These and other steroid-metabolizing enzymes present in the remaining subcellular fractions are also described briefly. To measure 5fi-androstane-3,17-dione production by the mouse adrenal soluble fraction, all 5B6 products first had to be oxidized to 5,B-androstane-3,17-dione, and the recovery of radioactivity between the substrate androst-4-ene-3,17-dione and product 5,6-androstane3,17-dione of 96.1 ± 3.2% validated this technique. C,9-steroid 5,6-reductase has a pH optimum of 6.5 and at low substrate concentrations the Km and Vm.x. for 5,6 reduction of [7a-3H]androst-4-ene-3,17-dione was 2.22 x 10-6 + 0.48 x 10-6 M and 450± 53 pmol/min per mg of protein respectively. At high substrate concentration, inhibition of the reaction occurred, which was shown to be due to increasing product concentration.