Effect of membrane lipid environment on the activity of bovine adrenal 3-oxo- Δ5-steroid isomerase

Abstract

The 3-oxo- Δ 5-steroid isomerase (EC 5.3.3.1) activity from bovine adrenal cortex microsomes can be extracted in soluble form by the use of appropriate detergents, although recovery of enzyme activity is low ( ca. 2%). Activity is restored upon removal of detergent and reconstitution of the enzyme into phospholipid vesicles. Both K m and V max of 3-oxo- Δ 5-steroid isomerase of intact microsomes increase as the pH is raised from 7.5 to 9.5, with a particularly sharp increase (6- to 8-fold) above pH 8.5. The kinetic parameters of a detergent-solubilized isomerase preparation show little increase from pH 7.5 to 9.0, but isomerase reconstituted into artificial phospholipid vesicles demonstrates a 6- to 10-fold increase in both K m and V max over this pH range. Addition of Ca ++ (1 mM) enhances the pH dependence of both K m and V max of the membrane-bound isomerase, causing a slight rise in V max K m .