Reconstitution of the steroidogenic pathway from cholesterol to aldosterone in liposome membranes

A steroidogenic pathway from cholesterol to aldosterone was reconstituted in liposome membranes using cytochromes P-450scc, P-450C21 and P-450(11) beta, and 3 beta-hydroxysteroid dehydrogenase/ delta 5-delta 4 isomerase (3 beta HSD/I) with their electron transfer systems. All of the enzymes were purified from bovine adrenocortical mitochondria and microsomes. The cholesterol metabolism in the liposomal reconstituted system was compared with that in the combined organella system composed of bovine adrenocortical mitochondria and microsomes, where the activity of P-450(17) alpha,lyase was inhibited by bifonazole. The metabolic activities in these two systems were similar except for aldosterone production. Aldosterone was produced in the liposomal system but not in the combined organella system. 4-fold increase in the amount of P-450scc in the liposomal system enhanced the activity of 3 beta HSD/I, P-450C21 and 11 beta-hydroxylase of P-450(11) beta but decreased 18-hydroxycorticosterone and aldosterone production by P-450(11) beta, supporting our previous findings describing the regulation mechanism of aldosterone synthesis (Kominami, S., Harada, D. and Takemori, S. (1994) Biochim. Biophys. Acta 1192, 234). It was demonstrated using the liposomal reconstituted system that the increase in the amount of one enzyme did not only increase the metabolizing activity of that enzyme but also affect other enzyme in various ways.