Topological studies of cytochromes P-450scc and P-45011 beta in bovine adrenocortical inner mitochondrial membranes. Effects of controlled tryptic digestion

• Published in: The Journal of biological chemistry Vol. 254; no. 20; pp. 10443 - 10448
• Main Authors: Churchill, P F, Kimura, T
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 25.10.1979
• Subjects: Adrenal Cortex - ultrastructure; Animals; Cattle; Cytochrome P-450 Enzyme System - analysis; Dithionite; Hydroxycholesterols - pharmacology; Intracellular Membranes - drug effects; Intracellular Membranes - ultrastructure; Mitochondria - drug effects; Mitochondria - ultrastructure; Pregnenolone - pharmacology; Spectrophotometry; Trypsin
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/s0021-9258(19)86728-5

The topology of the steroid hydroxylase complexes in bovine adrenocortical mitochondria were studied by using controlled digestion with trypsin of purified inner mitochondrial membranes. Inhibition of steroid hydroxylase activity by trypsin was only observed in inner mitochondrial membranes which had been disrupted by various techniques. The steroid hydroxylase activity of intact inner membranes was not inhibited by trypsin. The effect of tryptic digestion was monitored by measuring 11 beta-hydroxylase and cholesterol side chain cleavage activities, as well as cytochrome P-450 reduction. The effect of trypsin on the steroid-induced difference spectra using pregnenolone, 20 alpha-hydroxycholesterol, and deoxycorticosterone was also measured. The results were similar regardless of which procedure was utilized and strongly suggest that both cytochrome P-45011 beta and cytochrome P-450scc are located on the matrix side of the mitochondrial inner membrane.