Phosphorylation of purified mitochondrial cytochromes P-450 (cholesterol desmolase and 11 beta-hydroxylase) from bovine adrenal cortex.
Two key steroidogenic mitochondrial cytochromes P-450 (cholesterol side-chain cleavage (scc) and 11 beta-hydroxylation (11 beta)) were purified from bovine adrenal cortex and examined as potential phosphorylatable substrates using purified cAMP-dependent protein kinase subunit (C) and A type (CKA) and G type (CKG) cAMP-independent casein kinases. Of the two cytochromes P-450, only P-450 11 beta was able to incorporate phosphate from ATP in the presence of C (Km = 7.5 microM), whereas CKA and CKG were ineffective. Phosphorylation of P-450 11 beta (maximum incorporation of 1 mole of 32P per mole of cytochrome, only on serine residues) did not modify the enzymatic activity of an 11 beta-hydroxylation system reconstituted in vitro from purified components, when adrenodoxin was in excess in the reaction. However, kinetic studies showed that P-450 11 beta phosphorylation strikingly increases the P-450 11 beta-adrenodoxin affinity in a phosphorylation-dependent manner. This would result in a net increase in 11 beta-hydroxylase activity under in vivo conditions where adrenodoxin availability is limited. Possible significance of these observations in the regulation of differentiated adrenocortical functions remains to be further examined.
D000302:Adrenal Cortex / Q000201:enzymology*; D000324:Adrenocorticotropic Hormone / Q000494:pharmacology; D000818:Animals; D002417:Cattle; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D066298:In Vitro Techniques; D007700:Kinetics; D008928:Mitochondria / Q000201:enzymology; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D013252:Steroid 11-beta-Hydroxylase / Q000378:metabolism
D000324:Adrenocorticotropic Hormone; D003577:Cytochrome P-450 Enzyme System; D013252:Steroid 11-beta-Hydroxylase; D002786:Cholesterol Side-Chain Cleavage Enzyme; D011494:Protein Kinases