The catalytic cycle of cytochrome P-450scc and intermediates in the conversion of cholesterol to pregnenolone.
Cytochrome P-450scc as isolated is a cholesterol-depleted low-spin haemoprotein; addition of cholesterol results in formation of a high-spin complex. Cytochrome P-450scc--cholesterol is a one-electron acceptor on titration with NADPH. Cytochrome P-450scc--cholesterol can be anaerobically reduced to the ferrous state which, on oxygenation, forms an oxygenated cytochrome P-450scc--cholesterol complex. This oxygenated complex in the absence of adrenodoxin autoxidises to ferric cytochrome P-450scc--cholesterol without oxidation of cholesterol. The decay of the oxygenated complex is first-order, k = 9.3 X 10(-3) S-1 at 4 degrees C. The rate of autoxidation is influenced by pH, ionic strength and the chemical nature of bound sterol. The activation energy of autoxidation is 75 kJ mol-1. Addition of equimolar amounts of adrenodoxin to cytochrome P-450scc--cholesterol followed by stoichiometric reduction under anaerobic conditions and subsequent oxygenation, allows single catalytic turnover cycles of cytochrome P-450scc to be observed. This has led to detection of intermediates in the conversion of cholesterol to pregnenolone and a precursor/product sequence of cholesterol----22-hydroxycholesterol----20,22-dihydroxy-cholesterol ----pregnenolone has been established. Addition of oxidised adrenodoxin to oxygenated cytochrome P-450scc--cholesterol results in formation of 22-hydroxycholesterol.
D000302:Adrenal Cortex / Q000201:enzymology; D000818:Animals; D002384:Catalysis; D002417:Cattle; D002784:Cholesterol / Q000378:metabolism*; D002849:Chromatography, Gas; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D066298:In Vitro Techniques; D008928:Mitochondria / Q000201:enzymology; D010084:Oxidation-Reduction; D011284:Pregnenolone / Q000096:biosynthesis*; D013053:Spectrophotometry
D011284:Pregnenolone; D003577:Cytochrome P-450 Enzyme System; D002784:Cholesterol