The transfer of cholesterol and hydroxycholesterol derivatives from liposome to soluble cytochrome P-450 scc.
We have studied the cholesterol-binding reaction with purified steroid-free cytochrome P-450scc. By mixing an aqueous solution of cholesterol-, pregnenolone- and progesterone-free cytochrome with cholesterol-containing liposomes, the low to high spin conversion of the hemoprotein was observed spectrophotometrically and by electron spin resonance spectroscopy. When the binding rates were compared at a fixed molar heme: cholesterol ratio of 1 : 1, 20 mol% cholesterol-dioleoylglycerophosphocholine liposomes react with the cytochrome at a faster rate than 50 mol% cholesterol-dimyristoylglycerophosphocholine liposomes, indicating that the availability of cholesterol molecules in an unsaturated membrane is better than in a saturated membrane. When the number of cholesterol-dioleoylglycerophosphocholine liposomes was increased, the cholesterol-binding rates increased markedly. These results imply that the collision of the liposomes with the soluble hemoprotein molecules plays an important role in the binding reaction under our experimental conditions. The binding reaction was found to be temperature-dependent with a refractive temperature at near 20 degrees C. From the comparison of the binding abilities among cholesterol derivatives tested, wer conclude that the alpha-face of the A-B transfused rings and the portion of the hydrocarbon side-chain of cholesterol are important for the binding. Additionally, polar derivatives had faster rates than non-polar steroids in structurally homologous series.
D000302:Adrenal Cortex / Q000378:metabolism; D000325:Adrenodoxin / Q000378:metabolism; D000818:Animals; D002417:Cattle; D002784:Cholesterol / Q000378:metabolism*; D003577:Cytochrome P-450 Enzyme System / Q000302:isolation & purification / Q000378:metabolism*; D005287:Ferredoxin-NADP Reductase / Q000378:metabolism; D006888:Hydroxycholesterols / Q000378:metabolism*; D007700:Kinetics; D008081:Liposomes; D008928:Mitochondria / Q000378:metabolism; D011485:Protein Binding; D012995:Solubility; D013053:Spectrophotometry
D006888:Hydroxycholesterols; D008081:Liposomes; D000325:Adrenodoxin; D003577:Cytochrome P-450 Enzyme System; D002784:Cholesterol; D005287:Ferredoxin-NADP Reductase