The side-chain cleavage of cholesterol sulfate--III. The effect of adrenodoxin, membrane phospholipids and Tween 80 on the kinetics of oxidation of the sterol sulfate by a reconstituted cholesterol desmolase system.
This paper reports the Km values of a reconstituted cholesterol side-chain cleavage system for cholesterol sulfate, cholesterol, and adrenodoxin, determined under several experimental conditions. The Km values for adrenodoxin change depending on whether cholesterol or its sulfate is used as the substrate. Moreover, the Km values for both of the substrates and for adrenodoxin are greatly modulated by both membrane phospholipids, isolated from adrenal mitochondria, and Tween 80, 0.002%. In the absence of detergents or phospholipids, the enzyme system shows a high affinity for cholesterol sulfate, but is inhibited when high concentrations of the sterol sulfate are added to the incubation mixture. Raising the concentration of adrenodoxin in the assay mixture prevents the substrate inhibition. When cholesterol sulfate is incorporated into micelles containing the phospholipids, the enzyme system does not display substrate inhibition, and the kinetics of cleavage of the sterol sulfate are relatively independent of the concentration of adrenodoxin in the assay mixture. In the absence of phospholipids, the apparent kinetics of cleavage of cholesterol and its sulfate are quite different from each other, but when incorporated into micelles containing phospholipids, the kinetics of cleavage of the two substrates are similar to each other.
D000302:Adrenal Cortex / Q000378:metabolism; D000325:Adrenodoxin / Q000494:pharmacology*; D000818:Animals; D002417:Cattle; D002788:Cholesterol Esters / Q000378:metabolism*; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000378:metabolism*; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D007700:Kinetics; D008563:Membrane Lipids / Q000502:physiology*; D008928:Mitochondria / Q000378:metabolism; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000378:metabolism*; D010743:Phospholipids / Q000494:pharmacology* / Q000502:physiology; D011136:Polysorbates / Q000494:pharmacology*
D002788:Cholesterol Esters; D008563:Membrane Lipids; D010743:Phospholipids; D011136:Polysorbates; D000325:Adrenodoxin; D003577:Cytochrome P-450 Enzyme System; D010088:Oxidoreductases; D002786:Cholesterol Side-Chain Cleavage Enzyme; C007045:cholesteryl sulfate