BRANCHED PHOSPHATIDYLCHOLINES STIMULATE ACTIVITY OF CYTOCHROME P450SCC (CYP11A1) IN PHOSPHOLIPID-VESICLES BY ENHANCING CHOLESTEROL BINDING,MEMBRANE INCORPORATION, AND PROTEIN EXCHANGE

Phosphatidylcholines (PCs) with branched fatty acyl chains substituted in the two positions of the main chains (branched PCs) have been show n to be potent activators of the side chain cleavage activity of cytoc hrome P450SCC (CYP11A1) (Schwarz, D.,, Kisselev, P.,, Wessel, R.,, Jue ptner, O., and Schmid, R., D., (1996) J., Biol, Chem, 271, 12840-12846 ), The present study reports on the effect of a series of branched PC on cholesterol binding, membrane integration, and protein exchange in large unilamellar vesicles prepared by an extrusion technique, Enzyme kinetics using vesicles as well as optical titration using a micelle s ystem with the detergent Tween 20 demonstrate that activation is corre lated with the fraction of P450SCC in the high spin form, The potency of branched PCs both to activate the enzyme and to induce spin state c hanges increases with increasing lengths of both the branched and main fatty acyl chains, We found that the extent as well as the rate of in tegration of P450SCC into vesicle membranes studied by gel chromatogra phy and stopped flow kinetics were increased by branched PC, Finally, it is demonstrated by measurement of the enzymatic activity in primary and secondary vesicles that branched PCs are potent in retaining a ve ry rapid exchange of P450SCC between vesicles, in contrast to cardioli pin, that partially inhibits this exchange process, The data suggest t hat different properties of P450SCC in membrane systems including chol esterol binding, membrane integration, and protein exchange are affect ed by branched PCs and probably by other phospholipids, too, and there fore must be considered in an explanation of the observed high stimula tion of activity.