Structural Origins of Cholesterol Accelerated Lipid Flip-Flop Studied by Sum-Frequency Vibrational Spectroscopy

The unique structure of cholesterol and its role in modulating lipid translocation (flip-flop) were examined using sum-frequency vibrational spectroscopy (SFVS). Two structural analogues of cholesterol-cholestanol and cholestene-were examined to explore the influence of ring rigidity and amphiphilicity on controlling distearoylphosphochdline (DSPC) flip-flop. Kinetic rates for DSPC flip-flop were determined as a function of sterol concentration and temperature. All three sterols increased the rate of DSPC flip-flop in a concentration-dependent manner following the order cholestene > cholestanol > cholesterol. Rates of DSPC flip-flop were used to calculate the thermodynamic activation free energy barrier (Delta G double dagger) in the presence of cholesterol, cholestanol) and cholestene. The acyl chain gauche content of DSPC, mean lipid area, and membrane compressibility were correlated to observed trends in Delta G double dagger. Delta G double dagger for DSPC flip-flop showed a strong positive correlation with the molar compression modulus (K*) of the membrane, influenced by the type and concentration of the sterol added. Interestingly, cholesterol is distinctive in maintaining invariant membrane compressibility over the range of 2-10 mol %. The results in this study demonstrate that the compression modulus of a membrane plays a significant role in moderating Delta G double dagger and the kinetics of native, protein-free, lipid translocation in membranes.