Transmembrane cholesterol migration in planar lipid membranes measured with Vibrio cholerae cytolysin as molecular tool

The rate of transbilayer movement (flip-flop) of cholesterol was estimated using planar bilayers with defined initial asymmetry, formed by the opposing monolayers technique. Vibrio cholerae cytolysin (VCC) was utilized as a molecular tool for measuring the cholesterol concentration in the cis leaflet of asymmetric bilayers. To quantify cholesterol flip-flop in planar lipid bilayers, a mathematical model was developed. It considers both the lateral diffusion rate of cholesterol within each monolayer and the flip-flop rate. The difference in initial and steady-state cholesterol contents in bilayer leaflets was used as a start point. Assuming the lateral diffusion coefficient to be of 1 × 10⁻⁸ cm² s⁻¹, the characteristic time of cholesterol flip-flop at 25 ± 2 °C was estimated as <10 s.