Measurements of cholesterol chemical potential constrain models of cholesterol-phosphatidylcholine interactions.

Abstract

Bilayer membranes composed of cholesterol and phospholipids exhibit diverse forms of non-ideal mixing. In particular, many previous studies document macroscopic liquid-liquid phase separation as well as nanometer-scale heterogeneity in membranes of phosphatidylcholine (PC) and cholesterol. Here, we present experimental measurements of cholesterol chemical potential (μch) in binary membranes containing dioleoyl PC (DOPC), 1-palmitoyl-2-oleoyl PC (POPC), or dipalmitoyl PC (DPPC), and in ternary membranes of DOPC and DPPC, adapting a calibrated experimental protocol developed to measure μch in cells (Ayuyan and Cohen, Biophys. J. 114:904-918). μch is the thermodynamic quantity that dictates the availability of cholesterol to bind other factors, and notably must be equal between coexisting phases of a phase-separated mixture. It is simply related to concentration under conditions of ideal mixing, but is found to be far from ideal for the majority of lipid mixtures investigated. Experimental measurements of μch are used to constrain thermodynamic models of membrane interactions. Measurements are consistent with models involving cholesterol-phospholipid complexes only if complexes are more weakly bound than is assumed in previous reports. Experimental measurements are also well described by regular solution theory and lattice models of pairwise interactions between components. These findings reinforce that μch depends on membrane composition overall, providing avenues for cells to alter the availability of cholesterol without varying cholesterol concentration.