Asymmetric Structural Features in Single Supported Lipid Bilayers Containing Cholesterol and GM1 Resolved with Synchrotron X-Ray Reflectivity

The cell membrane comprises numerous protein and lipid molecules capable of asymmetric organization between leaflets and liquid-liquid phase separation. We use single supported lipid bilayers (SLBs) to model cell membranes, and study how cholesterol and asymmetrically oriented ganglioside receptor GM1 affect membrane structure using synchrotron x-ray reflectivity. Using mixtures of cholesterol, sphingomyelin, and 1,2-dioleoyl-sn-glycero-3-phosphocholine, we characterize the structure of liquid-ordered and liquid-disordered SLBs in terms of acyl-chain density, headgroup size, and leaflet thickness. SLBs modeling the liquid-ordered phase are 10 Å thicker and have a higher acyl-chain electron density (〈ρchain〉 = 0.33 e−/Å3) compared to SLBs modeling the liquid-disordered phase, or pure phosphatidylcholine SLBs (〈ρchain〉 = 0.28 e−/Å3). Incorporating GM1 into the distal bilayer leaflet results in membrane asymmetry and thickening of the leaflet of 4–9 Å. The structural effect of GM1 is more complex in SLBs of cholesterol/sphingomyelin/1,2-dioleoyl-sn-glycero-3-phosphocholine, where the distal chains show a high electron density (〈ρchain〉 = 0.33 e−/Å3) and the lipid diffusion constant is reduced by ∼50%, as measured by fluorescence microscopy. These results give quantitative information about the leaflet asymmetry and electron density changes induced by receptor molecules that penetrate a single lipid bilayer.