Order Differences between Coexisting Liquid Phases Driven by Lipid Unsaturation Determine Phase Separation in Biomimetic Membranes

Abstract

Biological membranes contain a broad variety of lipid species whose individual physicochemical properties and collective interactions ultimately determine membrane organization. A key aspect of the organization of cellular membranes is their lateral subdivision into domains of distinct structure and composition. The most widely studied membrane domains are lipid rafts, which are the biological manifestation of the capacity of sterol-containing membranes to form liquid-ordered phases. Detailed studies of biomimetic membrane mixtures have yielded wide-ranging insights into the physical principles behind lipid rafts; however, these simplified models do not fully capture the diversity and complexity of the mammalian lipidome, most notably in their exclusion of polyunsaturated lipids. Here, we assess the role of lipid tail unsaturation as a driving force for phase separation using coarse-grained molecular dynamics simulations validated by model membrane experiments. The clear trends in our observations and good qualitative agreements between simulations and experiments support the conclusions that highly unsaturated lipids promote liquid-liquid phase separation by enhancing the differences in lipid chain order, domain thickness, cholesterol preference, and diffusivity between the coexisting domains. To determine which of these parameters is the key driver of phase separation, we tuned lipid compositions to independently vary membrane order and thickness. We find that interdomain order differences driven by acyl chain unsaturation are the key determinants of domain formation in model membranes. These observations begin to define the important role of non-canonical biological lipids on the physical properties of membranes, showing that lipid polyunsaturation is a driving force for liquid-liquid phase separation.