Engineering synthetic cells with intramembrane domains possessing distinct bilayer asymmetries

Our understanding of how membrane asymmetry governs biological function is limited by the lack of techniques to produce model membranes which can reliably and accurately mimic cellular membrane asymmetry. Not only in terms of asymmetric lipid distribution, but also how that asymmetry can be confined to specific lateral locations across the membrane. Here we present an inverted emulsion method that can be used to produce synthetic cells with symmetric and asymmetric bilayers, as well as phase separation where the intermembrane domains possess distinct bilayer asymmetries. We assess the degree of lipid asymmetry using protein-lipid interaction and quenching assays. Surprisingly, the synthetic cells with asymmetric and phase separated membranes displayed pronounced curvature of the domains and resulted in membrane budding and division. Overall, this work develops biomimetic membranes with lipid compositions akin to natural biomembranes - an essential element in the development of functional synthetic cells.