Cooperative Regulation of Membrane Domain Registration by Lipid Headgroup Properties and Cholesterol Dynamics

The lateral organization of lipid domains and their coupling across the two leaflets are central to the structure of plasma membranes, yet the molecular determinants of domain registration and anti-registration remain incompletely understood. Here, we employ coarse-grained molecular dynamics simulations to investigate how lipid headgroup properties and cholesterol (CHOL) concentration regulate domain registration and anti-registration in symmetric model bilayers. Using a free energy perturbation (FEP)-like headgroup-interpolation approach to continuously interpolate between headgroup representations characteristic of phosphatidylethanolamine (PE) and phosphatidylcholine (PC) lipids at a fixed tail composition, we show that headgroup properties and CHOL concentration act cooperatively through their effects on lipid packing and membrane curvature. Simulations with restrained CHOL flip-flop further demonstrate that transbilayer CHOL exchange actively strengthens interleaflet coupling and promotes domain registration. These results identify headgroup-dependent packing, CHOL-induced curvature, and CHOL flip-flop as cooperative determinants of domain organization in model membranes.