BPS2026 – α-Synuclein interactions with lipid membranes: Cooperative binding and membrane remodeling

α-Synuclein (αSyn) is a small neuronal protein localized in vivo at presynaptic termini, whose interactions with lipid membranes have been associated with both physiological and pathological functions. While the disease-related roles of αSyn have been extensively studied, its physiological functions are less well understood. Recently, it was shown that αSyn binds to anionic lipid membranes in a cooperative manner, meaning that the binding of one protein is favored at membrane patches where another protein is already bound. However, the molecular origin of this strong cooperativity remains unknown. In this study, we systematically investigate αSyn cooperative binding to lipid membranes under a broad range of conditions, using a multi-technique approach including confocal microscopy, fluorescence correlation spectroscopy, and circular dichroism. First, we interrogate the role of electrostatics by independently varying pH and ionic strength, and by studying one of the five known missense mutations in αSyn, H50Q. Our results show that αSyn binding to anionic lipid membranes is cooperative under all conditions investigated, indicating that cooperativity is a robust property of the system. We further find that αSyn forms a more dilute layer at the membrane as pH and ionic strength increase, leading us to rule out direct protein-protein interactions as the primary driving force of cooperativity. Instead, we highlight the central role of membrane-mediated interactions in this phenomenon. Second, we examine αSyn’s ability to remodel lipid membranes. Using fluorescence microscopy and scattering techniques, complemented by cryo-EM, we observe both deformation and fission of lipid vesicles upon protein binding. Altogether, our findings demonstrate strong positive cooperativity under all conditions investigated, rule out direct protein-protein interactions as the main driver, and connect this phenomenon to membrane remodeling, potentially linking it to αSyn’s physiological roles at synapses.