Synaptobrevin JMD–TMD Regions Induce Robust Lipid Disorder, Membrane Remodeling and PIP2-Dependent Cluster Formation

Synaptobrevin (Syb/VAMP) is the vesicular SNARE that drives synaptic vesicle fusion. Before fusion, Syb is located in the vesicle membrane. After fusion, Syb is translocated to the presynaptic plasma membrane and exposed to a different lipid environment. How its membrane-proximal juxtamembrane and transmembrane domains (JMD–TMDs) organ-ize these different surrounding membrane environments remains incompletely under-stood. Here, we used MARTINI coarse-grained molecular dynamics simulations to in-vestigate Syb1/2 membrane organization and collective behavior. Syb1 and Syb2 induced similar and robust local lipid disorder, whereas the spatial organization of lipid disorder and membrane mismatch depended more strongly on membrane lipid composition. Higher order Syb clustering was promoted by the presence of PIP₂ in the membrane, which is decreased in its absence and insensitive to PS depletion. Consistently, Syb cluster formation was associated with pronounced PIP₂ redistribution and enrichment, while PS showed substantially weaker enrichment and decreased association with Syb in the presence of PIP₂. Clustering spatially overlapped otherwise localized lipid-disordered regions without proportionally amplifying their magnitude. Together, these results dis-tinguish robust local membrane disorder from composition-dependent geometric re-modeling and identify PIP₂ as a prominent correlate of higher-order Syb organization that may contribute to post-fusion Syb reorganization before endocytic retrieval.