Abstract
Recent advances in the structural biology of disease-relevant α-synuclein fibrils have revealed a variety of structures, yet little is known about the process of fibril aggregate formation. Characterization of intermediate species that form during aggregation is crucial; however, this has proven very challenging because of their transient nature, heterogeneity, and low population. Here, we investigate the aggregation of α-synuclein bound to negatively charged phospholipid small unilamellar vesicles. Through a combination of kinetic and structural studies, we identify key time points in the aggregation process that enable targeted isolation of prefibrillar and early fibrillar intermediates. By using solid-state nuclear magnetic resonance, we show the gradual buildup of structural features in an α-synuclein fibril filament, revealing a segmental folding process. We identify distinct membrane-binding domains in α-synuclein aggregates, and the combined data are used to present a comprehensive mechanism of the folding of α-synuclein on lipid membranes.