Molecular Role of Ca2+ and Hard Divalent Metal Cations on Accelerated Fibrillation and Interfibrillar Aggregation of alpha-Synuclein

alpha-Synuclein (alpha Syn) is an intrinsically disordered protein, the aggregation of which is highly related to the pathology of diverse alpha-synucleinopathies. Various hard divalent metal cations have been shown to affect alpha Syn aggregation. Especially, Ca2+ is suggested to be a crucial ion due to its physiological relevance to alpha-synucleinopathies. However, the molecular origin of alpha Syn aggregation mediated by the metal ions is not fully elucidated. In this study, we revealed that hard divalent metal ions had almost identical influences on alpha Syn aggregation. Based on these similarities, the molecular role of Ca2+ was investigated as a representative metal ion. Herein, we demonstrated that binding of multiple Ca2+ ions induces structural transition of alpha Syn monomers to extended conformations, which promotes rapid alpha Syn fibrillation. Additionally, we observed that Ca2+ induced further interfibrillar aggregation via electrostatic and hydrophobic interactions. Our results from multiple biophysical methods, including ion mobility-mass spectrometry (IM-MS), synchrotron small-angle X-ray scattering (SAXS), transmission electron microscopy (TEM), provide detailed information on the structural change of alpha Syn and the aggregation process mediated by Ca2+. Overall, our study would be valuable for understanding the influence of Ca2+ on the aggregation of alpha Syn during the pathogenesis of alpha-synucleinopathies.