Real-time investigation of alpha-synuclein liquid-liquid phase separation by time-resolved small-angle X-ray scattering and computer simulations

Parkinson's disease (PD) affects 10 million people worldwide and is characterized by the abnormal accumulation of protein aggregates in neuronal cells. Such aggregates are mainly composed of an intrinsically disordered protein called alpha-synuclein (α-syn). Recent evidence suggests that the nucleation event driving α-syn aggregation might be initiated by liquid-liquid phase separation (LLPS). Due to a lack of techniques able to look at the early-stage LLPS the link between α-syn aggregation and LLPS is missing. Current strategies mainly focus on temporal scales where LLPS is already in place (seconds, minutes, hours) and hence there is the need to access the temporal scale of sub-second to shed light on this mechanism. The goal of Elsα (Early-stage LLPS of α-syn) is to elucidate the molecular mechanisms underlying α-syn LLPS and disentangle the contributions at play. To this purpose, I will use an innovative multi-disciplinary approach that combines a cutting-edge time-resolved small-angle X-ray scattering technique with computer simulations. The first enables real-time investigation of LLPS formation with an unprecedented temporal resolution of up to milliseconds. The second will complement the experimental outcomes with a molecular view of the phenomena. By integrating experimental and computational methods, I will identify key factors initiating α-syn LLPS, disentangle their contributions, construct comprehensive phase diagrams for α-syn under various conditions, and eventually unravel the early-stage molecular events in LLPS formation. This research promises to significantly advance our understanding of PD pathogenesis, potentially leading to novel early-phase screening methods and more effective therapeutic strategies for PD neurodegenerative disorder. Further, It will lead to high-impact publications that will increase my visibility in the scientific community boosting my career on the path to becoming a young group leader.