ROLE OF N-TERMINUS IN ⍺-SYNUCLEIN CONFORMATIONS

Introduction α-Synuclein (Syn) is an intrinsically disordered protein, whose aggregation into Lewy bodies (LB) is a defining feature of Parkinson disease1 . Approximately 15–20% of Syn species in LB are truncated at the N- and/or C- terminus2 . While C-terminal truncations are known to enhance aggregation, the impact of N-terminal truncations is less characterized, despite the importance of the N-terminus for membrane binding and long-range intramolecular interactions3 . Materials and Methods To examine the role of the extreme N-terminus, we studied the Syn 7–140 variant. Membrane binding and remodeling were assessed using dynamic light scattering (DLS), circular dichroism (CD), and hydrogen– deuterium exchange mass spectrometry (HDX-MS). Aggregation kinetics were monitored by thioflavin T fluorescence. Native massspectrometry characterized the conformational ensembles of full- length Syn and Syn 7– 140 at time zero and during early aggregation. Results Deletion of residues 1–6 preserves net charge, enabling assessment of ensemble effects without altering electrostatics. Syn 7–140 binds lipid membranes with similar affinity as full-length Syn but adopts amore dynamic membrane-associated α-helical structure. Unlike full-length Syn, which induces liposome clustering, Syn 7–140 does not. Aggregation of Syn 7–140 is slower, with a decrease in elongation rate. Native MS reveals that full- length Syn samples interchanging conformations, whereas Syn 7–140 populates compact states at early stages. Under aggregating conditions, both proteins shift toward extended conformers, although this transition occurs later for Syn 7–140. Conclusions These findings identify the extreme N-terminus as a key regulator of Syn conformational plasticity, linking long- range intramolecular interactions to membrane remodeling and aggregation kinetics.