α-Synuclein Ensemble Structure Under Different Conditions Studied By NMR

The paper mainly focused on study of α-synuclein (αS) protein, and its monomer is intrinsically disordered. αS filamentous aggregates are the major component of Lewy bodies, which is the pathological hallmark of Parkinson’s disease (PD). The abnormal aggregation and fibrillation of αS is always to be one of the main etiopathogenisis of PD. Therefore, investigating the structure properties and aggregation mechanism of αS is important to understand its pathological roles in PD.αS can form into β-sheet-rich amyloid like fibrils, and the aggregation mechanism is still unclear. Studies have shown that αS aggregation kinetics and aggregate morphology are highly sensitive to the solution conditions. Therefore, the studies of αS initial ensemble structures under different conditions are of great important significance.Due to the highly flexible nature of disorder proteins, Nuclear Magnetic Resonance (NMR) Spectroscopy is especially well-suited to probe their structures and dynamics. Here, we mainly used NMR Spectroscopy to study αS ensemble structures under different conditions, and proposed the possible mechanism of αS aggregation.Topic One:a-synuclein-trivalent metal ions interaction:binding sites, conformation and fibrillationTo investigate what causes αS aggregation is important to understand its pathological roles in PD. Various metal ions, including iron and copper, have been implicated in the pathogenesis of PD. Studies have showed that αS can interact with many divalent metal ions in vitro, however, few studies have been performed to investigate the interaction between trivalent metal ions and αS and their effect on αS aggregation. The study of the interaction between divalent and trivalent metal ions with αS is vitally important to understand the mechanism of αS aggregation. Here we used NMR Spectroscopy to determine the trivalent metal ions (lanthanides, aluminum) binding sites in αS. Compared to divalent cations, trivalent metal ions accelerate αS fibrillation much faster in vitro. Based on the interaction information, we proposed the mechanism by which trivalent metal ions accelerated αS fibrillation in vitro.Topic two:The salts effects on a-synuclein initial ensemble structure, fibrillation pathways and kineticsStudies have shown aS formed into different aggregate morphologies in the presence or absence of 150 mM sodium chloride (NaCl) solution, and the aggregates exhibit diverse physiological properties and cytotoxicity. Here, we mainly used NMR Spectroscopy to characterize aS initial ensemble structures and dynamics in the presence or absence of 150 mM NaCl. We found that aS exhibits distinct conformations and aggregation kinetics in these two solutions. aS adopts a more compact and rigid ensemble structure which has faster fibrillation kinetics in the absence of NaCl. Using 19F NMR, we observed conformation selection during aS assembly for the first time. The dynamics results suggested that aS initial ensemble structure may be potentially related to the aggregate structures. Based on the ensemble structure and dynamics, we proposed possible molecular mechanisms how the initial conformation impacts aggregation pathways and kinetics.Topic three:Crowding effects on a-synuclein initial ensemble structure and fibrillation kineticsThe intracellular milieu contains upwards of 400 g of macromolecules per litre. This crowding is thought to have a larger influence on intrinsically disordered proteins, whose chains are expanded, than on compact globular proteins, because increasing the excluded volume favours compaction. We used NMR Spectroscopy, Circular Dichroism and Fluorescence Spectroscopies to characterize the structure and fibrillation of aS, an intrinsically disordered protein implicated in Parkinson’s disease, using Ficoll 70, its monomer Sucrose and Bull Serum Albumin (BSA) as crowding agents. Surprisingly, macromolecular excluded volume effects caused by crowding environments are not the main reason that inducing aS compaction. Based on aS structures and dynamics, we proposed the mechanism by which macromolecular crowding environments accelerated aS fibrillation.