Accelerated\nAmyloid Aggregation Dynamics of Intrinsically\nDisordered Proteins in Heavy Water

We explored the influence of D<sub>2</sub>O on the fibrillation\nkinetics and structural dynamics of amyloid intrinsically disordered\nproteins (IDPs), including α-synuclein, amyloid-β 1–42,\nand K18. Our findings revealed that fibrillation of IDPs was accelerated\nin D<sub>2</sub>O compared to that in H<sub>2</sub>O, exhibiting faster\nkinetics in contrast to the structured protein, insulin. Structural\ninvestigations using electrospray ionization ion mobility mass spectrometry\nand small-angle X-ray scattering combined with molecular dynamics\nsimulations demonstrated that IDPs did not show significant structural\nchanges that could influence accelerated fibrillation in D<sub>2</sub>O. Umbrella sampling of protein protofibrils verified that an increased\nlevel of hydrogen bonding of D<sub>2</sub>O and enhanced hydrophobic\ninteractions stabilized β-sheet structured fibrils in D<sub>2</sub>O. These findings indicate that stabilizing β-sheet\nfibrils and a more hydrophobic microenvironment in D<sub>2</sub>O\nresult in enhanced and faster fibrillation of IDPs. The study highlights\nthe importance of considering D<sub>2</sub>O’s differential\nimpact on protein interactions when conducting structural and kinetic\nanalyses, particularly for native peptides and proteins.