Evidence for Intramolecular Antiparallel Beta-Sheet Structure in Alpha-Synuclein Fibrils from a Combination of Two-Dimensional Infrared Spectroscopy and Atomic Force Microscopy

The aggregation of the intrinsically disordered protein alpha-synuclein (alpha S) into amyloid fibrils is thought to play a central role in the pathology of Parkinson's disease. Using a combination of techniques (AFM, UV-CD, XRD, and amide-I 1D- and 2D-IR spectroscopy) we show that the structure of aS fibrils varies as a function of ionic strength: fibrils aggregated in low ionic-strength buffers ([NaCl] <= 25 mM) have a significantly different structure than fibrils grown in higher ionic-strength buffers. The observations for fibrils aggregated in low-salt buffers are consistent with an extended conformation of aS molecules, forming hydrogen-bonded intermolecular beta-sheets that are loosely packed in a parallel fashion. For fibrils aggregated in high-salt buffers (including those prepared in buffers with a physiological salt concentration) the measurements are consistent with aS molecules in a more tightly-packed, antiparallel intramolecular conformation, and suggest a structure characterized by two twisting stacks of approximately five hydrogen-bonded intermolecular beta-sheets each. We find evidence that the high-frequency peak in the amide-I spectrum of aS fibrils involves a normal mode that differs fundamentally from the canonical high-frequency antiparallel beta-sheet mode. The high sensitivity of the fibril structure to the ionic strength might form the basis of differences in alpha S-related pathologies.