Structural and Motional Investigations of Polyglutamine-Containing Amyloid Fibrils by Magic-Angle-Spinning Solid-State NMR

More than 20 human diseases are associated with amyloid fibril formation, one of which is Huntington's Disease (HD). HD, which poses a risk for ∼200,000 Americans, is a neurodegenerative disease that degrades cognitive and motor functions and is one of ten disorders to feature polyglutamine (polyQ) domain expansion. The self-aggregation of expanded polyQ segments in the huntingtin protein (htt) leads to fibril formation. Flanking regions of the polyQ domain affect the kinetics of this aggregation. As protein aggregation appears to correlate to disease onset, we aim to understand the structure of the mature fibrils, a critical step toward resolving structural changes along the fibril formation pathway.Fibrils of site-specific isotopically labeled peptides mimicking polyQ-containing N-terminal fragments of the htt protein were studied by magic-angle-spinning (MAS) solid-state NMR (ssNMR). We probed the conformation of both the polyQ amyloid core and its flanking regions by multidimensional MAS ssNMR, finding an a-helical conformation for residues in the very N-terminus (httNT), which is known to initiate the aggregation process. Structural models of the aggregation pathway and fibril were assessed based on structural data, relaxation times, and order parameters from MAS ssNMR measurements. Fibrils featuring mutations in the linker region connecting the httNT helix to the amyloid core were also investigated. Earlier animal studies have shown these mutations to reduce toxicity, and EM shows significant changes in fibril morphology. The site-specific effects of these and other mutations, on both the httNT and the polyQ core, were explored through structural and dynamics measurements. The characterization of the mutants, which mimic post-translational Ser phosphorylation, suggests biophysical connections between the stability and molecular structure of species along the aggregation pathway and the observed changes in toxicity.