Structural Basis of Huntingtin Fibril Polymorphism Revealed by Cryogenic Electron Microscopy of Exon 1 HTT Fibrils.

The lack of detailed insight into the structure of aggregates formed by the huntingtin protein (HTT) has hampered the efforts to develop therapeutics and diagnostics targeting pathology formation in the brain of patients with Huntington's disease. To address this knowledge gap, we investigated the structural properties of in vitro-generated fibrils from exon1 of the huntingtin protein by cryogenic electron microscopy and single-particle analyses. We show that wildtype and mutant exon1 of the huntingtin protein form nonhelical fibrils with a polyglutamine amyloid core composed of β-hairpins with unique characteristics that have not been previously observed with other amyloid filaments. The stacks of β-hairpins form long planar β-sheets (protofilaments) which combine inter- and intra-molecular interactions, with variable stacking angles and occasional out-of-register states of individual β-hairpins. These features and the propensity of protofilaments to undergo lateral association result in a high degree of fibril polymorphisms, including fibrils composed of varying numbers of protofilaments. Our results allow us to speculate on how the flanking domains are organized around the polyglutamine core of the fibril and provide insight into how they might affect the huntingtin fibril structure and polymorphism. The removal of the first 17 amino acids at the N-terminus resulted in surprising intra-fibril structural heterogeneity and reduced fibril's propensity to lateral associations. Overall, this work provides valuable insights that could help guide future mechanistic studies to elucidate the sequence and structural determinants of huntingtin aggregation, as well as for cryo-EM and structural studies of fibrils derived from huntingtin protein and other disease-associated polyglutamine-containing proteins.