Selective observation of semi-rigid non-core residues in dynamically complex mutant huntingtin protein fibrils

Abstract

Many amyloid-forming proteins, which are normally intrinsically disordered, undergo a disorder-to-order transition to form fibrils with a rigid β-sheet core flanked by disordered domains. Solid-state NMR (ssNMR) and cryogenic electron microscopy (cryoEM) excel at resolving the rigid structures within amyloid cores but studying the disordered domains on fibril surfaces remains more challenging. This challenge is exemplified by a fragment of mutant huntingtin exon 1 (HttEx1), which self-assembles into pathogenic neuronal inclusions in Huntington disease (HD). The mutant protein’s expanded polyglutamine (polyQ) segment forms a highly rigid fibril core with dynamic surface residues and disordered ‘flanking’ segments. Here we deploy dynamically sensitive magic angle spinning ssNMR experiments and variable temperature studies to elucidate how solvent dynamics impact the fibrils’ segmental dynamics, and permit the identification of surface-facing (polyQ) residues. A novel implementation of dynamic spectral editing (DYSE) ssNMR is described and applied, in order to reveal the weak signals of surface-facing glutamine residues despite the strong background of rigid core signals. This type of ‘intermediate motion selection’ (IMS) experiment, based on cross-polarization (CP) ssNMR, is complementary to more traditional DYSE methods that highlight highly flexible or highly rigid protein segments. Integration of the IMS-DYSE element in standard CP-based ssNMR experiments permits the observation of previously invisible fiber surface signals and may be extended to a wider variety of biomolecular assemblies and (bio)materials. In pathogenic protein amyloid fibrils the detection of surfaces of the (polyQ) amyloid core is relevant to e.g. the development of fluorescent dyes and positron emission tomography tracers that selectively bind these surfaces.HighlightsMutant huntingtin exon 1 fibrils feature a broad range of molecular dynamics.Molecular motion is coupled to dynamics of the fiber-associated water solvent.Dynamics-based spectral editing ssNMR reveals mobile surface residues.Intermediate-motion selection uses dipolar-dephasing of rigid sites.Surface-facing glutamines outside the fiber core observed and identified.