Structural Characterization of Polyglutamine Fibrils by Solid-State NMR Spectroscopy

Abstract

Protein aggregation via polyglutamine stretches occurs in a number of severe neurodegenerative diseases such as Huntington's disease. We have investigated fibrillar aggregates of polyglutamine peptides below, at, and above the toxicity limit of around 37 glutamine residues using solid-state NMR and electron microscopy. Experimental data are consistent with a dry fibril core of at least 70-80 Å in width for all constructs. Solid-state NMR dipolar correlation experiments reveal a largely β-strand character of all samples and point to tight interdigitation of hydrogen-bonded glutamine side chains from different sheets. Two approximately equally frequent populations of glutamine residues with distinct sets of chemical shifts are found, consistent with local backbone dihedral angles compensating for β-strand twist or with two distinct sets of side-chain conformations. Peptides comprising 15 glutamine residues are present as single extended β-strands. Data obtained for longer constructs are most compatible with a superpleated arrangement with individual molecules contributing β-strands to more than one sheet and an antiparallel assembly of strands within β-sheets.

Atomic structure and hierarchical assembly of a cross-β amyloid fibril

“…For example, the linewidths of A108Cβ, A109Cβ, L110Cδ, and L111Cδ are ∼1.0-1.2 ppm. Similar doubling of some peaks has also been detected for polyglutamine (poly-Q) fibrils, although in this case this phenomenon is likely to arise from an antiparallel β-sheet structure, which is also a packing geometry leading to residues alternately pointing into (buried) and out (solvent exposed) of the fibril (28,29). The intensities of the peaks that exhibit doubling in SI Appendix, Fig.…”

“…The intensities of the peaks that exhibit doubling in SI Appendix, Fig. S5 are not 1:1, unlike the peaks in poly-Q spectra (28,29), because here, the solvent exposed side chains are more likely to be dynamic than those buried in the dry interface between the β-sheets.…”

Huntingtin exon 1 fibrils feature an interdigitated β-hairpin–based polyglutamine core

“…S1 shows that these NMR signals are indistinguishable from those of "simple" polyQ fibrils without htt flanking domains (11,(15)(16)(17). Previous studies have noted that these chemical shifts are indicative of β-sheet rather than α-helical or random coil structure.…”

“…S1). A much smaller set of signals (type "c") is observed for Gln outside the amyloid core (16,17) (SI Appendix, Fig. S1).…”

“…We previously used MAS ssNMR to elucidate the domain structure of aggregated htt N-terminal fragments, in which we identified the rigid amyloid core (15,16). This polyQ-based amyloid core was found to have an unusual spectral signature that is also shared by other polyQ aggregates (11,15,17). However, until now, no ssNMR-based structural constraints on the htt exon1 amyloid core were available.…”