Abstract
All amyloid fibrils contain a cross-β fold. How this structure differs in fibrils formed from proteins associated with different diseases remains unclear. Here, we combine cryo-EM and MAS-NMR to determine the structure of an amyloid fibril formed in vitro from β2-microglobulin (β2m), the culprit protein of dialysis-related amyloidosis. The fibril is composed of two identical protofilaments assembled from subunits that do not share β2m’s native tertiary fold, but are formed from similar β-strands. The fibrils share motifs with other amyloid fibrils, but also contain unique features including π-stacking interactions perpendicular to the fibril axis and an intramolecular disulfide that stabilises the subunit fold. We also describe a structural model for a second fibril morphology and show that it is built from the same subunit fold. The results provide insights into the mechanisms of fibril formation and the commonalities and differences within the amyloid fold in different protein sequences.
Highlights
All amyloid fibrils contain a cross-β fold
Despite the first identification of the cross-β motif more than 50 years ago[8], the structure of amyloid eluded high-resolution structural definition for all but the smallest of peptide assemblies[9]. This raised the question of how many structures conform to the canonical cross-β fold; how different sequences can assemble into this same fold family; and how the structure of amyloid fibrils generated in vitro relate to their counterparts formed in situ
Recent developments in magic angle spinning (MAS)-NMR and cryo-EM have seen an end to this impasse, with high-resolution structures of fibrils formed from Aβ42 and α-synuclein in vitro, and tau fibrils ex vivo being reported in the last year[10,11,12,13,14]
Summary
We observed a weak peak that is not connected to any other residues and is believed to stem from S20. The β2m subunits have an ordered, L-shaped core formed by residues 22–85 (Fig. 3d) This conformation is consistent with the MAS-NMR data, including a total of 1157 unique distance constraints (Fig. 3e; Supplementary Figure 3; Supplementary Tables 2–5). The ‘leg’ (residues Thr71–Arg[68] and Phe22–Ser[33], including β-strands 1 and 6) contains the intramolecular disulfide bond (Cys25–Cys80) that is found in the native protein and is required for fibril formation in vitro[51] and in vivo[52] This region is further stabilised by hydrophobic interactions between Phe[30], Val[27], and Tyr[78] (Fig. 5c). The distance between Tyr[67] rings across the a b d e
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