Fig. 5.

Five diverse biophysical techniques were integrated to determine unambiguously the structures of each of the motifs that make up the TTR(105–115) amyloid fibrils. Spanning five orders of magnitude, the overlapping length scales of MAS NMR (0.1–10 Å), X-ray diffraction (3–100 Å), cryo-EM (8–1,000 Å), AFM (30–1,000 Å), and STEM (80–1,000 Å) enabled us to derive self-consistent, high-precision structural restraints on the secondary (β-strand and -sheet; distance restraints of <6 Å), tertiary (protofilament; distance restraints of 4.5–37 Å), and quaternary structure (filament and fibril; distance restraints of 16–1,000 Å) of the TTR(105–115) amyloid fibrils. (A) Histogram of STEM MPL measurements of TTR(105–115) fibrils, which reveals three populations of fibrils, with a best fit (gray solid line) being the sum of three Gaussian curves with values of 2.5 ± 0.3 kDa/Å (orange solid line), 3.3 ± 0.3 kDa/Å (yellow solid line), and 4.1 ± 0.3 kDa/Å (purple solid line). The orange, yellow, and purple dashed lines refer to the number of TTR(105–115) peptides per 4.67-Å repeat in the doublet (8 peptides), triplet (12 peptides), and quadruplet (16 peptides) fibrils, respectively. (B) Comparison of the high-resolution experimental X-ray diffraction pattern from TTR(105–115) fibrils (34) (Left) and the simulated X-ray diffraction pattern for TTR(105–115) fibrils (Right). The fibril axis is vertical, with the incident beam directed orthogonally to this axis. The meridional reflection at 4.67 Å and the equatorial reflection at 8.86 Å are characteristic of cross-β structure. (C) High-resolution AFM image of fibrils (pink and purple) and filaments (green) formed by TTR(105–115). (Scale bar, 1 μm.) Fibrils (pink and purple) have heights ranging from 70 to 160 Å and pitches of 950 ± 100 Å. The filament has an average height of 38.7 ± 4.4 Å. (D) Hierarchy of atomic-resolution motifs involved in the self-assembly of the amyloid fibrils and their polymorphism.