Amyloid-β (Aβ) fibrils exhibit self-propagating, molecular-level polymorphisms that may contribute to variations in clinical and pathological characteristics of Alzheimer’s disease (AD). We report the molecular structure of a specific fibril polymorph, formed by 40-residue Aβ peptides (Aβ40), that is derived from cortical tissue of an AD patient by seeded fibril growth. The structure is determined from cryogenic electron microscopy (cryoEM) images, supplemented by mass-per-length (MPL) measurements and solid-state NMR (ssNMR) data. Previous ssNMR studies with multiple AD patients had identified this polymorph as the most prevalent brain-derived Aβ40 fibril polymorph from typical AD patients. The structure, which has 2.8-Å resolution according to standard criteria, differs qualitatively from all previously described Aβ fibril structures, both in its molecular conformations and its organization of cross-β subunits. Unique features include twofold screw symmetry about the fibril growth axis, despite an MPL value that indicates three Aβ40 molecules per 4.8-Å β-sheet spacing, a four-layered architecture, and fully extended conformations for molecules in the central two cross-β layers. The cryoEM density, ssNMR data, and MPL data are consistent with β-hairpin conformations for molecules in the outer cross-β layers. Knowledge of this brain-derived fibril structure may contribute to the development of structure-specific amyloid imaging agents and aggregation inhibitors with greater diagnostic and therapeutic utility.

Protein domains without the usual distribution of amino acids, called low complexity (LC) domains, can be prone to self-assembly into amyloid-like fibrils. Self-assembly of LC domains that are nearly devoid of hydrophobic residues, such as the 214-residue LC domain of the RNA-binding protein FUS, is particularly intriguing from the biophysical perspective and is biomedically relevant due to its occurrence within neurons in amyotrophic lateral sclerosis, frontotemporal dementia, and other neurodegenerative diseases. We report a high-resolution molecular structural model for fibrils formed by the C-terminal half of the FUS LC domain (FUS-LC-C, residues 111-214), based on a density map with 2.62 Å resolution from cryo-electron microscopy (cryo-EM). In the FUS-LC-C fibril core, residues 112-150 adopt U-shaped conformations and form two subunits with in-register, parallel cross-β structures, arranged with quasi-21 symmetry. All-atom molecular dynamics simulations indicate that the FUS-LC-C fibril core is stabilized by a plethora of hydrogen bonds involving sidechains of Gln, Asn, Ser, and Tyr residues, both along and transverse to the fibril growth direction, including diverse sidechain-to-backbone, sidechain-to-sidechain, and sidechain-to-water interactions. Nuclear magnetic resonance measurements additionally show that portions of disordered residues 151-214 remain highly dynamic in FUS-LC-C fibrils and that fibrils formed by the N-terminal half of the FUS LC domain (FUS-LC-N, residues 2-108) have the same core structure as fibrils formed by the full-length LC domain. These results contribute to our understanding of the molecular structural basis for amyloid formation by FUS and by LC domains in general.

Amyloid- (A) fibrils exhibit self-propagating, molecular-level polymorphisms that may underlie variations in clinical and pathological characteristics of Alzheimer's disease. We report the molecular structure of a specific brain-derived polymorph that has been identified as the most prevalent polymorph of 40-residue A fibrils in cortical tissue of Alzheimer's disease patients. This structure, developed from cryo-electron microscopy and supported by solid state NMR data, differs qualitatively from all previously described A fibril structures, both in its molecular conformation and its organization of cross- subunits. Knowledge of this brain-derived fibril structure may contribute to the development of structure-specific amyloid imaging agents and aggregation inhibitors with greater diagnostic and therapeutic utility.To derive a three-dimensional (3D) density map from the cryoEM images, we used RELION software for helical reconstruction (30,31), modified to include correlations of particle orientations about the fibril growth direction for particles from the same fibril segment (see Supporting Text). Calculations were performed without additional symmetry, with two-fold rotational symmetry in the repeat unit, and with 21 screw symmetry. The density map with the highest final resolution (2.77 Å) was obtained with near 21 symmetry, generated by a rise of 2.45 Å and twist of -180.34 between repeats ( Fig. 2A). A density map without additional symmetry and Fourier-shell correlation plots are shown in Fig. S2.The density in Fig. 2A consists of four cross- layers. Amino acid sidechains in the two inner layers are well resolved, allowing a unique fitting of residues 13-40 into the density (27). Residues 13-22 form an N-terminal extended segment with one continuous -strand. Residues 30-40 form a C-terminal extended segment comprised of -strands in residues 30-32, 34-36, and 38-39, defined by their intermolecular hydrogen bonding patterns. Glycine residues at positions 33 and 37 adopt non--strand conformations, disrupting these patterns. All -strand segments in the inner layers participate in in-register parallel -sheets. The N-terminal and C-terminal extended segments are separated by an irregular conformation in central residues 23-29, apparently stabilized in part by electrostatic interactions involving oppositely charged sidechains of D23 and K28 (Fig. 2B). Residues 1-12 are not sufficiently ordered to contribute to the density.Conformations within N-terminal, central, and C-terminal segments in Fig. 2A resemble conformations of the same segments in previously described A fibril structures (6,14,25), but the overall structural arrangement is qualitatively different. In previous structures, non--strand conformations at certain residues allow each A molecule to fold onto itself, resulting in roughly U-shaped (6, 7, 11, 25), S-shaped (12, 13, 15-17), or C-shaped (14, 26) conformations that are stabilized by interactions among hydrophobic sidechains within a single cross- subunit. In contrast, each A40 mol...