Amyloid β (Aβ) peptides are a primary component of fibrils and oligomers implicated in the etiology of Alzheimer’s disease (AD). However, the intrinsic flexibility of these peptides has frustrated efforts to investigate the secondary and tertiary structure of Aβ monomers, whose conformational landscapes directly contribute to the kinetics and thermodynamics of Aβ aggregation. In this work, de novo replica exchange molecular dynamics (REMD) simulations on the μs/replica timescale are used to characterize the structural ensembles of Aβ42, Aβ40, and M35-oxidized Aβ42, three physiologically relevant isoforms with substantially different aggregation properties. J-coupling data calculated from the REMD trajectories were compared to corresponding NMR-derived values acquired through two different pulse sequences, revealing that all simulations converge on the order of hundreds of ns/replica toward ensembles that yield good agreement with experiment. Though all three Aβ species adopt highly heterogeneous ensembles, these are considerably more structured compared to simulations on shorter timescales. Prominent in the C-terminus are antiparallel β-hairpins between L17-A21, A30-L36, and V39-I41, similar to oligomer and fibril intrapeptide models, that expose these hydrophobic side chains to solvent and may serve as hotspots for self-association. Compared to reduced Aβ42, the absence of a second β-hairpin in Aβ40 and the sampling of alternate β topologies by M35-oxidized Aβ42 may explain the reduced aggregation rates of these forms. A persistent V24-K28 bend motif, observed in all three species, is stabilized by buried backbone to side chain hydrogen bonds with D23 and a cross-region salt bridge between E22 and K28, highlighting the role of the familial AD-linked E22 and D23 residues in Aβ monomer folding. These characterizations help illustrate the conformational landscapes of Aβ monomers at atomic resolution and provide insight into the early stages of Aβ aggregation pathways.

The high Aβ42/Aβ40 production ratio is a hallmark of familial Alzheimer’s disease, which can be caused by mutations in the amyloid precursor protein (APP). The C-terminus of Aβ is generated by γ-secretase cleavage within the transmembrane domain of APP (APPTM), a process that is primed by an initial ε-cleavage at either T48 or L49, resulting in subsequent production of Aβ42 or Aβ40, respectively. Here we solve the dimer structures of wild-type APPTM (AAPTM WT) and mutant APPTM (FAD mutants V44M) with solution NMR. The right-handed APPTM helical dimer is mediated by GXXXA motif. From the NMR structural and dynamic data, we show that the V44M and V44A mutations can selectively expose the T48 site by weakening helical hydrogen bonds and increasing hydrogen–deuterium exchange rate (kex). We propose a structural model in which FAD mutations (V44M and V44A) can open the T48 site γ-secretase for the initial ε-cleavage, and consequently shift cleavage preference towards Aβ42.

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