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NMR-based site-resolved profiling of ?-amyloid misfolding reveals structural transitions from pathologically relevant spherical oligomer to fibril.

ABSTRACT: Increasing evidence highlights the central role of neurotoxic oligomers of the 42-residue-long ?-amyloid (A?42) in Alzheimer's disease (AD). However, very limited information is available on the structural transition from oligomer to fibril, particularly for pathologically relevant amyloids. To the best of our knowledge, we present here the first site-specific structural characterization of A?42 misfolding, from toxic oligomeric assembly yielding a similar conformation to an AD-associated A?42 oligomer, into a fibril. Transmission EM (TEM) analysis revealed that a spherical amyloid assembly (SPA) of A?42 with a 15.6 ± 2.1-nm diameter forms in a ?30-?m A?42 solution after a ?10-h incubation at 4 °C, followed by a slow conversion into fibril at ?180 h. Immunological analysis suggested that the SPA has a surface structure similar to that of amylospheroid (ASPD), a patient-derived toxic A? oligomer, which had a diameter of 10-15 nm in negative-stain TEM. Solid-state NMR analyses indicated that the SPA structure involves a ?-loop-? motif, which significantly differed from the triple-? motif observed for the A?42 fibril. The comparison of the 13C chemical shifts of SPA with those of the fibril prepared in the above conditions and interstrand distance measurements suggested a large conformational change involving rearrangements of intermolecular ?-sheet into in-register parallel ?-sheet during the misfolding. A comparison of the SPA and ASPD 13C chemical shifts indicated that SPA is structurally similar to the ASPD relevant to AD. These observations provide insights into the architecture and key structural transitions of amyloid oligomers relevant for AD pathology.

SUBMITTER: Xiao Y

PROVIDER: S-EPMC6956526 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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NMR-based site-resolved profiling of β-amyloid misfolding reveals structural transitions from pathologically relevant spherical oligomer to fibril.

Xiao Yiling Y Matsuda Isamu I Inoue Masafumi M Sasahara Tomoya T Hoshi Minako M Ishii Yoshitaka Y

The Journal of biological chemistry 20191126 2

Increasing evidence highlights the central role of neurotoxic oligomers of the 42-residue-long β-amyloid (Aβ42) in Alzheimer's disease (AD). However, very limited information is available on the structural transition from oligomer to fibril, particularly for pathologically relevant amyloids. To the best of our knowledge, we present here the first site-specific structural characterization of Aβ42 misfolding, from toxic oligomeric assembly yielding a similar conformation to an AD-associated Aβ42 o ...[more]

PMID: 31771980

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Project description:Deposits of fibrils formed by disease-specific proteins are the molecular hallmark of such diverse human disorders as Alzheimer's disease, type II diabetes, or rheumatoid arthritis. Amyloid fibril formation by structurally and functionally unrelated proteins exhibits many generic characteristics, most prominently the cross β-sheet structure of their mature fibrils. At the same time, amyloid formation tends to proceed along one of two separate assembly pathways yielding either stiff monomeric filaments or globular oligomers and curvilinear protofibrils. Given the focus on oligomers as major toxic species, the very existence of an oligomer-free assembly pathway is significant. Little is known, though, about the structure of the various intermediates emerging along different pathways and whether the pathways converge towards a common or distinct fibril structures. Using infrared spectroscopy we probed the structural evolution of intermediates and late-stage fibrils formed during in vitro lysozyme amyloid assembly along an oligomeric and oligomer-free pathway. Infrared spectroscopy confirmed that both pathways produced amyloid-specific β-sheet peaks, but at pathway-specific wavenumbers. We further found that the amyloid-specific dye thioflavin T responded to all intermediates along either pathway. The relative amplitudes of thioflavin T fluorescence responses displayed pathway-specific differences and could be utilized for monitoring the structural evolution of intermediates. Pathway-specific structural features obtained from infrared spectroscopy and Thioflavin T responses were identical for fibrils grown at highly acidic or at physiological pH values and showed no discernible effects of protein hydrolysis. Our results suggest that late-stage fibrils formed along either pathway are amyloidogenic in nature, but have distinguishable structural fingerprints. These pathway-specific fingerprints emerge during the earliest aggregation events and persist throughout the entire cascade of aggregation intermediates formed along each pathway.

Project description:Amyloid ? (A?) 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 A? monomers, whose conformational landscapes directly contribute to the kinetics and thermodynamics of A? aggregation. In this work, de novo replica exchange molecular dynamics (REMD) simulations on the microseconds-per-replica timescale are used to characterize the structural ensembles of A?42, A?40, and M35-oxidized A?42, 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 nanoseconds-per-replica toward ensembles that yield good agreement with experiment. Though all three A? species adopt highly heterogeneous ensembles, these are considerably more structured compared to simulations on shorter timescales. Prominent in the C-terminus are antiparallel ?-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 A?42, the absence of a second ?-hairpin in A?40 and the sampling of alternate ? topologies by M35-oxidized A?42 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 A? monomer folding. These characterizations help illustrate the conformational landscapes of A? monomers at atomic resolution and provide insight into the early stages of A? aggregation pathways.

Dataset Information

NMR-based site-resolved profiling of ?-amyloid misfolding reveals structural transitions from pathologically relevant spherical oligomer to fibril.

Publications

NMR-based site-resolved profiling of β-amyloid misfolding reveals structural transitions from pathologically relevant spherical oligomer to fibril.

Similar Datasets

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