High-resolution NMR characterization of low abundance oligomers of amyloid-β without purification

Abstract

Alzheimer’s disease is characterized by the misfolding and self-assembly of the amyloidogenic protein amyloid-β (Aβ). The aggregation of Aβ leads to diverse oligomeric states, each of which may be potential targets for intervention. Obtaining insight into Aβ oligomers at the atomic level has been a major challenge to most techniques. Here, we use magic angle spinning recoupling 1H-1H NMR experiments to overcome many of these limitations. Using 1H-1H dipolar couplings as a NMR spectral filter to remove both high and low molecular weight species, we provide atomic-level characterization of a non-fibrillar aggregation product of the Aβ1-40 peptide using non-frozen samples without isotopic labeling. Importantly, this spectral filter allows the detection of the specific oligomer signal without a separate purification procedure. In comparison to other solid-state NMR techniques, the experiment is extraordinarily selective and sensitive. A resolved 2D spectra could be acquired of a small population of oligomers (6 micrograms, 7% of the total) amongst a much larger population of monomers and fibers (93% of the total). By coupling real-time 1H-1H NMR experiments with other biophysical measurements, we show that a stable, primarily disordered Aβ1-40 oligomer 5–15 nm in diameter can form and coexist in parallel with the well-known cross-β-sheet fibrils.

Amylin–Aβ oligomers at atomic resolution using molecular dynamics simulations: a link between Type 2 diabetes and Alzheimer's disease

“…60 Previous ssNMR studies have shown α-helical structures of Amylin and not cross-β structure. 61,62 Recent ssNMR studies presented unstructured Aβ oligomers 63 and Amylin oligomers that form large micelles, 64 which may be a general phenomenon for natively unstructured Amylin. We did not apply these α-helical and the unstructured amylin, or the unstructured Aβ oligomers in the current study, because of the lack of the PDB coordinates.…”

Pathogenic Mutations Induce Partial Structural Changes in the Native β-Sheet Structure of Transthyretin and Accelerate Aggregation

“…Solid-state NMR has proven to be a powerful tool for detailed structural characterization of non-crystalline amyloid at atomic resolution. 43–55 However, structural analyses of the tertiary β-structure within this 127-residue TTR amyloid are a daunting task. We recently demonstrated that specific labeling schemes, which generate isolated 13 CO- 13 Cα dipolar-coupled spin pairs in the native β-sheets (Figure 5a), are of great use for probing the native β-sheet structure in amyloid state.…”

An Account of Amyloid Oligomers: Facts and Figures Obtained from Experiments and Simulations

“…They characterized low‐abundance oligomers of Aβ in the presence of other monomers and fibrils, without purification. With the aid of radio‐frequency‐driven 2D 1 H, 1 H experiments based on dipolar recoupling, a 5 to 10 nm small oligomeric species of Aβ40 was identified as a dynamic and disordered intermediate along the fibril pathway …”