Nanoscale Infrared Spectroscopy Identifies Structural Heterogeneity in Individual Amyloid Fibrils and Prefibrillar Aggregates

Abstract

Amyloid plaques are one of the central manifestations of Alzheimer’s disease pathology. Aggregation of the amyloid beta (Aβ) protein from amorphous oligomeric species to mature fibrils has been extensively studied. However, structural heterogeneities in prefibrillar species, and how that affects the structure of later-stage aggregates are not yet well understood. The integration of infrared spectroscopy with atomic force microscopy (AFM-IR) allows for identifying the signatures of individual nanoscale aggregates by spatially resolving spectra. We use AFM-IR to demonstrate that amyloid oligomers exhibit significant structural variations as evidenced in their infrared spectra. This heterogeneity is transmitted to and retained in protofibrils and fibrils. We show that amyloid fibrils do not always conform to their putative ordered structure and structurally different domains exist in the same fibril. We further demonstrate that these structural heterogeneities manifest themselves as a lack of β sheet structure in amyloid plaques in Alzheimer’s tissue using infrared imaging.

Nanoscale Infrared Spectroscopy Identifies Parallel to Antiparallel β-Sheet Transformation of Aβ Fibrils

“…The 2 h fibrils in fact show spectral variations along the same fibril, which clearly indicates that we do not observe distinct polymorphs. We have recently reported similar observations in Aβ42, 23 and these results point to heterogeneity being a key structural facet of early-stage amyloid aggregates. Both types of oligomers exhibit the same spectral bands or substructures but differ significantly in terms of their relative intensities and, thus, in terms of the overall secondary structure.…”

“…AFM provides nanoscale morphological information on the aggregates, whereas IR spectroscopy reveals the secondary structure of them at the individual aggregate level. 22,23 We have applied AFM−IR to investigate the structural reorganization of the Aβ 16−22 peptide, which is the smallest peptide spanning the hydrophobic core that forms fibrils. 15 The late-stage fibrils of Aβ 16−22 have been previously shown to have antiparallel β structure; we show that early-stage aggregates, including both prefibrillar aggregates and fibrils, adopt an ordered parallel β structure, which transforms into an antiparallel β structure upon fibril maturation.…”

Nanoscale infrared spectroscopy identifies parallel to antiparallel beta sheet transformation of Aβ fibrils

“…The 2h fibrils in fact show spectral variations along the same fibril, which clearly indicates that we do not observe distinct polymorphs. We have recently reported similar observations in Aβ42 13 , and these results point to heterogeneity being a key structural facet of early-stage amyloid aggregates. Both types of oligomers exhibit the same spectral bands or substructures, but differ significantly in terms of their relative intensities and thus in terms of the overall secondary structure.…”

“…In this report, we use atomic force microscopy (AFM) combined with infrared spectroscopy (AFM-IR) to address this issue. AFM provides nanoscale morphological information of the aggregates, whereas IR spectroscopy reveals the secondary structure of them at individual aggregate level 12, 13 . We have applied AFM-IR to investigate the structural reorganization of the Aβ 16-22 peptide, which is the smallest peptide spanning the hydrophobic core that forms fibrils 10 .…”

Applications of Single-Molecule Vibrational Spectroscopic Techniques for the Structural Investigation of Amyloid Oligomers

“…Waeytens et al [ 91 ] studied Aβ fibrils and oligomers by AFM-IR, reporting a detrimental effect of ZnSe substrate surface on the sample, drawing no further conclusion about the structural composition of the studied aggregates. Banerjee et al investigated aggregates of tau [ 92 ] and Aβ42 [ 93 ] by AFM-IR. While tau oligomers were not discussed in the main text, tau fibrils as well as Aβ42 oligomers and fibrils were reported to exhibit unexpected β-structure signatures in some cases, indicating a significant variability in structure on the single-molecule level, in contrast to common presumption.…”