Exploration of Insulin Amyloid Polymorphism Using Raman Spectroscopy and Imaging

Abstract

Our results revealed three Raman markers distinguishing amyloid fibril polymorphisms caused by salt and temperature effects; the relative proportion of protein secondary structures (α-helix and β-sheet), the ratio of tyrosine doublet, and the conformational differences of disulfide bonds.The lower values of tyrosine doublet in the case with salts were interpreted as the anions rob the hydration water from proteins which induced protein misfolding. Using these parameters, Raman images captured their higher order structural differences in situ without labeling. The images of hydrogen bonds strength variations due to tyrosine doublet is believed to include significant novelty. The present results imply the potential of Raman imaging for use as a diagnostic imaging tool for tissues with amyloid-induced diseases.

Unraveling the NaCl Concentration Effect on the First Stages of α-Synuclein Aggregation

“…More generally, the monomeric structures of several disease-related amyloidogenic proteins other than α-synuclein are also known to be affected by NaCl concentration, which in turn leads to altered aggregation pathways. Those proteins include Aβ(1–40), , IAPP, β 2 -microglobulin, , insulin, or the antimicrobial peptide uperin 3.5 . The suggested underlying mechanisms seem to be highly diverse, ranging from the aforementioned Debye–Hückel screening, specific ion–protein interactions and solvent-structure changes to the induction of specific secondary structure elements.…”

Conformational fingerprinting of tau variants and strains by Raman spectroscopy

“…60 It has been established that unique tau conformers exist in different tauopathies, [4][5][6][7][8]13,61 and that conformers from disease differ to those formed in vitro, 62,63 possibly due to differences in the available cofactor, 13 PTM, 19 local pH or ion environment. 23,25 Emerging evidence suggests that different populations of soluble tau aggregates may exist between different AD patients and that the presence of a given population may correlate with disease onset and progression. 20 We have shown that changes in the physiochemical environment can lead to the formation of distinct tau conformers, reinforcing the possibility that different conformers may also occur in different disease settings.…”

“… 24 Recently Raman spectroscopy was used to identify conformational polymorphism of insulin amyloid fibrils in different buffer conditions. 25 It is yet to be determined whether fibril polymorphism, aggregation cofactor, PTM or primary sequence have an impact on the Raman spectrum of tau aggregates.…”

Amyloid fibrils in superstructures – local ordering revealed by polarization analysis of two-photon excited autofluorescence

“…33 Moreover, in situ Raman imaging of amyloid aggregates by M. Ishigaki et al confirmed the presence of various polymorphs within insulin spherulite-like structures, based on the analysis of the relative proportion of β-sheet structures, the intensity ratio of the tyrosine doublet and the signal intensity arising from the disulfide bonds. 34 A drastic drop of the ThT fluorescence intensity near the core suggests that even though mature fibrils may be present there, the dye is not capable of properly staining them. We hypothesize that the aforementioned limitation may arise from the lack of accessibility of ThT molecules to fibrils formed de novo inside the core, or due to distortion of the binding sites of polymorphs formed in the initial state of superstructure growth.…”