Characterization of Homogeneous and Heterogeneous Amyloid-β42 Oligomer Preparations with Biochemical Methods and Infrared Spectroscopy Reveals a Correlation between Infrared Spectrum and Oligomer Size

Abstract

Soluble oligomers of the amyloid-β(1-42) (Aβ42) peptide, widely considered to be among the relevant neurotoxic species involved in Alzheimer’s disease, were characterized with a combination of biochemical and biophysical methods. Homogeneous and stable Aβ42 oligomers were prepared by treating monomeric solutions of the peptide with detergents. The prepared oligomeric solutions were analyzed with blue native and sodium dodecyl sulfate polyacrylamide gel electrophoresis, as well as with infrared (IR) spectroscopy. The IR spectra indicated a well-defined β-sheet structure of the prepared oligomers. We also found a relationship between the size/molecular weight of the Aβ42 oligomers and their IR spectra: The position of the main amide I′ band of the peptide backbone correlated with oligomer size, with larger oligomers being associated with lower wavenumbers. This relationship explained the time-dependent band shift observed in time-resolved IR studies of Aβ42 aggregation in the absence of detergents, during which the oligomer size increased. In addition, the bandwidth of the main IR band in the amide I′ region was found to become narrower with time in our time-resolved aggregation experiments, indicating a more homogeneous absorption of the β-sheets of the oligomers after several hours of aggregation. This is predominantly due to the consumption of smaller oligomers in the aggregation process.

Lithium ions display weak interaction with amyloid-beta (Aβ) peptides and have minor effects on their aggregation

“…This split band pattern is indicative of an anti-parallel β-sheet conformation (Barth, 2007;Cerf et al, 2009;Sarroukh et al, 2013). Earlier studies in our laboratory have shown a relationship between the position and width of this main band, and the size and homogeneity of the Aβ 42 oligomers (Vosough & Barth, 2021). The lower band position of the larger oligomers is in line with this relationship and our previous results, and confirms the different sizes of the oligomers produced at the two SDS concentrations.…”

“…The lower band position of the larger oligomers is in line with this relationship and our previous results, and confirms the different sizes of the oligomers produced at the two SDS concentrations. We have recently observed that a number of transition metal ions induce significant effects on the main band position for Aβ 42 oligomers (Vosough & Barth, 2021). Because the spectra for Aβ 42 oligomers formed with different amounts of LiCl generally superimpose on the IR spectra for the Li + -free oligomers, with no shifts observed for the main band, it appears that Li + ions have no significant effect on the oligomers' size or secondary structure.…”

“…In 0.2% (6.9 mM) SDS, small oligomers with a molecular weight (MW) around 16-20 kDa are formed. These oligomers appear to contain a large fraction of tetramers (Vosough & Barth, 2021). In 0.05% (1.7 mM) SDS, larger oligomers with MWs around 55-60 kD are formed (Barghorn et al, 2005).…”

“…The peptide fractions were flash-frozen in liquid nitrogen, covered with argon gas on top in 1.5 mL Eppendorf tubes, and stored at -80°C until used. Sodium dodecyl sulfate (SDS)-stabilized Aβ 42 oligomers of two well-defined sizes (approximately tetramers and dodecamers) were prepared according to a previously published protocol (Barghorn et al, 2005), but in D 2 O, at 4-fold lower peptide concentration and without the original dilution step (Vosough & Barth, 2021). The reaction mixtures (100 µM Aβ 42 in PBS and containing 0.05% or 0.2% SDS) were incubated together with 0-10 mM LiCl at 37°C for 24 hours, and then flash-frozen in liquid nitrogen and stored at -20°C for later analysis.…”

“…Homogeneous solutions of 80 µM Aβ 42 oligomers (Vosough & Barth, 2021) prepared in presence and absence of 0-10 mM Li + ions were analyzed with blue native polyacrylamide gel electrophoresis (BN-PAGE) using the Invitrogen system. 4-16% Bis-Tris Novex gels (ThermoFisher Scientific, USA) were loaded with 10 µL of Aβ 42 oligomer samples alongside the Amersham High Molecular Weight calibration kit for native electrophoresis (GE Healthcare, USA).…”