Fig. 3.

Probing brain-derived Aβ40 fibrils for population A with solid-state NMR. Two-dimensional ¹³C DARR NMR measurements of fibrils derived from sCAA and fCAA-Dutch patients. The fibrils were produced via templated growth of brain amyloid using specifically ¹³C-labeled Aβ40 to probe inter-residue contacts modeled in the cryo-EM structures. (a) F4-F20 interaction. A scheme of the backbone trace of population A (left) highlighting the close intramolecular contact of F4 and F20. Region of the 2D DARR NMR spectrum (right) exhibiting cross peaks between 1-¹³C F4 and ring-¹³C-F20 (or F19). The 1D row extracted from the 2D spectrum passes through the diagonal resonance of 1-¹³C F4 and highlights the cross peak between 1-¹³C F4 and ring-¹³C-F20 (or F19). The ¹³C labeling scheme chosen for these measurements generally provides well-resolved resonances such that cross-peaks can be assigned to specific residue contacts. One exception is that the F19 and F20 ¹³C labels overlap but are predicted to interact with either G33 or F4, respectively. The presence of the F4-F19/20 cross peak (square) is consistent with the close distance between F4 and F20 in the cryo-EM structure in both the sCAA and fCAA-Dutch patient samples. We also observe a cross peak between F19 and G33 (not shown), which is consistent with the population A structure. Note that the cross peak intensities in DARR NMR spectra are often not symmetric (see Ohashi and Takegoshi (2006)). (b) L17-M35 interaction. A scheme of the backbone trace of population A (left) highlights the close intermolecular contact of L17 and M35, while the 2D DARR NMR spectrum (right) exhibits cross peaks between 2-¹³C L17 and 5-¹³C M35. The 1D row extracted from the 2D spectrum passes through the diagonal resonance of 5-¹³C M35 and highlights the cross peak between 5-¹³C M35 and 2-¹³C L17. Cross peaks (asterisks) are also observed between 5-¹³C M35 and 2-¹³C G33, residues that are close together in the Aβ sequence. However, on the basis of the two structures presented here, these cross peaks may arise from the β-hairpins populating the outer densities in population B (see below). The population A structure indicates a register shift with respect to a previously published similar structure, leading to a flip of the C-terminal segment. This shift results in a close proximity between L17 and M35 consistent with the presence of observed cross-peaks in the DARR NMR spectra of the sCAA and fCAA-Dutch patient fibrils.