Fig. 4: NMR spectroscopy of His-NT2RepCT and NT and fibrillation propensity profile of NT.

a 2D ¹⁵N-HSQC spectra of a 10 mg/ml His-NT2RepCT solution before (blue) and after (red) incubation at 37 °C for 19 h. Isolated cross-peaks in the red spectrum and F24, G136, polyA in the blue spectrum are assigned using one-letter amino acid symbols and residue numbers. The inset shows signal intensity versus time for selected residues from the NT, 2Rep and CT domains. b Solid-state radio-frequency driven recoupling (RFDR) spectra of His-NT2RepCT hydrogels. Cα/Cβ correlations of residues observed in the RFDR spectrum determined by comparison with the chemical shifts of model peptides and values obtained from statistical data^82,83 and their secondary structure are indicated. SSB – spinning side band. c 1D ¹⁵N-HSQC spectra of a 10 mg/ml NT solution during incubation at 37 °C for 36 h. The inset shows bulk intensity versus time. d Solid-state RFDR spectra of NT hydrogels. Cα/Cβ correlations of the residues observed in the RFDR spectrum and their secondary structure are indicated. e Fibrillation propensity profile of NT^45,79 according to the Zipper database (https://services.mbi.ucla.edu/zipperdb/). Rosetta energies in kcal/mol of moving windows of hexapeptide steric zippers are shown. Red bars indicate hexapeptides with high fibrillation propensities (Rosetta energies below −23 kcal/mol; below dotted line). Green bars indicate Rosetta energies above the threshold which hence are segments that are unlikely to form steric zippers. Segments containing proline are omitted from the analysis (no bars). The square indicates a predicted amyloidogenic region according to the Waltz algorithm⁸¹, (https://waltz.switchlab.org). Amino acid residue sequence of NT on top with residue types found in β secondary structure (as determined by solid-state NMR spectroscopy) indicated in red. The positions of the five α-helices of NT are indicated (H1-H5)²⁸.