Fig. 2: I1 has inter- and intra- molecular (AP) β-strands and is a tetramer.

A ¹⁵N-¹⁵N correlation spectrum of I1 with uniformly ¹⁵N- labelled αS shows that there are AP segments (pink labels) which feature intra- and inter-molecular cross-peaks. Cross-peaks from PIR segments (green labels), correlations in loops (grey labels) and helices (black labels) are also observed. On the right are one-dimensional traces of cross-peaks comparing uniformly ¹⁵N- labelled I1 (black) and 1:1 ¹⁵N: unlabelled (50% dilute) I1 (blue). Intra-molecular cross-peaks are not affected by the 50% dilution (V63-V55 contact) whereas the intensity of inter-molecular cross-peaks is reduced 2-fold in the diluted spectrum (blue) (V66-V52 and K45-T44 contacts). B Contacts indicative of AP segments are indicated with pink lines on a backbone trace. C Ratios of cross-peak intensity between the fully ¹⁵N-labeled (black) and 50% ¹⁵N-labeled (blue) spectra. Color legend is the same as in panel (A). D Histograms of the number of photobleaching steps (purple) and number of polarization states per aggregate (pink) for I1 at a fluorophore labelling efficiency (p) of 25%. The grey line shows the best fit to the photobleaching and polarCOLD data with parameters p = 27% and m (no. of monomers per oligomer) =4 (Supplementary Fig. 4I). Error bars represent standard error of the mean calculated from 1421 particles (over 5 different fields of view (FOV)) for the photobleaching experiment and 1023 particles (over 3 different FOV) for the polarCOLD experiment. Probabilities for individual FOVs are shown in circles. Source data are provided as a Source Data file. E–H Examples of super-resolved polarCOLD images of different particles show the projection of aggregates with 1 (E), 2 (F), 3 (G) and 4 (H) dye molecules onto the imaging plane. The center of each spot displays the position of the fluorophore and its width represents the localization precision. The latter can vary due to the available signal-to-noise in each case, determined by the photophysics heterogeneity of fluorophores (Supplementary Fig. 4H). The number of particles with states in panel E-F can be determined from the polarCOLD histogram in panel D and the source data provided in the Source Data file.