Supplementary Figure 9: Mutation analysis of TDP-43 fibril structures.

a, b, Residues tested in mutagenesis study in (top) SegA-sym and (bottom) SegB A315E. Residues that reduce pathological fragment TDP-CTF aggregation when mutated are shown in magenta and those that do not influence TDP-CTF aggregation when mutated are shown in green. Arg293 was mutated to glutamate and others were mutated to tryptophan. In (b), the models of both structures are shown in sticks and semitransparent surface mode. Notice that Ala324, Leu330, Gln331 and Met337 in SegA-sym and Gln303 in SegB A315E (also see Supplementary Figure 10) are in a tightly packed environment that does not tolerant a tryptophan mutation, consistent with our previous mutagenesis study where these mutations delayed the aggregation of the pathogenic TDP-43 fragment. In contrast, the Ala326 in SegA-sym and Gly304 in SegB A315E are in a relatively loose environment that is able to accommodate the tryptophan mutations, consistent with the result that A326W and G304W did not delay aggregation of the pathogenic TDP-43 fragment. For the G304W mutation, a plausible model of the R-shaped fold with G304W is shown in a separated panel. G304 from the inner chain and outer chain of SegB A315E were computationally mutated to tryptophan residues and phenix.real_space_refine was used to generate a model with acceptable rotamers and Ramachandran angles (Supplementary Table 3 and Supplementary Data Set 1). The mutated model is shown in color and the non-mutated model is shown in gray. This model helps explain the tolerance of the G304W mutation in our previous study. c, Synthetic TDP-43 peptide 274–313 and 314–353 were reported to seed TDP-43 aggregation in cells, and the overlapping region of these two peptides in SegA-sym (left, 314–347) and SegB A315E (right, 288–313) structures are shown in color, whereas missing residues from the synthetic peptides are shown in grey. Notice that these two peptides preserve most of the dagger- and R-shaped folds making it likely that they adopt the same structures we report here. d, Positions of known hereditary mutations. The mutations compatible with the dagger- or R-shaped fold are colored black; the mutations potentially disruptive are colored red; the mutations favorable are bolded black.