Extended Data Fig. 2: Cryo-EM structure determination of TNFR1-DD, TRADD-DD and RIPK1-DD ternary super complex.
From: Electric dipole moment drives the dynamics of the TNFR1 complex I signalosome
a, Flow chart for the first attempt of cryo-EM structure determination of co-expressed TNFR1-DD/TRADD-DD/RIPK1-DD ternary complex, which failed to yield high resolution density map. b, Flow charts for the purification of co-expressed TNFR1-DD/TRADD-DD/RIPK1-DD complexes without and with K596E RIPK1-DD treatment. c-d, Size-exclusion chromatographs and SDS-PAGE analysis of complexes purified as in b. e, Size-exclusion chromatographs of GB1-tagged WT RIPK1-DD and K596E RIPK1-DD protein, whose molecular weight was determined by size-exclusion chromatography coupled with multi-angle static light scattering. f, Distribution of the length of filaments of TNFR1-DD/TRADD-DD/RIPK1-DD complexes purified as procedure in b. Data are presented as mean ± SEM for 100 filaments in each group. g, Flow chart for the cryo-EM structure determination of co-expressed TNFR1-DD/TRADD-DD/RIPK1-DD ternary complex treated with K596E RIPK1-DD. h, The FSC curve calculated between two half maps of the refined final density volume. Results are representative of three independent experiments for (c-f). The P-value in f is derived from a two-tailed Welch’s t test for comparison between two groups and presented as * (P < 0.05), ** (P < 0.01), *** (P < 0.001), **** (P < 0.0001).
