Extended Data Fig. 4: Cryo-EM processing of the BRISC-inhibitor co-complex.
From: Molecular glues that inhibit deubiquitylase activity and inflammatory signaling
Figures a–c correspond to the BRISC-FX-171-C cryo-EM dataset. Figures d–f correspond to the BRISC-JMS-175-2 dataset. a, d, Representative micrographs (from 16,750 (FX-171-C) and 7,771 movies (JMS-175-2)) and 2D class averages. b, e, Image processing workflow. Green maps indicate selected classes used for 3D refinement. c, f, Left, cryo-EM density maps after 3D refinement for the final reconstructions used for model building. Asterisks indicate BLUE compound binding sites. Right, final maps with corresponding Euler angular distribution with rod heights proportional to the number of particles in each direction. FSC curves with resolution calculated using the gold standard FSC cut-off at 0.143 frequency. g, Mask used for focused refinement of the BRISC-FX-171-C map. h, Chemical structure of FX-171-C fitted into EM density before (left) and after (right) focused refinement. Cryo-EM density visualised using the surface zone tool in ChimeraX; left, radius 2.04, right, radius 2.60. i, Mask applied during refinement of BRISC-JMS-175-2 map. j, Chemical structure of JMS-175-2 fitted into EM density before (left) and after (right) focused refinement. Cryo-EM density visualised using the surface zone tool in ChimeraX; left, radius 2.20, right, radius 2.41. k, Overlay of two BRISC dimers aligned on one BRISC molecule (grey) for comparison. Models are represented as surfaces. Orange, model fitted to the BRISC-FX-171-C structure shown in c, yellow, BRISC models rigid-body fitted in the cryo-EM density of the asymmetric dimer shown in Extended Data Fig. 2g. The yellow molecule is shifted relative to the orange molecule. l, Models described in k, focussed on the small molecule binding site highlighting the shift in the BRCC45’ α6 and α10 helices.
