Extended Data Fig. 5: Three-dimensional reconstruction and model building of the DICER(D991G/H992G)–26S-GU complex.
From: DICER cleavage fidelity is governed by 5′-end binding pockets
a, Overview of cryo-EM data collection and processing. Movies were motion-corrected, CTF-estimated, and subjected to particle picking, 2D/3D classification, non-uniform refinement, and CryoTEN map sharpening. The final reconstruction used 787,381 particles and reached 3.29 Å. b, Representative micrograph and selected 2D class averages of the dicing-state complex. c, Front and side views of the 3D density map showing the overall protein-RNA architecture. d, GS-FSC curve. The global resolution at the 0.143 criterion is 3.29 Å. e, Local-resolution estimates, ranging from ~2.5 Å in rigid regions to ~10 Å in flexible portions; front and side views illustrate spatial variation. f, Particle orientation heat map showing the distribution of particle orientations used in the 3D reconstruction. g, Map-model fit. The sharpened density (mesh) overlaid with the atomic model, with domains labelled: Platform, PAZ, Connector helix, RIIIDa, RIIIDb, and dsRBD; RNA (26S-GU, light blue). h, Close-up of the 5′-terminal G1 adjacent to the boundary loop and the G-pocket mutations (G991, G992; positions of D991/H992 mutated to Gly). i, Docking of the duplex relative to the boundary loop (red) and the DC21/DC22 pockets (open circles). The 26S-GU RNA is shown in blue. j, View of the 5′-end highlighting G1 and U62 positioned near the boundary loop. k, 3′-end pocket in the PAZ domain showing A64 and its contacts with surrounding residues.
