Extended Data Fig. 5: Extraordinary cullin–RING conformational changes in catalytic architecture juxtaposing the substrate and the active site of ubiquitylation.
From: NEDD8 nucleates a multivalent cullin–RING–UBE2D ubiquitin ligation assembly
a, Side-by-side comparison of relative RING-domain locations in different CRL complexes after superposition of the C/R domains from the original CUL1–RBX1 structure (PDB: 1LDJ, ‘pre-neddylation’—which data herein show is dynamic—although the crystal structure probably captured the conformation that enabled CAND1 binding and substrate receptor exchange)7, the structure representing the neddylation reaction (PDB: 4P5O)39, and a structure of a neddylated CUL5–RBX1 domain (PDB: 3DQV, labelled ‘post-neddylation’, which revealed the potential for conformational changes in the neddylated CUL WHB- and RBX1 RING-domains3, and data herein shows is dynamic), and the structure presented here showing how the neddylated CUL1 WHB domain and RBX1 RING domain are harnessed in a catalytic architecture for ‘active ubiquitylation’. Trp35 of RBX1 is highlighted to show how it serves as a multifunctional platform for either the RING domain in different orientations, or for the E2-linked NEDD8 during neddylation39. b, Superposition of the structures shown in a, highlighting different relative positions of the RING domain. c, Comparison of the relative locations of the CUL WHB domain in different structures after superimposing their C/R domains (not shown). d, Cryo-EM density from the neddylated CRL1β-TRCP–UBE2D~Ub–substrate intermediate complex, showing patchiness of the region corresponding to CUL1 ‘helix-29’7. This CUL1 region connecting the C/R and WHB domains is visible only as patchy density, whereas in previous cullin crystals this forms the rod-like helix-29 continuing into the WHB domain7. It seems that helix-29 of CUL1 dissolves into a flexible tether, which rationalizes the previously observed proteolytic sensitivity of this region in a neddylated CUL1–RBX1 complex3, and enables the displacement and rotation required for placing the ensuing WHB domain and its linked NEDD8 at the centre of the ubiquitylation complex.
