Fig. 3: UBE2R2’s UBC domain assists the synergy loop to recruit the acceptor ubiquitin.
From: Mechanism of millisecond Lys48-linked poly-ubiquitin chain formation by cullin-RING ligases
a, Ribbon diagram of the chain formation complex highlighting the UBE2R2–acceptor ubiquitin (UBA) interface and the positions of key residues and their side chains mediating the interaction. Cryo-EM density from the composite map is shown. b, Bar graphs comparing the Km values for the indicated proteins of unanchored UBA for UBE2R2 in the presence of neddylated CUL2–RBX1. Compensatory mutations in UBE2R2 and UBA re-establish the interface, reducing Km values to near WT. Bars showing a ‘>’ reflect reactions in which saturation of UBE2R2 with UBA was not possible (the top concentration in the dilution series is shown). The value of each bar represents the estimated value for Km based on n = 3 technical replicates. c, Schematic showing the assay used to estimate the rates of chain extension onto ubiquitin-primed CRL substrates by UBE2R-family E2s (also see Extended Data Fig. 5d for an illustration of quench flow operation). SR, substrate receptor; S, substrate; R, RBX1, UBD, donor ubiquitin. d, Bar graph comparing the rates (kobs) of donor ubiquitin transfer from UBE2R2 to ubiquitin-primed Sil1 peptide substrate for the indicated proteins. The value of each bar represents the estimated value of kobs based on n = 3 technical replicates.
