FIGURE 1.

A region beyond the AO7 RING finger is required for high-affinity UbcH5B binding. A, schematic of human AO7 (above). Components of the AO7 clamp are indicated in white (L, Linker). Sequence of AO7RE (below) with Zn²⁺-coordinating residues and structural features noted in “Results”; disordered regions are indicated with dashed lines. B, binding of in vitro transcribed and translated ³⁵S-labeled UbcH5B to N-terminal GST fusions of AO7 prebound to GS. GST fusions were provided in excess relative to UbcH5B, and each was present in equimolar amounts as determined by Coomassie Blue. C, UbcH5B-Green modified with FlAsH was added to increasing amounts of AO7 from which GST had been cleaved. Binding was assessed by MST.