Priming and extending: how UbcH5 and Cdc34 act to assemble ubiquitin chains on a SCF substrate

Abstract

I will discuss a mechanistic model of polyubiquitination by the SCFβTrCP2 E3 ubiquitin (Ub) ligase using human IκBα as substrate. Biochemical reconstitution experiments revealed that the polyubiquitination of IκBα began with the action of the UbcH5 E2 Ub conjugating enzyme, transferring a single Ub to IκBα K21/K22 rapidly and efficiently. Subsequently, the Cdc34 E2 functioned in the formation of polyubiquitin chains. Our recent work shows that an Ub fused at IκBα K21 acts as a receptor, directing Cdc34 for rapid and efficient K48‐linked Ub chain synthesis that depends on SCFβTrCP2 and the substrate's N terminus. The IκBα‐linked fusion Ub appears to mediate direct contacts with Cdc34 and the SCF's RING sub‐complex. Taken together, these results suggest a role for the multifaceted interactions between the K21/K22‐linked receptor Ub, the SCF's RING complex, and Cdc34~S~Ub in establishing the optimal orientation of the receptor Ub to drive conjugation.