Structural Snapshots of 26S Proteasome Reveal Tetraubiquitin-Induced Conformations

The 26S proteasome is the ATP-dependent protease responsible for regulating the proteome of eukaryotic cells through degradation of mainly ubiquitin-tagged substrates. In order to understand how proteasome responds to ubiquitin signal, we resolved an ensemble of cryo-EM structures of proteasome in the presence of K48-Ub4, with three of them resolved at near-atomic resolution. We identified a conformation with stabilized ubiquitin receptors and a previously unreported orientation of the lid, assigned as a Ub-accepted state C1-b. We determined another structure C3-b with localized K48-Ub4 to the toroid region of Rpn1, assigned as a substrate-processing state. Our structures indicate that tetraUb induced conformational changes in proteasome could initiate substrate degradation. We also propose a CP gate-opening mechanism involving the propagation of the motion of the lid to the gate through the Rpn6-α2 interaction. Our results enabled us to put forward a model of a functional cycle for proteasomes induced by tetraUb and nucleotide. [Display omitted] •TetraUb transduces proteasome conformations from lid through base to gate opening•C1-b has stabilized Ub receptors and a previously unreported lid orientation•C3-b has a novel tetraUb density localized to the toroid region of Rpn1•TetraUb, Rad23 (UBL domain), and Ubp6/USP14 can co-occupy the proteasome Cells contain millions of proteasomes to degrade abnormal proteins to maintain homeostasis. Understanding the degradation mechanism will benefit therapeutic intervention. Ding et al. report a series of proteasome structures representing the functional cycle of proteasome in action and put forward a mechanistic model of proteasome degradation induced by tetraUb.