A Single α Helix Drives Extensive Remodeling of the Proteasome Lid and Completion of Regulatory Particle Assembly

Summary Most short-lived eukaryotic proteins are degraded by the proteasome. A proteolytic core particle (CP) capped by regulatory particles (RPs) constitutes the 26S proteasome complex. RP biogenesis culminates with the joining of two large subcomplexes, the lid and base. In yeast and mammals, the lid appears to assemble completely before attaching to the base, but how this hierarchical assembly is enforced has remained unclear. Using biochemical reconstitutions, quantitative cross-linking/mass spectrometry, and electron microscopy, we resolve the mechanistic basis for the linkage between lid biogenesis and lid-base joining. Assimilation of the final lid subunit, Rpn12, triggers a large-scale conformational remodeling of the nascent lid that drives RP assembly, in part by relieving steric clash with the base. Surprisingly, this remodeling is triggered by a single Rpn12 α helix. Such assembly-coupled conformational switching is reminiscent of viral particle maturation and may represent a commonly used mechanism to enforce hierarchical assembly in multisubunit complexes.
Graphical Abstract
Highlights • First in vitro reconstitution of RP assembly with completely recombinant components • Electron microscopy and cross-linking reveal massive remodeling of a lid precursor • Remodeling of the lid relieves steric clash with the RP base to promote RP assembly • Lid remodeling can be triggered by a single C-terminal α helix in the Rpn12 subunit
A single alpha helix from the final subunit that incorporates into the proteasomal lid triggers a large-scale conformational switch that enables subsequent assembly of the lid and base, suggesting a general paradigm for hierarchical assembly of macromolecular complexes similar to that of virus particles.