Structure, Dynamics and Function of the 26S Proteasome.

The 26S proteasome is the most complex ATP-dependent protease machinery, of ~2.5 MDa mass, ubiquitously found in all eukaryotes. It selectively degrades ubiquitin-conjugated proteins and plays fundamentally indispensable roles in regulating almost all major aspects of cellular activities. To serve as the sole terminal "processor" for myriad ubiquitylation pathways, the proteasome evolved exceptional adaptability in dynamically organizing a large network of proteins, including ubiquitin receptors, shuttle factors, deubiquitinases, AAA-ATPase unfoldases, and ubiquitin ligases, to enable substrate selectivity and processing efficiency and to achieve regulation precision of a vast diversity of substrates. The inner working of the 26S proteasome is among the most sophisticated, enigmatic mechanisms of enzyme machinery in eukaryotic cells. Recent breakthroughs in three-dimensional atomic-level visualization of the 26S proteasome dynamics during polyubiquitylated substrate degradation elucidated an extensively detailed picture of its functional mechanisms, owing to progressive methodological advances associated with cryogenic electron microscopy (cryo-EM). Multiple sites of ubiquitin binding in the proteasome revealed a canonical mode of ubiquitin-dependent substrate engagement. The proteasome conformation in the act of substrate deubiquitylation provided insights into how the deubiquitylating activity of RPN11 is enhanced in the holoenzyme and is coupled to substrate translocation. Intriguingly, three principal modes of coordinated ATP hydrolysis in the heterohexameric AAA-ATPase motor  were discovered to regulate intermediate functional steps of the proteasome, including ubiquitin-substrate engagement, deubiquitylation, initiation of substrate translocation and processive substrate degradation. The atomic dissection of the innermost working of the 26S proteasome opens up a new era in our understanding of the ubiquitin-proteasome system and has far-reaching implications in health and disease.