In the proteasome (), polypeptides are digested to short peptides, 90% of which range between 2 and 10 residues in length (). Nearly all are digested in seconds to amino acids by cytosolic peptidases, but in mammals some serve as precursors for antigenic peptides displayed on MHC-class I molecules. Because proteolysis is irreversible, the consequences for cells can be severe if proteasomes destroy proteins non-selectively or function non-processively and release partially degraded polypeptides. Therefore, proteasomes have evolved intricate mechanisms to avoid such failures and to ensure efficient, selective degradation.

Genetic and biochemical studies have greatly advanced our understanding of the multiple steps in proteasomal degradation: the binding of ubiquitylated proteins, their deubiquitylation, and their ATP-driven unfolding and translocation into the 20S chamber for proteolysis (). Although the roles of many of its 60-odd subunits and associated proteins are still unclear, dramatic progress has been made recently through cryo-EM in capturing the dynamism of the 26S complex. The goal of this article is not to summarize these various advances, as has been done in many valuable reviews (; ; ; ; ; ; ). Instead, we build on many of these discoveries to try to understand the inherent logic of proteasome function and how its catalytic and regulatory features promote efficient and selective proteolysis. This analysis also highlights important gaps in our understanding that merit further study.

The ability of the proteasome to recognize both a Ub chain and a loosely folded region provides the fundamental basis for how it determines which proteins to degrade and which to spare. This critical life-or-death decision can be explained by the discovery of two types of conjugate binding: 1) an initial, reversible step in which the Ub chain undergoes high affinity binding to receptors on the 26S particle, and 2) a subsequent tighter-binding step that depends on the ubiquitylated protein's structure and requires ATP hydrolysis (). This sequence and the “dwell-time” of the substrate on the 26S provide an opportunity for competing processes to determine the protein's fate. On one hand, the proteasome's multiple de-ubiquitylation enzymes (DUBs) () shorten the substrate's dwell-time and promote the release of some, perhaps many, of the ubiquitylated proteins that initially bind (). However, if the substrate becomes tightly bound through its loosely folded domain, the six proteasome ATPases are activated (), and the substrate becomes committed to the steps leading to its destruction —further deubiquitylation, unfolding, processive translocation, and hydrolysis to small peptides in the 20S core particle.

In addition to these inherent forms of regulation, cells contain many mechanisms that modulate 26S activity (). Several protein kinases can rapidly enhance 26S activities and stimulate the breakdown of ubiquitylated proteins in vivo. Moreover, the association of activating proteins and complexes seem to enable the proteasome to serve specialized roles e.g. when proteolysis increases in proteotoxic stress () or muscle atrophy ().