Structural basis for the activation of 20S proteasomes by 11S regulators

Abstract

Most of the non-lysosomal proteolysis that occurs in eukaryotic cells is performed by a nonspecific and abundant barrel-shaped complex called the 20S proteasome. Substrates access the active sites, which are sequestered in an internal chamber, by traversing a narrow opening (alpha-annulus) that is blocked in the unliganded 20S proteasome by amino-terminal sequences of alpha-subunits. Peptide products probably exit the 20S proteasome through the same opening. 11S regulators (also called PA26 (ref. 4), PA28 (ref. 5) and REG) are heptamers that stimulate 20S proteasome peptidase activity in vitro and may facilitate product release in vivo. Here we report the co-crystal structure of yeast 20S proteasome with the 11S regulator from Trypanosoma brucei (PA26). PA26 carboxy-terminal tails provide binding affinity by inserting into pockets on the 20S proteasome, and PA26 activation loops induce conformational changes in alpha-subunits that open the gate separating the proteasome interior from the intracellular environment. The reduction in processivity expected for an open conformation of the exit gate may explain the role of 11S regulators in the production of ligands for major histocompatibility complex class I molecules.

Concurrent Translocation of Multiple Polypeptide Chains through the Proteasomal Degradation Channel

“…Crystal structures of the yeast proteasome core particle in complex with the caps show that the axial pore has a diameter of ϳ13 Å (11,12,14). The degradation channel in the archebacterial proteasome has restrictions that are smaller than 20 Å as judged by the crystal structure (6) and the observation that a gold particle with a diameter of ϳ 20Å attached to a substrate protein could not enter the archebacterial proteasome (42).…”

“…At its narrowest point, the opened degradation channel is ϳ13 Å wide (11,12). This constriction is too small for folded proteins to fit through it, and substrate proteins must unfold to gain access to the proteolytic sites.…”

“…The channel opens when the regulatory caps bind to the core (12). The caps consist of 17 subunits, six of which have ATPase activity, and contact the core particle through the ATPase subunits (13,14).At its narrowest point, the opened degradation channel is ϳ13 Å wide (11,12). This constriction is too small for folded proteins to fit through it, and substrate proteins must unfold to gain access to the proteolytic sites.…”

Structural Insights into the Conformational Diversity of ClpP from Bacillus subtilis

“…HslU, ClpA/ClpX and the 19S regulatory cap utilize a domain(s) for recognizing substrate proteins and delivering these in an unfolded form, generated by ATP hydrolysis, to the matching proteases (Bochtler et al, 1999;Park and Song, 2008;Park et al, 2007;Sauer and Baker, 2011;Song and Eck, 2003). Structural studies of HslVU complex, ClpP in complex with activator acyldepsipeptide (ADEP), the 20S proteasome in complex with the 11S proteasome activator PA28, and the Cterminal peptide of proteasome-activating nucleosidase (PAN) have revealed the activation mechanism of the protease components (Lee et al, 2010a;Li et al, 2010;Rabl et al, 2008;Sousa et al, 2000;Stadtmueller and Hill, 2011;Whitby et al, 2000). As already mentioned, the cylindrical shape of the protease component has two substrate entry pores at both ends doubly capped by two AAA+ ATPases.…”

Novel proteases: common themes and surprising features

“…Although this structure corresponds to only one half of DegP, it suggests that substrate access to the active sites is also regulated by the PDZ domains. The structure of the yeast 20S proteasome in complex with the 11S regulator from Trypanosoma brucei [12] shows how egress of substrates may be regulated. The 11S regulators bind to the apical parts of the 20S proteasome, inducing a conformational change in the α subunits of the 20S proteasome that creates a more open conformation.…”