The 1.9 Å Structure of a Proteasome-11S Activator Complex and Implications for Proteasome-PAN/PA700 Interactions

Förster, A.; Masters, E.I.; Whitby, Frank G.; Robinson, Howard; Hill, Christopher P.

doi:10.1016/j.molcel.2005.04.016

Cited by 231 publications

(398 citation statements)

References 51 publications

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“…Near-axial contacts may coordinate substrate translocation through the central pore of the unfoldase ring and into the degradation chamber of the peptidase. Crystal structures of 20S with the heptameric ATP-independent activator PA26 also show peripheral interactions between C-terminal tails and pockets in the α ring as well as interactions between an internal activation loop in PA26 and the near-axial gating residues of the α ring (17), further supporting the possibility that bipartite recognition is a universal feature of functional interactions with the 20S peptidase.…”

Section: Discussionmentioning

confidence: 84%

Bipartite determinants mediate an evolutionarily conserved interaction between Cdc48 and the 20 S peptidase

Barthelme

Sauer

2013

Proc. Natl. Acad. Sci. U.S.A.

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Proteasomes are essential and ubiquitous ATP-dependent proteases that function in eukarya, archaea, and some bacteria. These destructive but critically important proteolytic machines use a 20S core peptidase and a hexameric ATPase associated with a variety of cellular activities (AAA+) unfolding ring that unfolds and spools substrates into the peptidase chamber. In archaea, 20S can function with the AAA+ Cdc48 or proteasome-activating nucleotidase (PAN) unfoldases. Both interactions are stabilized by C-terminal tripeptides in AAA+ subunits that dock into pockets on the 20S periphery. Here, we provide evidence that archaeal Cdc48 also uses a distinct set of near-axial interactions to bind 20S and propose that similar dual determinants mediate PAN-20S interactions and Rpt 1-6 -20S interactions in the 26S proteasome. Current dogma holds that the Rpt 1-6 unfolding ring of the 19S regulatory particle is the only AAA+ partner of eukaryotic 20S. By contrast, we show that mammalian Cdc48, a key player in cell-cycle regulation, membrane fusion, and endoplasmic-reticulum-associated degradation, activates mammalian 20S and find that a mouse Cdc48 variant supports protein degradation in combination with 20S. Our results suggest that eukaryotic Cdc48 orthologs function directly with 20S to maintain intracellular protein quality control. AAA+ protease | p97A AA+ (ATPase associated with a variety of cellular activities) enzymes use cycles of ATP binding and hydrolysis to exert mechanical forces on substrates and power a diverse array of biological functions (1). In all cells, protein-unfolding machines of the AAA+ family form hexameric rings that cooperate with associated peptidases to execute intracellular protein degradation (2). The proteolytic sites of AAA+ proteases are sequestered within the interior of a barrel-shaped, self-compartmentalized peptidase, accessible only through narrow axial entrance tunnels that exclude folded proteins. Peptidase access is controlled by the AAA+ unfoldase, which docks with the peptidase, pulls the native structure of target proteins apart, and then translocates the polypeptide through its axial channel and into the degradation chamber.The 20S peptidase is the degradation module for proteasomes in all domains of life, but functions with different AAA+ unfoldases, including the Rpt 1-6 unfolding ring of the eukaryotic 26S proteasome, the double-ring Cdc48 or single-ring proteasomeactivating nucleotidase (PAN) enzymes in archaeal proteasomes, and the Mpa unfoldase in Actinobacteria (Fig. 1A) (3-6). In all organisms, the 20S peptidase has a four-ring α 7 β 7 β 7 α 7 structure and uses N-terminal residues of the α subunits to gate entrance of substrates into its degradation chamber (Fig. 1B) (5, 7-9).The subunits of the AAA+ ring have C-terminal tripeptides that dock into conserved pockets on the peptidase α ring and help stabilize an open-gate conformation to allow 20S degradation of proteins or large peptides (Fig. 1B) (10). These tripeptides generally match a hydrophobic, tyrosine, any resi...

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Section: Discussionmentioning

confidence: 84%

Bipartite determinants mediate an evolutionarily conserved interaction between Cdc48 and the 20 S peptidase

Barthelme

Sauer

2013

Proc. Natl. Acad. Sci. U.S.A.

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“…2B). We then introduced a cysteine (L745C) at the X position of the C-terminal HbYX tripeptide of Cdc48*, because these residues are hypothesized to dock into a binding cleft in the 20S α ring (8,20). We introduced another cysteine (K66C) into the hydrophobic cleft of the α-subunit of 20S*.…”

Section: Resultsmentioning

confidence: 99%

“…Residues suitable for cross-linking Cdc48 to 20S were identified by analyzing PA26-bound 20S structures (PDB ID codes 3JRM, 1YA7, and 3IPM) using the program Disulfide by Design (20,(29)(30)(31). For EM structural analyses, we cross-linked Cdc48 */ΔN/L745C to 20S */K66C and crosslinked Cdc48 */L745C to 20S* bearing αK33C and βT1A mutations.…”

Section: Methodsmentioning

confidence: 99%

Architecture and assembly of the archaeal Cdc48⋅20S proteasome

Barthelme¹,

Chen²,

Grabenstatter

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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ATP-dependent proteases maintain protein quality control and regulate diverse intracellular functions. Proteasomes are primarily responsible for these tasks in the archaeal and eukaryotic domains of life. Even the simplest of these proteases function as large complexes, consisting of the 20S peptidase, a barrel-like structure composed of four heptameric rings, and one or two AAA+ (ATPase associated with a variety of cellular activities) ring hexamers, which use cycles of ATP binding and hydrolysis to unfold and translocate substrates into the 20S proteolytic chamber. Understanding how the AAA+ and 20S components of these enzymes interact and collaborate to execute protein degradation is important, but the highly dynamic nature of prokaryotic proteasomes has hampered structural characterization. Here, we use electron microscopy to determine the architecture of an archaeal Cdc48·20S proteasome, which we stabilized by site-specific cross-linking. This complex displays coaxial alignment of Cdc48 and 20S and is enzymatically active, demonstrating that AAA+ unfoldase wobbling with respect to 20S is not required for function. In the complex, the N-terminal domain of Cdc48, which regulates ATP hydrolysis and degradation, packs against the D1 ring of Cdc48 in a coplanar fashion, constraining mechanisms by which the N-terminal domain alters 20S affinity and degradation activity.AAA+ protease | dynamic wobbling model | p97/VCP P roteasomes are large macromolecular complexes that degrade misfolded or damaged proteins to maintain cellular homeostasis and quality control in all domains of life. In addition, selective proteasomal turnover of regulatory proteins is often a critical element of signaling cascades that allow cells to respond to changing conditions and environmental stress (1, 2). Proteasomes consist of the self-compartmentalized 20S peptidase and one or two AAA+ (ATPase associated with a variety of cellular activities) family ring hexamers. The α 7 β 7 β 7 α 7 ring topology of the 20S enzyme ensures that the proteolytic active sites, which reside in the β-subunits, are sequestered and can only cleave substrates that enter the chamber through narrow axial pores formed by the α-subunits (3). As a consequence, ATP-dependent unfolding of natively folded protein substrates by the AAA+ ring and subsequent polypeptide translocation through a narrow axial channel and into the 20S chamber are required for degradation (4).Although the 20S peptidase is the degradation module in all proteasomes, the associated AAA+ unfolding machines differ. For example, homohexamers of Mpa/Arc in actinobacteria and PAN in archaea serve as proteasomal motors, whereas a heterohexameric Rpt 1-6 ring in the 19S particle is the motor of the eukaryotic 26S proteasome (2, 5). Characteristic of type-I AAA+ enzymes, Mpa/Arc, PAN, and Rpt 1-6 subunits contain a single AAA+ module for ATP binding and hydrolysis (6). By contrast, as found in type-II AAA+ enzymes, the homohexameric Cdc48 motor in the recently discovered archaeal Cdc48·20S proteasome cont...

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“…Therefore, the means by which the 20S proteasome gate is opened by either the 19S ATPases or 11S activators appears to be quite different. Indeed, while the 11S activators induce gate opening without a major conformational change to the 20S proteasome α-subunits, 14 the 19S ATPases do so with a large 4° rotation of the entire α-ring. 15 It is conceivable, therefore, that β-sheet-rich PrP acts to inhibit the 20S proteasome by stabilizing its un-rotated, closed-gate conformation, which would reduce gate opening by the 19S ATPases, but not gate opening without α-subunit rotation by the 11S activators.…”

Section: Misfolded Prp and A Novel Mechanism Of Proteasome Inhibitionmentioning

confidence: 99%

Misfolded PrP and a novel mechanism of proteasome inhibition

André

Tabrizi

2012

Prion

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The 1.9 Å Structure of a Proteasome-11S Activator Complex and Implications for Proteasome-PAN/PA700 Interactions

Cited by 231 publications

References 51 publications

Bipartite determinants mediate an evolutionarily conserved interaction between Cdc48 and the 20 S peptidase

Bipartite determinants mediate an evolutionarily conserved interaction between Cdc48 and the 20 S peptidase

Architecture and assembly of the archaeal Cdc48⋅20S proteasome

Misfolded PrP and a novel mechanism of proteasome inhibition

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