scholarly journals Conformational landscape of the p28-bound human proteasome regulatory particle

2016 ◽  
Author(s):  
Ying Lu ◽  
Jiayi Wu ◽  
Shuobing Chen ◽  
Shuangwu Sun ◽  
Yong-Bei Ma ◽  
...  
Keyword(s):  
Core Particle ◽  
Conformational Landscape ◽  
Regulatory Particle ◽  
Aaa Atpases

SUMMARYThe proteasome holoenzyme is activated by its regulatory particle (RP) consisting of two subcomplexes, the lid and the base. A key event in base assembly is the formation of a heterohexameric ring of AAA-ATPases, which is guided by at least four RP assembly chaperones in mammals: PAAF1, p28/gankyrin, p27/PSMD9 and S5b. We determined a cryo-EM structure of the human RP in complex with its assembly chaperone p28 at 4.5-Å resolution. The Rpn1-p28-AAA subcomplex in the p28-bound RP is highly dynamic and was resolved to subnanometer resolution in seven states, which recapitulate the conformational landscape of the complex. Surprisingly, the p28-bound AAA ring does not form a channel in the free RP. Instead, it spontaneously samples multiple ‘open’ and ‘closed’ topologies. Our analysis suggests that p28 guides the proteolytic core particle to select certain conformation of the ATPase ring for RP engagement in the last step of the chaperone-mediated proteasome assembly.

scholarly journals Maturation of the proteasome core particle induces an affinity switch that controls regulatory particle association

2015 ◽  
Vol 6 (1) ◽  
Author(s):  
Prashant S. Wani ◽  
Michael A. Rowland ◽  
Alex Ondracek ◽  
Eric J. Deeds ◽  
Jeroen Roelofs
Keyword(s):  
Core Particle ◽  
Regulatory Particle ◽  
Particle Association

scholarly journals Proteasomes: unfoldase-assisted protein degradation machines

2019 ◽  
Vol 401 (1) ◽  
pp. 183-199 ◽  
Author(s):  
Parijat Majumder ◽  
Wolfgang Baumeister
Keyword(s):  
Stress Response ◽  
Molecular Machines ◽  
Molecular Architecture ◽  
Core Particle ◽  
Protein Substrates ◽  
Macromolecular Assemblies ◽  
Current State ◽  
Intracellular Proteins ◽  
Domains Of Life ◽  
Aaa Atpases

Abstract Proteasomes are the principal molecular machines for the regulated degradation of intracellular proteins. These self-compartmentalized macromolecular assemblies selectively degrade misfolded, mistranslated, damaged or otherwise unwanted proteins, and play a pivotal role in the maintenance of cellular proteostasis, in stress response, and numerous other processes of vital importance. Whereas the molecular architecture of the proteasome core particle (CP) is universally conserved, the unfoldase modules vary in overall structure, subunit complexity, and regulatory principles. Proteasomal unfoldases are AAA+ ATPases (ATPases associated with a variety of cellular activities) that unfold protein substrates, and translocate them into the CP for degradation. In this review, we summarize the current state of knowledge about proteasome – unfoldase systems in bacteria, archaea, and eukaryotes, the three domains of life.

scholarly journals Ubiquitinated proteins promote the association of proteasomes with the deubiquitinating enzyme Usp14 and the ubiquitin ligase Ube3c

2017 ◽  
Vol 114 (17) ◽  
pp. E3404-E3413 ◽  
Author(s):  
Chueh-Ling Kuo ◽  
Alfred Lewis Goldberg
Keyword(s):  
Ubiquitin Ligase ◽  
Mammalian Cells ◽  
Proteasome Activity ◽  
Deubiquitinating Enzyme ◽  
Core Particle ◽  
Ubiquitinated Proteins ◽  
Protein Ubiquitination ◽  
Cell Extracts ◽  
Regulatory Particle ◽  
In Cells

In mammalian cells, the 26S proteasomes vary in composition. In addition to the standard 28 subunits in the 20S core particle and 19 subunits in each 19S regulatory particle, a small fraction (about 10–20% in our preparations) also contains the deubiquitinating enzyme Usp14/Ubp6, which regulates proteasome activity, and the ubiquitin ligase, Ube3c/Hul5, which enhances proteasomal processivity. When degradation of ubiquitinated proteins in cells was inhibited, levels of Usp14 and Ube3c on proteasomes increased within minutes. Conversely, when protein ubiquitination was prevented, or when purified proteasomes hydrolyzed the associated ubiquitin conjugates, Usp14 and Ube3c dissociated rapidly (unlike other 26S subunits), but the inhibitor ubiquitin aldehyde slowed their dissociation. Recombinant Usp14 associated with purified proteasomes preferentially if they contained ubiquitin conjugates. In cells or extracts, adding Usp14 inhibitors (IU-1 or ubiquitin aldehyde) enhanced Usp14 and Ube3c binding further. Thus, in the substrate- or the inhibitor-bound conformations, Usp14 showed higher affinity for proteasomes and surprisingly enhanced Ube3c binding. Moreover, adding ubiquitinated proteins to cell extracts stimulated proteasome binding of both enzymes. Thus, Usp14 and Ube3c cycle together on and off proteasomes, and the presence of ubiquitinated substrates promotes their association. This mechanism enables proteasome activity to adapt to the supply of substrates.

scholarly journals Structural Defects in the Regulatory Particle-Core Particle Interface of the Proteasome Induce a Novel Proteasome Stress Response

2011 ◽  
Vol 286 (42) ◽  
pp. 36652-36666 ◽  
Author(s):  
Soyeon Park ◽  
Woong Kim ◽  
Geng Tian ◽  
Steven P. Gygi ◽  
Daniel Finley
Keyword(s):  
Stress Response ◽  
Structural Defects ◽  
Core Particle ◽  
Regulatory Particle

scholarly journals Conformational Landscape of the p28-Bound Human Proteasome Regulatory Particle

2017 ◽  
Vol 67 (2) ◽  
pp. 322-333.e6 ◽  
Author(s):  
Ying Lu ◽  
Jiayi Wu ◽  
Yuanchen Dong ◽  
Shuobing Chen ◽  
Shuangwu Sun ◽  
...  
Keyword(s):  
Conformational Landscape ◽  
Regulatory Particle

Chaperone-driven proteasome assembly

2008 ◽  
Vol 36 (5) ◽  
pp. 807-812 ◽  
Author(s):  
Rina Rosenzweig ◽  
Michael H. Glickman
Keyword(s):  
26S Proteasome ◽  
Biologically Active ◽  
Core Particle ◽  
Regulatory Particle ◽  
Particle Attachment

Assembly of the 34-subunit, 2.5 MDa 26S proteasome is a carefully choreographed intricate process. It starts with formation of a seven-membered α-ring that serves as a template for assembly of the complementary β-ring-forming ‘half-proteasomes’. Dimerization results in a latent 20S core particle that can serve further as a platform for 19S regulatory particle attachment and formation of the biologically active 26S proteasome for ubiquitin-dependent proteolysis. Both general and dedicated proteasome assembly chaperones regulate the efficiency and outcome of critical steps in proteasome biogenesis, and in complex association.

scholarly journals Assembly manual for the proteasome regulatory particle: the first draft

2010 ◽  
Vol 38 (1) ◽  
pp. 6-13 ◽  
Author(s):  
Soyeon Park ◽  
Geng Tian ◽  
Jeroen Roelofs ◽  
Daniel Finley
Keyword(s):  
Biochemical Study ◽  
Release Mechanism ◽  
Substrate Selection ◽  
Core Particle ◽  
Genetic Studies ◽  
Macromolecular Assembly ◽  
Assembly Pathway ◽  
Ongoing Work ◽  
The Common ◽  
Regulatory Particle

The proteasome is the most complex protease known, with a molecular mass of approx. 3 MDa and 33 distinct subunits. Recent studies reported the discovery of four chaperones that promote the assembly of a 19-subunit subcomplex of the proteasome known as the regulatory particle, or RP. These and other findings define a new and highly unusual macromolecular assembly pathway. The RP mediates substrate selection by the proteasome and injects substrates into the CP (core particle) to be degraded. A heterohexameric ring of ATPases, the Rpt proteins, is critical for RP function. These ATPases abut the CP and their C-terminal tails help to stabilize the RP–CP interface. ATPase heterodimers bound to the chaperone proteins are early intermediates in assembly of the ATPase ring. The four chaperones have the common feature of binding the C-domains of Rpt proteins, apparently a remarkable example of convergent evolution; each chaperone binds a specific Rpt subunit. The C-domains are distinct from the C-terminal tails, but are proximal to them. Some, but probably not all, of the RP chaperones appear to compete with CP for binding of the Rpt proteins, as a result of the proximity of the tails to the C-domain. This competition may underlie the release mechanism for these chaperones. Genetic studies in yeast point to the importance of the interaction between the CP and the Rpt tails in assembly, and a recent biochemical study in mammals suggests that RP assembly takes place on pre-assembled CP. These results do not exclude a parallel CP-independent pathway of assembly. Ongoing work should soon clarify the roles of both the CP and the four chaperones in RP assembly.

scholarly journals Affinity Switch during Proteasome Core Particle Maturation that Regulates Pba1–Pba2 and Regulatory Particle Association

2015 ◽  
Vol 29 (S1) ◽  
Author(s):  
Prashant Wani ◽  
Michael Rowland ◽  
Alex Ondracek ◽  
Eric Deeds ◽  
Jeroen Roelofs
Keyword(s):  
Core Particle ◽  
Regulatory Particle ◽  
Particle Association

scholarly journals The Regulatory Particle of the Saccharomyces cerevisiae Proteasome

1998 ◽  
Vol 18 (6) ◽  
pp. 3149-3162 ◽  
Author(s):  
Michael H. Glickman ◽  
David M. Rubin ◽  
Victor A. Fried ◽  
Daniel Finley
Keyword(s):  
Active Sites ◽  
Sequence Similarity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Core Particle ◽  
Deubiquitinating Enzymes ◽  
Electrophoretic Variants ◽  
Regulatory Particle ◽  
The Individual ◽  
Limited Sequence

ABSTRACT The proteasome is a multisubunit protease responsible for degrading proteins conjugated to ubiquitin. The 670-kDa core particle of the proteasome contains the proteolytic active sites, which face an interior chamber within the particle and are thus protected from the cytoplasm. The entry of substrates into this chamber is thought to be governed by the regulatory particle of the proteasome, which covers the presumed channels leading into the interior of the core particle. We have resolved native yeast proteasomes into two electrophoretic variants and have shown that these represent core particles capped with one or two regulatory particles. To determine the subunit composition of the regulatory particle, yeast proteasomes were purified and analyzed by gradient sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Resolution of the individual polypeptides revealed 17 distinct proteins, whose identities were determined by amino acid sequence analysis. Six of the subunits have sequence features of ATPases (Rpt1 to Rpt6). Affinity chromatography was used to purify regulatory particles from various strains, each of which expressed one of the ATPases tagged with hexahistidine. In all cases, multiple untagged ATPases copurified, indicating that the ATPases assembled together into a heteromeric complex. Of the remaining 11 subunits that we have identified (Rpn1 to Rpn3 and Rpn5 to Rpn12), 8 are encoded by previously described genes and 3 are encoded by genes not previously characterized for yeasts. One of the previously unidentified subunits exhibits limited sequence similarity with deubiquitinating enzymes. Overall, regulatory particles from yeasts and mammals are remarkably similar, suggesting that the specific mechanistic features of the proteasome have been closely conserved over the course of evolution.

scholarly journals Structural basis for dynamic regulation of the human 26S proteasome

2016 ◽  
Vol 113 (46) ◽  
pp. 12991-12996 ◽  
Author(s):  
Shuobing Chen ◽  
Jiayi Wu ◽  
Ying Lu ◽  
Yong-Bei Ma ◽  
Byung-Hoon Lee ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Ground State ◽  
Atp Hydrolysis ◽  
26S Proteasome ◽  
Structural Basis ◽  
Core Particle ◽  
Dynamic Regulation ◽  
Aaa Atpase ◽  
The Core ◽  
Aaa Atpases

The proteasome is the major engine of protein degradation in all eukaryotic cells. At the heart of this machine is a heterohexameric ring of AAA (ATPases associated with diverse cellular activities) proteins that unfolds ubiquitylated target proteins that are concurrently translocated into a proteolytic chamber and degraded into peptides. Using cryoelectron microscopy, we determined a near–atomic-resolution structure of the 2.5-MDa human proteasome in its ground state, as well as subnanometer-resolution structures of the holoenzyme in three alternative conformational states. The substrate-unfolding AAA-ATPase channel is narrowed by 10 inward-facing pore loops arranged into two helices that run in parallel with each other, one hydrophobic in character and the other highly charged. The gate of the core particle was unexpectedly found closed in the ground state and open in only one of the alternative states. Coordinated, stepwise conformational changes of the regulatory particle couple ATP hydrolysis to substrate translocation and regulate gating of the core particle, leading to processive degradation.

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