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- PDB-8qym: Human 20S proteasome assembly intermediate structure 3 -

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Basic information

Entry
Database: PDB / ID: 8qym
TitleHuman 20S proteasome assembly intermediate structure 3
Components
  • (Proteasome assembly chaperone ...) x 2
  • (Proteasome subunit alpha type- ...) x 7
  • (Proteasome subunit beta type- ...) x 3
  • Proteasome maturation protein
KeywordsHYDROLASE / Complex
Function / homology
Function and homology information


cerebellar granule cell precursor proliferation / purine ribonucleoside triphosphate binding / regulation of endopeptidase activity / protein folding chaperone complex / Regulation of ornithine decarboxylase (ODC) / proteasome core complex / Cross-presentation of soluble exogenous antigens (endosomes) / : / proteasome core complex assembly / Somitogenesis ...cerebellar granule cell precursor proliferation / purine ribonucleoside triphosphate binding / regulation of endopeptidase activity / protein folding chaperone complex / Regulation of ornithine decarboxylase (ODC) / proteasome core complex / Cross-presentation of soluble exogenous antigens (endosomes) / : / proteasome core complex assembly / Somitogenesis / myofibril / immune system process / proteasome binding / mitotic spindle assembly checkpoint signaling / NF-kappaB binding / chaperone-mediated protein complex assembly / proteasome assembly / proteasome endopeptidase complex / proteasome core complex, beta-subunit complex / proteasome core complex, alpha-subunit complex / threonine-type endopeptidase activity / sarcomere / proteasome complex / Regulation of activated PAK-2p34 by proteasome mediated degradation / proteolysis involved in protein catabolic process / Autodegradation of Cdh1 by Cdh1:APC/C / APC/C:Cdc20 mediated degradation of Securin / Asymmetric localization of PCP proteins / SCF-beta-TrCP mediated degradation of Emi1 / NIK-->noncanonical NF-kB signaling / Ubiquitin-dependent degradation of Cyclin D / AUF1 (hnRNP D0) binds and destabilizes mRNA / TNFR2 non-canonical NF-kB pathway / Vpu mediated degradation of CD4 / Assembly of the pre-replicative complex / Degradation of DVL / Ubiquitin Mediated Degradation of Phosphorylated Cdc25A / Cdc20:Phospho-APC/C mediated degradation of Cyclin A / Dectin-1 mediated noncanonical NF-kB signaling / negative regulation of inflammatory response to antigenic stimulus / lipopolysaccharide binding / Hh mutants are degraded by ERAD / Degradation of AXIN / Activation of NF-kappaB in B cells / Degradation of GLI1 by the proteasome / Hedgehog ligand biogenesis / G2/M Checkpoints / Defective CFTR causes cystic fibrosis / Negative regulation of NOTCH4 signaling / GSK3B and BTRC:CUL1-mediated-degradation of NFE2L2 / P-body / Autodegradation of the E3 ubiquitin ligase COP1 / Vif-mediated degradation of APOBEC3G / Regulation of RUNX3 expression and activity / Hedgehog 'on' state / Degradation of GLI2 by the proteasome / GLI3 is processed to GLI3R by the proteasome / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / MAPK6/MAPK4 signaling / APC/C:Cdh1 mediated degradation of Cdc20 and other APC/C:Cdh1 targeted proteins in late mitosis/early G1 / response to virus / Degradation of beta-catenin by the destruction complex / Oxygen-dependent proline hydroxylation of Hypoxia-inducible Factor Alpha / ABC-family proteins mediated transport / response to organic cyclic compound / CDK-mediated phosphorylation and removal of Cdc6 / CLEC7A (Dectin-1) signaling / SCF(Skp2)-mediated degradation of p27/p21 / Regulation of expression of SLITs and ROBOs / nuclear matrix / FCERI mediated NF-kB activation / Regulation of PTEN stability and activity / Interleukin-1 signaling / Orc1 removal from chromatin / Regulation of RAS by GAPs / Separation of Sister Chromatids / Regulation of RUNX2 expression and activity / The role of GTSE1 in G2/M progression after G2 checkpoint / UCH proteinases / KEAP1-NFE2L2 pathway / Antigen processing: Ubiquitination & Proteasome degradation / Downstream TCR signaling / positive regulation of NF-kappaB transcription factor activity / RUNX1 regulates transcription of genes involved in differentiation of HSCs / Neddylation / ER-Phagosome pathway / regulation of inflammatory response / secretory granule lumen / proteasome-mediated ubiquitin-dependent protein catabolic process / endopeptidase activity / ficolin-1-rich granule lumen / postsynapse / molecular adaptor activity / nuclear body / Ub-specific processing proteases / ribosome / nuclear speck / intracellular membrane-bounded organelle / centrosome / ubiquitin protein ligase binding
Similarity search - Function
Proteasome assembly chaperone 1 / Proteasome assembly chaperone 4 / Proteasome maturation factor Ump1 / Proteasome maturation factor UMP1 / Proteasome assembly chaperone 2, eukaryotic / Proteasome assembly chaperone 2 / Proteasome assembly chaperone 2 superfamily / PAC2 family / Proteasome subunit alpha 1 / Proteasome beta subunit, C-terminal ...Proteasome assembly chaperone 1 / Proteasome assembly chaperone 4 / Proteasome maturation factor Ump1 / Proteasome maturation factor UMP1 / Proteasome assembly chaperone 2, eukaryotic / Proteasome assembly chaperone 2 / Proteasome assembly chaperone 2 superfamily / PAC2 family / Proteasome subunit alpha 1 / Proteasome beta subunit, C-terminal / Proteasome beta subunits C terminal / Proteasome subunit beta 2 / Proteasome beta 3 subunit / Proteasome subunit alpha6 / Proteasome subunit alpha5 / Proteasome beta-type subunits signature. / Peptidase T1A, proteasome beta-subunit / Proteasome beta-type subunit, conserved site / Proteasome subunit A N-terminal signature / Proteasome alpha-type subunits signature. / Proteasome alpha-subunit, N-terminal domain / Proteasome subunit A N-terminal signature Add an annotation / : / Proteasome alpha-type subunit / Proteasome alpha-type subunit profile. / Proteasome B-type subunit / Proteasome beta-type subunit profile. / Proteasome subunit / Proteasome, subunit alpha/beta / Nucleophile aminohydrolases, N-terminal
Similarity search - Domain/homology
Proteasome subunit alpha type-7 / Proteasome assembly chaperone 1 / Proteasome subunit alpha type-1 / Proteasome subunit alpha type-2 / Proteasome subunit alpha type-3 / Proteasome subunit alpha type-4 / Proteasome subunit alpha type-5 / Proteasome subunit beta type-3 / Proteasome subunit beta type-2 / Proteasome subunit alpha type-6 ...Proteasome subunit alpha type-7 / Proteasome assembly chaperone 1 / Proteasome subunit alpha type-1 / Proteasome subunit alpha type-2 / Proteasome subunit alpha type-3 / Proteasome subunit alpha type-4 / Proteasome subunit alpha type-5 / Proteasome subunit beta type-3 / Proteasome subunit beta type-2 / Proteasome subunit alpha type-6 / Proteasome assembly chaperone 2 / Proteasome subunit beta type-7 / Proteasome maturation protein
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.73 Å
AuthorsSchulman, B.A. / Hanna, J.W. / Harper, J.W. / Adolf, F. / Du, J. / Rawson, S.D. / Walsh Jr, R.M. / Goodall, E.A.
Funding support Germany, United States, 4items
OrganizationGrant numberCountry
Max Planck Society Germany
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS) United States
National Institutes of Health/National Institute on Aging (NIH/NIA) United States
Aligning Science Across Parkinsons (ASAP) United States
Citation
Journal: Nat Struct Mol Biol / Year: 2024
Title: Visualizing chaperone-mediated multistep assembly of the human 20S proteasome.
Authors: Frank Adolf / Jiale Du / Ellen A Goodall / Richard M Walsh / Shaun Rawson / Susanne von Gronau / J Wade Harper / John Hanna / Brenda A Schulman /
Abstract: Dedicated assembly factors orchestrate the stepwise production of many molecular machines, including the 28-subunit proteasome core particle (CP) that mediates protein degradation. Here we report ...Dedicated assembly factors orchestrate the stepwise production of many molecular machines, including the 28-subunit proteasome core particle (CP) that mediates protein degradation. Here we report cryo-electron microscopy reconstructions of seven recombinant human subcomplexes that visualize all five chaperones and the three active site propeptides across a wide swath of the assembly pathway. Comparison of these chaperone-bound intermediates and a matching mature CP reveals molecular mechanisms determining the order of successive subunit additions, as well as how proteasome subcomplexes and assembly factors structurally adapt upon progressive subunit incorporation to stabilize intermediates, facilitate the formation of subsequent intermediates and ultimately rearrange to coordinate proteolytic activation with gated access to active sites. This work establishes a methodologic approach for structural analysis of multiprotein complex assembly intermediates, illuminates specific functions of assembly factors and reveals conceptual principles underlying human proteasome biogenesis, thus providing an explanation for many previous biochemical and genetic observations.
#1: Journal: bioRxiv / Year: 2024
Title: Visualizing chaperone-mediated multistep assembly of the human 20S proteasome.
Authors: Frank Adolf / Jiale Du / Ellen A Goodall / Richard M Walsh / Shaun Rawson / Susanne von Gronau / J Wade Harper / John Hanna / Brenda A Schulman /
Abstract: Dedicated assembly factors orchestrate stepwise production of many molecular machines, including the 28-subunit proteasome core particle (CP) that mediates protein degradation. Here, we report cryo- ...Dedicated assembly factors orchestrate stepwise production of many molecular machines, including the 28-subunit proteasome core particle (CP) that mediates protein degradation. Here, we report cryo-EM reconstructions of seven recombinant human subcomplexes that visualize all five chaperones and the three active site propeptides across a wide swath of the assembly pathway. Comparison of these chaperone-bound intermediates and a matching mature CP reveals molecular mechanisms determining the order of successive subunit additions, and how proteasome subcomplexes and assembly factors structurally adapt upon progressive subunit incorporation to stabilize intermediates, facilitate the formation of subsequent intermediates, and ultimately rearrange to coordinate proteolytic activation with gated access to active sites. The structural findings reported here explain many previous biochemical and genetic observations. This work establishes a methodologic approach for structural analysis of multiprotein complex assembly intermediates, illuminates specific functions of assembly factors, and reveals conceptual principles underlying human proteasome biogenesis.
History
DepositionOct 26, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Feb 21, 2024Provider: repository / Type: Initial release
Revision 1.1Mar 6, 2024Group: Database references / Category: citation_author / Item: _citation_author.name
Revision 1.2Apr 17, 2024Group: Database references / Category: citation / citation_author
Revision 1.3Aug 28, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID / _em_admin.last_update
Revision 1.4Nov 13, 2024Group: Data collection / Structure summary
Category: em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Proteasome subunit alpha type-2
B: Proteasome subunit alpha type-4
C: Proteasome subunit alpha type-7
D: Proteasome subunit alpha type-5
E: Proteasome subunit alpha type-1
F: Proteasome subunit alpha type-3
G: Proteasome subunit alpha type-6
H: Proteasome maturation protein
I: Proteasome assembly chaperone 1
J: Proteasome assembly chaperone 2
K: Proteasome subunit beta type-7
L: Proteasome subunit beta type-3
M: Proteasome subunit beta type-2


Theoretical massNumber of molelcules
Total (without water)354,75713
Polymers354,75713
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Proteasome subunit alpha type- ... , 7 types, 7 molecules ABCDEFG

#1: Protein Proteasome subunit alpha type-2 / Macropain subunit C3 / Multicatalytic endopeptidase complex subunit C3 / Proteasome component C3


Mass: 25927.535 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMA2, HC3, PSC3 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P25787
#2: Protein Proteasome subunit alpha type-4 / Macropain subunit C9 / Multicatalytic endopeptidase complex subunit C9 / Proteasome component C9 / ...Macropain subunit C9 / Multicatalytic endopeptidase complex subunit C9 / Proteasome component C9 / Proteasome subunit L


Mass: 29525.842 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMA4, HC9, PSC9 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P25789
#3: Protein Proteasome subunit alpha type-7 / Proteasome subunit RC6-1 / Proteasome subunit XAPC7


Mass: 27929.891 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMA7, HSPC / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: O14818
#4: Protein Proteasome subunit alpha type-5 / Macropain zeta chain / Multicatalytic endopeptidase complex zeta chain / Proteasome zeta chain


Mass: 26462.016 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMA5 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P28066
#5: Protein Proteasome subunit alpha type-1


Mass: 29621.662 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMA1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P25786
#6: Protein Proteasome subunit alpha type-3 / Macropain subunit C8 / Multicatalytic endopeptidase complex subunit C8 / Proteasome component C8


Mass: 28469.252 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMA3, HC8, PSC8 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P25788
#7: Protein Proteasome subunit alpha type-6 / 27 kDa prosomal protein / p27K / Macropain iota chain / Multicatalytic endopeptidase complex iota ...27 kDa prosomal protein / p27K / Macropain iota chain / Multicatalytic endopeptidase complex iota chain / Proteasome iota chain


Mass: 27432.459 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMA6, PROS27 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P60900

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Protein , 1 types, 1 molecules H

#8: Protein Proteasome maturation protein / Proteassemblin / Protein UMP1 homolog / hUMP1 / Voltage-gated K channel beta subunit 4.1


Mass: 15804.993 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: POMP, C13orf12, UMP1, HSPC014, HSPC036, PNAS-110 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9Y244

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Proteasome assembly chaperone ... , 2 types, 2 molecules IJ

#9: Protein Proteasome assembly chaperone 1 / PAC-1 / Chromosome 21 leucine-rich protein / C21-LRP / Down syndrome critical region protein 2


Mass: 32891.887 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMG1, C21LRP, DSCR2, PAC1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: O95456
#10: Protein Proteasome assembly chaperone 2


Mass: 29449.080 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMG2 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q969U7

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Proteasome subunit beta type- ... , 3 types, 3 molecules KLM

#11: Protein Proteasome subunit beta type-7


Mass: 35405.301 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: Where the identity of amino acids cannot be confidently assign, these are modelled as unknown (UNK)
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMB7 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q99436
#12: Protein Proteasome subunit beta type-3 / Proteasome chain 13 / Proteasome component C10-II / Proteasome theta chain


Mass: 22972.896 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMB3 / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: P49720, proteasome endopeptidase complex
#13: Protein Proteasome subunit beta type-2 / Macropain subunit C7-I / Multicatalytic endopeptidase complex subunit C7-I / Proteasome component C7-I


Mass: 22864.277 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PSMB2 / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: P49721, proteasome endopeptidase complex

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Details

Has ligand of interestN
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Human 20S proteasome assembly intermediate map 3 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE-PROPANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2600 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 66.9 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM softwareName: SerialEM / Category: image acquisition
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.73 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 217491 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00621305
ELECTRON MICROSCOPYf_angle_d0.59528947
ELECTRON MICROSCOPYf_dihedral_angle_d4.5533013
ELECTRON MICROSCOPYf_chiral_restr0.0463418
ELECTRON MICROSCOPYf_plane_restr0.0053690

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