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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-4860 | |||||||||
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Title | Human 20S-PA200 Proteasome Complex | |||||||||
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![]() | proteasome / PA200 / activator / HYDROLASE | |||||||||
Function / homology | ![]() spermatoproteasome complex / sperm DNA condensation / peptidase activator activity / purine ribonucleoside triphosphate binding / regulation of endopeptidase activity / Regulation of ornithine decarboxylase (ODC) / proteasome core complex / Cross-presentation of soluble exogenous antigens (endosomes) / Somitogenesis / proteasomal ubiquitin-independent protein catabolic process ...spermatoproteasome complex / sperm DNA condensation / peptidase activator activity / purine ribonucleoside triphosphate binding / regulation of endopeptidase activity / Regulation of ornithine decarboxylase (ODC) / proteasome core complex / Cross-presentation of soluble exogenous antigens (endosomes) / Somitogenesis / proteasomal ubiquitin-independent protein catabolic process / immune system process / myofibril / proteasome binding / NF-kappaB binding / proteasome endopeptidase complex / proteasome core complex, beta-subunit complex / proteasome core complex, alpha-subunit complex / threonine-type endopeptidase activity / negative regulation of inflammatory response to antigenic stimulus / lysine-acetylated histone binding / : / sarcomere / proteasome complex / Regulation of activated PAK-2p34 by proteasome mediated degradation / ciliary basal body / 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 / CDK-mediated phosphorylation and removal of Cdc6 / Ubiquitin Mediated Degradation of Phosphorylated Cdc25A / Dectin-1 mediated noncanonical NF-kB signaling / Cdc20:Phospho-APC/C mediated degradation of Cyclin A / Hh mutants are degraded by ERAD / lipopolysaccharide binding / SCF(Skp2)-mediated degradation of p27/p21 / Degradation of AXIN / Degradation of GLI1 by the proteasome / Activation of NF-kappaB in B cells / Hedgehog ligand biogenesis / Defective CFTR causes cystic fibrosis / Negative regulation of NOTCH4 signaling / GSK3B and BTRC:CUL1-mediated-degradation of NFE2L2 / G2/M Checkpoints / Vif-mediated degradation of APOBEC3G / Autodegradation of the E3 ubiquitin ligase COP1 / Hedgehog 'on' state / Regulation of RUNX3 expression and activity / P-body / Degradation of GLI2 by the proteasome / GLI3 is processed to GLI3R by the proteasome / Orc1 removal from chromatin / 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 / ABC-family proteins mediated transport / Oxygen-dependent proline hydroxylation of Hypoxia-inducible Factor Alpha / response to organic cyclic compound / CLEC7A (Dectin-1) signaling / Regulation of expression of SLITs and ROBOs / FCERI mediated NF-kB activation / nuclear matrix / Regulation of PTEN stability and activity / Interleukin-1 signaling / 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 / regulation of inflammatory response / Downstream TCR signaling / RUNX1 regulates transcription of genes involved in differentiation of HSCs / Neddylation / positive regulation of NF-kappaB transcription factor activity / peptidase activity / ER-Phagosome pathway / endopeptidase activity / proteasome-mediated ubiquitin-dependent protein catabolic process / secretory granule lumen / response to oxidative stress / ficolin-1-rich granule lumen / postsynapse / nuclear body / ribosome / Ub-specific processing proteases / nuclear speck / cadherin binding / intracellular membrane-bounded organelle Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.0 Å | |||||||||
![]() | Toste Rego A / da Fonseca PCA | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Characterization of Fully Recombinant Human 20S and 20S-PA200 Proteasome Complexes. Authors: Ana Toste Rêgo / Paula C A da Fonseca / ![]() Abstract: Proteasomes are essential in all eukaryotic cells. However, their function and regulation remain considerably elusive, particularly those of less abundant variants. We demonstrate the human 20S ...Proteasomes are essential in all eukaryotic cells. However, their function and regulation remain considerably elusive, particularly those of less abundant variants. We demonstrate the human 20S proteasome recombinant assembly and confirmed the recombinant complex integrity biochemically and with a 2.6 Å resolution cryo-EM map. To assess its competence to form higher-order assemblies, we prepared and analyzed recombinant human 20S-PA200, a poorly characterized nuclear complex. Its 3.0 Å resolution cryo-EM structure reveals the PA200 unique architecture; the details of its intricate interactions with the proteasome, resulting in unparalleled proteasome α ring rearrangements; and the molecular basis for PA200 allosteric modulation of the proteasome active sites. Non-protein cryo-EM densities could be assigned to PA200-bound inositol phosphates, and we speculate regarding their functional role. Here we open extensive opportunities to study the fundamental properties of the diverse and distinct eukaryotic proteasome variants and to improve proteasome targeting under different therapeutic conditions. | |||||||||
History |
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Structure visualization
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 675.4 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 36.8 KB 36.8 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 19.2 KB | Display | ![]() |
Images | ![]() | 98.2 KB | ||
Filedesc metadata | ![]() | 10.2 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6reyMC ![]() 4877C ![]() 6rgqC M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Voxel size | X=Y=Z: 0.81 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
+Entire : Human 20S-PA200 Proteasome complex
+Supramolecule #1: Human 20S-PA200 Proteasome complex
+Supramolecule #2: 20S Proteasome
+Supramolecule #3: PA200 activator
+Macromolecule #1: Proteasome subunit alpha type-6
+Macromolecule #2: Proteasome subunit alpha type-2
+Macromolecule #3: Proteasome subunit alpha type-4
+Macromolecule #4: Proteasome subunit alpha type-7
+Macromolecule #5: Proteasome subunit alpha type-5
+Macromolecule #6: Proteasome subunit alpha type-1
+Macromolecule #7: Proteasome subunit alpha type-3
+Macromolecule #8: Proteasome subunit beta type-6
+Macromolecule #9: Proteasome subunit beta type-7
+Macromolecule #10: Proteasome subunit beta type-3
+Macromolecule #11: Proteasome subunit beta type-2
+Macromolecule #12: Proteasome subunit beta type-5
+Macromolecule #13: Proteasome subunit beta type-1
+Macromolecule #14: Proteasome subunit beta type-4
+Macromolecule #15: Proteasome activator complex subunit 4
+Macromolecule #16: INOSITOL HEXAKISPHOSPHATE
+Macromolecule #17: [(1~{S},2~{R},3~{R},4~{S},5~{S},6~{R})-2-[oxidanyl(phosphonooxy)p...
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.1 mg/mL | ||||||||||||
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Buffer | pH: 7.4 Component:
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Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR | ||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 295.15 K / Instrument: FEI VITROBOT MARK IV | ||||||||||||
Details | The sample was homogeneous |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number real images: 717 / Average exposure time: 60.0 sec. / Average electron dose: 45.7 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal magnification: 95000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Initial model | PDB ID: Chain - Source name: PDB / Chain - Initial model type: experimental model |
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Details | The model of the human recombinant 20S proteasome was built based on PDB: 5LE5 using real-space refinement in Coot and Phenix |
Refinement | Space: REAL / Protocol: FLEXIBLE FIT |
Output model | ![]() PDB-6rey: |
-Atomic model buiding 2
Details | the model for the human PA200 derived from Phyre2 and I-Tasser. These PA200 models were used as initial guides for the assignment of secondary structure using real-space refinement in Coot and Phenix, followed by correction of the sequence register using the amino acid side chain densities clearly resolved in our cryo-EM map. Connecting loops and all other regions were built ab initio from the cryo-EM density. |
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Refinement | Protocol: AB INITIO MODEL |
Output model | ![]() PDB-6rey: |