+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-10586 | |||||||||
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Title | Rat 20S proteasome | |||||||||
Map data | endogenous rat liver 20S proteasome, local resolution filtered | |||||||||
Sample |
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Keywords | PROTEASOME / HYDROLASE | |||||||||
Function / homology | Function and homology information Cross-presentation of soluble exogenous antigens (endosomes) / SCF-beta-TrCP mediated degradation of Emi1 / Regulation of ornithine decarboxylase (ODC) / Assembly of the pre-replicative complex / GSK3B and BTRC:CUL1-mediated-degradation of NFE2L2 / Oxygen-dependent proline hydroxylation of Hypoxia-inducible Factor Alpha / Autodegradation of Cdh1 by Cdh1:APC/C / APC/C:Cdc20 mediated degradation of Securin / APC/C:Cdh1 mediated degradation of Cdc20 and other APC/C:Cdh1 targeted proteins in late mitosis/early G1 / Cdc20:Phospho-APC/C mediated degradation of Cyclin A ...Cross-presentation of soluble exogenous antigens (endosomes) / SCF-beta-TrCP mediated degradation of Emi1 / Regulation of ornithine decarboxylase (ODC) / Assembly of the pre-replicative complex / GSK3B and BTRC:CUL1-mediated-degradation of NFE2L2 / Oxygen-dependent proline hydroxylation of Hypoxia-inducible Factor Alpha / Autodegradation of Cdh1 by Cdh1:APC/C / APC/C:Cdc20 mediated degradation of Securin / APC/C:Cdh1 mediated degradation of Cdc20 and other APC/C:Cdh1 targeted proteins in late mitosis/early G1 / Cdc20:Phospho-APC/C mediated degradation of Cyclin A / SCF(Skp2)-mediated degradation of p27/p21 / Autodegradation of the E3 ubiquitin ligase COP1 / Asymmetric localization of PCP proteins / Degradation of DVL / Hedgehog ligand biogenesis / Dectin-1 mediated noncanonical NF-kB signaling / Degradation of GLI1 by the proteasome / Hedgehog 'on' state / TNFR2 non-canonical NF-kB pathway / NIK-->noncanonical NF-kB signaling / UCH proteinases / CDK-mediated phosphorylation and removal of Cdc6 / G2/M Checkpoints / Ubiquitin Mediated Degradation of Phosphorylated Cdc25A / Ubiquitin-dependent degradation of Cyclin D / The role of GTSE1 in G2/M progression after G2 checkpoint / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / RUNX1 regulates transcription of genes involved in differentiation of HSCs / Regulation of RUNX3 expression and activity / GLI3 is processed to GLI3R by the proteasome / Activation of NF-kappaB in B cells / Degradation of beta-catenin by the destruction complex / Degradation of AXIN / Regulation of RAS by GAPs / Orc1 removal from chromatin / Neddylation / AUF1 (hnRNP D0) binds and destabilizes mRNA / MAPK6/MAPK4 signaling / Regulation of PTEN stability and activity / KEAP1-NFE2L2 pathway / Separation of Sister Chromatids / Antigen processing: Ubiquitination & Proteasome degradation / ABC-family proteins mediated transport / Ub-specific processing proteases / proteasome core complex / Neutrophil degranulation / immune system process / myofibril / 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 / response to organonitrogen compound / sarcomere / proteasome complex / ciliary basal body / proteolysis involved in protein catabolic process / lipopolysaccharide binding / P-body / response to virus / response to organic cyclic compound / nuclear matrix / peptidase activity / positive regulation of NF-kappaB transcription factor activity / postsynapse / proteasome-mediated ubiquitin-dependent protein catabolic process / endopeptidase activity / response to oxidative stress / ribosome / centrosome / synapse / ubiquitin protein ligase binding / proteolysis / RNA binding / nucleoplasm / identical protein binding / nucleus / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Rattus norvegicus (Norway rat) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.7 Å | |||||||||
Authors | Deshmukh FK / Polkinghorn CR | |||||||||
Funding support | Israel, 2 items
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Citation | Journal: ACS Cent Sci / Year: 2020 Title: Comparative Structural Analysis of 20S Proteasome Ortholog Protein Complexes by Native Mass Spectrometry. Authors: Shay Vimer / Gili Ben-Nissan / David Morgenstern / Fanindra Kumar-Deshmukh / Caley Polkinghorn / Royston S Quintyn / Yury V Vasil'ev / Joseph S Beckman / Nadav Elad / Vicki H Wysocki / Michal Sharon / Abstract: Ortholog protein complexes are responsible for equivalent functions in different organisms. However, during evolution, each organism adapts to meet its physiological needs and the environmental ...Ortholog protein complexes are responsible for equivalent functions in different organisms. However, during evolution, each organism adapts to meet its physiological needs and the environmental challenges imposed by its niche. This selection pressure leads to structural diversity in protein complexes, which are often difficult to specify, especially in the absence of high-resolution structures. Here, we describe a multilevel experimental approach based on native mass spectrometry (MS) tools for elucidating the structural preservation and variations among highly related protein complexes. The 20S proteasome, an essential protein degradation machinery, served as our model system, wherein we examined five complexes isolated from different organisms. We show that throughout evolution, from the archaeal prokaryotic complex to the eukaryotic 20S proteasomes in yeast () and mammals (rat - , rabbit - and human - HEK293 cells), the proteasome increased both in size and stability. Native MS structural signatures of the rat and rabbit 20S proteasomes, which heretofore lacked high-resolution, three-dimensional structures, highly resembled that of the human complex. Using cryoelectron microscopy single-particle analysis, we were able to obtain a high-resolution structure of the rat 20S proteasome, allowing us to validate the MS-based results. Our study also revealed that the yeast complex, and not those in mammals, was the largest in size and displayed the greatest degree of kinetic stability. Moreover, we also identified a new proteoform of the PSMA7 subunit that resides within the rat and rabbit complexes, which to our knowledge have not been previously described. Altogether, our strategy enables elucidation of the unique structural properties of protein complexes that are highly similar to one another, a framework that is valid not only to ortholog protein complexes, but also for other highly related protein assemblies. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_10586.map.gz | 146.5 MB | EMDB map data format | |
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Header (meta data) | emd-10586-v30.xml emd-10586.xml | 32.5 KB 32.5 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_10586_fsc.xml | 14.2 KB | Display | FSC data file |
Images | emd_10586.png | 203.6 KB | ||
Masks | emd_10586_msk_1.map | 244.1 MB | Mask map | |
Filedesc metadata | emd-10586.cif.gz | 8.4 KB | ||
Others | emd_10586_half_map_1.map.gz emd_10586_half_map_2.map.gz | 192.3 MB 192.4 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-10586 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-10586 | HTTPS FTP |
-Validation report
Summary document | emd_10586_validation.pdf.gz | 462.7 KB | Display | EMDB validaton report |
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Full document | emd_10586_full_validation.pdf.gz | 461.9 KB | Display | |
Data in XML | emd_10586_validation.xml.gz | 20 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-10586 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-10586 | HTTPS FTP |
-Related structure data
Related structure data | 6tu3MC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_10586.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | endogenous rat liver 20S proteasome, local resolution filtered | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.86 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_10586_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: Rat 20S proteasome, Unfiltered half map 2
File | emd_10586_half_map_1.map | ||||||||||||
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Annotation | Rat 20S proteasome, Unfiltered half map 2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Rat 20S proteasome, Unfiltered half map 1
File | emd_10586_half_map_2.map | ||||||||||||
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Annotation | Rat 20S proteasome, Unfiltered half map 1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
+Entire : Rat liver 20S proteasome
+Supramolecule #1: Rat liver 20S proteasome
+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
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 14 mg/mL |
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Buffer | pH: 7.4 / Component - Concentration: 160.0 mM / Component - Formula: K3PO4 / Component - Name: Potassium phosphate |
Grid | Model: C-flat-2/2 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.052000000000000005 kPa Details: Grids were glow discharged 30 minutes before applying the sample. |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: blot for 3 seconds. |
Details | Purified endogenous rat liver 20S proteasome |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Details | Preliminary grid screening was performed on Talos Arctica |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 2234 / Average exposure time: 1.5 sec. / Average electron dose: 37.0 e/Å2 Details: Images were collected in movie-mode at 45 frames per second |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Illumination mode: OTHER / Imaging mode: DIFFRACTION / Cs: 2.7 mm / Nominal defocus max: 1.6 µm / Nominal defocus min: 0.6 µm / Nominal magnification: 105000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Refinement | Protocol: FLEXIBLE FIT |
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Output model | PDB-6tu3: |