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Yorodumi- PDB-6ef3: Yeast 26S proteasome bound to ubiquitinated substrate (4D motor state) -
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-Basic information
Entry | Database: PDB / ID: 6ef3 | |||||||||
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Title | Yeast 26S proteasome bound to ubiquitinated substrate (4D motor state) | |||||||||
Components |
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Keywords | MOTOR PROTEIN / 26S Proteasome / ATPase / AAA+ / Protease / Ubiquitin | |||||||||
Function / homology | Function and homology information Metalloprotease DUBs / peroxisome fission / proteasome storage granule assembly / proteasome regulatory particle assembly / mitotic cell cycle phase transition / cytosolic proteasome complex / proteasome regulatory particle, lid subcomplex / protein-containing complex localization / proteasome-activating activity / mitochondrial fission ...Metalloprotease DUBs / peroxisome fission / proteasome storage granule assembly / proteasome regulatory particle assembly / mitotic cell cycle phase transition / cytosolic proteasome complex / proteasome regulatory particle, lid subcomplex / protein-containing complex localization / proteasome-activating activity / mitochondrial fission / proteasome regulatory particle, base subcomplex / metal-dependent deubiquitinase activity / nonfunctional rRNA decay / cyclin-dependent protein serine/threonine kinase regulator activity / proteasome core complex assembly / nuclear outer membrane-endoplasmic reticulum membrane network / Cross-presentation of soluble exogenous antigens (endosomes) / TNFR2 non-canonical NF-kB pathway / proteasomal ubiquitin-independent protein catabolic process / Ubiquitin Mediated Degradation of Phosphorylated Cdc25A / Regulation of PTEN stability and activity / peptide catabolic process / KEAP1-NFE2L2 pathway / proteasome binding / CDK-mediated phosphorylation and removal of Cdc6 / Neddylation / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / Orc1 removal from chromatin / MAPK6/MAPK4 signaling / proteasome storage granule / Antigen processing: Ubiquitination & Proteasome degradation / endopeptidase activator activity / ribosomal large subunit export from nucleus / protein deubiquitination / proteasome assembly / positive regulation of RNA polymerase II transcription preinitiation complex assembly / proteasome endopeptidase complex / proteasome core complex, beta-subunit complex / proteasome core complex, alpha-subunit complex / Ub-specific processing proteases / threonine-type endopeptidase activity / ERAD pathway / Neutrophil degranulation / proteasome complex / nucleotide-excision repair / positive regulation of transcription elongation by RNA polymerase II / modification-dependent protein catabolic process / positive regulation of protein catabolic process / protein tag activity / metallopeptidase activity / peroxisome / ribosome biogenesis / ribosomal large subunit assembly / ubiquitin-dependent protein catabolic process / cytoplasmic translation / proteasome-mediated ubiquitin-dependent protein catabolic process / cytosolic large ribosomal subunit / endopeptidase activity / ubiquitinyl hydrolase 1 / cysteine-type deubiquitinase activity / protein ubiquitination / structural constituent of ribosome / chromatin remodeling / protein domain specific binding / cell division / mRNA binding / ubiquitin protein ligase binding / endoplasmic reticulum membrane / ATP hydrolysis activity / mitochondrion / ATP binding / identical protein binding / nucleus / metal ion binding / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Saccharomyces cerevisiae S288c (yeast) Homo sapiens (human) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.17 Å | |||||||||
Authors | de la Pena, A.H. / Goodall, E.A. / Gates, S.N. / Lander, G.C. / Martin, A. | |||||||||
Funding support | United States, 2items
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Citation | Journal: Science / Year: 2018 Title: Substrate-engaged 26 proteasome structures reveal mechanisms for ATP-hydrolysis-driven translocation. Authors: Andres H de la Peña / Ellen A Goodall / Stephanie N Gates / Gabriel C Lander / Andreas Martin / Abstract: The 26 proteasome is the primary eukaryotic degradation machine and thus is critically involved in numerous cellular processes. The heterohexameric adenosine triphosphatase (ATPase) motor of the ...The 26 proteasome is the primary eukaryotic degradation machine and thus is critically involved in numerous cellular processes. The heterohexameric adenosine triphosphatase (ATPase) motor of the proteasome unfolds and translocates targeted protein substrates into the open gate of a proteolytic core while a proteasomal deubiquitinase concomitantly removes substrate-attached ubiquitin chains. However, the mechanisms by which ATP hydrolysis drives the conformational changes responsible for these processes have remained elusive. Here we present the cryo-electron microscopy structures of four distinct conformational states of the actively ATP-hydrolyzing, substrate-engaged 26 proteasome. These structures reveal how mechanical substrate translocation accelerates deubiquitination and how ATP-binding, -hydrolysis, and phosphate-release events are coordinated within the AAA+ (ATPases associated with diverse cellular activities) motor to induce conformational changes and propel the substrate through the central pore. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6ef3.cif.gz | 1010.9 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6ef3.ent.gz | 804.3 KB | Display | PDB format |
PDBx/mmJSON format | 6ef3.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6ef3_validation.pdf.gz | 1.5 MB | Display | wwPDB validaton report |
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Full document | 6ef3_full_validation.pdf.gz | 1.5 MB | Display | |
Data in XML | 6ef3_validation.xml.gz | 143.8 KB | Display | |
Data in CIF | 6ef3_validation.cif.gz | 226.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ef/6ef3 ftp://data.pdbj.org/pub/pdb/validation_reports/ef/6ef3 | HTTPS FTP |
-Related structure data
Related structure data | 9045MC 9042C 9043C 9044C 6ef0C 6ef1C 6ef2C M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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-Components
-Proteasome subunit beta type- ... , 8 types, 8 molecules 1234567n
#1: Protein | Mass: 23573.604 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PRE3, YJL001W, J1407 / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P38624, proteasome endopeptidase complex |
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#2: Protein | Mass: 28299.889 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PUP1, YOR157C / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P25043, proteasome endopeptidase complex |
#3: Protein | Mass: 22627.842 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PUP3, YER094C / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P25451, proteasome endopeptidase complex |
#4: Protein | Mass: 22545.676 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PRE1, YER012W / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P22141, proteasome endopeptidase complex |
#5: Protein | Mass: 31670.539 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PRE2, DOA3, PRG1, YPR103W, P8283.10 / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P30656, proteasome endopeptidase complex |
#6: Protein | Mass: 26905.076 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PRE7, PRS3, PTS1, YBL041W, YBL0407 / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P23724, proteasome endopeptidase complex |
#7: Protein | Mass: 29471.289 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PRE4, YFR050C / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P30657, proteasome endopeptidase complex |
#21: Protein/peptide | Mass: 1789.039 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Gene: PSB7 / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 / References: UniProt: P30657*PLUS |
-Proteasome subunit alpha type- ... , 6 types, 6 molecules ABCDEF
#8: Protein | Mass: 28033.830 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: SCL1, PRC2, PRS2, YGL011C / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P21243, proteasome endopeptidase complex |
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#9: Protein | Mass: 27191.828 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PRE8, PRS4, YML092C / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P23639, proteasome endopeptidase complex |
#10: Protein | Mass: 28748.230 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PRE9, PRS5, YGR135W / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P23638, proteasome endopeptidase complex |
#11: Protein | Mass: 28478.111 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PRE6, YOL038W / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P40303, proteasome endopeptidase complex |
#12: Protein | Mass: 28649.086 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PUP2, DOA5, YGR253C, G9155 / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P32379, proteasome endopeptidase complex |
#13: Protein | Mass: 25634.000 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PRE5, YMR314W, YM9924.06 / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P40302, proteasome endopeptidase complex |
-Protein , 4 types, 4 molecules GLru
#14: Protein | Mass: 31575.068 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: PRE10, PRC1, PRS1, YOR362C, O6650 / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 References: UniProt: P21242, proteasome endopeptidase complex |
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#19: Protein | Mass: 49480.137 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: RPT4, CRL13, PCS1, SUG2, YOR259C / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 / References: UniProt: P53549 |
#22: Protein | Mass: 34442.281 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: RPN11, MPR1, YFR004W / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 / References: UniProt: P43588, ubiquitinyl hydrolase 1 |
#24: Protein | Mass: 14583.077 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: RPL40A, UBI1, YIL148W / Production host: Escherichia coli B (bacteria) / References: UniProt: P0CH08 |
-26S proteasome regulatory subunit ... , 5 types, 5 molecules HIJKM
#15: Protein | Mass: 52054.891 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: RPT1, CIM5, YTA3, YKL145W / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 / References: UniProt: P33299 |
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#16: Protein | Mass: 48898.160 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: RPT2, YHS4, YTA5, YDL007W, D2920 / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 / References: UniProt: P40327 |
#17: Protein | Mass: 45342.742 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: RPT6, CIM3, CRL3, SUG1, TBPY, TBY1, YGL048C / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 / References: UniProt: Q01939 |
#18: Protein | Mass: 47953.676 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: RPT3, YNT1, YTA2, YDR394W, D9509.14 / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 / References: UniProt: P33298 |
#20: Protein | Mass: 48315.727 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae S288c (yeast) / Strain: ATCC 204508 / S288c / Gene: RPT5, YTA1, YOR117W, O3258, YOR3258W / Production host: Saccharomyces cerevisiae W303 (yeast) / Strain (production host): YYS40 / References: UniProt: P33297 |
-Protein/peptide , 1 types, 1 molecules s
#23: Protein/peptide | Mass: 4068.414 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli BL21 (bacteria) / Strain (production host): BL21 / References: UniProt: P07818*PLUS |
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-Non-polymers , 2 types, 6 molecules
#25: Chemical | ChemComp-ATP / #26: Chemical | |
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-Details
Has protein modification | Y |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
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Source (recombinant) |
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Buffer solution | pH: 7.6 | ||||||||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 25 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES Details: 26S proteasomes were diluted to a concentration of 20 micromolar in a buffer with an ATP regeneration system, and 6 mM ortho-phenanthroline. This solution was mixed with an equal volume of ...Details: 26S proteasomes were diluted to a concentration of 20 micromolar in a buffer with an ATP regeneration system, and 6 mM ortho-phenanthroline. This solution was mixed with an equal volume of 50 micromolar ubiquitinated model substrate | ||||||||||||||||||||||||||||||||||||
Specimen support | Details: unspecified | ||||||||||||||||||||||||||||||||||||
Vitrification | Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Humidity: 90 % / Chamber temperature: 277 K Details: specimens were manually blotted with Whatman #1 filter paper |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS / Details: images were acquired in nanoprobe mode |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 29000 X / Nominal defocus max: -2500 nm / Nominal defocus min: -1000 nm / Calibrated defocus min: -1500 nm / Calibrated defocus max: -3000 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 6.25 sec. / Electron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 11656 |
Image scans | Sampling size: 5 µm / Width: 3710 / Height: 3838 / Movie frames/image: 25 / Used frames/image: 1-25 |
-Processing
EM software |
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Image processing | Details: Camera was operated in counting mode | ||||||||||||||||||||||||||||||||||||||||||||||||
CTF correction | Details: CTF correction was performed by Relion during reconstruction Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 579361 Details: Particles were selected using the Relion template-based particle picker | ||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4.17 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 48335 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT / Space: REAL | ||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 5MPC Accession code: 5MPC / Source name: PDB / Type: experimental model |