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Yorodumi- PDB-8f66: Thermoplasma acidophilum 20S proteasome - L81Y mutation in alpha ... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8f66 | ||||||
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Title | Thermoplasma acidophilum 20S proteasome - L81Y mutation in alpha subunit | ||||||
Components |
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Keywords | HYDROLASE / Protease / threonine protease / endopeptidase activity | ||||||
Function / homology | Function and homology information proteasome endopeptidase complex / proteasome core complex, beta-subunit complex / proteasome core complex, alpha-subunit complex / threonine-type endopeptidase activity / proteasomal protein catabolic process / ubiquitin-dependent protein catabolic process / endopeptidase activity / cytoplasm Similarity search - Function | ||||||
Biological species | Thermoplasma acidophilum (acidophilic) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.28 Å | ||||||
Authors | Chuah, J. / Smith, D. | ||||||
Funding support | United States, 1items
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Citation | Journal: Commun Biol / Year: 2023 Title: High resolution structures define divergent and convergent mechanisms of archaeal proteasome activation. Authors: Janelle J Y Chuah / Matthew S Rexroad / David M Smith / Abstract: Considering the link between neurodegenerative diseases and impaired proteasome function, and the neuro-protective impact of enhanced proteasome activity in animal models, it's crucial to understand ...Considering the link between neurodegenerative diseases and impaired proteasome function, and the neuro-protective impact of enhanced proteasome activity in animal models, it's crucial to understand proteasome activation mechanisms. A hydrophobic-tyrosine-any residue (HbYX) motif on the C-termini of proteasome-activating complexes independently triggers gate-opening of the 20S core particle for protein degradation; however, the causal allosteric mechanism remains unclear. Our study employs a structurally irreducible dipeptide HbYX mimetic to investigate the allosteric mechanism of gate-opening in the archaeal proteasome. High-resolution cryo-EM structures pinpoint vital residues and conformational changes in the proteasome α-subunit implicated in HbYX-dependent activation. Using point mutations, we simulated the HbYX-bound state, providing support for our mechanistic model. We discerned four main mechanistic elements triggering gate-opening: 1) back-loop rearrangement adjacent to K66, 2) intra- and inter- α subunit conformational changes, 3) occupancy of the hydrophobic pocket, and 4) a highly conserved isoleucine-threonine pair in the 20S channel stabilizing the open and closed states, termed the "IT switch." Comparison of different complexes unveiled convergent and divergent mechanism of 20S gate-opening among HbYX-dependent and independent activators. This study delivers a detailed molecular model for HbYX-dependent 20S gate-opening, enabling the development of small molecule proteasome activators that hold promise to treat neurodegenerative diseases. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8f66.cif.gz | 1.1 MB | Display | PDBx/mmCIF format |
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PDB format | pdb8f66.ent.gz | 911.6 KB | Display | PDB format |
PDBx/mmJSON format | 8f66.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8f66_validation.pdf.gz | 1.6 MB | Display | wwPDB validaton report |
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Full document | 8f66_full_validation.pdf.gz | 1.6 MB | Display | |
Data in XML | 8f66_validation.xml.gz | 156.7 KB | Display | |
Data in CIF | 8f66_validation.cif.gz | 216.6 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/f6/8f66 ftp://data.pdbj.org/pub/pdb/validation_reports/f6/8f66 | HTTPS FTP |
-Related structure data
Related structure data | 28876MC 8f6aC 8f7kC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 25879.465 Da / Num. of mol.: 14 / Mutation: L81Y Source method: isolated from a genetically manipulated source Source: (gene. exp.) Thermoplasma acidophilum (acidophilic) / Gene: psmA, Ta1288 / Production host: Escherichia coli BL21(DE3) (bacteria) References: UniProt: P25156, proteasome endopeptidase complex #2: Protein | Mass: 23169.811 Da / Num. of mol.: 14 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Thermoplasma acidophilum (acidophilic) / Gene: psmB, Ta0612 / Production host: Escherichia coli BL21(DE3) (bacteria) References: UniProt: P28061, proteasome endopeptidase complex |
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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
Component | Name: Thermoplasma acidophilum 20S proteasome - alphaL81Y mutant Type: COMPLEX / Entity ID: #2, #1 / Source: RECOMBINANT |
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Molecular weight | Value: 0.7 MDa / Experimental value: NO |
Source (natural) | Organism: Thermoplasma acidophilum (acidophilic) |
Source (recombinant) | Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) |
Buffer solution | pH: 7.4 |
Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: TFS KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2400 nm / Nominal defocus min: 1200 nm |
Image recording | Electron dose: 50 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: FEI FALCON III (4k x 4k) |
-Processing
Software | Name: PHENIX / Version: 1.20rc1_4392: / Classification: refinement | ||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 2.28 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 131453 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT | ||||||||||||||||||||||||
Refine LS restraints |
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