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Yorodumi- PDB-9b5j: Ubiquitin E1-Ub-E2 tetrahedral transthiolation intermediate mimic... -
+Open data
-Basic information
Entry | Database: PDB / ID: 9b5j | |||||||||
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Title | Ubiquitin E1-Ub-E2 tetrahedral transthiolation intermediate mimic (doubly Ub-loaded) - cluster 5 map and model (Ub(A)-AMP) | |||||||||
Components |
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Keywords | LIGASE / UBIQUITIN / E1 / E2 / UBA1 / UBC4 / TRANSTHIOESTERIFICATION / THIOESTER / TRANSTHIOLATION / TETRAHEDRAL INTERMEDIATE / ADENYLATION / INHIBITOR / NUCLEUS / PHOSPHOPROTEIN / UBL CONJUGATION PATHWAY / UBL / ATP ATP-BINDING / AMP / NUCLEOTIDE-BINDING / ISOPEPTIDE BOND | |||||||||
Function / homology | Function and homology information E3 ubiquitin ligases ubiquitinate target proteins / Peroxisomal protein import / nuclear SCF ubiquitin ligase complex / Synthesis of active ubiquitin: roles of E1 and E2 enzymes / Antigen processing: Ubiquitination & Proteasome degradation / E1 ubiquitin-activating enzyme / ubiquitin activating enzyme activity / SREBP signaling pathway / positive regulation of mitotic metaphase/anaphase transition / SCF-dependent proteasomal ubiquitin-dependent protein catabolic process ...E3 ubiquitin ligases ubiquitinate target proteins / Peroxisomal protein import / nuclear SCF ubiquitin ligase complex / Synthesis of active ubiquitin: roles of E1 and E2 enzymes / Antigen processing: Ubiquitination & Proteasome degradation / E1 ubiquitin-activating enzyme / ubiquitin activating enzyme activity / SREBP signaling pathway / positive regulation of mitotic metaphase/anaphase transition / SCF-dependent proteasomal ubiquitin-dependent protein catabolic process / E2 ubiquitin-conjugating enzyme / ubiquitin conjugating enzyme activity / ubiquitin ligase complex / modification-dependent protein catabolic process / protein polyubiquitination / protein tag activity / ubiquitin-protein transferase activity / ribosome biogenesis / ubiquitin-dependent protein catabolic process / cytosolic large ribosomal subunit / cytoplasmic translation / protein ubiquitination / structural constituent of ribosome / DNA damage response / ubiquitin protein ligase binding / nucleolus / magnesium ion binding / ATP hydrolysis activity / ATP binding / nucleus / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Schizosaccharomyces pombe 972h- (yeast) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.86 Å | |||||||||
Authors | Kochanczyk, T. / Lima, C.D. | |||||||||
Funding support | United States, 2items
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Citation | Journal: To be published Title: Structural basis for transthiolation intermediates in the ubiquitin pathway Authors: Kochanczyk, T. / Hann, Z.S. / Lux, M.C. / Reyes, A.M.V.D. / Ji, C. / Tan, D.S. / Lima, C.D. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 9b5j.cif.gz | 254.9 KB | Display | PDBx/mmCIF format |
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PDB format | pdb9b5j.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 9b5j.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 9b5j_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 9b5j_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 9b5j_validation.xml.gz | 50 KB | Display | |
Data in CIF | 9b5j_validation.cif.gz | 74.1 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/b5/9b5j ftp://data.pdbj.org/pub/pdb/validation_reports/b5/9b5j | HTTPS FTP |
-Related structure data
Related structure data | 44214MC 9b5cC 9b5dC 9b5eC 9b5fC 9b5gC 9b5hC 9b5iC 9b5kC 9b5lC 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
-Protein , 4 types, 4 molecules ABCD
#1: Protein | Mass: 111764.047 Da / Num. of mol.: 1 / Fragment: residues 13-1012 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Schizosaccharomyces pombe 972h- (yeast) Strain: 972 / ATCC 24843 / Gene: ptr3, SPBC1604.21c, SPBC211.09 / Plasmid: pSMT3 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: O94609, E1 ubiquitin-activating enzyme |
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#2: Protein | Mass: 8568.769 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Schizosaccharomyces pombe 972h- (yeast) Strain: 972 / ATCC 24843 / Gene: uep1, ubi2, SPAC1805.12c / Plasmid: pET3C / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P0CH07 |
#3: Protein | Mass: 17043.336 Da / Num. of mol.: 1 / Mutation: C21S,C107S Source method: isolated from a genetically manipulated source Source: (gene. exp.) Schizosaccharomyces pombe 972h- (yeast) Strain: 972 / ATCC 24843 / Gene: ubc4, SPBC119.02 / Plasmid: pET29b+ / Production host: Escherichia coli BL21(DE3) (bacteria) References: UniProt: P46595, E2 ubiquitin-conjugating enzyme |
#4: Protein | Mass: 8769.948 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Schizosaccharomyces pombe 972h- (yeast) Strain: 972 / ATCC 24843 / Gene: uep1, ubi2, SPAC1805.12c / Plasmid: pTXB1 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P0CH07 |
-Non-polymers , 2 types, 2 molecules
#5: Chemical | ChemComp-AMP / |
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#6: Chemical | ChemComp-A1AIV / Mass: 84.120 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C4H8N2 / Feature type: SUBJECT OF INVESTIGATION |
-Details
Has ligand of interest | 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
Component | Name: Covalent E1-Ub-E2 transthiolation intermediate mimic complex with second Ub bound to E1 (doubly Ub-loaded E1-Ub-E2 complex) Type: COMPLEX / Entity ID: #1-#3 / Source: RECOMBINANT | ||||||||||||||||||||||||||||||
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Molecular weight | Experimental value: NO | ||||||||||||||||||||||||||||||
Source (natural) | Organism: Schizosaccharomyces pombe 972h- (yeast) / Strain: 972 / ATCC 24843 | ||||||||||||||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli BL21(DE3) (bacteria) | ||||||||||||||||||||||||||||||
Buffer solution | pH: 7.2 Details: 20 mM Tris-HCl, 100 mM NaCl, 5 mM MgCl2, 1 mM ATP, 0.05% CHAPSO | ||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3 | ||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 800 nm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 4 sec. / Electron dose: 72 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.86 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 122776 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL | ||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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