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Yorodumi- PDB-8oiu: Cryo-EM reconstruction of the native 24-mer E2o core of the 2-oxo... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8oiu | ||||||
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Title | Cryo-EM reconstruction of the native 24-mer E2o core of the 2-oxoglutarate dehydrogenase complex of C. thermophilum at 3.35 A resolution | ||||||
Components | Dihydrolipoyllysine-residue succinyltransferase | ||||||
Keywords | TRANSFERASE / Dihydrolipoyl Succinyltransferase / E2 / Oxoglutarate / a-Ketoglutarate | ||||||
Function / homology | Function and homology information L-lysine catabolic process to acetyl-CoA via saccharopine / dihydrolipoyllysine-residue succinyltransferase / dihydrolipoyllysine-residue succinyltransferase activity / oxoglutarate dehydrogenase complex / tricarboxylic acid cycle / mitochondrion / cytosol Similarity search - Function | ||||||
Biological species | Thermochaetoides thermophila DSM 1495 (fungus) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.35 Å | ||||||
Authors | Skalidis, I. / Tueting, C. / Kyrilis, F.L. / Hamdi, F. / Kastritis, P.L. | ||||||
Funding support | 1items
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Citation | Journal: Commun Biol / Year: 2023 Title: Structural analysis of an endogenous 4-megadalton succinyl-CoA-generating metabolon. Authors: Ioannis Skalidis / Fotis L Kyrilis / Christian Tüting / Farzad Hamdi / Toni K Träger / Jaydeep Belapure / Gerd Hause / Marta Fratini / Francis J O'Reilly / Ingo Heilmann / Juri Rappsilber ...Authors: Ioannis Skalidis / Fotis L Kyrilis / Christian Tüting / Farzad Hamdi / Toni K Träger / Jaydeep Belapure / Gerd Hause / Marta Fratini / Francis J O'Reilly / Ingo Heilmann / Juri Rappsilber / Panagiotis L Kastritis / Abstract: The oxoglutarate dehydrogenase complex (OGDHc) participates in the tricarboxylic acid cycle and, in a multi-step reaction, decarboxylates α-ketoglutarate, transfers succinyl to CoA, and reduces NAD+. ...The oxoglutarate dehydrogenase complex (OGDHc) participates in the tricarboxylic acid cycle and, in a multi-step reaction, decarboxylates α-ketoglutarate, transfers succinyl to CoA, and reduces NAD+. Due to its pivotal role in metabolism, OGDHc enzymatic components have been studied in isolation; however, their interactions within the endogenous OGDHc remain elusive. Here, we discern the organization of a thermophilic, eukaryotic, native OGDHc in its active state. By combining biochemical, biophysical, and bioinformatic methods, we resolve its composition, 3D architecture, and molecular function at 3.35 Å resolution. We further report the high-resolution cryo-EM structure of the OGDHc core (E2o), which displays various structural adaptations. These include hydrogen bonding patterns confining interactions of OGDHc participating enzymes (E1o-E2o-E3), electrostatic tunneling that drives inter-subunit communication, and the presence of a flexible subunit (E3BPo), connecting E2o and E3. This multi-scale analysis of a succinyl-CoA-producing native cell extract provides a blueprint for structure-function studies of complex mixtures of medical and biotechnological value. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8oiu.cif.gz | 59.4 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8oiu.ent.gz | 41 KB | Display | PDB format |
PDBx/mmJSON format | 8oiu.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8oiu_validation.pdf.gz | 1.2 MB | Display | wwPDB validaton report |
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Full document | 8oiu_full_validation.pdf.gz | 1.2 MB | Display | |
Data in XML | 8oiu_validation.xml.gz | 23.2 KB | Display | |
Data in CIF | 8oiu_validation.cif.gz | 31.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/oi/8oiu ftp://data.pdbj.org/pub/pdb/validation_reports/oi/8oiu | HTTPS FTP |
-Related structure data
Related structure data | 16900MC 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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-Components
#1: Protein | Mass: 45987.004 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Thermochaetoides thermophila DSM 1495 (fungus) References: UniProt: G0SAX9, dihydrolipoyllysine-residue succinyltransferase |
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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: Native 24-mer core of Oxoglutarate Dehydrogenase Complex Type: COMPLEX / Entity ID: all / Source: NATURAL |
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Molecular weight | Value: 3 MDa / Experimental value: NO |
Source (natural) | Organism: Thermochaetoides thermophila DSM 1495 (fungus) |
Buffer solution | pH: 7.4 |
Buffer component | Conc.: 200 mM / Name: Ammonium acetate / Formula: NH4CH2COOH |
Specimen | Conc.: 0.3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 277 K Details: For plunging, blot force 0 and blotting time of 4 sec were applied. |
-Electron microscopy imaging
Microscopy | Model: TFS GLACIOS |
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Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: OTHER |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 92000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: OTHER / Temperature (max): 103.15 K / Temperature (min): 77.15 K |
Image recording | Electron dose: 30 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of real images: 21381 |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 1300 / Details: Template picking was directly applied | ||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: O (octahedral) | ||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.35 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 69028 / Algorithm: SIMULTANEOUS ITERATIVE (SIRT) Details: Particles were symmetry expanded for the final reconstruction, then local reconstruction was performed without symmetry application. Num. of class averages: 50 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL Details: Initial fitting was performed in ChimeraX v1.2, then real-space refined in PHENIX v1.19 | ||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 7Q5Q Pdb chain-ID: all / Accession code: 7Q5Q / Source name: PDB / Type: experimental model |