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Yorodumi- PDB-8v5d: Human mitochondrial DNA polymerase catalytic subunit, PolG, in an... -
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-Basic information
Entry | Database: PDB / ID: 8v5d | ||||||||||||
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Title | Human mitochondrial DNA polymerase catalytic subunit, PolG, in an APO conformation | ||||||||||||
Components | DNA polymerase subunit gamma-1 | ||||||||||||
Keywords | DNA BINDING PROTEIN/DNA / DNA BINDING PROTEIN / DNA polymerase / mitochondrial DNA replication / DNA polymerase catalytic subunit / DNA BINDING PROTEIN-DNA complex | ||||||||||||
Function / homology | Function and homology information gamma DNA polymerase complex / mitochondrial DNA replication / single-stranded DNA 3'-5' DNA exonuclease activity / DNA replication proofreading / Hydrolases; Acting on ester bonds; Exodeoxyribonucleases producing 5'-phosphomonoesters / DNA metabolic process / mitochondrial nucleoid / Lyases; Carbon-oxygen lyases; Other carbon-oxygen lyases / 5'-deoxyribose-5-phosphate lyase activity / base-excision repair, gap-filling ...gamma DNA polymerase complex / mitochondrial DNA replication / single-stranded DNA 3'-5' DNA exonuclease activity / DNA replication proofreading / Hydrolases; Acting on ester bonds; Exodeoxyribonucleases producing 5'-phosphomonoesters / DNA metabolic process / mitochondrial nucleoid / Lyases; Carbon-oxygen lyases; Other carbon-oxygen lyases / 5'-deoxyribose-5-phosphate lyase activity / base-excision repair, gap-filling / 3'-5' exonuclease activity / base-excision repair / DNA-templated DNA replication / protease binding / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / mitochondrial matrix / intracellular membrane-bounded organelle / chromatin binding / protein-containing complex / mitochondrion / DNA binding Similarity search - Function | ||||||||||||
Biological species | Homo sapiens (human) | ||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3 Å | ||||||||||||
Authors | Riccio, A.A. / Brannon, A.J. / Krahn, J.M. / Bouvette, J. / Borgnia, J.M. / Copeland, W.C. | ||||||||||||
Funding support | United States, 3items
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Citation | Journal: Nucleic Acids Res / Year: 2024 Title: Coordinated DNA polymerization by Polγ and the region of LonP1 regulated proteolysis. Authors: Amanda A Riccio / Asia J Brannon / Juno M Krahn / Jonathan Bouvette / Jason G Williams / Mario J Borgnia / William C Copeland / Abstract: The replicative mitochondrial DNA polymerase, Polγ, and its protein regulation are essential for the integrity of the mitochondrial genome. The intricacies of Polγ regulation and its interactions ...The replicative mitochondrial DNA polymerase, Polγ, and its protein regulation are essential for the integrity of the mitochondrial genome. The intricacies of Polγ regulation and its interactions with regulatory proteins, which are essential for fine-tuning polymerase function, remain poorly understood. Misregulation of the Polγ heterotrimer, consisting of (i) PolG, the polymerase catalytic subunit and (ii) PolG2, the accessory subunit, ultimately results in mitochondrial diseases. Here, we used single particle cryo-electron microscopy to resolve the structure of PolG in its apoprotein state and we captured Polγ at three intermediates within the catalytic cycle: DNA bound, engaged, and an active polymerization state. Chemical crosslinking mass spectrometry, and site-directed mutagenesis uncovered the region of LonP1 engagement of PolG, which promoted proteolysis and regulation of PolG protein levels. PolG2 clinical variants, which disrupted a stable Polγ complex, led to enhanced LonP1-mediated PolG degradation. Overall, this insight into Polγ aids in an understanding of mitochondrial DNA replication and characterizes how machinery of the replication fork may be targeted for proteolytic degradation when improperly functioning. | ||||||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8v5d.cif.gz | 185.7 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8v5d.ent.gz | 139.2 KB | Display | PDB format |
PDBx/mmJSON format | 8v5d.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8v5d_validation.pdf.gz | 1.1 MB | Display | wwPDB validaton report |
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Full document | 8v5d_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | 8v5d_validation.xml.gz | 41.7 KB | Display | |
Data in CIF | 8v5d_validation.cif.gz | 59.9 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/v5/8v5d ftp://data.pdbj.org/pub/pdb/validation_reports/v5/8v5d | HTTPS FTP |
-Related structure data
Related structure data | 42982MC 8v54C 8v55C 8v5rC 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: 138723.438 Da / Num. of mol.: 1 / Mutation: D198A, E200A Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: POLG / Production host: unidentified baculovirus / References: UniProt: P54098 |
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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: mitochondrial DNA polymerase catalytic subunit, PolG / Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES |
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Molecular weight | Value: 0.120 MDa / Experimental value: YES |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: unidentified baculovirus / Plasmid: pET21a |
Buffer solution | pH: 8 |
Specimen | Conc.: 1.2 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: 2500 nm / Nominal defocus min: 1000 nm |
Image recording | Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
-Processing
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
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3D reconstruction | Resolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 122659 / Symmetry type: POINT | ||||||||||||||||||||||||
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