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Yorodumi- PDB-9bh6: Human DNA polymerase theta helicase domain tetramer in the apo form -
+Open data
-Basic information
Entry | Database: PDB / ID: 9bh6 | |||||||||
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Title | Human DNA polymerase theta helicase domain tetramer in the apo form | |||||||||
Components | DNA polymerase theta | |||||||||
Keywords | DNA BINDING PROTEIN / DNA repair / TMEJ / MMEJ | |||||||||
Function / homology | Function and homology information single-stranded DNA endodeoxyribonuclease activity / HDR through MMEJ (alt-NHEJ) / single-stranded DNA helicase activity / double-strand break repair via alternative nonhomologous end joining / replication fork processing / site of DNA damage / 5'-deoxyribose-5-phosphate lyase activity / somatic hypermutation of immunoglobulin genes / negative regulation of double-strand break repair via homologous recombination / error-prone translesion synthesis ...single-stranded DNA endodeoxyribonuclease activity / HDR through MMEJ (alt-NHEJ) / single-stranded DNA helicase activity / double-strand break repair via alternative nonhomologous end joining / replication fork processing / site of DNA damage / 5'-deoxyribose-5-phosphate lyase activity / somatic hypermutation of immunoglobulin genes / negative regulation of double-strand break repair via homologous recombination / error-prone translesion synthesis / DNA helicase activity / base-excision repair / protein homooligomerization / RNA-directed DNA polymerase / RNA-directed DNA polymerase activity / double-strand break repair / site of double-strand break / DNA helicase / damaged DNA binding / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / DNA repair / DNA damage response / chromatin binding / Golgi apparatus / magnesium ion binding / ATP hydrolysis activity / nucleoplasm / ATP binding / identical protein binding / cytosol Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å | |||||||||
Authors | Zerio, C.J. / Lander, G.C. | |||||||||
Funding support | United States, 2items
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Citation | Journal: To Be Published Title: Human polymerase theta helicase positions DNA microhomologies for double-strand break repair Authors: Zerio, C.J. / Bai, Y. / Sosa-Alvarado, B.A. / Guzi, T. / Lander, G.C. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 9bh6.cif.gz | 598.3 KB | Display | PDBx/mmCIF format |
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PDB format | pdb9bh6.ent.gz | 486.1 KB | Display | PDB format |
PDBx/mmJSON format | 9bh6.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 9bh6_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 9bh6_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 9bh6_validation.xml.gz | 85.4 KB | Display | |
Data in CIF | 9bh6_validation.cif.gz | 127.7 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/bh/9bh6 ftp://data.pdbj.org/pub/pdb/validation_reports/bh/9bh6 | HTTPS FTP |
-Related structure data
Related structure data | 44534MC 9bh7C 9bh8C 9bh9C 9bhaC 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: 99671.344 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: POLQ, POLH / Plasmid: pET-Duet-1 / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): Rosetta References: UniProt: O75417, DNA helicase, DNA-directed DNA polymerase, RNA-directed DNA polymerase |
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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: Human DNA polymerase theta helicase domain / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT | ||||||||||||||||||||
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Molecular weight | Value: 0.4 MDa / Experimental value: NO | ||||||||||||||||||||
Source (natural) | Organism: Homo sapiens (human) | ||||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli (E. coli) / Strain: Rosetta(DE3) / Plasmid: pET-Duet-1 | ||||||||||||||||||||
Buffer solution | pH: 7.5 | ||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 0.9 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||
Specimen support | Details: Grids were glow discharged under vacuum for 30 s at 15 mA in a Pelco easiGlow 91000 Glow Discharge Cleaning System. Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 | ||||||||||||||||||||
Vitrification | Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 277 K Details: 3 microliters of sample was applied to the surface of the grid, blotted with Whatman 1 filter paper until 2 seconds after the liquid spot on the filter paper stopped spreading, and the grid ...Details: 3 microliters of sample was applied to the surface of the grid, blotted with Whatman 1 filter paper until 2 seconds after the liquid spot on the filter paper stopped spreading, and the grid was plunged into a liquid ethane pool cooled by liquid nitrogen using a manual plunge freezer. |
-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 magnification: 105000 X / Calibrated magnification: 60024 X / Nominal defocus max: 1000 nm / Nominal defocus min: 1000 nm / Calibrated defocus min: 300 nm / Calibrated defocus max: 2000 nm / Cs: 2.7 mm / C2 aperture diameter: 150 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 77 K / Temperature (min): 70 K |
Image recording | Average exposure time: 0.9 sec. / Electron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 3906 |
Image scans | Sampling size: 5 µm / Width: 5760 / Height: 4092 |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 1474207 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: D2 (2x2 fold dihedral) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 208545 / Algorithm: FOURIER SPACE Details: We performed local refinement of a single protomer and then multiplied the output protomer map to fit the D2 symmetric tetramer map. Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 54.43 / Protocol: FLEXIBLE FIT / Space: REAL / Target criteria: Cross-correlation coefficient Details: We used ISOLDE to flexibly fit one protomer into the locally refined protomer map. The output protomer model was multiplied and we used ISOLDE to fit four protomers into the tetramer map. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 5A9J Pdb chain-ID: A / Accession code: 5A9J / Chain residue range: 67-892 / Pdb chain residue range: 67-892 / Source name: PDB / Type: experimental model |