+Open data
-Basic information
Entry | Database: PDB / ID: 8gj3 | ||||||
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Title | E. coli clamp loader on primed template DNA | ||||||
Components |
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Keywords | DNA BINDING PROTEIN/DNA / clamp loader / DNA clamp / AAA+ / ATPase / DNA BINDING PROTEIN-DNA complex | ||||||
Function / homology | Function and homology information DNA polymerase III, clamp loader complex / DNA polymerase III complex / replisome / DNA polymerase processivity factor activity / 3'-5' exonuclease activity / ribonucleoside triphosphate phosphatase activity / response to radiation / DNA-templated DNA replication / DNA replication / DNA-directed DNA polymerase ...DNA polymerase III, clamp loader complex / DNA polymerase III complex / replisome / DNA polymerase processivity factor activity / 3'-5' exonuclease activity / ribonucleoside triphosphate phosphatase activity / response to radiation / DNA-templated DNA replication / DNA replication / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / ATP hydrolysis activity / DNA binding / ATP binding / identical protein binding Similarity search - Function | ||||||
Biological species | Escherichia coli K-12 (bacteria) Escherichia coli (E. coli) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.8 Å | ||||||
Authors | Oakley, A.J. / Xu, Z.-Q. / Dixon, N.E. | ||||||
Funding support | Australia, 1items
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Citation | Journal: To Be Published Title: Structural characterisation of the complete cycle of sliding clamp loading in E. coli Authors: Xu, Z.-Q. / Oakley, A.J. / Dixon, N.E. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8gj3.cif.gz | 890.2 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8gj3.ent.gz | 582.8 KB | Display | PDB format |
PDBx/mmJSON format | 8gj3.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/gj/8gj3 ftp://data.pdbj.org/pub/pdb/validation_reports/gj/8gj3 | HTTPS FTP |
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-Related structure data
Related structure data | 40084MC 8giyC 8gizC 8gj0C 8gj1C 8gj2C 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
-DNA polymerase III subunit ... , 4 types, 6 molecules ABCDEF
#1: Protein | Mass: 38745.574 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: holA, b0640, JW0635 / Production host: Escherichia coli BL21 (bacteria) / References: UniProt: P28630, DNA-directed DNA polymerase | ||||
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#2: Protein | Mass: 71228.648 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: dnaX, dnaZ, dnaZX, b0470, JW0459 / Production host: Escherichia coli BL21 (bacteria) / References: UniProt: P06710, DNA-directed DNA polymerase #3: Protein | | Mass: 36980.484 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: holB, b1099, JW1085 / Production host: Escherichia coli BL21 (bacteria) / References: UniProt: P28631, DNA-directed DNA polymerase #4: Protein | | Mass: 15188.276 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: holD, b4372, JW4334 / Production host: Escherichia coli BL21 (bacteria) / References: UniProt: P28632, DNA-directed DNA polymerase |
-DNA chain , 2 types, 2 molecules XY
#5: DNA chain | Mass: 4856.191 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
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#6: DNA chain | Mass: 7969.154 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
-Non-polymers , 4 types, 13 molecules
#7: Chemical | #8: Chemical | #9: Chemical | #10: Chemical | ChemComp-ZN / |
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-Details
Has ligand of interest | N |
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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: E. coli clamp loader on primed template DNA / Type: COMPLEX Details: Clamp loader complex composed of DNA polymerase III delta tau(3) delta' chi and psi subunits. Clamp composed of DNA polymerase III beta(2) subunits. Entity ID: #1-#6 / Source: MULTIPLE SOURCES | |||||||||||||||||||||||||||||||||||||||||||||
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Molecular weight | Value: 0.333 MDa / Experimental value: NO | |||||||||||||||||||||||||||||||||||||||||||||
Source (natural) | Organism: Escherichia coli (E. coli) / Strain: K-12 | |||||||||||||||||||||||||||||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli (E. coli) / Strain: BL21 | |||||||||||||||||||||||||||||||||||||||||||||
Buffer solution | pH: 7.6 Details: 30 mM Tris-HCl pH 7.6, 5 mM MgCl2, 2 mM ADP, 0.5 mM AlCl3, 5 mM NaF, 5 mM dithiothreitol, 0.25 mM EDTA, 2% glycerol. | |||||||||||||||||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES Details: 30 microL of 6.0 microM delta/tau3/delta' complex was mixed with chi/psi complex, beta, and p/t DNA (annealed DNA oligonucleotides) at a molar ratio of 1:1.3 and dialysed at 4 degrees C ...Details: 30 microL of 6.0 microM delta/tau3/delta' complex was mixed with chi/psi complex, beta, and p/t DNA (annealed DNA oligonucleotides) at a molar ratio of 1:1.3 and dialysed at 4 degrees C against 250 mL of 30 mM Tris-HCl pH 7.6, 5 mM MgCl2, 2 mM ADP, 0.5 mM AlCl3, 5 mM NaF, 5 mM dithiothreitol, 0.25 mM EDTA, 2% glycerol. | |||||||||||||||||||||||||||||||||||||||||||||
Specimen support | Details: Used a Zepto Low-pressure plasma systems (PLASMA CLEANER) (Diener Electronic) at 10% power for 120 seconds. 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: 279 K Details: 3 uL of sample was applied onto a Ultrafoil Au R1.2/1.3 grid. Blot for 4.5 s with no extra force before plunging into liquid ethane. |
-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 FIELDBright-field microscopy / Nominal defocus max: 1300 nm / Nominal defocus min: 400 nm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 6 sec. / Electron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2909 |
EM imaging optics | Energyfilter name: GIF Bioquantum / Details: unfiltered mode |
Image scans | Width: 4096 / Height: 4096 / Movie frames/image: 60 / Used frames/image: 1-60 |
-Processing
Software |
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EM software |
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CTF correction | Details: CTF refinement per-particle was performed in RELION before final 3D reconstruction. Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 645000 | ||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 308000 / Algorithm: BACK PROJECTION / Num. of class averages: 2 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT | ||||||||||||||||||||||||||||||||||||||||
Atomic model building |
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Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 124.83 Å2 | ||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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