+Open data
-Basic information
Entry | Database: PDB / ID: 8gj2 | ||||||
---|---|---|---|---|---|---|---|
Title | E. coli clamp loader with closed clamp on primed template DNA | ||||||
Components |
| ||||||
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 / Hda-beta clamp complex / bacterial-type DNA replication / replication inhibiting complex / DNA polymerase III complex / replisome / regulation of DNA-templated DNA replication initiation / DNA strand elongation involved in DNA replication / DNA polymerase processivity factor activity / error-prone translesion synthesis ...DNA polymerase III, clamp loader complex / Hda-beta clamp complex / bacterial-type DNA replication / replication inhibiting complex / DNA polymerase III complex / replisome / regulation of DNA-templated DNA replication initiation / DNA strand elongation involved in DNA replication / DNA polymerase processivity factor activity / error-prone translesion synthesis / negative regulation of DNA-templated DNA replication initiation / 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 / DNA damage response / ATP hydrolysis activity / protein homodimerization activity / DNA binding / ATP binding / identical protein binding / cytosol Similarity search - Function | ||||||
Biological species | Escherichia coli K-12 (bacteria) Escherichia coli (E. coli) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.6 Å | ||||||
Authors | Oakley, A.J. / Xu, Z.-Q. / Dixon, N.E. | ||||||
Funding support | Australia, 1items
| ||||||
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 |
|
-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
---|
-Downloads & links
-Download
PDBx/mmCIF format | 8gj2.cif.gz | 1.2 MB | Display | PDBx/mmCIF format |
---|---|---|---|---|
PDB format | pdb8gj2.ent.gz | 815.4 KB | Display | PDB format |
PDBx/mmJSON format | 8gj2.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/gj/8gj2 ftp://data.pdbj.org/pub/pdb/validation_reports/gj/8gj2 | HTTPS FTP |
---|
-Related structure data
Related structure data | 40083MC 8giyC 8gizC 8gj0C 8gj1C 8gj3C M: map data used to model this data C: citing same article (ref.) |
---|---|
Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
|
---|---|
1 |
|
-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 | ||||
---|---|---|---|---|---|
#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 Details: DNA polymerase III subunit psi / 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 |
-Protein , 1 types, 2 molecules HI
#5: Protein | Mass: 40630.508 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: dnaN, b3701, JW3678 / Production host: Escherichia coli BL21 (bacteria) / References: UniProt: P0A988 |
---|
-DNA chain , 2 types, 2 molecules XY
#6: DNA chain | Mass: 7097.624 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
---|---|
#7: DNA chain | Mass: 10063.474 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
-Non-polymers , 4 types, 13 molecules
#8: Chemical | #9: Chemical | #10: Chemical | #11: Chemical | ChemComp-ZN / |
---|
-Details
Has ligand of interest | N |
---|
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: E. coli clamp loader with closed clamp 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-#7 / Source: MULTIPLE SOURCES | |||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Molecular weight | Value: 0.419 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 |
| |||||||||||||||||||||||||||||||||||||||||||||
Specimen | Conc.: 1.5 mg/ml / 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 |
---|---|
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 magnification: 165000 X / Nominal defocus max: 1300 nm / Nominal defocus min: 400 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / 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: 60 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 1703 |
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 |
| ||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
EM software |
| ||||||||||||||||||||||||||||||||||||||||
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: 600000 | ||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 452000 / Algorithm: BACK PROJECTION / Num. of class averages: 2 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL | ||||||||||||||||||||||||||||||||||||||||
Atomic model building |
| ||||||||||||||||||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 100.76 Å2 | ||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
|