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Yorodumi- PDB-1nke: A BACILLUS DNA POLYMERASE I PRODUCT COMPLEX BOUND TO A CYTOSINE-T... -
+Open data
-Basic information
Entry | Database: PDB / ID: 1nke | |||||||||
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Title | A BACILLUS DNA POLYMERASE I PRODUCT COMPLEX BOUND TO A CYTOSINE-THYMINE MISMATCH AFTER A SINGLE ROUND OF PRIMER EXTENSION, FOLLOWING INCORPORATION OF DCTP. | |||||||||
Components |
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Keywords | transferase/DNA / DNA POLYMERASE I / DNA REPLICATION / KLENOW FRAGMENT / PROTEIN-DNA COMPLEX / DNA MISMATCH / transferase-DNA COMPLEX | |||||||||
Function / homology | Function and homology information 3'-5' exonuclease activity / DNA-templated DNA replication / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / DNA repair / DNA binding Similarity search - Function | |||||||||
Biological species | Geobacillus stearothermophilus (bacteria) | |||||||||
Method | X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.8 Å | |||||||||
Authors | Johnson, S.J. / Beese, L.S. | |||||||||
Citation | Journal: Cell(Cambridge,Mass.) / Year: 2004 Title: Structures of mismatch replication errors observed in a DNA polymerase. Authors: Johnson, S.J. / Beese, L.S. #1: Journal: Proc.Natl.Acad.Sci.USA / Year: 2003 Title: Processive DNA synthesis observed in a polymerase crystal suggests a mechanism for the prevention of frameshift mutations. Authors: Johnson, S.J. / Taylor, J.S. / Beese, L.S. #2: Journal: Nature / Year: 1998 Title: Visualizing DNA Replication in a Catalytically Active Bacillus DNA Polymerase Crystal Authors: Kiefer, J.R. / Mao, C. / Braman, J.C. / Beese, L.S. #3: Journal: Structure / Year: 1997 Title: Crystal Structure of a Thermostable Bacillus DNA Polymerase I Large Fragment at 2.1 A Resolution Authors: Kiefer, J.R. / Mao, C. / Hansen, C.J. / Basehore, S.L. / Hogrefe, H.H. / Braman, J.C. / Beese, L.S. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 1nke.cif.gz | 160.6 KB | Display | PDBx/mmCIF format |
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PDB format | pdb1nke.ent.gz | 118.4 KB | Display | PDB format |
PDBx/mmJSON format | 1nke.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/nk/1nke ftp://data.pdbj.org/pub/pdb/validation_reports/nk/1nke | HTTPS FTP |
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-Related structure data
Related structure data | 1njwC 1njxC 1njyC 1njzC 1nk0C 1nk4C 1nk5C 1nk6C 1nk7C 1nk8C 1nk9C 1nkbC 1nkcC 2bdpS C: citing same article (ref.) S: Starting model for refinement |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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Unit cell |
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Details | Exists as a monomer. One molecule per asymmetric unit. |
-Components
-DNA chain , 2 types, 2 molecules BC
#1: DNA chain | Mass: 3319.175 Da / Num. of mol.: 1 / Source method: obtained synthetically |
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#2: DNA chain | Mass: 4914.190 Da / Num. of mol.: 1 / Source method: obtained synthetically |
-Protein / Sugars , 2 types, 3 molecules A
#3: Protein | Mass: 66172.844 Da / Num. of mol.: 1 Fragment: BACILLUS FRAGMENT (ANALOGOUS TO THE E. COLI KLENOW FRAGMENT) Source method: isolated from a genetically manipulated source Source: (gene. exp.) Geobacillus stearothermophilus (bacteria) Plasmid: PET-30A(+) / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3)PLYSS / References: UniProt: P52026, DNA-directed DNA polymerase |
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#4: Polysaccharide |
-Non-polymers , 4 types, 448 molecules
#5: Chemical | ChemComp-DCP / | ||||
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#6: Chemical | ChemComp-SO4 / #7: Chemical | ChemComp-MG / | #8: Water | ChemComp-HOH / | |
-Details
Compound details | THE POLYMERASE |
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-Experimental details
-Experiment
Experiment | Method: X-RAY DIFFRACTION / Number of used crystals: 1 |
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-Sample preparation
Crystal | Density Matthews: 2.76 Å3/Da / Density % sol: 55.11 % | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Crystal grow | Temperature: 290 K / Method: vapor diffusion, hanging drop / pH: 5.8 Details: AMMONIUM SULFATE, MAGNESIUM SULFATE, MPD, MES, pH 5.80, VAPOR DIFFUSION, HANGING DROP, temperature 290K | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Components of the solutions |
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Crystal grow | *PLUS pH: 5.8 / Method: vapor diffusion, hanging dropDetails: Johnson, S.J., (2003) Proc.Natl.Acad.Sci.USA, 100, 3895. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Components of the solutions | *PLUS
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-Data collection
Diffraction | Mean temperature: 100 K |
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Diffraction source | Source: SYNCHROTRON / Site: APS / Beamline: 14-BM-C / Wavelength: 0.9 Å |
Detector | Type: ADSC QUANTUM 4 / Detector: CCD / Date: Apr 2, 2002 |
Radiation | Monochromator: Bent Ge(III) monochromator / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray |
Radiation wavelength | Wavelength: 0.9 Å / Relative weight: 1 |
Reflection | Resolution: 1.8→43.07 Å / Num. all: 80698 / Num. obs: 80698 / % possible obs: 97.8 % / Observed criterion σ(F): -3 / Observed criterion σ(I): -3 / Redundancy: 5.4 % / Biso Wilson estimate: 19.1 Å2 / Rsym value: 0.064 / Net I/σ(I): 21.9 |
Reflection shell | Resolution: 1.8→1.86 Å / Redundancy: 4.6 % / Mean I/σ(I) obs: 2.5 / Num. unique all: 8556 / Rsym value: 0.474 / % possible all: 95.2 |
Reflection | *PLUS Highest resolution: 1.8 Å / Lowest resolution: 50 Å / Num. measured all: 446710 / Rmerge(I) obs: 0.064 |
Reflection shell | *PLUS Rmerge(I) obs: 0.474 |
-Processing
Software |
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Refinement | Method to determine structure: MOLECULAR REPLACEMENT Starting model: PDB ENTRY 2BDP Resolution: 1.8→43.07 Å / Rfactor Rfree error: 0.004 / Data cutoff high absF: 2156189.35 / Data cutoff high rms absF: 2156189.35 / Data cutoff low absF: 0 / Isotropic thermal model: RESTRAINED / Cross valid method: THROUGHOUT / σ(F): 2 / Stereochemistry target values: Engh & Huber Details: THE BACILLUS POLYMERASE WAS CO- CRYSTALLIZED WITH A DNA CONTAINING A 10 BASE PAIR DUPLEX AND A 5 BASE 5' TEMPLATE OVERHANG. THE CRYSTAL WAS SUBSEQUENTLY PLACED INTO A STABILIZING SOLUTION ...Details: THE BACILLUS POLYMERASE WAS CO- CRYSTALLIZED WITH A DNA CONTAINING A 10 BASE PAIR DUPLEX AND A 5 BASE 5' TEMPLATE OVERHANG. THE CRYSTAL WAS SUBSEQUENTLY PLACED INTO A STABILIZING SOLUTION CONTAINING DCTP. THE RESULTING PRODUCT COMPLEX CONTAINED 11 BASE PAIRS OF DUPLEX DNA AND A 4 BASE 5' TEMPLATE OVERHANG. THE FOUR BASES IN THE OVERHANG ARE DISORDERED AND ARE NOT INCLUDED IN THE MODEL. A SINGLE BASE 3' OVERHANG ON THE TEMPLATE STRAND (CHAIN C) ASSURED THAT THE DNA DUPLEX WAS NOT BOUND BACKWARDS BY THE POLYMERASE DURING CRYSTALLIZATION. ELECTRON DENSITY WAS OBSERVED FOR ALL PROTEIN SIDE CHAINS EXCEPT LYSINE 298, WHICH WAS MODELLED TO THE BETA CARBON.ALTERNATE SIDE CHAIN CONFORMATIONS ARE INCLUDED FOR RESIDUES 615,658,797,830. THE MAGNESIUM AT POSITION 920 WAS ASSIGNED DUE TO THE LOW REFINED B-FACTOR AND COMPARISON WITH A RELATED KLENTAQ POLYMERASE STRUCTURE (3KTQ). HOWEVER, THE RESOLUTION OF THE STRUCTURE PREVENTS A DEFINITIVE ASSIGNMENT BETWEEN WATER AND MAGNESIUM.
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Solvent computation | Solvent model: FLAT MODEL / Bsol: 43.6188 Å2 / ksol: 0.384351 e/Å3 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 29.1 Å2
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Refine analyze |
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Refinement step | Cycle: LAST / Resolution: 1.8→43.07 Å
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Refine LS restraints |
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LS refinement shell | Resolution: 1.8→1.86 Å / Rfactor Rfree error: 0.016 / Total num. of bins used: 10
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Xplor file |
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Refinement | *PLUS Highest resolution: 1.8 Å / Lowest resolution: 50 Å / % reflection Rfree: 5 % | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Solvent computation | *PLUS | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | *PLUS | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints | *PLUS
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LS refinement shell | *PLUS Rfactor Rfree: 0.272 |