4TQS

Ternary complex of Y-family DNA polymerase Dpo4 with (5'S)-8,5'-Cyclo-2'-deoxyguanosine and dCTP


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.06 Å
  • R-Value Free: 0.290 
  • R-Value Work: 0.228 
  • R-Value Observed: 0.231 

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Ligand Structure Quality Assessment 


This is version 2.1 of the entry. See complete history


Literature

Kinetic and Structural Mechanisms of (5'S)-8,5'-Cyclo-2'-deoxyguanosine-Induced DNA Replication Stalling.

Xu, W.Ouellette, A.M.Wawrzak, Z.Shriver, S.J.Anderson, S.M.Zhao, L.

(2015) Biochemistry 54: 639-651

  • DOI: https://doi.org/10.1021/bi5014936
  • Primary Citation of Related Structures:  
    4TQR, 4TQS

  • PubMed Abstract: 

    The (5'S)-8,5'-cyclo-2'-deoxyguanosine (S-cdG) lesion is produced from reactions of DNA with hydroxyl radicals generated from ionizing radiation or endogenous oxidative metabolisms. An elevated level of S-cdG has been detected in Xeroderma pigmentosum, Cockayne syndrome, breast cancer patients, and aged mice. S-dG blocks DNA replication and transcription in vitro and in human cells and produces mutant replication and transcription products in vitro and in vivo. Major cellular protection against S-dG includes nucleotide excision repair and translesion DNA synthesis. We used kinetic and crystallographic approaches to elucidate the molecular mechanisms of S-cdG-induced DNA replication stalling using model B-family Sulfolobus solfataricus P2 DNA polymerase B1 (Dpo1) and Y-family S. solfataricus P2 DNA polymerase IV (Dpo4). Dpo1 and Dpo4 inefficiently bypassed S-cdG with dCTP preferably incorporated and dTTP (for Dpo4) or dATP (for Dpo1) misincorporated. Pre-steady-state kinetics and crystallographic data mechanistically explained the low-efficiency bypass. For Dpo1, S-cdG attenuated Kd,dNTP,app and kpol. For Dpo4, the S-cdG-adducted duplex caused a 6-fold decrease in Dpo4:DNA binding affinity and significantly reduced the concentration of the productive Dpo4:DNA:dCTP complex. Consistent with the inefficient bypass, crystal structures of Dpo4:DNA(S-cdG):dCTP (error-free) and Dpo4:DNA(S-cdG):dTTP (error-prone) complexes were catalytically incompetent. In the Dpo4:DNA(S-cdG):dTTP structure, S-cdG induced a loop structure and caused an unusual 5'-template base clustering at the active site, providing the first structural evidence of the previously suggested template loop structure that can be induced by a cyclopurine lesion. Together, our results provided mechanistic insights into S-cdG-induced DNA replication stalling.


  • Organizational Affiliation

    Department of Chemistry and Biochemistry and ‡Science of Advanced Materials Program, Central Michigan University , Mount Pleasant, Michigan 48859, United States.


Macromolecules

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Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
DNA polymerase IV
A, B
358Saccharolobus solfataricus P2Mutation(s): 0 
Gene Names: dbhdpo4SSO2448
EC: 2.7.7.7
UniProt
Find proteins for Q97W02 (Saccharolobus solfataricus (strain ATCC 35092 / DSM 1617 / JCM 11322 / P2))
Explore Q97W02 
Go to UniProtKB:  Q97W02
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupQ97W02
Sequence Annotations
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  • Reference Sequence

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Entity ID: 2
MoleculeChains LengthOrganismImage
DNA (5'-D(*GP*GP*GP*GP*GP*AP*AP*GP*GP*AP*TP*T)-3')C,
E [auth P]
12synthetic construct
Sequence Annotations
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  • Reference Sequence

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Entity ID: 3
MoleculeChains LengthOrganismImage
DNA (5'-D(P*TP*CP*AP*CP*(2LF)P*GP*AP*AP*TP*CP*CP*TP*TP*CP*CP*CP*CP*C)-3')D,
F [auth T]
18synthetic construct
Sequence Annotations
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  • Reference Sequence
Small Molecules
Ligands 3 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
DCP
Query on DCP

Download Ideal Coordinates CCD File 
G [auth A],
J [auth B]
2'-DEOXYCYTIDINE-5'-TRIPHOSPHATE
C9 H16 N3 O13 P3
RGWHQCVHVJXOKC-SHYZEUOFSA-N
DOC
Query on DOC

Download Ideal Coordinates CCD File 
M [auth C],
N [auth P]
2',3'-DIDEOXYCYTIDINE-5'-MONOPHOSPHATE
C9 H14 N3 O6 P
RAJMXAZJKUGYGW-POYBYMJQSA-N
MG
Query on MG

Download Ideal Coordinates CCD File 
H [auth A],
I [auth A],
K [auth B],
L [auth B]
MAGNESIUM ION
Mg
JLVVSXFLKOJNIY-UHFFFAOYSA-N
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.06 Å
  • R-Value Free: 0.290 
  • R-Value Work: 0.228 
  • R-Value Observed: 0.231 
  • Space Group: P 1 21 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 52.12α = 90
b = 184.242β = 110.09
c = 52.124γ = 90
Software Package:
Software NamePurpose
PHENIXrefinement
HKL-2000data scaling
Cootmodel building
MOLREPphasing
PDB_EXTRACTdata extraction

Structure Validation

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Ligand Structure Quality Assessment 


Entry History 

Deposition Data

  • Released Date: 2015-01-14 
  • Deposition Author(s): Zhao, L.

Revision History  (Full details and data files)

  • Version 1.0: 2015-01-14
    Type: Initial release
  • Version 1.1: 2015-01-21
    Changes: Database references
  • Version 1.2: 2015-02-04
    Changes: Database references
  • Version 1.3: 2017-11-22
    Changes: Database references, Derived calculations, Other, Refinement description, Source and taxonomy, Structure summary
  • Version 2.0: 2019-10-30
    Changes: Advisory, Atomic model, Data collection, Derived calculations, Refinement description
  • Version 2.1: 2023-12-27
    Changes: Data collection, Database references, Derived calculations