6FWK

The crystal structure of Pol2CORE-M644G in complex with DNA and an incoming nucleotide


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.50 Å
  • R-Value Free: 0.263 
  • R-Value Work: 0.223 
  • R-Value Observed: 0.225 

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Literature

Structural consequence of the most frequently recurring cancer-associated substitution in DNA polymerase epsilon.

Parkash, V.Kulkarni, Y.Ter Beek, J.Shcherbakova, P.V.Kamerlin, S.C.L.Johansson, E.

(2019) Nat Commun 10: 373-373

  • DOI: https://doi.org/10.1038/s41467-018-08114-9
  • Primary Citation of Related Structures:  
    6FWK, 6G0A, 6I8A

  • PubMed Abstract: 

    The most frequently recurring cancer-associated DNA polymerase ε (Pol ε) mutation is a P286R substitution in the exonuclease domain. While originally proposed to increase genome instability by disrupting exonucleolytic proofreading, the P286R variant was later found to be significantly more pathogenic than Pol ε proofreading deficiency per se. The mechanisms underlying its stronger impact remained unclear. Here we report the crystal structure of the yeast orthologue, Pol ε-P301R, complexed with DNA and an incoming dNTP. Structural changes in the protein are confined to the exonuclease domain, with R301 pointing towards the exonuclease site. Molecular dynamics simulations suggest that R301 interferes with DNA binding to the exonuclease site, an outcome not observed with the exonuclease-inactive Pol ε-D290A,E292A variant lacking the catalytic residues. These results reveal a distinct mechanism of exonuclease inactivation by the P301R substitution and a likely basis for its dramatically higher mutagenic and tumorigenic effects.


  • Organizational Affiliation

    Department of Medical Biochemistry and Biophysics, Umeå University, Umeå, SE-90187, Sweden.


Macromolecules

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Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
DNA polymerase epsilon catalytic subunit AA,
D [auth B]
1,191Saccharomyces cerevisiae S288CMutation(s): 1 
Gene Names: POL2DUN2YNL262WN0825
EC: 2.7.7.7 (PDB Primary Data), 3.1.11 (UniProt)
UniProt
Find proteins for P21951 (Saccharomyces cerevisiae (strain ATCC 204508 / S288c))
Explore P21951 
Go to UniProtKB:  P21951
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP21951
Sequence Annotations
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  • Reference Sequence

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Entity ID: 2
MoleculeChains LengthOrganismImage
DNA (5'-D(P*TP*AP*AP*CP*CP*GP*CP*GP*TP*TP*(DOC))-3')B [auth P],
E [auth C]
11synthetic construct
Sequence Annotations
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  • Reference Sequence

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Entity ID: 3
MoleculeChains LengthOrganismImage
DNA (5'-D(P*TP*CP*TP*TP*GP*AP*AP*CP*GP*CP*GP*GP*TP*TP*A)-3')C [auth T],
F [auth D]
15synthetic construct
Sequence Annotations
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  • Reference Sequence
Small Molecules
Ligands 3 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
DTP
Query on DTP

Download Ideal Coordinates CCD File 
G [auth A],
K [auth B]
2'-DEOXYADENOSINE 5'-TRIPHOSPHATE
C10 H16 N5 O12 P3
SUYVUBYJARFZHO-RRKCRQDMSA-N
FE
Query on FE

Download Ideal Coordinates CCD File 
J [auth A],
O [auth B]
FE (III) ION
Fe
VTLYFUHAOXGGBS-UHFFFAOYSA-N
CA
Query on CA

Download Ideal Coordinates CCD File 
H [auth A],
I [auth A],
L [auth B],
M [auth B],
N [auth B]
CALCIUM ION
Ca
BHPQYMZQTOCNFJ-UHFFFAOYSA-N
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.50 Å
  • R-Value Free: 0.263 
  • R-Value Work: 0.223 
  • R-Value Observed: 0.225 
  • Space Group: P 1 2 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 153.961α = 90
b = 70.342β = 112.82
c = 158.965γ = 90
Software Package:
Software NamePurpose
PHENIXrefinement
XDSdata reduction
XSCALEdata scaling
PHASERphasing
PDB_EXTRACTdata extraction

Structure Validation

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Entry History & Funding Information

Deposition Data


Funding OrganizationLocationGrant Number
Swedish Research CouncilSweden--
CancerfondenSweden--
Knut and Alice Wallenberg FoundationSweden--

Revision History  (Full details and data files)

  • Version 1.0: 2019-01-30
    Type: Initial release
  • Version 1.1: 2019-02-06
    Changes: Data collection, Database references
  • Version 1.2: 2024-05-08
    Changes: Advisory, Data collection, Database references, Derived calculations