3ISC

Binary complex of human DNA polymerase beta with an abasic site (THF) in the gapped DNA


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.00 Å
  • R-Value Free: 0.227 
  • R-Value Work: 0.186 
  • R-Value Observed: 0.186 

wwPDB Validation   3D Report Full Report


This is version 1.3 of the entry. See complete history


Literature

DNA polymerase beta substrate specificity: side chain modulation of the "A-rule".

Beard, W.A.Shock, D.D.Batra, V.K.Pedersen, L.C.Wilson, S.H.

(2009) J Biol Chem 284: 31680-31689

  • DOI: https://doi.org/10.1074/jbc.M109.029843
  • Primary Citation of Related Structures:  
    3ISB, 3ISC, 3ISD

  • PubMed Abstract: 

    Apurinic/apyrimidinic (AP) sites are continuously generated in genomic DNA. Left unrepaired, AP sites represent noninstructional premutagenic lesions that are impediments to DNA synthesis. When DNA polymerases encounter an AP site, they generally insert dAMP. This preferential insertion is referred to as the A-rule. Crystallographic structures of DNA polymerase (pol) beta, a family X polymerase, with active site mismatched nascent base pairs indicate that the templating (i.e. coding) base is repositioned outside of the template binding pocket thereby diminishing interactions with the incorrect incoming nucleotide. This effectively produces an abasic site because the template pocket is devoid of an instructional base. However, the template pocket is not empty; an arginine residue (Arg-283) occupies the space vacated by the templating nucleotide. In this study, we analyze the kinetics of pol beta insertion opposite an AP site and show that the preferential incorporation of dAMP is lost with the R283A mutant. The crystallographic structures of pol beta bound to gapped DNA with an AP site analog (tertrahydrofuran) in the gap (binary complex) and with an incoming nonhydrolyzable dATP analog (ternary complex) were solved. These structures reveal that binding of the dATP analog induces a closed polymerase conformation, an unstable primer terminus, and an upstream shift of the templating residue even in the absence of a template base. Thus, dATP insertion opposite an abasic site and dATP misinsertions have common features.


  • Organizational Affiliation

    Laboratory of Structural Biology, NIEHS, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.


Macromolecules

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Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
DNA polymerase beta335Homo sapiensMutation(s): 0 
Gene Names: POLB
EC: 2.7.7.7 (PDB Primary Data), 4.2.99 (PDB Primary Data)
UniProt & NIH Common Fund Data Resources
Find proteins for P06746 (Homo sapiens)
Explore P06746 
Go to UniProtKB:  P06746
PHAROS:  P06746
GTEx:  ENSG00000070501 
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP06746
Sequence Annotations
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  • Reference Sequence

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Entity ID: 2
MoleculeChains LengthOrganismImage
5'-D(P*GP*TP*CP*GP*G)-3'B [auth D]5synthetic construct
Sequence Annotations
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  • Reference Sequence

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Entity ID: 3
MoleculeChains LengthOrganismImage
5'-D(*GP*CP*TP*GP*AP*TP*GP*CP*GP*C)-3'C [auth P]10synthetic construct
Sequence Annotations
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  • Reference Sequence

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Entity ID: 4
MoleculeChains LengthOrganismImage
5'-D(*CP*CP*GP*AP*CP*(3DR)P*GP*CP*GP*CP*AP*TP*CP*AP*GP*C)-3'D [auth T]16synthetic construct
Sequence Annotations
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  • Reference Sequence
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.00 Å
  • R-Value Free: 0.227 
  • R-Value Work: 0.186 
  • R-Value Observed: 0.186 
  • Space Group: P 1 21 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 54.41α = 90
b = 79.3β = 105.8
c = 54.95γ = 90
Software Package:
Software NamePurpose
HKL-2000data collection
CNSrefinement
HKL-2000data reduction
HKL-2000data scaling
CNSphasing

Structure Validation

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Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2009-09-15
    Type: Initial release
  • Version 1.1: 2011-07-13
    Changes: Version format compliance
  • Version 1.2: 2019-02-06
    Changes: Data collection, Database references, Refinement description, Source and taxonomy, Structure summary
  • Version 1.3: 2023-09-06
    Changes: Data collection, Database references, Derived calculations, Refinement description