DNA lesion alters global conformational dynamics of Y-family DNA polymerase during catalysis

J Biol Chem. 2012 Apr 13;287(16):13040-7. doi: 10.1074/jbc.M112.345835. Epub 2012 Feb 23.

Abstract

A major product of oxidative damage to DNA, 8-oxo-7,8-dihydro-2'-deoxyguanine (8-oxoG), can lead to genomic mutations if it is bypassed unfaithfully by DNA polymerases in vivo. However, our pre-steady-state kinetic studies show that DNA polymerase IV (Dpo4), a prototype Y-family enzyme from Sulfolobus solfataricus, can bypass 8-oxoG both efficiently and faithfully. For the first time, our stopped-flow FRET studies revealed that a DNA polymerase altered its synchronized global conformational dynamics in response to a DNA lesion. Relative to nucleotide incorporation into undamaged DNA, three of the four domains of Dpo4 undertook different conformational transitions during 8-oxoG bypass and the subsequent extension step. Moreover, the rapid translocation of Dpo4 along DNA induced by nucleotide binding was significantly hindered by the interactions between the embedded 8-oxoG and Dpo4 during the extension step. These results unprecedentedly demonstrate that a Y-family DNA polymerase employs different global conformational dynamics when replicating undamaged and damaged DNA.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Bacterial Proteins* / chemistry
  • Bacterial Proteins* / genetics
  • Bacterial Proteins* / metabolism
  • Catalysis
  • DNA Damage / physiology
  • DNA Polymerase beta* / chemistry
  • DNA Polymerase beta* / genetics
  • DNA Polymerase beta* / metabolism
  • DNA, Bacterial / metabolism
  • Fluorescence Resonance Energy Transfer
  • Guanine / analogs & derivatives
  • Guanine / metabolism
  • Oxidative Stress / physiology*
  • Protein Conformation
  • Protein Structure, Tertiary
  • Sulfolobus solfataricus / enzymology*
  • Sulfolobus solfataricus / genetics*

Substances

  • 7,8-dihydro-8-oxoguanine
  • Bacterial Proteins
  • DNA, Bacterial
  • Guanine
  • DNA Polymerase beta