Comparison of the in vitro replication of the 7-(2-oxoheptyl)-1,N2-etheno-2'-deoxyguanosine and 1,N2-etheno-2'-deoxyguanosine lesions by Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4)

Chem Res Toxicol. 2010 Aug 16;23(8):1330-41. doi: 10.1021/tx100082e.


Oligonucleotides were synthesized containing the 7-(2-oxoheptyl)-etheno-dGuo adduct, which is derived from the reaction of dGuo and the lipid peroxidation product 4-oxo-2-nonenal. The in vitro replication of 7-(2-oxoheptyl)-etheno-dGuo by the model Y-family polymerase Sulfolobus solfataricus P2 DNA Polymerase IV (Dpo4) was examined in two sequences. The extension products were sequenced using an improved LC-ESI-MS/MS protocol developed in our laboratories, and the results were compared to that of the 1,N(2)-etheno-dGuo adduct in the same sequence contexts. Both etheno adducts were highly miscoding when situated in 5'-TXG-3' local sequence contexts with <4% of the extension products being derived from error-free bypass. The major extension products resulted from the misinsertion of Ade opposite the adduct and a one-base deletion. The major extension products from replication of the etheno lesions in a 5'-CXG-3' local sequence context were the result of misinsertion of Ade, a one-base deletion, and error-free bypass. Other minor extension products were also identified. The 7-(2-oxoheptyl)-etheno-dGuo lesion resulted in a larger frequency of misinsertion of Ade, whereas the 1,N(2)-etheno-dGuo gave more of the one-base deletion product. Conformational studies of duplex DNA containing the 7-(2-oxoheptyl)-etheno-dGuo in a 5'-TXG-3' sequence context by NMR indicated the presence of a pH-dependent conformational transition, likely involving the glycosyl bond at the adducted guanosine; the pK(a) for this transition was lower than that observed for the 1,N(2)-epsilon-dGuo lesion. However, the 7-(2-oxoheptyl)-etheno-dGuo lesion, the complementary Cyt, and both flanking base pairs remained disordered at all pH values, which is attributed to the presence of the hydrophobic heptyl group of the 7-(2-oxoheptyl)-etheno-dGuo lesion. The altered pK(a) value and the structural disorder at the 7-(2-oxoheptyl)-etheno-dGuo lesion site, as compared to the same sequence containing the 1,N(2)-etheno-dGuo, may contribute to higher frequency of misinsertion of Ade.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural

MeSH terms

  • Chromatography, High Pressure Liquid
  • DNA Polymerase beta / metabolism*
  • Deoxyadenosines / metabolism*
  • Guanine / analogs & derivatives*
  • Guanine / chemistry
  • Guanine / metabolism
  • Molecular Structure
  • Oligonucleotides / biosynthesis*
  • Oligonucleotides / chemistry*
  • Oligonucleotides / metabolism
  • Spectrometry, Mass, Electrospray Ionization
  • Sulfolobus solfataricus / enzymology*


  • 7-(2-oxoheptyl)-1,N2-etheno-2'-deoxyguanine
  • Deoxyadenosines
  • Oligonucleotides
  • Guanine
  • 1,N(6)-ethenodeoxyadenosine
  • DNA Polymerase beta