Long-patch base excision DNA repair of 2-deoxyribonolactone prevents the formation of DNA-protein cross-links with DNA polymerase beta

J Biol Chem. 2005 Nov 25;280(47):39095-103. doi: 10.1074/jbc.M506480200. Epub 2005 Sep 27.


Oxidized abasic sites are a major form of DNA damage induced by free radical attack and deoxyribose oxidation. 2-Deoxyribonolactone (dL) is a C1'-oxidized abasic site implicated in DNA strand breakage, mutagenesis, and formation of covalent DNA-protein cross-links (DPCs) with repair enzymes such as DNA polymerase beta (polbeta). We show here that mammalian cell-free extracts incubated with Ape1-incised dL substrates under non-repair conditions give rise to DPCs, with a major species dependent on the presence of polbeta. DPC formation was much less under repair than non-repair conditions, with extracts of either polbeta-proficient or -deficient cells. Partial base excision DNA repair (BER) reconstituted with purified enzymes demonstrated that Flap endonuclease 1 (FEN1) efficiently excises a displaced oligonucleotide containing a 5'-terminal dL residue, as would be produced during long-patch (multinucleotide) BER. Simultaneous monitoring of dL repair and dL-mediated DPC formation demonstrated that removal of the dL residue through the combined action of strand-displacement DNA synthesis by polbeta and excision by FEN1 markedly diminished DPC formation with the polymerase. Analysis of the patch size distribution associated with DNA repair synthesis in cell-free extracts showed that the processing of dL residues is associated with the synthesis of >or=2 nucleotides, compared with predominantly single nucleotide replacement for regular abasic sites. Our observations reveal a cellular repair process for dL lesions that avoids formation of DPCs that would threaten the integrity of DNA and perhaps cell viability.

Publication types

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

MeSH terms

  • Animals
  • Base Sequence
  • Cell-Free System
  • Cells, Cultured
  • Cross-Linking Reagents
  • DNA / genetics
  • DNA / metabolism*
  • DNA Polymerase beta / metabolism*
  • DNA Repair / physiology*
  • Flap Endonucleases / metabolism
  • HeLa Cells
  • Humans
  • In Vitro Techniques
  • Mice
  • Proteins / metabolism*
  • Sugar Acids / metabolism*


  • Cross-Linking Reagents
  • Proteins
  • Sugar Acids
  • 2,4,5-trihydroxypentanoic acid gamma-lactone
  • DNA
  • DNA Polymerase beta
  • Flap Endonucleases
  • FEN1 protein, human