In vitro replication and repair of DNA containing a C2'-oxidized abasic site

Biochemistry. 2004 Dec 7;43(48):15217-22. doi: 10.1021/bi048360c.


Abasic lesions are unable to form Watson-Crick hydrogen bonds with nucleotides. Nonetheless, polymerase and repair enzymes distinguish between various oxidized abasic lesions, as well as from nonoxidized abasic sites (AP). The C2-AP lesion is produced when DNA is exposed to gamma-radiolysis. Its effects on polymerases and repair enzymes are unknown. A recently reported method for the chemical synthesis of oligonucleotides containing C2-AP at a defined site was utilized for studying the activity of Klenow exo(-) and repair enzymes on templates containing the lesion. The C2-AP lesion has a similar effect on Klenow exo(-) as do AP and C4-AP sites. Deoxyadenosine is preferentially incorporated opposite C2-AP, but extension of the primer past the lesion is strongly blocked. C2-AP is incised less efficiently by exonuclease III and endonuclease IV than are other abasic lesions. Furthermore, although a Schiff base between C2-AP and endonuclease III can be chemically trapped, the location of the 3'-phosphate alpha with respect to the aldehyde prevents beta-elimination associated with the lyase activity of type I base excision repair enzymes. The interactions of the C2'-oxidized abasic site with Klenow exo(-) and repair enzymes suggest that the lesion will be mutagenic and that it will be removed by strand displacement synthesis and flap endonuclease processing via a long patch repair mechanism.

Publication types

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

MeSH terms

  • Apurinic Acid / chemical synthesis
  • Apurinic Acid / genetics
  • Apurinic Acid / metabolism*
  • DNA Polymerase I / chemistry
  • DNA Polymerase I / metabolism
  • DNA Primers / chemical synthesis
  • DNA Primers / metabolism
  • DNA Repair*
  • DNA Replication*
  • DNA, Bacterial / chemical synthesis
  • DNA, Bacterial / genetics
  • DNA, Bacterial / metabolism*
  • Deoxyribonuclease IV (Phage T4-Induced) / chemistry
  • Deoxyribonuclease IV (Phage T4-Induced) / metabolism
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / metabolism
  • Exodeoxyribonucleases / chemistry
  • Exodeoxyribonucleases / metabolism
  • Nucleic Acid Heteroduplexes / chemical synthesis
  • Nucleic Acid Heteroduplexes / genetics
  • Nucleic Acid Heteroduplexes / metabolism
  • Nucleic Acid Hybridization
  • Oligonucleotides / chemical synthesis
  • Oligonucleotides / genetics
  • Oligonucleotides / metabolism
  • Oxidation-Reduction
  • Polynucleotides / chemical synthesis
  • Polynucleotides / genetics
  • Polynucleotides / metabolism*
  • Schiff Bases / chemistry
  • Schiff Bases / metabolism
  • Templates, Genetic


  • DNA Primers
  • DNA, Bacterial
  • Escherichia coli Proteins
  • Nucleic Acid Heteroduplexes
  • Oligonucleotides
  • Polynucleotides
  • Schiff Bases
  • apyrimidinic acid
  • Apurinic Acid
  • DNA Polymerase I
  • Exodeoxyribonucleases
  • exodeoxyribonuclease III
  • Deoxyribonuclease IV (Phage T4-Induced)
  • endonuclease IV, E coli