title: The major human AP endonuclease (Ape1) is involved in the nucleotide incision repair pathway.
abstract: In nucleotide incision repair (NIR), an endonuclease nicks oxidatively damaged DNA in a DNA glycosylase-independent manner, providing the correct ends for DNA synthesis coupled to the repair of the remaining 5'-dangling modified nucleotide. This mechanistic feature is distinct from DNA glycosylase-mediated base excision repair. Here we report that Ape1, the major apurinic/apyrimidinic endonuclease in human cells, is the damage- specific endonuclease involved in NIR. We show that Ape1 incises DNA containing 5,6-dihydro-2'-deoxyuridine, 5,6-dihydrothymidine, 5-hydroxy-2'-deoxyuridine, alpha-2'-deoxyadenosine and alpha-thymidine adducts, generating 3'-hydroxyl and 5'-phosphate termini. The kinetic constants indicate that Ape1-catalysed NIR activity is highly efficient. The substrate specificity and protein conformation of Ape1 is modulated by MgCl2 concentrations, thus providing conditions under which NIR becomes a major activity in cell-free extracts. While the N-terminal region of Ape1 is not required for AP endonuclease function, we show that it regulates the NIR activity. The physiological relevance of the mammalian NIR pathway is discussed.
authors.affiliation: Groupe Réparation de l'ADN', UMR 8113 CNRS, LBPA-ENS Cachan, Institut Gustave Roussy, 39, rue Camille Desmoulins, 94805 Villejuif Cedex, France.
authors.forename: Laurent
authors.initials: L
authors.lastname: Gros
authors.forename: Alexander A
authors.initials: AA
authors.lastname: Ishchenko
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Ide
authors.forename: Rhoderick H
authors.initials: RH
authors.lastname: Elder
authors.forename: Murat K
authors.initials: MK
authors.lastname: Saparbaev
mesh_terms: D002457:Cell Extracts; D004249:DNA Damage; D004260:DNA Repair; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000737:chemistry / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D009711:Nucleotides / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D011487:Protein Conformation / Q000187:drug effects; D013379:Substrate Specificity
chemical_list: D002457:Cell Extracts; D009711:Nucleotides; D009838:Oligodeoxyribonucleotides; C085108:APEX1 protein, human; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D008274:Magnesium