3'-phosphodiesterase and 3'-->5' exonuclease activities of yeast Apn2 protein and requirement of these activities for repair of oxidative DNA damage.
In Saccharomyces cerevisiae, the AP endonucleases encoded by the APN1 and APN2 genes provide alternate pathways for the removal of abasic sites. Oxidative DNA-damaging agents, such as H(2)O(2), produce DNA strand breaks which contain 3'-phosphate or 3'-phosphoglycolate termini. Such 3' termini are inhibitory to synthesis by DNA polymerases. Here, we show that purified yeast Apn2 protein contains 3'-phosphodiesterase and 3'-->5' exonuclease activities, and mutation of the active-site residue Glu59 to Ala in Apn2 inactivates both these activities. Consistent with these biochemical observations, genetic studies indicate the involvement of APN2 in the repair of H(2)O(2)-induced DNA damage in a pathway alternate to APN1, and the Ala59 mutation inactivates this function of Apn2. From these results, we conclude that the ability of Apn2 to remove 3'-end groups from DNA is paramount for the repair of strand breaks arising from the reaction of DNA with reactive oxygen species.
Sealy Center for Molecular Science, University of Texas Medical Branch, Galveston, Texas 77555-1061, USA.
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D000409:Alanine / Q000737:chemistry; D001483:Base Sequence; D001665:Binding Sites; D019757:Carbon-Oxygen Lyases / Q000737:chemistry / Q000502:physiology*; D004249:DNA Damage; D004260:DNA Repair; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D043223:Deoxyribonuclease IV (Phage T4-Induced); D004305:Dose-Response Relationship, Drug; D043211:Exodeoxyribonuclease V; D005090:Exodeoxyribonucleases / Q000378:metabolism*; D018698:Glutamic Acid / Q000737:chemistry; D005982:Glutathione Transferase / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D009154:Mutation; D009708:Nucleotidases / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D029701:Saccharomyces cerevisiae Proteins; D013379:Substrate Specificity; D013997:Time Factors
D029701:Saccharomyces cerevisiae Proteins; D018698:Glutamic Acid; D006861:Hydrogen Peroxide; D005982:Glutathione Transferase; D005090:Exodeoxyribonucleases; D043211:Exodeoxyribonuclease V; D043223:Deoxyribonuclease IV (Phage T4-Induced); D009708:Nucleotidases; C040470:3'-nucleotidase; D019757:Carbon-Oxygen Lyases; C468953:APN2 protein, S cerevisiae; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D000409:Alanine; D010100:Oxygen