Abstract
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.
Publication types
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Research Support, U.S. Gov't, Non-P.H.S.
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Alanine / chemistry
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Base Sequence
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Binding Sites
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Carbon-Oxygen Lyases / chemistry
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Carbon-Oxygen Lyases / physiology*
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DNA Damage*
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DNA Repair*
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DNA-(Apurinic or Apyrimidinic Site) Lyase
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Deoxyribonuclease IV (Phage T4-Induced)
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Dose-Response Relationship, Drug
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Exodeoxyribonuclease V
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Exodeoxyribonucleases / metabolism*
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Glutamic Acid / chemistry
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Glutathione Transferase / metabolism
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Hydrogen Peroxide / pharmacology
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Hydrogen-Ion Concentration
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Molecular Sequence Data
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Mutation
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Nucleotidases / metabolism*
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Oxygen / metabolism*
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Saccharomyces cerevisiae / enzymology
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Saccharomyces cerevisiae Proteins*
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Substrate Specificity
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Time Factors
Substances
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Saccharomyces cerevisiae Proteins
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Glutamic Acid
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Hydrogen Peroxide
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Glutathione Transferase
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Exodeoxyribonucleases
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Exodeoxyribonuclease V
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Deoxyribonuclease IV (Phage T4-Induced)
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Nucleotidases
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3'-nucleotidase
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Carbon-Oxygen Lyases
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APN2 protein, S cerevisiae
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DNA-(Apurinic or Apyrimidinic Site) Lyase
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Alanine
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Oxygen