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 A PN2 genes provide alternate pathways for the removal of abasic sites. Oxida tive DNA-damaging agents, such as H2O2, produce DNA strand breaks which con tain 3'-phosphate or 3'-phosphoglycolate termini. Such 3' termini are inhib itory to synthesis by DNA polymerases, Here, we show that purified yeast Ap n2 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, gene tic studies indicate the involvement of APN2 in the repair of H2O2-induced DNA damage in a pathway alternate to APN1, and the Ala59 mutation inactivat es 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 str and breaks arising from the reaction of DNA with reactive oxygen species.