Repair of oxidative DNA damage in Drosophila melanogaster: identification and characterization of dOgg1, a second DNA glycosylase activity for 8-hydroxyguanine and formamidopyrimidines

In Drosophila, the S3 ribosomal protein has been shown to act as a DNA glyc osylase/AP lyase capable of releasing 8-hydroxyguanine (8-OH-Gua) in damage d DNA. Here we describe a second Drosophila protein (dOgg1) with 8-OH-Gua a nd abasic (AP) site DNA repair activities. The Drosophila OGG1 gene codes f or a protein of 327 amino acids, which shows 33 and 37% identity with the y east and human Ogg1 proteins, respectively. The DNA glycosylase activity of purified dOgg1 was investigated using gamma -irradiated DNA and gas chroma tography/ isotope dilution mass spectrometry (GC/IDMS). The dOgg1 protein e xcises 8-OH-Gua and 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyGua) f rom gamma -irradiated DNA. with k(cat)/K-M values of 21.0 x 10(-5) and 11.2 x 10(-5) (min(-1) nM(-1)), respectively. Enzymatic assays using oligodeoxy ribonucleotides containing a single lesion show that dOgg1 displays a marke d preference for DNA duplexes containing 8-OH-Gua, 8-OH-Ade or an AP site p laced opposite a cytosine, The cleavage of the 8-OH-Gua-containing strand r esults from the excision of the damaged base followed by a beta -eliminatio n reaction at the 3'-side of the resulting AP site. Cleavage of 8-OH-Gua.C duplex involves the formation of a reaction intermediate that is converted into a stable covalent adduct in the presence of sodium borohydre. dOgg1 co mplements the mutator phenotype of fpg mutY mutants of Escherichia coli. Wh ole-mount in situ hybridizations on tissues at different stages of Drosophi la development reveal that the dOGG1 messenger is expressed uniformly at a low level in cells in which mitotic division occurs. Therefore, Drosophila possesses two DNA glycosylase activities that can excise 8-OH-Gua and forma midopyrimidines from DNA, dOgg1 and the ribosomal protein S3.