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

Published

December 1, 2000

Author(s)

C. Dherin, M. Dizdaroglu, H. Doerflinger, S. Boiteux, J. P. Radicella

Abstract

In Drosophila, the S3 ribosomal protein has been shown to act as a DNA glycosylase/AP lyase capable of releasing 8-hydroxyguanine (8-OH-Gua) in damaged DNA. Here we describe a second Drosophila protein (dOggl) with 8-OH-Gua and abasic (AP) site DNA repair activities. The Drosophila OGG1 gene codes for a protein of 327 amino acids, which shows 33 and 37% identity with the yeast and human Ogg1 proteins, respectively. The DNA glycosylase activity of purified dOgg1 was investigated using γ-irradiated DNA and gas chromatography/isotope dilution mass spectrometry (GC/IDMS). The dOgg1 protein excises 8-OH-Gua and 2.6-diamino-4-hydroxy-5-formamidopyrimidine (FapyGua) from γ-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 oligodeoxyribonucleotides containing a single lesion show that dOgg1 displays a marked preference for DNA duplexes containing 8-OH-Gua, 8-OH-Ade or AP site placed opposite a cytosine. The cleavage of the 8-OH-Gua-containing strand results from the excision of the damaged DNA base followed by a Β-elimination 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 complements the mutator phenotype of fpg mutY mutant of Escherichia coli. Whole-mount in situ hybridizations on tissues at different stages of Drosophila 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 formamidopyrimidines from DNA, dOgg1 and the ribosomal Protein S3.

Citation

Nucleic Acids Research

Volume

28

Issue

No. 23

Pub Type

Journals

Keywords

8-hydroxyguanine, base-excision repair, drosophila melanogaster, formamidopyrimidine, gas chromatography/mass spectrometry, oxidative DNA damage

Citation

Dherin, C. , Dizdaroglu, M. , Doerflinger, H. , Boiteux, S. and Radicella, J. (2000), Repair of oxidative DNA damage in Drosophila melanogaster: identification and characterization of dOgg1, a second DNA glycosylase activity for 8-hydroxyguanine and formamidopyrimidines, Nucleic Acids Research (Accessed December 26, 2024)