Clostridium acetobutylicum 8-oxoguanine DNA glycosylase (Ogg) differs from eukaryotic Oggs with respect to opposite base discrimination

During repair of damaged DNA, the oxidized base 8-oxoguanine (8-oxoG) is removed by 8-oxoguanine-DNA glycosylase (Ogg) in eukaryotes and most archaea, whereas in most bacteria it is removed by formamidopyrimidine-DNA glycosylase (Fpg). We report the first characterization of a bacterial Ogg, Clostridium acetobutylicum Ogg (CacOgg). Like human OGG1 and Escherichia coli Fpg (EcoFpg), CacOgg excised 8-oxoguanine. However, unlike hOGG1 and EcoFpg, CacOgg showed little preference for the base opposite the damage during base excision and removed 8-oxoguanine from single-stranded DNA. Thus, our results showed unambiguous qualitative functional differences in vitro between CacOgg and both hOGG1 and EcoFpg. CacOgg differs in sequence from the eukaryotic enzymes at two sequence positions, M132 and F179, which align with amino acids (R154 and Y203) in human OGG1 (hOGG1) found to be involved in opposite base interaction. To address the sequence basis for functional differences with respect to opposite base interactions, we prepared three CacOgg variants, M132R, F179Y, and M132R/F179Y. All three variants showed a substantial increase in specificity for 8-oxoG center dot C relative to 8-oxoG center dot A. While we were unable to definitively associate these qualitative functional differences with differences in selective pressure between eukaryotes, Clostridia, and other bacteria, our results are consistent with the idea that evolution of Ogg function is based on kinetic control of repair.