8-hydroxyguanine(7,8-dihydro-8-oxoguanine) DNA glycosylase and AP lyase activities of hOGG1 protein and their substrate specificity

Abstract Recently we cloned a structural human homolog ( hOGG1 ) of the yeast OGG1 ( yOGG1 ) gene that is involved in the excision repair of 8-hydroxyguanine (also known as 7,8-dihydro-8-oxoguanine; oh 8 Gua). hOGG1 protein shares 38% amino acid identity with yOGG1 protein. In this paper, we define the substrate specificity of oh 8 Gua DNA glycosylase and AP lyase activities of the hOGG1 protein. The oh 8 Gua released from oh 8 Gua containing DNA was measured by analysis with HPLC coupled with electrochemical detector (ECD) and cleavage sites in the DNA were identified by cleavage assay using gel electrophoresis. GST-hOGG1 protein possessed the oh 8 Gua DNA glycosylase/AP lyase activity and weak δ -elimination activity. oh 8 Gua opposite the C in duplex oligonucleotide was most efficiently released by GST-hOGG1 protein and oh 8 Gua opposite the T was also released, while oh 8 Gua opposite the G or A was very slowly done. The rank order of DNA cleavage efficiency was the same as that of oh 8 Gua glycosylase activity. Glycosylase/AP lyase activities and their substrate specificities of the GST-hOGG1 protein was similar to GST-yOGG1 protein but different from MutM protein. These results indicate that the dominant function of hOGG1 protein is a oh 8 Gua glycosylase reaction by specifically recognizing oh 8 Gua and pyrimidine opposite the oh 8 Gua and δ -elimination reaction in the same manner as yOGG1 protein. Thus, the hOGG1 gene is a functional human homolog of the yOGG1 gene on oh 8 Gua excision repair in spite of the low structural identity at amino acid level between hOGG1 and yOGG1 proteins.