8-hydroxyguanine (7,8-dihydro-8-oxoguanine) DNA glycosylase and AP lyase activities of hOGG1 protein and their substrate specificity.
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; oh8Gua), hOGG1 protein shares 38% amino acid identity with yOGG1 protein. In this paper, we define the substrate specificity of oh8Gua DNA glycosylase and AP lyase activities of the hOGG1 protein. The oh8Gua released from oh8Gua 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 oh8Gua DNA glycosylase/AP lyase activity and weak delta-elimination activity, oh8Gua opposite the C in duplex oligonucleotide was most efficiently released by GST-hOGG1 protein and oh8Gua opposite the T was also released, while oh8Gua opposite the G or A was very slowly done. The rank order of DNA cleavage efficiency was the same as that of oh8Gua 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 oh8Gua glycosylase reaction by specifically recognizing oh8Gua and pyrimidine opposite the oh8Gua and delta-elimination reaction in the same manner as yOGG1 protein. Thus, the hOGG1 gene is a functional human homolog of the yOGG1 gene on oh8Gua excision repair in spite of the low structural identity at amino acid level between hOGG1 and yOGG1 proteins.
Shinmura
K
K
Biology Division, National Cancer Center Research Institute, Tokyo, Japan.
Kasai
H
H
Sasaki
A
A
Sugimura
H
H
Yokota
J
J
D000080242:8-Hydroxy-2'-Deoxyguanosine; D019757:Carbon-Oxygen Lyases / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D004260:DNA Repair / Q000502:physiology*; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D043382:DNA-Formamidopyrimidine Glycosylase; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000378:metabolism; D043223:Deoxyribonuclease IV (Phage T4-Induced); D006801:Humans; D009699:N-Glycosyl Hydrolases / Q000378:metabolism*; D009692:Nucleic Acid Heteroduplexes / Q000032:analysis; D009838:Oligodeoxyribonucleotides / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013379:Substrate Specificity
D009692:Nucleic Acid Heteroduplexes; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D000080242:8-Hydroxy-2'-Deoxyguanosine; D004247:DNA; D043223:Deoxyribonuclease IV (Phage T4-Induced); D009699:N-Glycosyl Hydrolases; D043382:DNA-Formamidopyrimidine Glycosylase; D019757:Carbon-Oxygen Lyases; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D003849:Deoxyguanosine