In isolated DNA, formamidopyrimidine-DNA glycosylase-sensitive sites determined by electrophoresis correspond to the amount of 8-oxo-7,8-dihydro-2'-deoxyguanosine by HPLC-ECD.
Several methods have been developed for determining the amount of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) in DNA. In the present study, we compared an electrophoretic method that uses formamidopyrimidine-DNA glycosylase (FPG protein) with a HPLC-ECD method. Firstly, we produced 8-oxodG in lambda DNA with methylene blue and visible light and cleaved it in one-half of the modified DNA enzymatically with FPG protein. Then, we determined the number of FPG protein-sensitive sites by electrophoresis (Y) and the number of 8-oxodGs by HPLC-ECD (X) per 10(5)dG of isolated DNA. Simple regression analysis of the data showed Y=1.07X+1.52 to be the most likely relationship. The correlation coefficient was 0.97. The values obtained by the two methods were very similar. This result is noteworthy because the number of FPG protein-sensitive sites determined by other methods have not yet come close to the number obtained by HPLC-ECD. Thus, this method might be more quantitative than other methods that measure FPG protein-sensitive sites. Another reason this electrophoresis method might be more useful than HPLC-ECD is that we can determine some other types of oxidative DNA damage well, by changing the DNA glycosylase.
Yoshida
R
R
National Institute of Industrial Health, 6-21-1, Nagao, Tama-ku, Kawasaki, 214-8585, Kanagawa, Japan. yoshidar@niih.go.jp
Ogawa
Y
Y
Takayanagi
S
S
D000080242:8-Hydroxy-2'-Deoxyguanosine; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000737:chemistry*; D043382:DNA-Formamidopyrimidine Glycosylase; D003849:Deoxyguanosine / Q000031:analogs & derivatives / Q000032:analysis*; D004586:Electrophoresis / Q000379:methods; D009699:N-Glycosyl Hydrolases / Q000378:metabolism*; D013379:Substrate Specificity
D000080242:8-Hydroxy-2'-Deoxyguanosine; D004247:DNA; D009699:N-Glycosyl Hydrolases; D043382:DNA-Formamidopyrimidine Glycosylase; D003849:Deoxyguanosine