A simplified assay for O6-methylguanine-DNA methyltransferase activity and its application to human neoplastic and non-neoplastic tissues

A rapid assay of O⁶-MeG-DNA methyltransferase activity is described.Following incubation of cell extracts with O⁶-[³H]MeG-containingDNA, remaining radioactive DNA was hydrolyzed in trichloroaceticacid and separated from methylated radioactive protein by filtrationor centrifugation. Transfer of radioactive methyl from DNA toprotein was proportional to the amount of protein added, andwas not linear with time. More than 90% of the radioactivityprecipitated after acid hydrolyses was in S-methyl cysteineresidues. The method was used to measure O⁶-MeG-DNA methyltransferaseactivity in extracts of 24 neoplastic tissues from human organs.Although five tumor tissues had 28–84% lower activityof O⁶-MeG-DNA methyltransferase than the corresponding normaltissue from the same patient, higher or similar levels of activitywere found more frequently. Thus, a lack of O⁶-MeG-DNA methyltransferaseactivity in human tumors appears not to be a frequent event.The DNA repair enzyme uracil-DNA glycosylase was also measuredin the same extracts. Most frequently the level of uracil-DNAglycosylase activity was essentially similar in tumors and normaltissues but significantly higher or lower levels were also observed.