The protective role of thiol reducing agents in the in vitro inhibition of rat liver DNA methylase by direct acting carcinogens.

The direct-acting carcinogens N-acetoxy-N-acetyl-2-aminofluorene (AcAAF), methyl nitrosourea (MNU), and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) were tested for their ability to inhibit highly purified, rat liver DNA methylase in vitro. Fifty percent inhibition of DNA methylase activity was achieved with 4.3 mM AcAAF, 47 mM MNU and 2.8 mM MNNG. When the enzyme was reassayed in the presence and absence of dithiothreitol, it was shown that DNA methylase was protected by increasing amounts of the thiol reducing agent. When other thiol reducing agents were tested for their ability to protect DNA methylase from carcinogen damage, a differential protective ability was observed. Dithiothreitol, beta-mercaptoethanol, and reduced glutathione were effective in protecting DNA methylase from carcinogen inhibition, while the effect of cysteine was intermediary and the effect of ergothioneine was minimal. These results may be related to the hypomethylation of DNA observed in several cancers, suggesting that the carcinogens achieve this effect at least in part by inhibiting crucial sulfhydryl group(s) in the methylase molecule. These data also suggest that various intracellular thiols may play an important role in protecting DNA-modifying enzymes from carcinogen damage.