Counteracting the mutagenic effect of hydrolytic deamination of DNA 5-methylcytosine residues at high temperature: DNA mismatch N-glycosylase Mig.Mth of the thermophilic archaeon Methanobacterium thermoautotrophicum THF

Abstract

Spontaneous hydrolytic deamination of DNA 5-methylcytosine residues gives rise to T/G mismatches which are pre-mutagenic lesions requiring DNA repair. For fundamental reasons, the significance of this and other processes lowering genetic fidelity must be accentuated at elevated temperatures, making thermophilic organisms attractive objects for studying how cells cope with thermal noise threatening the integrity of their genetic information. Gene mig of Methanobacterium thermo-autotrophicum THF, an anaerobic archaeon with an optimal growth temperature of 65°C, was isolated and its product (Mig.Mth; EC3.2.2-) shown to be a T/G-selective DNA thymine N-glycosylase with the properties required for counteracting the mutagenic effect of hydrolytic 5-meC deamination. The enzyme acts on T/G and U/G oppositions with similar efficiency; G/G, A/G, T/C and U/C are minor substrates; no other opposition of common nucleobases is attacked and no removal of U from single-stranded DNA is observed. Substrate preferences are modulated by sequence context. Together with the results presented here, one example of an enzyme directed against the hydrolytic deamination damage of 5-meC is known from each of the three phylogenetic kingdoms; entry into the repair pathway is glycosylytic in the eukaryotic and the archaeal case, whereas the eubacterial repair starts with an endonucleolytic DNA incision.