Correlation between sequence-dependent glycosylase repair and the thermal stability of oligonucleotide duplexes containing 1,N-6-ethenoadenine

Previous experiments on DNA sequence context reported that base modificatio n, replication, and repair are affected by the nature of neighbor bases. We now report that repair by mammalian alkylpurine-DNA-N-glycosylases (APNG) of 15-mer oligonucleotides with a central 1,N-6-ethenoadenine (epsilon A), flanked by 5' and 3' tandem bases, is also highly sequence dependent. Oligo nucleotides with the central sequences -GG epsilon AGG- or -CC epsilon ACC- are repaired 3-5-fold more efficiently than those containing -AA epsilon A AA- or -TT epsilon ATT- when using human or mouse APNG, Melting curves of t he same duplexes showed that oligomers with G.C/C.G neighbors were less den atured than those with A.T/T.A neighbors at 37 degrees C. This sequence-dep endent difference in denaturation correlates with the relative thermodynami c stability of oligomers with G.C/C.G or A.T/T.A neighbors. The dependence of repair on thermal stability was confirmed by enzyme reactions performed over 0-45 degrees C. Under these conditions, repair of epsilon A flanked by G.C/C.G was dramatically increased at 37 degrees C with continuous increas e up to 45 degrees C, in contrast to that with flanking A.T/T.A pairs, whic h was in agreement with the degree of denaturation of these duplexes, These results indicate that the thermodynamic stability conferred by base pairs flanking epsilon A plays an essential role in maintaining the integrity of the duplex structure which is necessary for repair.