Inhibitory Effects of Mismatch Binding Molecules on the Repair Reaction of Uracil-Containing DNA.

Abstract

The stable R-loop formed during transcription induces enzyme-mediated deamination of cytosine, and the uracil in the DNA produced activates the base excision repair (BER) pathway. DNA cleavage involved in the BER pathway is thought to be one of the possible causes of trinucleotide repeat instability. Here, we performed an in vitro assay to investigate the effect of a DNA-binding small molecule, naphthyridine carbamate dimer (NCD), on BER enzyme reactions. The gel electrophoretic mobility shift assay (EMSA) and thermal melting analysis revealed the binding of NCD to a 5'-XGG-3'/5'-XGG-3' triad (X = C or U or apurinic/apyrimidinic site), which is a mimic of a BER enzyme substrate. Polyacrylamide gel electrophoresis (PAGE) of the reaction products of these substrates with hSMUG1 and APE1 enzymes in the presence of NCD showed that NCD interfered with the repair reaction in the 5'-XGG-3'/5'-XGG-3' triad. These findings would broaden the potential of small molecules in modulating trinucleotide repeat instability.