Proton exchange and local stability in a DNA triple helix containing a G.TA triad

Recognition of a thymine-adenine base pair in DNA by triplex-forming oligon ucleotides can be achieved by a guanine through the formation of a G.TA tri ad within the parallel triple helix motif. In the present work, we provide the first characterization of the stability of individual base pairs and ba se triads in a DNA triple helix containing a G.TA triad. The DNA investigat ed is the intramolecular triple helix formed by the 32mer d(AGATAGAACCCCTTC TATCTTATATCTGTCTT). The exchange rates of imino protons in this triple heli x have been measured by nuclear magnetic resonance spectroscopy using magne tization transfer from water and real-time exchange. The exchange rates are compared with those in a homologous DNA triple helix in which the G.TA tri ad is replaced by a canonical C+.GC triad. The results indicate that, in th e G.TA triad, the stability of the Watson-Crick TA base pair is comparable with that of AT base pairs in canonical T.AT triads. However, the presence of the G.TA triad destabilizes neighboring triads by 0.6-1.8 kcal/mol at 1 degreesC. These effects extend to triads that are two positions removed fro m the site of the G.TA triad. Therefore, the lower stability of DNA triple helices containing G.TA triads originates, in large part, from the energeti c effects of the G.TA triad upon the stability of canonical triads located in its vicinity.