Importance of isothermal titration calorimetry for the detection of the direct binding of metal ions to mismatched base pairs in duplex DNA.

Abstract

Metal ion-nucleic acid interactions contribute substantially to the structure and biological activity of nucleic acids and have a wide range of potential applications in nanotechnology. In this study, we examined the interactions between metal ions and mismatched base pairs in duplex DNA to reveal the underlying molecular mechanism. UV melting analyses showed that the melting temperature (Tm) of a 21-base pair duplex DNA with each of the C-A, C-C and C-T mismatched base pairs increased upon the addition of Ag+. However, isothermal titration calorimetry (ITC) demonstrated that Ag+ only bound to the C-C mismatched base pair of the duplex DNA to form C-Ag-C bonds, without binding to the C-A and C-T mismatches. These results showed that Tm increased even when metal ions did not bind to the mismatched base pairs of the duplex DNA. Although the increase in Tm upon the addition of the metal ions is often used to detect metal ion binding to mismatched base pairs of duplex DNA, these results indicated that UV melting analyses are unable to detect the direct binding of metal ions to the mismatched base pairs. Because ITC analyses directly detect the heat derived from metal ion binding to mismatched base pairs of duplex DNA, we concluded that this may be an effective detection approach.