Substantial difference between temperature dependencies of dsDNA predenaturation process obtained by voltammetry and spectroscopy

Abstract

The electrooxidation of double stranded DNA has been studied in a wide temperature range by cyclic and square wave voltammetries and the results were compared with UV-Vis and CD spectroscopies. A twofold increase of dsDNA voltammetric peaks related to the conformation changes preceding the denaturation process was found to begin at a temperature lower than the well known increase of dsDNA absorbance upon denaturation (the melting point) by circa 20 °C. Since the dsDNA voltammetric peaks are related directly to the electrooxidation of guanine and adenine, early conformational changes in guanine–cytosine (G–C) and adenine–thymine (A–T) pairs in dsDNA are possible reasons for the increase of the voltammetric peaks. The above observations prove that the voltammetric methods are much more sensitive to the changes in guanine and adenine base-pairs conformations and indicate increased exposition of guanine and adenine in the reversible stages that precede the denaturation process of dsDNA. Interestingly, two distinct ranges of change in ellipticity in CD spectra correlate well to the changes obtained by voltammetry and UV spectroscopy, respectively. We also checked the influence of an intercalator on the dsDNA denaturation (temperature) plots. The spectroscopic measurements indicated an increase in the stability of the structure of the double stranded helix upon the addition of the intercalator, while voltammetry showed also that the exposition of guanine and adenine for the electrooxidation drops substantially in the region preceding the denaturation process.