Melting Behavior of DNA Complexes With Joined Triple and Duplex Motifs

Abstract

One focus of our laboratory is to understand how sequence, duplex and triplex stabilities, and solution conditions affect the melting behavior of complex DNA structures. We used a combination of UV and circular dichroism (CD) spectroscopies and differential scanning calorimetry (DSC) techniques to obtain a full thermodynamic description of the melting behavior of seven DNA complexes involving joined triplex and duplex motifs. Six of these complexes are formed intramolecularly while the seventh forms an intermolecular complex. The circular dichroism spectra at low temperatures indicated that all complexes maintained the “B” conformation. UV and DSC melting curves of each complex show biphasic or triphasic transitions. However, the transition temperatures, TMs, of the intramolecular complexes remained constant with increasing strand concentration, while the TM of the intermolecular complex did not. This confirms their molecularity. Deconvolution of the DSC thermograms allowed us to determine standard thermodynamic profiles for the transitions of each complex. For each transition, the favorable folding free energy terms result from the characteristic compensation of a favorable enthalpy and unfavorable entropy contribution. The magnitude of these thermodynamic parameters (and associated TMs) indicate that the overall folding of each complex depends on several factors: a) the extent of the favorable heat contributions (formation of base pair and base triplet stacks that are compensated with both the ordering of the oligonucleotide and the putative uptake of protons and ions; b) inclusion of the more stable C+GC base triplets; c) stabilizing the duplex stem of the complex; d) complex molecularity; and e) solution conditions, such as pHand salt concentration. Overall, the melting behavior of each complex corresponds to the initial disruption of the triplex motif (removal of the third strand) followed by the partial or full unfolding of the duplex stem.