The stability of DNA duplex and triplex structures not only depends on molecular forces such as base pairing or tripling or electrostatic interactions but also is sensitive to its aqueous environment. This paper presents data on the melting of Escherichia coli and poly(dA).poly(dT) duplex DNA and on the poly(dT).poly(dA). poly(dT) triplex in a variety of media to assess the contributions from the osmotic status and salt content of the media. The effects of volume exclusion on the stability of the DNA structures are also studied. From thermal transition measurements in the presence of low-molecular weight osmotic stressors, the number of water molecules released upon melting is found to be four waters per base pair for duplex melting and one water for the conversion of triplex to single-strand and duplex. The effects of Na+ counterion binding are also determined in ethylene glycol solutions so that the variation of counterion binding with water activity is evaluated. The data show that there is a modest decrease in the extent of counterion binding for both duplex and triplex as water activity decreases. Finally, using larger polyethylene glycol cosolutes, the effects on melting of volume exclusion by the solutes are assessed, and the results correlated with simple geometric models for the excluded volume. These results point out that DNA stability is sensitive to important conditions in the environment of the duplex or triplex, and thus, conformation and reactivity can be influenced by these solution conditions.

G-quadruplex formation in the sequences 5′-(TTAGGG)n and 5′(TTAGGG)nTT (n=4,8,12) was studied using circular dichroism, sedimentation velocity, differential scanning calorimetry and molecular dynamics simulations. Sequences containing 8 and 12 repeats formed higher-order structures with two and three contiguous quadruplexes, respectively. Plausible structures for these sequences were determined by molecular dynamics simulations followed by experimental testing of predicted hydrodynamic properties by sedimentation velocity. These structures featured folding of the strand into contiguous quadruplexes with mixed hybrid conformations. Thermodynamic studies showed the strands folded spontaneous to contain the maximum number contiguous quadruplexes. For the sequence 5′(TTAGGG)12TT, more than 90% of the strands contained completely folded structures with three quadruplexes. Statistical mechanical-based deconvolution of thermograms for three quadruplex structures showed that each quadruplex melted independently with unique thermodynamic parmameters. Thermodynamic analysis revealed further that quadruplexes in higher-ordered structures were destabilized relative to their monomeric counterparts, with unfavorable coupling free energies. Quadruplex stability thus depends critically on the sequence and structural context.

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