The A and B forms are relatively stable for RNA and DNA, respectively, under physiological conditions. They must not be too stable, however, because processes, such as DNA replication (see here) and transcription, cannot occur unless the double-helix structure can be opened up. Denaturation refers to the loss of secondary (or tertiary) structure over large regions of a polynucleotide. Forces favoring denaturation of polynucleotides include

1. The electrostatic repulsion of the negative charges on the phosphate groups.

2. The higher entropy of the denatured state (the denatured form has more possible conformations than the double helix form, so the denatured form has greater randomness).

Forces stabilizing secondary structure, on the other hand, include

1. The hydrogen bonds between A and T and between G and C base pairs

2. The van der Waals interactions between the planar bases, which stack upon each other in the double helix structure (Figure 4.15).

The free energy change in going from a double helix structure to two individual random coils is given by

G = H -TS

1. S is positive due to the increased entropy of the random coil, so the -TS term makes a negative contribution to the free energy, thus favoring denaturation.

2. The electrostatic repulsion of the phosphate groups makes a negative contribution to H, thus favoring denaturation, too.

3. Energy must be expended to break the hydrogen bonds between base pairs and the van der Waals interactions between the stacked bases, so these forces make a positive contribution to H (and G), thus favoring the double helix structure. These positive contributions exceed the negative contributions due to the phosphate groups.

Therefore, the stability of a duplex is a function of temperature. At high temperatures, denaturation is favored (the -TS term dominates), but at lower temperatures duplexes are favored.The "melting temperature," Tm, corresponds to the temperature where G = 0. Tm is related to the ratio of (G+C) / (A+T) because G-C base pairs have more hydrogen bonds (3) than A-T base pairs (2) and thus have a more positive H (Figure 4.32).

Denaturation of DNA can be followed spectrophotometrically. Duplex DNA has bases packed into a helical configuration, causing them to absorb less light at 260 nm than a random coil (Figure 4.31). This phenomenon is called hypochromism. As followed in this manner, denaturation (also called strand melting) can be seen to occur over a fairly narrow temperature range. This means that the process is a cooperative transition; that is, it occurs all at once, not bit by bit or base pair by base pair. It would be very difficult for a single base to pop out of the stacked, hydrogen-bonded structure of the double helix.