The nature of stacking interations in polynucleotides. Molecular states in Oligo- and polyribocytidylic acids by relaxation analysis.

The dynamics of the helix-coil transition of single-stranded poly(C) (polyribocytidylate) and CpC (cytidyly(3'-5')cytosine) was investigated by an improved cable temperature-jump technique. The single-strand relaxation was characterized by following the ultraviolet (uv) absorbance changes at 248 and 280 nm. Poly(C) and CpC showed single relaxation processes with amplitudes corresponding to those expected from equilibrium melting curves. The relaxation time contants in the range of 25-100 ns were independent of the nucleotide concentration, but strongly dependent upon temperature. Using thermodynanic parameters obtained from circular dichroism (CD) and uv absorbance melting curves, the following rate constants k (at 20 degrees C, 1.05 M ionic strength, pH 7) and activation enthalpies EA were calculated for poly (C): helix formation kR = 1.11 X 10(-7) s-1 (EAR = 2.6 kcal); helix dissociation kD = 2.1 X 10(6) s-1 (EAD = 11.9 kcal). The rate constants obtained for CpC were higher by a factor of about 2 in kR and 12 in kD, whereas the activation enthalpies closely corresponded to those found for the polymer. In addition to the single-stranded helix-coil relaxation, poly(C) and CpC exhibit a relaxation process with a time constant below 25 ns and maximum amplitudes at wavelengths lambda greater than or equal to 285 nm. The same process is found in cytidine and is attributed to hydration equilibria. The hydration reaction can be considered to be in equilibrium during the entire time range of the helix-coil transition and thus the data obtained for the helix-coil transition can be described by a simple two-state model. The rate parameters indicate the existence of relatively high energy barriers in the helix-coil transition and provide strong evidence evidence against an oscillating dimer model. If there is an ensemble of substates for one of the states (as may be expected for the coil form), the energy difference between the populated substates is small compared with the energy difference between the major conformational states.