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Notes: We have studied the hydration of Na-DNA and Li-DNA fibers and films, measuring water contents, x-ray fiber diffraction patterns, low-frequency Raman spectra (below 100 cm-1), high-frequency Raman spectra (600-1000 cm-1), and swelling, as a function of relative humidity. Most samples gain weight equilibrium (though not conformational equilibrium) in one day. The volume occupied by a base pair as the DNA is hydrated (obtained from the x-ray and swelling data) shows anomalies for the case of Na-DNA in the region where the A-form occurs. Our Raman and x-ray data reproduce the well-known features of the established conformational transitions, but we find evidence in the Raman spectra and optical properties of a transition to what may be a disordered B-like conformation in Na-DNA below 40% relative humidity. We have studied the effects of crystallinity on the A to B transition. We find that the transition to the B-form is impeded in highly crystalline samples. In most samples, the transition occurs in three days (after putting the sample at 92% relative humidity) but in highly crystalline samples, the transition may take months. By comparing the high-frequency Raman spectra of highly ordered and disordered films, we show that the extent of crystallinity controls the amount of A-DNA formed when ethanol is used to dehydrate the films. We show that rapid dehydration (by laser heating) does not result in a B to A transition. A fiber that gives A-type x-ray reflections probably contains B-like material in noncrystalline regions. The low-frequency Raman spectrum is dominated by a band at about 25 cm-1 in both Na- and Li-DNA. Another band is seen near 35 cm-1 in Na-DNA at humidities where the sample is in the A-form. In contrast to earlier reports, we find that the Raman intensity does not depend on fiber orientation relative to the scattering vector. The “35-cm-1” band is largely depolarized (i.e. vertical polarization incident and horizontal polarization scattered, VH, or vice versa, HV) while the “25-cm-1” band appears in both VV, VH and HV polarizations. These bands are all weaker in HH polarization. The “25-cm-1” band may be due to a shearing motion of the phosphates and their associated counterions, while the “35-cm-1” band may be characteristic of A-DNA crystallites. We consider mass-loading, relaxational coupling to the hydration shell, and softening of interatomic potentials as possible explanations of the observed softening of the low-frequency Raman bands on hydration. Relaxation data suggest that the added water binds tightly (on these time scales) and a mass-loading model accounts for the observed softening rather well.We conclude that the A to B transition is not driven by softening of the “25-cm-1” band. Rather, it is most probably a consequence of crystal-packing forces, with the more regular A-form favored in crystals when these forces are strong.