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A high-angle neutron fibre diffraction study of the hydration of A-DNA has been performed using the single-crystal diffractometer D19 at the Institut Laue-Langevin (Grenoble, France). The sample was prepared using deuterated DNA extracted from E. Coli cells cultured on deuterated nutrients. In common with our previous neutron fibre diffraction studies of DNA, this work exploits the ability to isotopically replace H2O around the DNA by D2O. However this study benefitted additionally from the fact that the hydrogen atoms which are covalently bonded to carbon atoms in the DNA sugars and bases were replaced by deuterium so that incoherent scattering and absorption effects were minimised. Successive cycles of Fourier synthesis and Fourier difference synthesis allowed water peaks to be identified and their positional and occupancy parameters to be refined against the observed diffraction data. The results confirm the main hydration features noted in our earlier studies with a clear network of water running along the inside edge of the major groove linking successive OI phosphate oxygen atoms. The central core of water running along the axis of the double helix is very much clearer in this work. Additionally this study shows chains of ordered water lying in the centre of the major groove.

The solution conformation and internal motions of five superhelical DNAs between 2100 and 10200 base-pairs in length have been characterized by dynamic light scattering (DLS). Variations in the diffusion coefficients and rotational relaxation times with molecular weight are both indicative of an anisotropic extended structure of these DNAs; we therefore conclude that under our conditions the interwound superhelical structure prevails. The internal dynamics can be described by a superposition of rotational diffusion and internal relaxation. The latter process is characterized by the internal diffusion of persistence length size segments within the DNA chain and faster bending motions within these segments.

We have conducted photon correlation spectroscopy (PCS) studies on the plasmid pUC8 (2717 bp) in order to elucidate the internal dynamics of this superhelical DNA. We confirm that the first-order autocorrelation function of the scattered light from pUC8 solutions can be separated into two distinct exponential decay components, as first shown by Lewis et al. (R. Lewis, J.H. Huang and P. Pecora, Macromolecules 18 (1985) 944). A thorough analysis of the dependence on scattering vector K of the rates and amplitudes of the two components enables us to assign the slowly relaxing part to the center-of-mass diffusion of the DNA, while the faster component corresponds to rotational, bending and twisting motions of the superhelix. For larger K values the internal motions can be formally expressed in terms of an 'internal diffusion coefficient' Di, whose value of 2.0-2.5 X 10(-11) m2 s-1 is approximately equal to the translational diffusion coefficient predicted for a stiff DNA piece of the persistence length, 65 nm. Comparison of our measured Di values to those predicted from a recent theory of circular worm-like coils (K. Soda, Macromolecules 17 (1984) 2365) shows that the internal motions are faster than the theoretical values. One of the reasons for this discrepancy could be that the theory does not take into account torsional motions, which contribute significantly to the internal dynamics (J.C. Thomas, S.A. Allison, C.J. Appelof and J.M. Schurr, Biophys. Chem. 12 (1980) 177). At low K values, the fast relaxation of superhelical pUC8 is no longer proportional to K2, but reaches a constant value as K approaches zero. This behavior, not seen for the linearized DNA, can be interpreted in terms of rotational diffusion of a flexible rod-like molecule (T. Maeda and S. Fujime, Macromolecules 17 (1984) 2381) and supports an interwound rod-like structure for pUC8 DNA with an average end-to-end distance of 220 nm.