Method of the anomalous time dependence of viscosity for registration of changes in conformational state of Escherichia coli cell genome

The sensitivity of the anomalous time dependence of viscosity to the concentration of the DNA-protein complexes (DNA + histone-like proteins of bacteria or, in other words, the genome) such as chromatin and the conformations of these complexes in lysates of E. coli AB1157 cells were studied. A linear region of the anomalous viscosity time dependence on the concentration of E. coli cells was found in which the interactions between single DNA-protein complexes can be neglected. The response of the genome of E. coli to ethidium bromide at concentrations of 0.0003-3 mg/ml was studied. Significant differences in the effect of ethidium bromide on E. coli cells in the stationary and logarithmic growth phases were found. The effect of heating cell lysates, the molar concentration of NaCl in lysates, and the addition of proteins into lysates on the parameters of the anomalous viscosity time dependence was studied. It was shown that proteins do not contribute significantly to the effect of anomalous viscosity time dependence. The results obtained confirm that the method is sensitive to changes in the conformational state of the genome of E. coli cells.     

Position of caesium ions in crystalline B-form DNA determined by synchrotron radiation diffraction. Preliminary results

The diffraction patterns of CsDNA taken with copper X-irradiation are considerably impaired because of the strong X-ray absorption by caesium ions. The use of high power synchrotron radiation of wavelength λ = 1.2 Å has yielded photographs suitable for intensity measurements.The structure of phage T2 CsDNA at 76% relative humidity is isomorphous to the crystalline B-form of LiDNA, and the disposition of cations appears to conform to the 10-fold screw symmetry of B-DNA. The structure factor amplitudes of 20 reflections in the diffraction pattern of phage T2 CsDNA are noticeably different from those of the same reflections in the LiDNA diffraction pattern, and the positions of the Cs⁺ ions could thus be found. The best model has the cations located significantly close to dyad axes lying between the planes of successive nucleotide pairs. One of the two cations “belonging” to a nucleotide pair touches the surface of the narrow groove of the B-DNA double helix, while the other is on the wide groove side, rather far from its “own” DNA molecule.     

Structure of the B DNA cationic shell as revealed by an X-ray diffraction study of CsDNA. Sequence-specific cationic stabilization of B form DNA

Synchrotron radiation diffraction data for phage T2 CsDNA fibres have been used to determine the co-ordinates of the caesium ions in crystalline B form DNA. The R value is 0.16 for an optimized structure. The caesium ions are distributed equally between the narrow and wide grooves of B DNA and are located close to the dyad axes lying between the planes of adjacent base-pairs. On the wide-groove side the cations are separated from the nearest phosphate atoms by a hydration layer one to two water molecules thick. In the narrow groove the cations are directly co-ordinated to the base atoms and, for six out of ten possible DNA stacking types, form chelation complexes: O-2(Pyr)-Cs+-O-2(Pyr), O-2(Pyr)-Cs+-N-3(Pur) or N-3(Pur)-Cs+-N-3(Pur), which stabilize the B conformation. The steric properties of such complexes as estimated for different base sequences and for different ions are consistent with the structural behaviour of double-helical polynucleotides with different base sequences, as experimentally observed.