[The nature of radiation damage of the DNA molecule during gamma-irradiation of its solutions].
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Biomedical subjects
Publications and source records attributed to E V Frisman.
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It was shown that gamma-irradiation of water-ethanol and water-n-propanol solutions of DNA with doses of 10-30 Gy leads to a fall of the specific volume of the macromolecule upon the cooperative transition at a critical concentration of alcohol in solution at which the destroying of water structure by a nonelectrolyte occurs.
DNA--acriflavin complexes have been investigated by the methods of flow birefringence and viscometry. The intrinsic viscosity and the optic anysotropy of the complex increase with the increasing quantities of binding dye. Experimental data are treated on the basis of different models of binding. At high ionic strength (mu = 0,1) one type of binding takes place which is described by the intercalation model. In this case the thermodynamic rigidity of DNA-molecule within the complex is proportional to "r". In solutions of low ionic strength (mu = 0,001), two types of DNA-acriflavin binding occur: intercalation and external binding. At low ionic strength, the spectrophotometric titration technique is shown to give a reduced value of "r".
The hydrodynamic and optical properties (intrinsic viscosity and optical anisotropy of DNA) have been studied at the high ionic strength mu greater than or equal to 1 M. It has been shown that the effective volume of DNA molecule doesn't depend of mu when mu greater than or equal to 1 M. In these conditions the electrostatical interactions in DNA disappear. But thermodynamic excluded volume effects do not depend on mu and play also an important role in this range of mu (mu greater than or equal to 1 M). It has been concluded that the condensation of DNA in solutions of high salt concentration is the result of local denaturation of DNA. It has been shown that the optical anisotropy of DNA increases drastically at mu congruent to 2 M but the persistence length of DNA does not change under these conditions.
The interaction of DNA with divalent metal ions: Ba2+, Mg2+, Mn2+, Ni2+, Cu2+ in solutions at different ionic strengths mu was investigated. The combination of following methods: flow birefringence, viscometry, UV-spectroscopy and circular dichroism made possible to follow the state of the secondary and tertiary structure of the DNA molecule during its interaction with ions. The presence of divalent ions in solution affects the hydrodynamic properties of DNA only at low mu. At high mu the difference in the action of mono- and divalent ions disappears. The persistence length of DNA does not change during the experiment. It is shown that the Mg2+ and Ba2+ ions interact only with phosphate groups of DNA but Mn2+, Ni2+, Cu2+ ions interact also with the nitrogen bases of the macromolecule.
The DNA conformation was studied at different relation between Na+ and Me2+ (Mn2+ or Mg2+) ions in solution at the fixed total ionic strength mu. At low mu the intrinsic viscosity of DNA [eta] decreased to the limited fixed value with the increasing of Mn2+ or Mg2+ concentration (CMe2+). At higher mu greater than or equal to 0.1 M [eta] doesn't depend on CMe2+. The presence of Mn2+ in solution caused a decrease of the optical anisotropy of DNA and the value of epsilon 260 (p) independent on ionic strengths. In contrary, these parameters of DNA didn't change in solution with Mg2+-concentration. The observed differences in the effects of Mn2+ and Mg2+ on the optical properties of the macromolecule suggest that there are different modes of binding of these ions to DNA. It has been concluded, that Mn2+ interacts with bases and phosphate groups of DNA, but Mg2+--only with phosphates. The persistence length of DNA doesn't depend on Me2+ concentration under the conditions of the experiment (mu greater than or equal to 0.005 M).
The comparative studies of the formation of DNA-complexes with the acridines containing one and two chromophores were accomplished. It was shown that both of acridines were bonded with DNA by means of intercalation irrespective of the ionic strength of medium (mu). When mu = 0.1 the diacridine (1,6-bis(9-acridylamino)-hexan) behaves as an mono-intercalator. Under these conditions both of the ligands exert equal influence of the molecular parameters of DNA. When mu = 0.001 the binding mode of the diacridine with DNA depends on its concentration in a complex. If a number of diacridine molecules on a pair of nucleotides (r) falls in a region 0 less than r less than 0.2 its binding with DNA is accomplished via the bis-intercalation mode and accompanied by the structure distortion of the monomer remnant of the macromolecule. As r increases from 0.2 to 0.4 the gradual change of the binding mode of the diacridine with DNA from bis-intercalation to mono-intercalation takes place. Moreover the structure of nucleotides is reduced. When mu = 0.001 the behaviour of DNA complexes with mono-acridine is analogous to the observed one when mu = 0.1.
Effect of the temperature on the conformation of the native DNA molecule in solution of different electrolytes (LiCl, NaCl, KCl, CsCl, Gu-HCl) at ionic strengths mu = 5; 0.1; 0.01; 0.005 and temperatures ranging from 10 to 40 degrees C were studied by the methods of flow birefringence and viscometry. The experiments showed that the value of intrinsic viscosity [eta] of DNA increases at increase of temperatures in solutions of all the chlorides studied, excluding guanidine. The effect of temperature on the value of [eta] doesn't depend on the type of the cation at a fixed value of mu and is elevated when mu decreases. The observed alterations of the value of [eta] for DNA in water-salt solutions at different temperatures can be explained by an increase in the hydration of the alkaline ions at temperature increase. The experiments showed the specificity of the effect of different ions on the dimensions of the DNA molecule in solution. The data on optical anisotropy of the DNA molecule testify, that the thermodynamic rigidity of the latter doesn't depend on the temperature of solutions of different electrolytes in the temperature range studied.
The influence of acriflavine in the process of exposing DNA solution to gamma-radiation is studied. Acriflavine being actively bound to DNA is demonstrated not to protect DNA molecule from radiation damage. No radiation-induced variation in acriflavine-DNA binding degree is discovered. Acriflavine protective properties are revealed when the concentration of acriflavine is essentially high. This confirms our early results according to which only free ligands in solution protect DNA from radiation damage.
The hydrodynamical and optical properties of DNA were investigated in the wide-range of pH by the methods of streaming birefringence, viscometry and spectrophotometry for the different ionic strengths of environment. The measurements of the intrinsic viscosity as a function of pH allow us to determine the compactization of protonated DNA without the destruction of double-helical conformation. This transition is accompanied by a decrease in the optical anisotropy of DNA and the coefficient of molar extinction E260 (P). The increase of volume and persistence length of DNA was observed in the alkaline range of pH. Analyses of experimental data lead to an assumption that the predominant cause of these effects is the change of flexibility of DNA as a result of ionization of its bases. The data obtained were compared with those for polycationic molecules.
The influence of different alcohol-water solvents on the conformation of the native DNA molecule has been studied by the methods of flow birefringence and viscometry. The experiments have shown, that a certain content of the organic component induces a cooperative transition in the tertiary structure of the DNA molecule. The conformational transitions in the tertiary structure of DNA have been observed at low alcohol concentrations, corresponding to the desruption of the ordered spatial water structure and are caused by the alteration of the intermolecular interactions between the DNA and solvent molecules. The alcohol concentrations, corresponding to the observed transition, as well, as relative decrease of the intrinsic viscosity of DNA, depend on the chain length and branching of the hydrocarbon portion of the alcohols. The data on the optical anisotropy of the DNA molecule testify that the secondary structure of the latter doesn't change in a wide range of the alcohol concentrations.
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