2Institute of Agricultural Biotechnology, 127550 Moscow, Russia
* To whom correspondence should be addressed.
Received January 12, 2018; Revision received June 4, 2018
We studied the thermodynamics of melting of isolated rat liver nuclei with different degrees of chromatin condensation determined by the concentration of polyamines (PA) and the solution ionic strength, as well as the effect of the antibiotic distamycin A (DM) on melting. Differential scanning calorimetry (DSC) profiles of nuclear preparations contained three peaks that reflected melting of three main chromatin domains. The number of peaks did not depend on the degree of condensation; however, nuclei with more condensed chromatin had a higher total enthalpy. DM stabilized peaks II and III corresponding to the melting of relaxed and topologically strained DNA, respectively, but destabilized peak I corresponding to the melting of nucleosome core histones. At the saturating concentration (DM/DNA molar ratio = 0.1), DM increased Tm of peaks II and III by ~5°C and decreased Tm of peak I by ~2.5°C. Based on the dependence of ΔH on DM concentration, we established that at low DM/DNA ratio (≤0.03), when DM interacted predominantly with AT-rich DNA regions, the enthalpy of peak II decreased in parallel with the increase in the enthalpy of peak III, which indicated that DM induces structural transitions in the nuclear chromatin associated with the increase in torsional stress in DNA. An increase in free energy under saturation conditions was equal to the change in the free energy of DM interaction with DNA. However, the increase in the enthalpy of melting of the nuclei in the presence of DM was much greater than the enthalpy of titration of nuclei with DM. This indicates a significant increase in the strength of interaction between the two DNA strands apparently due, among other things, to changes in the torsional stress of DNA in the nuclei. Titration of the nuclei with increasing PA concentrations resulted in the decrease in the number of DM-binding sites and the non-monotonous dependence of the enthalpy and entropy contribution to the binding free energy on the PA content. We suggested that the observed differences in the thermodynamic parameters were due to the different width of the minor groove in the nuclear chromatin DNA, which depends on PA concentration.
KEY WORDS: nucleus, chromatin, distamycin A, differential scanning calorimetry, isothermal calorimetric titration, polyamines, DNA torsional stress