CALORIMETRIC INVESTIGATION OF ETHIDIUM AND NETROPSIN BINDING TO CHICKEN ERYTHROCYTE CHROMATIN

We have investigated the thermodynamic aspects of the ligand binding t o chromatin, using isothermal titration calorimetry. Two classical DNA ligands were used: an intercalator, ethidium bromide, and a minor gro ove binder, netropsin. Stoichiometry, affinity constant, and thermodyn amic parameters were determined at various salt concentrations and dif ferent temperatures. The effect of ionic strength was analyzed accordi ng to the Record theory applied to chromatin. We also compared the bin ding parameters on naked DNA, H1/H5-depleted chromatin, and chromatin. We demonstrated that the presence of histones on DNA still allows the ligand binding that takes place according to a simple one single-site model. For both ligand types, the thermodynamic driving force is enth alpic and the association is characterized by a somewhat weaker affini ty and more scattered ligand distribution than on naked DNA. The ligan d affinity is weakly altered by the salt-induced compaction of the chr omatin and the binding is accompanied by a release of one counterion p er ligand molecule. The temperature-dependent studies revealed the exi stence of a small heat capacity change associated with ligand binding to chromatin, together with an enthalpy-entropy compensation that main tains the free energy constant over the investigated temperature range .