Cooperative binding interaction of ethidium with allosteric DNA

The specific association of drugs with deoxyoligonucleotides, containing a B-Z junction between left-handed Z-DNA and right-handed B-DNA, was examined by fluorescence and circular dichroism (CD) technique. Ethidium was chosen for a simple DNA binding compound because it binds to right-handed DNA and hybrid B-Z forms containing a B-Z junction in a highly cooperative manner. The binding isotherms were analyzed by an allosteric model in order to describe the cooperativity of association. Binding of ethidium to the DNA that are initially in the hybrid B-Z forms showed over an order of magnitude higher affinity than other DNA which were entirely in the B-form. The conformational transitions of deoxyoligonucleotides containing a B-Z junction as a result of ethidium binding were monitored by CD and the influence of NaCl on the complex formation was also determined by the CD spectra. The singular value decomposition (SVD) analysis was used to characterize a family of CD spectra of the species in binding equilibria. The results of SVD analysis showed a strikingly complex thermodynamic equilibria of cooperative binding of drugs to the allosterically converted DNA forms. The results also showed that these DNA forms in low- and high-salt were different in the absence or presence of drug. These results demonstrate that DNA-binding-drugs can preferentially interact with specific DNA structures and that these interactions are accompanied by allosteric changes of DNA conformations.