The interaction of sequence-specific ligands with the nucleosome

The interaction of sequence specific ligands with DNA has been widely studied and the majority of this research has focused upon the binding of these drugs to free DNA. However, a therapeutic compound that targets DNA must interact with chromatin in vivo. Previous work with nucleosomes using reconstituted TyrT DNA, from E. coli, demonstrated that in the presence of sequence selective ligands the DNA appeared to rotate by 180° relative to the histone octamer. Since these studies utilised natural DNA, which contains many drug binding sites, only the gross effect of ligand binding could be observed. This work utilises DNA constructs containing drug-binding sites at defined rotational and translational positions, with respect to the histone octamer. Therefore it is possible to assess changes in nucleosome structure in the presence of a defined number of ligand molecules binding at a defined region of the DNA superhelix. The ligands used in this study are the minor groove binder Hoechst 33258 and the bis-intercalator echinomycin. It is observed that Hoechst molecules can bind to sites on the outer surface of the DNA superhelix without altering the structure of the core particle. Echinomycin does not appear to recognise targets in this rotational setting. The interaction of Hoechst and echinomycin with single target sites located on the inner surface of the DNA helix also has little effect up on the structure of the nucleosome. However, it has been observed that the binding of two or three Hoechst molecules to the inner surface appears to alter the rotational position of the DNA superhelix, with respect to the histone octamer, by 180°.