Binding of Hoechst 33258 and its derivatives to DNA.

In the present work, we employed UV-VIS spectroscopy, fluorescence methods, and circular dichroism spectroscopy (CD) to study the interaction of dye Hoechst 33258, Hoechst 33342, and their derivatives to poly[d(AT)].poly[d(AT)], poly(dA).poly(dT), and DNA dodecamer with the sequence 5'-CGTATATATACG-3'. We identified three types of complexes formed by Hoechst 33258, Hoechst 33342, and methylproamine with DNA, corresponding to the binding of each drug in monomer, dimer, and tetramer forms. In a dimer complex, two dye molecules are sandwiched in the same place of the minor DNA groove. Our data show that Hoechst 33258, Hoechst 33342, and methylproamine also form complexes of the third type that reflects binding of dye associates (probably tetramers) to DNA. Substitution of a hydrogen atom in the ortho position of the phenyl ring by a methyl group has a little effect on binding of monomers to DNA. However it reduces strength of binding of tetramers to DNA. In contrast, a Hoechst derivative containing the ortho-isopropyl group in the phenyl ring exhibits a low affinity to poly(dA).poly(dT) and poly[d(AT)].poly[d(AT)] and binds to DNA only in the monomer form. This can be attributed to a sterical hindrance caused by the ortho-isopropyl group for side-by-side accommodation of two dye molecules in the minor groove. Our experiments show that mode of binding of Hoechst 33258 derivatives and their affinity for DNA depend on substituents in the ortho position of the phenyl ring of the dye molecule. A statistical mechanical treatment of binding of Hoechst 33258 and its derivatives to a polynucleotide lattice is described and used for determination of binding parameters of Hoechst 33258 and its derivatives to poly[d(AT)].poly[d(AT)] and poly(dA).poly(dT).