Spectroscopic studies of the multiple binding modes of a trimethine-bridged cyanine dye with DNA

The interaction between DNA and a benzothiazole-quinoline cyanine dye with a trimethine bridge (TO-PRO-3) results in the formation of three noncovalent complexes. Unbound TO-PRO-3 has an absorption maximum (λ_max) of 632 nm, while the bound dyes (with calf thymus DNA) have electronic transitions with λ_max = 514nm (complex I), 584nm (complex II) and 642 nm (complex III). The blue shifts in the electronic transitions and the bisignate shape of the circular dichroism bands indicate that TO-PRO-3 aggregates with DNA. Complex I has a high dye:base pair stoichiometry, which does not depend on base sequence or base modifications. The bound dyes exhibit strong interdye coupling, based on studies with a short oligonucleotide and on enhanced resonance scattering. From thermal dissociation studies, the complex is weakly associated with DNA. Studies with poly(dGdC)₂ and poly(dIdC)₂ and competitive binding with distamycin demonstrate that complex II is bound in the minor groove. This complex stabilizes the helix against dissociation. For complex III, the slightly red-shifted electronic transition and the stoichiometry are most consistent with intercalation. Using poly(dAdT)₂, the complexes have the following dye mole fractions (X_dye): X_dye = 0.65 (complex I), 0.425 (complex II) and 0.34 (complex III).