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 (lambda(max)) of 632 nm, while the bound dyes (with calf thymus DNA) have electronic transitions with lambda(max) = 514 nm (complex I), 584 nm (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).
The structures of the noncovalent complexes of DNA with a thiazole orange dimer (TOTO) were investigated
in solution and when adsorbed to a mica substrate, using viscometry and atomic force microscopy (AFM),
respectively. From the viscosity measurements at low TOTO:base pair concentrations, the dye lengthens the
DNA helix by two times the base pair spacing, consistent with bisintercalation. Higher concentrations of
TOTO cause a significant decrease in the viscosity of the DNA solutions, as also observed for related
bifunctional ligands. The conformational changes induced by the dimer are compared to those of the monomer.
The contour lengths of the DNA-TOTO complexes adsorbed onto mica were directly measured by AFM.
These lengths and the size of the binding site are inconsistent with the bisintercalation binding mode observed
in solution, suggesting an alternative mode of binding on mica.
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