The binding of 2,5-bis(4-amidinophenyl)furan (APF) to calf thymus DNA, [poly(dA-dT)]2, and [poly(dG-dC)]2 has been studied with flow linear dichroism and circular dichroism spectroscopy. The electronic excited states of the APF chromophore were first characterized using experimental and quantum mechanical methods: it is shown that the low-energy absorption band (320-400 nm) originates from only a single electronic transition which is polarized along the long axis of the molecule, information that is crucial for the structural interpretation of the linear and circular dichroism spectra of the APF-DNA complexes. By contrast, in the unsymmetric analogue 4',6-diamidino-2-phenylindole (DAPI) two overlapping transitions, with somewhat divergent polarizations, both contribute to the first absorption band. Upon binding to DNA the spectroscopic behavior of APF strongly resembles that of DAPI. The linear dichroism data show that the drug binds to calf thymus DNA and [poly(dA-dT)]2 with an angle of 46 degrees +/- 2 degrees between its symmetry long axis and the DNA helix axis, confirming that APF, just like DAPI, is an AT-specific minor-groove binder. Upon binding to [poly(dG-dC)]2, however, the orientation of the long axis is parallel with the plane of the DNA bases, a geometry which excludes binding parallel to the grooves but could be consistent with intercalation. However, a short axis polarized transition is strongly inclined to the base plane and, furthermore, the persistence length of the polynucleotide is markedly reduced, observations that contradict classical intercalation.(ABSTRACT TRUNCATED AT 250 WORDS)