CD Spectroscopy

Although circular dichroism (CD) is not observed for achiral molecules, when they form complexes with DNA they are placed within a chiral environment and give

400 450 500 550 600 S50

Fig. 2.5.3. Spectrofluorimetric titration of acridizinium salt 5a with ct DNA (A), (poly[dA-dT]-poly[dA-dT]) (B), and (poly[dG-dC]-poly[dG-dC]) (C) in phosphate buffer (10 mM, pH 7.0); arrows indicate the decreasing emission intensity during the course of the titration.

400 450 500 550 600 S50

Wavelength / nm

Fig. 2.5.3. Spectrofluorimetric titration of acridizinium salt 5a with ct DNA (A), (poly[dA-dT]-poly[dA-dT]) (B), and (poly[dG-dC]-poly[dG-dC]) (C) in phosphate buffer (10 mM, pH 7.0); arrows indicate the decreasing emission intensity during the course of the titration.

an induced CD (ICD) signal [33], which results from non-degenerative coupling between the transition of the bound ligand and that of the nucleic-base transitions. The appearance of an ICD signal confirms the dye-DNA interaction and might provide further information about the position of a dye in its complex with DNA, because the intensity and the phase of the ICD signal depends on the position and the orientation of the chromophore relative to the DNA bases. An intercalator usually has a weak and negative ICD signal when its transition moment is polarized along the long axis of the binding pocket (i.e. parallel to the bisector of the base pairs). In contrast, relatively strong positive ICD bands appear when the transition moment is polarized perpendicular to the long axis of the binding pocket. Groove binders give even more intense ICD signals, usually with a positive band.

Thus aqueous solutions of the achiral acridizinium salts 5a and 5b alone have no CD activity, but an ICD is observed on addition of DNA to these salts; the spectra differ in phase and intensity, however (Figure 2.5.4). The shapes of the CD signals roughly resemble the broadened long-wavelength absorption spectrum of the corresponding acridizinium salts. The positive ICD of 5b (Figure 2.5.4B, spectrum 2)

Fig. 2.5.4. CD spectra of salts 5a (A) and 5b (B) with ct DNA; 1. without DNA; 2. [dye]/[DNA] = 0.1; 3. [dye]/[DNA] = 0.05.

results from orientation of the short molecular axis of the dye parallel to the binding pocket [34]. In contrast, the CD spectrum of the amino-substituted acridizi-nium salt 5a (Figure 2.5.4A, spectrum 2) in the presence of DNA gives negative signals for the S0-Sj transition at dye-to-DNA ratios smaller than 0.05; this is indicative of intercalative binding with the acridisinium long axis (which roughly resembles the polarization of the long-wavelength transition) oriented parallel to the long axis of the intercalation pocket. Nevertheless, at a higher dye-to-DNA ratio (0.1) a bi-signate signal pattern appears for the long-wavelength absorption of amine 5a (Figure 2.5.4A, spectrum 3); this might result from exciton coupling [35]. This exciton band presumably arises because of aggregation of the dye and association of these aggregates with the DNA backbone. Such exciton CD signals are usually significantly stronger than those of intercalated molecules and can overlap with the latter.

0 0

Post a comment