GROOVE BINDING OF A STYRYLCYANINE DYE TO THE DNA DOUBLE HELIX - THE SALT EFFECT

The cationic styryl dye DSMI binds to the double helical DNA with a high affinity. A red shift of the absorption spectrum and hypochromism accompany the binding of DSMI to calf thymus DNA. The fluorescence yields increase dramatically when DSMI binds to DNA, with no shifts in the emission maximum. These spectral changes are in contrast to the behavior observed with many fluorescent intercalators. Groove binding rather than intercalation of the dye was suggested to be the cause of these spectral changes, based on a variety of experimental results. Consistent with groove binding, the fluorescence enhancements were greater with poly(dA-dT) when compared with poly(dG-dC). The addition of salt or a strong electrolyte to the solution releases the DNA-bound dye cations from the groove and causes a decrease in the fluorescence yield. Therefore, when the dye binds to DNA, the fluorescence quenching with anionic quenchers was enhanced dramatically, in contrast to the expected protection of the probe fluorescence by the DNA helix. Binding of DSMI to DNA inhibits the non-radiative deactivation of the excited state, perhaps owing to the restriction imposed by the narrow minor groove of the DNA. The steric constraints imposed on the dye owing to the binding in the groove result in enhanced fluorescence yields and longer fluorescence lifetimes. Since the lifetimes of the DNA-bound dye were still in the nanosecond range, it is very likely that groove binding of the dye permits twisting in the excited state, which may be important in the excited state dynamics of this class of dyes. The large fluorescence enhancements observed when DSMI binds to the helix and the poor fluorescence yield of the dye in the absence of DNA suggests that DSMI should be quite useful in the staining of DNA gels and for the detection of very low concentrations of DNA.