Metachromatic reaction of proflavine bound to DNA and RNA. Spectrophotometric titration and quantitative optical parameters

Abstract

Experimental conditions were determined under which spectral changes in proflavine due to binding to nucleic acids can be quantitatively analyzed by starting with the maximally-saturated complex (1 dye per phosphate group) and keeping the amount of bound dye constant (negligible free dye) as the number of sites is increased. Calculation of various optical parameters shows that both a blue shift in frequency and decrease in oscillator strength occur in the spectra of 1- to-1 complexes of proflavine with native and denatured DNA and RNA, and that the extent of hypochromism is greater for denatured DNA and RNA than for native DNA. At ratios of sites-to-dye (P/D) greater than one, the red-shifted spectrum of the “strong-binding” complexes appears. The redistribution of dyes on excess sites is similar for denatured and native DNA up to P/D= 5. However, quantitative and qualitative differences in the spectra of these complexes occur at higher P/D. The data emphasize the multiplicity in geometry of binding, discussed by others in regard to native DNA, and elucidate the origin of several features of the induced optical activity of proflavine in these complexes.