Étude des transferts d'énergie dans le complexe DNA-bromure d'éthidium au moyen du déclin de l'anisotropie de fluorescence

Abstract

In a recent investigation ( Ph. Wahl et al., Proc. Natl. Acad. Sci. U.S., 65 (1971) 417), the decay of fluorescence anisotropy of the ethidium bromide-DNA complex has been studied for high values of P D (ratio of molar concentrations of DNA and bound ethidium bromide). Evidence for a brownian motion of oscillation has been established, the angular amplitude of which is 35°. In the present work we study the decay of anisotropy for different values of P D . It is observed that the slope of this decay is increased when P D decreases. We attributed this phenomenon, to the existence of energy transfers between molecules of ethidium bromide bound to the same molecule of DNA. To describe quantitatively this behaviour, we try to use Jablonsky's and Förster's theory. The expression of anisotropy decay depends, in particular, on the angular distribution of the molecules. We have considered successively an isotropic distribution, and a distribution in which chromophores have a direction perpendicular to the DNA molecule axis. This last distribution corresponds to the generally admitted hypothesis of intercalation of ethidium bromide between the DNA base pairs. The best fit has been obtained using Förster's theory with the chromophores perpendicular to the axis of DNA. The fit is good in the range of small concentrations of ethidium bromide. For higher concentrations the fit is good only in the initial part of the decay curve. The deviation can be explained by the fact that only transfers between two molecules are considered in the theory. No good fit can be obtained with the Förster's formula corresponding to an isotropic distribution of the chromophores. Furthermore, the shapes of the decay curves predicted by Jablonsky's theory differ markedly from the experimental curves. Our measurements suggest strongly, for the first time, that the decay of fluorescence anisotropy of a solution of chromophores is better described by a Förster type theory than by the Jablonsky theory.