Fluorescence anisotropy decay due to rotational brownian motion of ethidium intercalated in double strand DNA

Abstract

The transient fluorescence of solutions of ethidium bromide · DNA complexes has been measured by pulse fluorimetry at different temperatures and in solvents containing various amounts of sucrose. The molar ratio of ethidium to nucleotides was low. Under these conditions the anisotropy decay was due to the Brownian motion of ethidium molecules intercalated in the double strand DNA molecules. This anisotropy decay could be described by a sum of 3 exponential terms, with correlation times θ 1, θ 2, θ 3 which were linear functions of the ratio of the solvent viscosity to the absolute temperature ( η T ). The amplitude of the exponential term characterized by the shortest correlation time ( θ 1) has been found to depend on temperature while the ratio of the amplitude of the two other terms (characterized by θ 2 and θ 3) was independent of temperature. These results were interpreted as follows: θ 1 corresponds to a fast motion of the dye in its site. θ 2 and θ 3 describe a tortional motion of the ethidium bromide · DNA complex, involving several nucleotide pairs.