End-to-end distance distributions of flexible molecules from steady state fluorescence energy transfer and quenching-induced changes in the Förster distance

We describe a new method to recover the distribution of distances between two sites on flexible molecules, which requires only steady state measurements of fluorescence intensities. We used collisional quenching of the donor fluorescence to vary the characteristic Förster distance (R₀) for energy transfer. The measured transfer efficiencies for each value of R₀ provide different samples of the distance distribution. The R₀-dependent transfer efficiencies, when analyzed by non-linear least squares, were found to reliably determine the distance distribution, as shown by evaluation of the uncertainties in the distribution and by agreement with the frequency-domain method (Chem. Phys. Letters 138 (1987) 587).