RESOLUTION OF A DISTRIBUTION OF DISTANCES BY FLUORESCENCE ENERGY TRANSFER AND FREQUENCY-DOMAIN FLUOROMETRY.

We used frequency-domain fluorescence spectroscopy to examine the intensity decays of a fluorescent donor attached to an acceptor via a flexible alkyl chain. The intensity decay of the indole donor becomes markedly heterogeneous due to energy transfer to the dansyl acceptor. The measured dispersion of fluorescence decay times was used to recover the donor-to-acceptor distance distribution. The distance distribution was found to be characteristic of the molecule, and not the model used for data analysis. The ability to recover distance distributions in solution should be valuable in studies of biological macromolecules.