Theory of measurement of Förster-type energy transfer in macromolecules

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Abstract
The theoretical basis of an unconventional method for the determination of the amount of energy transferred between 2 fluorophores by the Forster mechanism is described. The method involves an internal comparison made by separation of the florophores in situ (i.e., in the optical cell), for example, by means of enzymic digestion; it eliminates several important sources of error and it simplifies calculation while making maximal use of the information contained in the fluorescence spectra. The validity of the method is demonstrated by determination of the known distance between 2 modifiable sites on the tRNA molecule, and its usefulness is exemplified by its applicaiton to triangulation of the ribosome of Escherichia coli.