Modulation of the spectral properties of the acceptor for quantitative determinations of FRET.

ELIZABETH JARES-ERIJMAN; LUCIANA GIORDANO; CARLA SPAGNUOLO; MASAHIRO IRIE; THOMAS M. JOVIN

Buenos Aires

Congreso; XIV International Biophysics Congress – IUPAB/SAB.; 2002

IUPAB-SAB

A new methodology based on the modulation of the fluorescent emission of the donor by a photochromic acceptor has been recently developed for the quantitative determination of fluorescence resonance energy transfer (FRET. In pcFRET, the fluorescent emission of the donor is modulated by reversible transformations of a photochromic acceptor. We designed several model systems consisting of a fluorescent donor and acceptor bound through a tether of given dimension. Light induced a reversible change in the structure and concomitantly in the absorption proprieties of the acceptor. Only the closed form of the photochromic compound has an absorption band overlapping the emission band of the donor. This provides the means for switching the process of FRET on and off, allowing direct and repeated evaluation of the relative changes in the donor fluorescence quantum yield due to FRET. Diarylethenes show excellent stability in aqueous media, no thermal back reactions and negligible fatigue process upon related cycles of UV visible irradiation.