Light-induced conformational changes and energy transfer in red fluorescent protein

Reversible conformational changes have been photo-induced in the red fluorescent protein DsRed at low temperature by wavelength-selective laser irradiation. We have found two new fluorescent forms: a shifted-red (SR-) and a new green (G′-) form that absorb and emit, respectively, ∼14 nm to the red and ∼80 nm to the blue of the ‘mature’ red (R-) form present in an un-illuminated sample of DsRed. Further, we have identified the 0–0 transitions of the various forms by spectral hole burning and estimated their ground-state energy differences and barrier heights by means of temperature-dependent excitation and fluorescence spectroscopy between 1.6 and 295 K. We have also proven that ‘downhill’ energy transfer takes place between these forms within the tetrameric structure of DsRed.