The Green Fluorescent Protein GFP or ESPT in a Box

At room temperature, wtGFP (wild-type GFP) exhibits two main absorption peaks with maxima at 398 nm (band A) and 478 nm (band B). Several groups have studied the excited state dynamics of wtGFP [58-63]. These led to the conclusion that the two visible absorption bands of the wtGFP correspond to protonated and deprotonated ground-state conformations, while upon photoexcitation the acid form A rapidly deprotonates to form the B form. In addition, ultrafast kinetics suggests the presence of a third intermediate I which involves a proton transfer without conformational relaxation to the B form [64]. 

Since the excited-state pKf of the GFP chromophore is less than zero already in the protein, then this chromophore, by analogy with the hydroxycamptothecin outlined above, also falls into the category of biological super photoacids. Isolated synthetic GFP and its derivatives do not fluoresce outside the y9-barrel at room temperature due to very effective internal conversion related to isomerization along the double bond [59-63]. Deep understanding of the photophysics of these dyes as well as synthesis of their fluorescence derivatives capable of ESPT are our immediate goals. We anticipate further studies in biological proton transfer will be facilitated by these powerful new photophysical tools. 

Support of this research by the U.S. National Science Foundation, under Grant GHE-0456892, is gratefully acknowledged. 

28 Boule, P. Richard, G. David-Oudjehani, K. Grabner, G. Proc. Indian Acad. Sci., Chem. Sci. 1997, 109, 509-519. 

T in Structure, Energetics and Reactivity on Aqueous Solutions, Gramer, G. J. Truhlar, D. G., Eds. AGS Books, Washington DG, 1994. 

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