Study of the Long-Time Fluorescence Tail of the Green Fluorescent Protein

Abstract

The time-resolved optical response of the wild-type green fluorescent protein (WT-GFP) in water and D2O was measured at room temperature by two optical techniques. The pump probe technique, with about 150 fs resolution, was used to measure the short-time response up to 150 ps. The short-time signals are similar to previously reported measurements. Time-correlated single photon counting (TCSPC) was used to measure the fluorescence of both the protonated and deprotonated emission bands of WT-GFP. The long-time fluorescence decay of the protonated form of WT-GFP decays nonexponentially. When this decay curve is multiplied by exp(t/τf) where τf is the lifetime of deprotonated form, the long-time tail decays as a power law of about t-3/2. Such a long-time fluorescence decay behavior represents the general emission decay pattern of excited photoacids in solutions and microemulsions and adds additional previously unrevealed information on the dynamics of this reaction. We attribute the long-time behavior to a diffusion-assisted geminate recombination process of the proton with the deprotonated species to reform the protonated chromophore.