Cis − Trans Photoisomerization of Fluorescent-Protein Chromophores

Scuola Normale Superiore, Italian Institute of Technology, Pisa, Italy.
The Journal of Physical Chemistry B (Impact Factor: 3.3). 09/2008; 112(34):10714-22. DOI: 10.1021/jp802419h
Source: PubMed


Photochromic variants of fluorescent proteins are opening the way to a number of opportunities for high-sensitivity regioselective studies in the cellular environment and may even lead to applications in information and communication technology. Yet, the detailed photophysical processes at the basis of photoswitching have not been fully clarified. In this paper, we used synthetic FP chromophores to clarify the photophysical processes associated with the photochromic behavior. In particular, we investigated the spectral modification of synthetic chromophore analogues of wild-type green fluorescent protein (GFP), Y66F GFP (BFPF), and Y66W GFP (CFP) upon irradiation in solutions of different polarities. We found that the cis-trans photoisomerization mechanism can be induced in all the chromophores. The structural assignments were carried out both by NMR measurements and DFT calculations. Remarkably, we determined for the first time the spectra of neutral trans isomers in different solvents. Finally, we calculated the photoconversion quantum yields by absorption measurements under continuous illumination at different times and by a nanosecond laser-flash photolysis method. Our results indicate that cis-trans photoisomerization is a general mechanism of FP chromophores whose efficiency is modulated by the detailed mutant-specific protein environment.

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Available from: Valerio Voliani, Jul 02, 2014
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    • "Our theoretical method was also put to the test by calculating the 3 J C,H for the cis and trans isomers of p-HBDI and the analog of B1 where the carbon atom in position 4 is replaced by a nitrogen atom. The experimental values of 3 J C,H for the latter compounds in both cis and trans form are available from previous reports (Prokofev and Karpeiskaya 1979; Voliani et al. 2008). Excellent agreement between experimental and theoretical values emerged for all compounds for both configurations (Table 3). "
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