Exciton dynamics in solid-state green fluorescent protein

Abstract

We study the decay characteristics of Frenkel excitons in solid-state enhanced green fluorescent protein (eGFP) dried from solution. We further monitor the changes of the radiative exciton decay over time by crossing the phase transition from the solved to the solid state. Complex interactions between protonated and deprotonated states in solid-state eGFP can be identified from temperature-dependent and time-resolved fluorescence experiments that further allow the determination of activation energies for each identified process.Keywords: fluorescent proteins; exciton dynamics; proton transferIn the last two decades, fluorescent proteins (FPs) have revolutionized many different areas of research. In particular biology, medicine and physiology are probably the uttermost affected. Their sublime photophysical and biochemical properties have created new eras of investigations such as live-cell 1 and super-resolution imaging 2 and have fundamentally changed research disciplines such as molecular biology and cell biology. FPs are most commonly used as biological markers or fluorescent probes in living organisms. FPs have therein enabled the investigation of nerve cell developments or cancer cell spreadings 3 . Those seminal advancements are first and foremost related to the ability to express FPs in any possible cell type without affecting intrinsic cell properties which is due to the fact that FP chromophores can be formed by autocatalytic cyclization without the need of a cofactor 4 . Combined with an excellent photophysical stability, FPs have become one of the most important tools in bioscience.Another interesting development has taken place in recent years in the research areas of semiconductor physics and organic photonics and might lead to a second revolution triggered by fluorescent proteins. The persistent seek for new emerging materials in optics and photonics has identified FPs in their solid state as promising bright and stable light emitters in photonic structures such as optical microcavities 6,7 . Especially their spectral characteristics (self-absorption, optical gain etc.) have triggered extensive research in using FPs as active material for photon 6,8 or polariton lasing experiments 9 . Those experiments revealed that microcavities filled with fluorescent protein show extremely low lasing thresholds and remarkable photostability compared to other fluorescent dyes 9 . Those advantages originate from the molecular structure of FPs, which is unique among organic dyes. The chromophore is basically surrounded by a protecting layer a) Electronic mail: christof.dietrich@physik.uni-wuerzburg.de of β-sheets, effectively forming a barrel (see Fig.1(a)). This barrel-like geometry ensures a strong reduction of concentration-induced luminescence quenching related to singlet-singlet annihilation and largely suppresses bimolecular quenching at high excitation densities 9 . It further maintains a fixed distance between neighboring chromophores in its solid state and thus supports efficient energy transfer between nex...

Time‐Resolved Studies of Energy Transfer in Thin Films of Green and Red Fluorescent Proteins

“…However, when the film dries, water evaporates, and thus the excited‐state proton transfers to the emissive B and I states is slowed down. This explanation is consistent with recent results on thin films with different water contents . (The difference in the absolute lifetime values between ref.…”

“…(The difference in the absolute lifetime values between ref. and our work is likely due to differences in sample fabrication. )…”

“…Recently, it was found that while the PLQY of fluorescent proteins is also very high in the solid state, it is lower than in solution, which indicates that these materials can potentially be further optimized once the exact photophysics are known. However, there is limited information on the temporal dynamics of thin films, with the exception of a very recent study on thin films of EGFP and an older report that used multifrequency phase fluorimetric data to determine the fluorescence lifetime of crystals of wild‐type GFP…”

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