title: Low-temperature switching by photoinduced protonation in photochromic fluorescent proteins.
abstract: We have studied the photoswitching behaviour of a number of photochromic fluorescent proteins at cryo-temperature. Spectroscopic investigations at the ensemble level showed that EYFP, Dronpa and IrisFP all exhibit reversible photoswitching at 100 K, albeit with a low quantum yield. The photophysics of the process were studied in more details in the case of EYFP. The data suggest that photoinduced protonation of the chromophore is responsible for off-switching at cryo-temperature, and thus is possible in the absence of significant conformational freedom. This finding is consistent with the hypothesis that chromophore protonation may precede large amplitude conformational changes such as cis-trans isomerisation during off-photoswitching at room temperature. However, our data suggest that low-barrier photoinduced protonation pathways may in fact compete with room-temperature off-switching reactions in photochromic fluorescent proteins. The occurrence of reversible photoswitching at low-temperature is of interest to envisage cryo-nanoscopy experiments using genetically encoded fluorophores.
authors.affiliation: IBS, Institut de Biologie Structurale Jean-Pierre Ebel, CEA, CNRS, Université Joseph Fourier, 41 rue Jules Horowitz, F-38027, Grenoble, France.
authors.forename: Aline Regis
authors.initials: AR
authors.lastname: Faro
authors.forename: Virgile
authors.initials: V
authors.lastname: Adam
authors.forename: Philippe
authors.initials: P
authors.lastname: Carpentier
authors.forename: Claudine
authors.initials: C
authors.lastname: Darnault
authors.forename: Dominique
authors.initials: D
authors.lastname: Bourgeois
authors.forename: Eve
authors.initials: E
authors.lastname: de Rosny
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry; D003080:Cold Temperature; D007536:Isomerism; D008164:Luminescent Proteins / Q000737:chemistry*; D011522:Protons; D011789:Quantum Theory; D013056:Spectrophotometry, Ultraviolet
chemical_list: D001426:Bacterial Proteins; D008164:Luminescent Proteins; D011522:Protons; C054562:yellow fluorescent protein, Bacteria