Ultraviolet fluorescence of coelenteramide and coelenteramide-containing fluorescent proteins. Experimental and theoretical study

Abstract

Coelenteramide-containing fluorescent proteins are products of bioluminescent reactions of marine coelenterates. They are called 'discharged photoproteins'. Their light-induced fluorescence spectra are variable, depending considerably on external conditions. Current work studies a dependence of light-induced fluorescence spectra of discharged photoproteins obelin, aequorin, and clytin on excitation energy. It was demonstrated that photoexcitation to the upper electron-excited states (260-300nm) of the discharged photoproteins initiates a fluorescence peak in the near UV region, in addition to the blue-green emission. To characterize the UV fluorescence, the light-induced fluorescence spectra of coelenteramide (CLM), fluorophore of the discharged photoproteins, were studied in methanol solution. Similar to photoproteins, the CLM spectra depended on photoexcitation energy; the additional peak (330nm) in the near UV region was observed in CLM fluorescence at higher excitation energy (260-300nm). Quantum chemical calculations by time depending method with B3LYP/cc-pVDZ showed that the conjugated pyrazine-phenolic fragment and benzene moiety of CLM molecule are responsible for the additional UV fluorescence peak. Quantum yields of CLM fluorescence in methanol were 0.028±0.005 at 270-340nm photoexcitation. A conclusion was made that the UV emission of CLM might contribute to the UV fluorescence of the discharged photoproteins. The study develops knowledge on internal energy transfer in biological structures - complexes of proteins with low-weight aromatic molecules.

Chemi- and Bioluminescence of Cyclic Peroxides

“…290,291 Even its UV fluorescence has been considered recently. 292 The different bioluminescence and fluorescence of obelin have prompted both experimental and theoretical investigations aimed at identifying which is/are its light emitter(s). Thanks to the seminal study by Shimomura and Teranishi who investigated fluorescence of coelenteramide and some analogues in various solvents, 293 several possible light emitters were proposed (Figure 36).…”

“…Other light emitters may come into the game and contribute significantly, albeit to a lower extent, to coelenteramide bioluminescence or fluorescence emission spectra. 298 It is worth mentioning the possible implication of UV components in the fluorescence spectrum of obelin, 292 which more generally opens the question of the role of the higher excited states in both bioluminescence and fluorescence mechanisms. 5.2.3.…”

Enzymatic Responses to Low-Intensity Radiation of Tritium

“…CLM-FPs are products of bioluminescent reactions of coelenterates that constitute a potential use in biomedicine as biomarkers [36][37][38]. The fluorescence characteristics of CLM-FPs were intensively studied [39][40][41][42][43][44][45][46]; they were found to be sensitive to external exposures: excitation energy [44], chemical agents [47,48], and radioactivity [49,50]. Recently, [51] suggested CLM-FPs for intracellular application as being the simplest toxicity bioassay.…”

Monitoring of Low-Intensity Exposures via Luminescent Bioassays of Different Complexity: Cells, Enzyme Reactions, and Fluorescent Proteins

“…These reactions are Ca 2+ dependent, and this is a basis for their biomedical application [42,43]. The biochemical and photophysical mechanisms of the bioluminescence reactions [44,45,46,47,48,49] and spectral characteristics of their products, CLM-containing proteins [50,51,52,53,54,55,56,57], are now under intensive investigation.…”