Ultravioletfluorescence of coelenteramide and coelenteramide-containingfluorescent proteins. Experimental and theoretical study

Abstract

Coelenteramide-containingfluorescent proteins are products of bioluminescent reactions of marine coelenter-ates. They re called‘discharged photoproteins’. Their light-inducedfluorescence spectra are variable, depending considerably on external conditions. Current work studies a dependence of light-inducedfluorescence spectra of discharged photoproteins obelin, aequorin, and clytin on excitation energy. It was demonstrated that photoexci-tation to the upper electron-excited states (260–300 nm) of the discharged photoproteins initiates afluorescence peak in the near UV region, in addition to the blue-green emission. To characterize the UVfluorescence, the light-inducedfluorescence 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 (330 nm) in the near UV region was observed in CLMfluorescence at higher excitation energy (260–300 nm). 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 UVfluorescence peak. Quantum yields of CLMfluorescence in methanol were 0.028 ± 0.005 at 270–340 nm pho-toexcitation. A conclusion was made that the UV emission of CLM might contribute to the UVfluorescence of the discharged photoproteins. The study develops knowledge on internal energy transfer in biological structures – complexes of proteins with low-weight aromatic molecules.