Chemiluminescence Associated with Singlet Oxygen Reactions with Amino Acids, Peptides and Proteins†

Abstract

Low level chemiluminescence (CL) is observed after protein oxidation mediated by singlet oxygen produced in Rose Bengal (RB) irradiation. This CL lasts for several minutes after the end of the photolysis. In this work, the mechanism of the process was assessed from the spectral characteristics of the CL and the effect of antioxidants (Trolox or ascorbate), Ebselen (a compound with peroxidase-like activity), azide (a singlet oxygen scavenger) and D2O, added prior to or after RB irradiation. It is concluded that most of the light emission is due to formation of excited states generated in the decomposition of peroxides and/or hydroperoxides accumulated during the photolysis. Experiments carried out in the presence of several amino acids (Cys, Met, His, Tyr and Trp) and di- and tripeptides suggest that peroxides (and/or hydroperoxides) of Trp residues are mainly responsible for the CL observed after singlet oxygen-mediated protein oxidation. The much weaker CL observed after the oxidation of proteins without Trp residues supports this conclusion. A comparison of the results obtained employing free Trp, Ala-Trp and Trp-Ala dipeptides, Ala-Trp-Ala tripeptide and Trp-containing proteins supports the conclusion that blocking the amino group of the Trp moiety strongly increases the efficiency of the chemiluminescent process, producing approximately 2.5x10(-8) photons per oxidized Trp group in Ala-Trp. A mechanism comprising two chemiluminescent oxidation pathways of Trp residues is proposed to explain the results.

Using spontaneous photon emission to image lipid oxidation patterns in plant tissues

“…However, compared to lipids, emission in the blue/green spectral region appeared to be enhanced relative to the emission at wavelengths >640 nm, reaching approximately 50% of total emission (Figure S1 in Supporting Information), in agreement with previous data showing that protein chemiluminescence occurs predominantly between 490 and 580 nm (Pollet et al. , 1998; Alarcon et al. , 2007).…”

“…The half time of the protein chemiluminescence decrease was about 20 min. Chemiluminescence associated with reaction between proteins, including BSA and lysozyme, and 1 O 2 has been previously reported to decay with a half‐time of several minutes (Alarcon et al. , 2007), in line with the rapid loss of oxidized protein chemiluminescence recorded with our imaging system.…”

“…, 2007). Oxidized proteins rather than oxidized free amino acids emit chemiluminescence with a more complex and significantly slower decay (Alarcon et al. , 2007).…”

Four‐membered cyclic peroxides: Carriers of chemical energy

“…In fact, the very existence of the bioluminescence phenomenon implies that the formation of electronically excited species in vivo does not require light. Notwithstanding, living organisms can also emit dim light resulting from peroxidation of biomolecules, such as phospholipids and proteins . This process, called ultraweak chemiluminescence or biophoton emission , reiterates the formation of electronically excited species in vivo .…”

Light emission from tryptophan oxidation by hypobromous acid

“…Examples are oxidation of melatonin catalyzed by peroxidases (25) and oxidation of tryptophan by HOCl (26), peroxynitrite (27) and singlet oxygen (28). In this regard, the indole moiety, which is ubiquitous in living organisms as a component of tryptophan in proteins, the neurotransmitter serotonin and the pineal neurohormone melatonin, deserves special attention, as oxidation of these biomolecules is frequently associated with chemiluminescence.…”