Analysis of Photoexcitation Energy Dependence in the Photoluminescence of Firefly Luciferin

Abstract

The whole pathways for photoluminescence, which include absorption, relaxation and emission, of firefly luciferin in aqueous solutions of different pH values with different photoexcitation energies were theoretically investigated by considering protonation/deprotonation. It is experimentally known that the color of fluorescence changes from green to red with a decrease in the photoexcitation energy. We confirmed with the theoretical analysis that the peak energy shift in the fluorescence spectra with varying photoenergies is due to a change in photoluminescence pathway. When the photoexcitation energy is decreased, the red emission from a monoanion form of firefly luciferin with carboxylate and phenolate groups and N-protonated thiazoline ring occurs irrespective of the pH values. However, because the species abundant in the solution and those excited by the photon depend on the solution pH, the pathway leading to the monoanion form changes with the solution pH.

Oxyluciferin Photoacidity: The Missing Element for Solving the Keto–Enol Mystery?

“…Thus, the ∼390 nm band was assigned to the + HNROH species, while the ∼590 nm band was assigned to the excited‐state zwitterion + HNRO − species 22. The experimental deduction that the protonation of a nitrogen heteroatom is responsible for the redshift in the absorption spectrum was corroborated by two theoretical computational studies 29. 30…”

Theoretical insights into the effect of pH values on oxidation processes in the emission of firefly luciferin in aqueous solution

“…The most important theoretical protocol for identifying a reaction pathway is the estimation and comparison of total energies for the reactant, transition state, and product . In aqueous solutions, luciferin exists as its carboxylate anion (LH − ) at pH 7–8 and dianion (L 2− ) at pH 9, whereas oxyluciferin exists as phenolate‐keto, phenolate‐enol, or enolate forms . Accordingly, we identify the emission process by comparison of the potential energy profiles, including the energies of intermediate chemical species such as LH‐AMP, Diox, and their deprotonated chemical species.…”

“…In previous studies, deprotonation in a general ‘AH’ molecule was described by the following calculation:…”

“…Because firefly bioluminescence is light emission caused by chemical reactions in proteins, the intensity of light signal increases when this quantum yield and/or reaction rate increases. This importance has motivated further studies on the mechanism of the chemical reaction in firefly bioluminescence . In 2006, the crystal structures of genjibotaru ( Luciola cruciata ) luciferase containing D‐luciferin bound to an adenosine monophosphate (AMP) analog or oxyluciferin were reported .…”

“…In 2006, the crystal structures of genjibotaru ( Luciola cruciata ) luciferase containing D‐luciferin bound to an adenosine monophosphate (AMP) analog or oxyluciferin were reported . Determination of these crystal structures stimulated theoretical studies on firefly bioluminescence …”