Protein structure and bioluminescent spectra for firefly bioluminescence

Abstract

Modern theory on general and specific effects of microenvironment on emission spectra was used for explanation of spectral differences for both natural and mutant forms of beetle luciferases, as well as for bioluminescence emitter oxyluciferin in model systems. For the analysis, both authors' and other published data were used. It was shown that active site mutations that resulted in spectral shifts of bioluminescence as a rule caused substantial decrease in the catalytic activity of the enzyme. At the same time, mutations in the conservative regions of the protein amino acid sequence that were in the periphery of the protein globe resulted in red shift of the bioluminescence spectra without affecting catalytic activity. Correlation was observed between the value of spectral shift and polarizability of the introduced amino acid residue: the higher the polarizability, the larger was the red shift of bioluminescence.

D-Luciferin, derivatives and analogues: synthesis and in vitro/in vivo luciferase-catalyzed bioluminescent activity

“…However, all the above explanations are only a partial view of a more complex "microenvironment mechanism" that has to take in consideration the protonation state of the O6′ atom, in turn related to the polarity of the hydrogen bond that can be formed with Arg 218. 42,43 Finally, the control of the resonance-based charge delocalization of the anionic keto-form of the oxyluciferin excited should be further stabilized. This could be achieved extending the π-electron system through a quinone-enolate species at a lower energy responsible for red light emission …”

Theoretical Study of the Relationships between Excited State Geometry Changes and Emission Energies of Oxyluciferin

“…[10][11][12][13][14][15][16] Several mechanisms have been proposed to explain this phenomenon. 1,2,4,6,11,12,14,[17][18][19][20][21][22][23][24][25] To the present, the detailed mechanism of the bioluminescence and its shifts in color remain unclear due to the complexity of the microenvironment in vivo.…”

A Dual‐Color Far‐Red to Near‐Infrared Firefly Luciferin Analogue Designed for Multiparametric Bioluminescence Imaging

“…Between l = 600-800 nm, the absorption of light by Hb decreases by a factor of approximately 50, resulting in less attenuation of red light. This wavelength range is within what is termed the "bio-optical window" and there has been much focus on engineering red-shifted Fluc enzymes that have maximum emission wavelengths in this range, [10][11][12][13][14][15] but these have peaked at wavelengths less than l = 645 nm.The most red-shifted luciferin analogues to date [16] are based upon amino derivatives (Figure 1 b), for example cyclic aminoluciferin (3 a: l max = 599 nm; 3 b: l max = 607 nm), [17] seleno-d-aminoluciferin (4: l max = 600 nm), [18] and a rationally designed 4-(dimethylamino)phenyl derivative conjugated to a thiazoline group (5: l max = 675 nm). [19] In particular cyclic aminoluciferin derivative 3 a has been shown to give improved bioluminescence imaging compared to luciferin (LH 2 ; 1) at dilute concentrations where the intracellular concentration of the luciferin or analogue is limiting.…”

“…Between l = 600-800 nm, the absorption of light by Hb decreases by a factor of approximately 50, resulting in less attenuation of red light. This wavelength range is within what is termed the "bio-optical window" and there has been much focus on engineering red-shifted Fluc enzymes that have maximum emission wavelengths in this range, [10][11][12][13][14][15] but these have peaked at wavelengths less than l = 645 nm.…”

“…[11][12][13] Current measurements and calculations suggest that color modulation is due to perturbing interactions in the microenvironment surrounding the anionic phenolate of excited-state oxyluciferin (2) in the Fluc active site. [22][23][24][25][26][27][28] Additionally, p-p overlap between the benzothiazole and thiazolone heterocycles in 2 also appears to be important.…”