PROTEIN STRUCTURE AND BIOLUMINESCENT SPECTRA FOR FIREFLY BIOLUMINESCENCE

摘要

Beetle luciferases isolated from various insects are characterized by identical structures of both substrates and emitter. The main difference is in color of bioluminescence that is determined by the properties of microenvironment of the emitter in the enzyme active site. Several mechanisms were proposed to explain the bioluminescence color difference for native and mutant luciferases. According to White bioluminescence spectral shifts are attributed to the changes in the polarity of oxyluciferin binding site similar to the effects of polarity of the solvent on spectral properties of fluorophore. However, this mechanism could not explain the existence of two forms of oxyluciferin (keton and enol) at different pH. According to other theory, the changes in bioluminescent spectra, especially at different pH, could be the result of keto-enol tautomerization of oxyluciferin. However, this mechanism could not explain the spectral shifts without changes in the shape that were observed for click-beetles and mutant luciferases. According to McCapra excited oxyluciferin could exist in two stereo conformations that have different energy and provide different color of emission. However, this mechanism could not explain green fluorescence of enol forms of oxyluciferin and its analogues. Besides, the calculations of molecular structure of oxyluciferin in ground and excited state revealed that planar conformations had minimal energy and thus were optimal for both ground and first singlet excited states. It seems that the mechanism of "twisted" conformation for c olor c hanges i n firefly b ioluminescence i s non feasible. In this paper theoretical consideration of environment effect on emission spectra is used for analysis of spectral changes in different natural and mutant forms of luciferases.