The bacterial luciferase of P. fischeri is known to catalyze the concomittant oxidation of $FMNH_2$ and long chain aliphatic aldehyde(RCHO) by use of molecular oxygen to FMN and the corresponding carboxylic acid(RCOOH) resulting a blue green luminiscence which has emission peak at 480-490 nm. We consider the reaction of $FMNH_2$ with molecular oxygen as the activation of kinetically inert dioxygen for the effective oxidation of real substrate that is, RCHO to RCOOH.
For the elucidation of the properties of the high energy intermediate(HEI) which is believe to be involved in the oxidation pathway of RCHO and also excitation step of the direct emitter, 6,7-dimethyl-8-ribityl lumazine(LMZ) which is the cofactor of the blue fluorescent protein(BFP) associated to the bacterial luciferase in vivo, the dependence of the initial maximum intensity of light production (Io) on the enzymatic activity in the stopped-flow assay system is demonstrated, and the substrate specificity of the enzyme for aldehyde, the various inhibition factors and the thermal stability is considered.
By comparison of this reaction with various type of monooxygenation, the possible structure of HEI is proposed. We examine the effects of the chemicals that affect the production and cleavage of the structure on the total bioluminescence reaction to confirm this proposition and to make clear the mechanism of the intermolecular energy transfer resulting the elevation of the LMZ to its electronic excited state coupled with conversion of HEI to RCOOH.