Kinetics of bacterial bioluminescence and the fluorescent transient.

The addition of FMNH(2), to Vibrio harveyi luciferase at 2°C in the presence of tetradecanal results in the formation of a highly fluorescent transient species with a spectral distribution indistinguishable from that of the bioluminescence. The bioluminescence reaches maximum intensity in 1.5 s and decays in a complex manner with exponential components of 10(-1) s(-1) , 7 x 10(-3)S(-1). and 7 x10(4)s(-1). The fluorescent transient rises exponentially at 7 x 10(-2)s(-1) and decays at 3 x 10 (4)s(-1) . The slowest bioluminescence component. comprising the bulk of the bioluminescence. decays at twice the rate of the fluorescent transient under all variations of reaction conditions: concentration of reactants.temperature 2 - 20°C. and aldehyde chain length - decana1, dodecanal and tetradecanal. The activation energy for both the slowest bioluminescence decay and the transient fluorescence decay is 80 kJ-mol(-1). An energy transfer scheme is proposed to explain the results where two distinct chemically energized species utilize the fluorescent transient as emitter for the slower bioluminescences, and for the faster process a fluorophore present in the protein preparation. Kinetic observations suggest that typical preparations of V. harveyi luciferase comprise 15% active protein.