Ultrafast transient fluorescence spectroscopy measurements.A, EAS spectra obtained by global analysis of the transient fluorescence data of WT. The result of the global fit shows a heterogenous decay of the flavin. B, EAS spectra obtained by global analysis of the transient fluorescence data of the D67N mutant. C, individual kinetics of the fluorescence decay observed at 513 nm. Using a monoexponential fit, we obtained a fluorescence lifetime of 40 ± 2 ps for WT and 25 ± 2 ps for D67N. EAS, evolutionary-associated spectra.