Studies on luciferase from Photobacterium phosphoreum. IX. Further studies on the spectroscopic characteristics of the enzyme-FMN intermediates.
The absorption and fluorometric changes of the reaction mixture of luciferase-FMNH2 complex with O2 were re-examined. Rapid formation (k2(app) = 2.0 s-1 at [O2] = 120 micrometer) of an intermediate with a single absorption maximum at 380 nm within the range of 350-550 nm, and a weak fluorescence at 520 nm (less than or equal to 10% of that of FMN when excited at 380 nm) was observed. The absorption and fluorescence spectra and decay rate of the intermediate were estimated by simulation using an analog computer. The decay rate (0.27 s-1 at 20 degrees C) was in agreement with that of an obligatory intermediate of the luminescent reaction previously determined by measuring aldehyde-initiated luminescence. The process of decay of X1 to FMN involved another intermediate X1' with spectroscopic characteristics rather similar to those of FMN.
D000447:Aldehydes / Q000378:metabolism; D005486:Flavin Mononucleotide / Q000378:metabolism*; D007700:Kinetics; D008156:Luciferases / Q000378:metabolism*; D010101:Oxygen Consumption; D010776:Photobacterium / Q000201:enzymology*; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry
D000447:Aldehydes; D005486:Flavin Mononucleotide; D008156:Luciferases