The impact of LuxF on light intensity in bacterial bioluminescence

Abstract

The enzymes involved in bacterial bioluminescence are encoded in the lux operon with a conserved gene order of luxCDABEG. Some photobacterial strains carry an additional gene, termed luxF, which produces the LuxF protein, whose function and influence on bacterial bioluminescence is still uncertain. The LuxF protein binds the flavin derivative 6-(3′-(R)-myristyl)-flavin mononucleotide (myrFMN), which is generated as a side product in the luciferase-catalyzed reaction. This study utilized an Escherichia coli (E. coli) based lux operon expression system where the lux operons of Photobacterium leiognathi subsp. mandapamensis 27561 or of Photobacterium leiognathi subsp. leiognathi 25521, namely luxCDAB(F)EG, were cloned into a single expression vector. Exclusion of luxF gene from the lux operon enabled novel insights into the role of LuxF protein in light emission. E. coli cultures harboring and expressing the genes of the lux operon including luxF gene emit more light than without luxF gene. Furthermore, isolation of the tightly bound flavin derivative revealed the presence of at least three different flavin derivatives. Analysis by UV/Vis absorption and NMR spectroscopy as well as mass spectrometry showed that the flavin derivatives bear fatty acids of various chain lengths. This distribution of FMN derivatives is vastly different to what was found in bioluminescent bacteria and indicates that the luciferase is supplied with a range of aldehyde substrates in E. coli.