title: Activities, kinetics and emission spectra of bacterial luciferase-fluorescent protein fusion enzymes.
abstract: A new approach to alter bacterial bioluminescence color was developed by fusing Vibrio harveyi luciferase with the coral Discosoma sp. fluorescent protein mOrange, a homolog of the Aequorea green fluorescent protein. Attachment of mOrange to the N- or C-terminus of luciferase α or β subunit, via a 5 or 10 residue linker, produced fully active fusion enzymes. However, only the fusion of mOrange to the N-terminus of luciferase α produced a new 560 nm emission. The differences in emission color by two such fusion enzymes from that of the wild-type luciferase (λ(max) 490 nm) were evident by eye or photographically with the aid of cut-off optical filters. In nonturnover reactions, light decay rates of fusion enzyme remained the same when monitored as the full-spectrum light or at 480 nm (from the luciferase emitter) or 570 nm (from mOrange). No 560 nm emission component was observed with a mixture of luciferase and free mOrange. These findings support that the 560 nm emission by the fusion enzyme was due to bioluminescence resonance energy transfer from luciferase to mOrange. We believe that the same approach could also alter the bacterial bioluminescence color by covalent attachment of other suitable fluorescent proteins or chromophores to luciferase.
authors.affiliation: Department of Biology and Biochemistry, University of Houston, Houston, TX, USA.
authors.forename: Dachuan
authors.initials: D
authors.lastname: Ke
authors.forename: Shiao-Chun
authors.initials: SC
authors.lastname: Tu
mesh_terms: D001483:Base Sequence; D017931:DNA Primers; D049452:Green Fluorescent Proteins / Q000378:metabolism*; D007700:Kinetics; D008156:Luciferases / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D014733:Vibrio / Q000201:enzymology*
chemical_list: D017931:DNA Primers; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D008156:Luciferases