PMID:3480518

Citation	Daubner, SC, Astorga, AM, Leisman, GB and Baldwin, TO (1987) Yellow light emission of Vibrio fischeri strain Y-1: purification and characterization of the energy-accepting yellow fluorescent protein. Proc. Natl. Acad. Sci. U.S.A. 84:8912-6
Abstract	A strain of luminous bacteria, Vibrio fischeri Y-1, emits yellow light rather than the blue-green emission typical of other luminous bacteria. The yellow emission has been postulated previously to result from energy transfer from an electronically excited species formed in the bacterial luciferase-catalyzed reaction to a secondary emitter protein, termed the yellow fluorescent protein (YFP). We report here the purification of YFP to homogeneity without loss of the chromophore. The protein was found to be a homodimer of Mr 22,000 subunits with one weakly bound FMN per subunit. The FMN-protein complex was stabilized by 10% (vol/vol) glycerol in the buffers, allowing purification of the active holo-YFP. The protein migrated as a single spot with an isoelectric point of approximately 6.5 on two-dimensional polyacrylamide gel electrophoresis and gave an N-terminal sequence of Met-Phe-Lys-Gly-Ile-Val-Glu-Gly-Ile-Gly-Ile-Ile-Glu-Lys-Ile. Addition of purified YFP to a reaction in which luciferase was supplied with FMNH2 (reduced FMN) by a NADH:FMN oxidoreductase resulted in a dramatic enhancement in the intensity of bioluminescence and an additional peak in the emission spectrum at about 534 nm. The resulting bimodal bioluminescence emission spectrum had peaks at 484 nm, apparently due to emission from the luciferase-flavin complex, and at 534 nm, corresponding to the fluorescence emission maximum of YFP. This bimodal spectrum closely matched the emission spectrum in vivo.
Links	PubMed PMC299661
Keywords	Bacterial Proteins/isolation & purification; Energy Transfer; Luciferases/metabolism; Luminescent Measurements; Luminescent Proteins/isolation & purification; Molecular Weight; Spectrum Analysis; Vibrio/physiology
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
ALIFS:LUXY		GO:0008218: bioluminescence	ECO:0000314:		P		Fig 4 shows the presence of YFP in luciferase-FMN oxidoreductase assay emits light at 484nm and 534nm.	complete CACAO 8836
ALIFS:LUXY	involved_in	GO:0008218: bioluminescence	ECO:0000314: direct assay evidence used in manual assertion		P		Seeded From UniProt	complete

References

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