Journal of Biological Chemistry · 2000 · 329 citations · 15 references
Natural SciencesBiologyZoologyMolecular ImagingFluorescence ImagingFluorescence MicroscopyFluorescence SpectroscopyPhotophysics Of Fluorescent ProbesPhotosynthetic PigmentsAnimal PigmentsColoration SignalsGreen Fluorescent Protein ReportersProtein Fluorescence
It is generally accepted that the colors displayed by living organisms are determined by low molecular weight pigments or chromoproteins that require a prosthetic group. The exception to this rule is green fluorescent protein (GFP) from Aequorea victoria that forms a fluorophore by self-catalyzed protein backbone modification. Here we found a naturally nonfluorescent homolog of GFP to determine strong purple coloration of tentacles in the sea anemone Anemonia sulcata. Under certain conditions, this novel chromoprotein produces a trace amount of red fluorescence (emission lambda(max) = 595 nm). The fluorescence demonstrates unique behavior: its intensity increases in the presence of green light but is inhibited by blue light. The quantum yield of fluorescence can be enhanced dramatically by single amino acid replacement, which probably restores the ancestral fluorescent state of the protein. Other fluorescent variants of the novel protein have emission peaks that are red-shifted up to 610 nm. They demonstrate that long wavelength fluorescence is attainable in GFP-like fluorescent proteins.
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Site-directed mutagenesis by overlap extension using the polymerase chain reaction
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Crystal Structure of the <b> <i>Aequorea victoria</i> </b> Green Fluorescent Protein
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