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Bioluminescence resonance energy transfer from aequorin to a fluorophore: an artificial jellyfish for applications in multianalyte detection
Authors:Sapna?K?Deo  Mara?Mirasoli  Email author" target="_blank">Sylvia?DaunertEmail author
Institution:(1) Department of Chemistry, University of Kentucky, Lexington, KY 40506-0055, USA
Abstract:In nature, the green light emission observed in the jellyfish Aequorea victoria is a result of a non-radiative energy transfer from the excited-state aequorin to the green fluorescent protein. In this work, we have modified the photoprotein aequorin by attaching selected fluorophores at a unique site on the protein. This will allow for in vitro transfer of bioluminescent energy from aequorin to the fluorophore thus creating an ldquoartificial jellyfishrdquo. The fluorophores are selected such that the excitation spectrum of the fluorophore overlaps with the emission spectrum of aequorin. By modifying aequorin with different fluorophores, bioluminescent labels with different emission maxima are produced, which will allow for the simultaneous detection of multiple analytes. By examining the X-ray crystal structure of the protein, four different sites for introduction of the unique cysteine residue were evaluated. Two fluorophores with differing emission maxima were attached individually to the mutants through the sulfhydryl group of the cysteine molecule. Two of the fluorophore-labeled mutants showed a peak corresponding to fluorophore emission thus indicating resonance energy transfer from aequorin to the fluorophore.
Keywords:Aequorin  Bioluminescence energy transfer  Site-directed mutagenesis
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