Targeted green-red photoconversion of EosFP, a fluorescent marker protein
Ivanchenko, S., Röcker, C., Oswald, F., Wiedenmann, J. and Nienhaus, G.U. (2005) Targeted green-red photoconversion of EosFP, a fluorescent marker protein. Journal of Biological Physics, 31, (3-4), 249-259. (doi:10.1007/s10867-005-0174-z).
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EosFP is a novel fluorescent protein from the stony coral Lobophyllia hemprichii. Its gene was cloned in Escherichia coli to express the tetrameric wild-type protein. The protein emits strong green fluorescence (516 nm) that shifts toward red (581 nm) upon near-ultraviolet irradiation at ∼390 nm due to a photo-induced modification that involves a break in the peptide backbone next to the chromophore. Using site-directed mutagenesis, dimeric (d1EosFP, d2EosFP) and monomeric (mEosFP) variants were produced with essentially unaltered spectroscopic properties. Here we present a spectroscopic characterization of EosFP and its variants, including room- and low-temperature spectra, fluorescence lifetime determinations, two-photon excitation and two-photon photoconversion. Furthermore, by transfection of a human cancer (HeLa) cell with a fusion construct of a mitochondrial targeting sequence and d2EosFP, we demonstrate how localized photoconversion of EosFP can be employed for resolving intracellular processes.
|Keywords:||fluorescent protein - fusion protein - two-photon excitation - fluorescence spectroscopy - photoconversion|
|Subjects:||Q Science > QH Natural history > QH301 Biology|
|Divisions:||University Structure - Pre August 2011 > School of Ocean & Earth Science (SOC/SOES)
|Date Deposited:||08 May 2008|
|Last Modified:||27 Mar 2014 18:34|
|RDF:||RDF+N-Triples, RDF+N3, RDF+XML, Browse.|
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