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. 2005 Dec;31(3-4):249–259. doi: 10.1007/s10867-005-0174-z

Targeted Green-Red Photoconversion of EosFP, a Fluorescent Marker Protein

Sergey Ivanchenko 1, Carlheinz Röcker 1, Franz Oswald 2, Jörg Wiedenmann 3, G Ulrich Nienhaus 1,4,✉
PMCID: PMC3456337  PMID: 23345897

Abstract

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.

Key words: fluorescent protein, fusion protein, two-photon excitation, fluorescence spectroscopy, photoconversion

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