Journal of Biological Physics

, Volume 31, Issue 3–4, pp 249–259 | Cite as

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

  • Sergey Ivanchenko
  • Carlheinz Röcker
  • Franz Oswald
  • Jörg Wiedenmann
  • G. Ulrich Nienhaus
Article

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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Copyright information

© Springer Science + Business Media, Inc. 2005

Authors and Affiliations

  • Sergey Ivanchenko
    • 1
  • Carlheinz Röcker
    • 1
  • Franz Oswald
    • 2
  • Jörg Wiedenmann
    • 3
  • G. Ulrich Nienhaus
    • 1
    • 4
  1. 1.Department of BiophysicsUniversity of UlmUlmGermany
  2. 2.Department of Internal Medicine IUniversity of UlmUlmGermany
  3. 3.Department of General Zoology and EndocrinologyUniversity of UlmUlmGermany
  4. 4.Department of PhysicsUniversity of Illinois at Urbana-ChampaignUrbana

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