Red fluorescent protein with reversibly photoswitchable absorbance for photochromic FRET

Chem Biol. 2010 Jul 30;17(7):745-55. doi: 10.1016/j.chembiol.2010.05.022.

Abstract

We have developed the first red fluorescent protein, named rsTagRFP, which possesses reversibly photoswitchable absorbance spectra. Illumination with blue and yellow light switches rsTagRFP into a red fluorescent state (ON state) or nonfluorescent state (OFF state), respectively. The ON and OFF states exhibit absorbance maxima at 567 and 440 nm, respectively. Due to the photoswitchable absorbance, rsTagRFP can be used as an acceptor for a photochromic Förster resonance energy transfer (pcFRET). The photochromic acceptor facilitates determination of a protein-protein interaction by providing an internal control for FRET. Using pcFRET with EYFP as a donor, we observed an interaction between epidermal growth factor receptor and growth factor receptor-binding protein 2 in live cells by detecting the modulation of both the fluorescence intensity and lifetime of the EYFP donor upon the ON-OFF photoswitching of the rsTagRFP acceptor.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Survival
  • ErbB Receptors / metabolism
  • Fluorescence Resonance Energy Transfer*
  • GRB2 Adaptor Protein / metabolism
  • HeLa Cells
  • Humans
  • Light*
  • Luminescent Proteins / chemistry*
  • Luminescent Proteins / genetics*
  • Mutagenesis
  • Mutation
  • Protein Engineering / methods*
  • Red Fluorescent Protein

Substances

  • GRB2 Adaptor Protein
  • Luminescent Proteins
  • ErbB Receptors