Photoswitchable Cyan Fluorescent Protein for Protein Tracking

Nat Biotechnol. 2004 Nov;22(11):1435-9. doi: 10.1038/nbt1025. Epub 2004 Oct 24.

Abstract

In recent years diverse photolabeling techniques using green fluorescent protein (GFP)-like proteins have been reported, including photoactivatable PA-GFP, photoactivatable protein Kaede, the DsRed 'greening' technique and kindling fluorescent proteins. So far, only PA-GFP, which is monomeric and gives 100-fold fluorescence contrast, could be applied for protein tracking. Here we describe a dual-color monomeric protein, photoswitchable cyan fluorescent protein (PS-CFP). PS-CFP is capable of efficient photoconversion from cyan to green, changing both its excitation and emission spectra in response to 405-nm light irradiation. Complete photoactivation of PS-CFP results in a 1,500-fold increase in the green-to-cyan fluorescence ratio, making it the highest-contrast monomeric photoactivatable fluorescent protein described to date. We used PS-CFP as a photoswitchable tag to study trafficking of human dopamine transporter in living cells. At moderate excitation intensities, PS-CFP can be used as a pH-stable cyan label for protein tagging and fluorescence resonance energy transfer applications.

Publication types

  • Evaluation Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Escherichia coli / cytology
  • Escherichia coli / metabolism*
  • Fluorescence Resonance Energy Transfer / methods*
  • Green Fluorescent Proteins / chemistry*
  • Green Fluorescent Proteins / radiation effects
  • Microscopy, Fluorescence / methods*
  • Molecular Sequence Data
  • Photochemistry / methods
  • Protein Transport / physiology*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / radiation effects

Substances

  • Cyan Fluorescent Protein
  • Recombinant Proteins
  • Green Fluorescent Proteins

Associated data

  • GENBANK/AY533824