Single-cell FRET imaging of transferrin receptor trafficking dynamics by Sfp-catalyzed, site-specific protein labeling

Chem Biol. 2005 Sep;12(9):999-1006. doi: 10.1016/j.chembiol.2005.07.006.


Fluorescence imaging of living cells depends on an efficient and specific method for labeling the target cellular protein with fluorophores. Here we show that Sfp phosphopantetheinyl transferase-catalyzed protein labeling is suitable for fluorescence imaging of membrane proteins that spend at least part of their membrane trafficking cycle at the cell surface. In this study, transferrin receptor 1 (TfR1) was fused to peptide carrier protein (PCP), and the TfR1-PCP fusion protein was specifically labeled with fluorophore Alexa 488 by Sfp. The trafficking of transferrin-TfR1-PCP complex during the process of transferrin-mediated iron uptake was imaged by fluorescence resonance energy transfer between the fluorescently labeled transferrin ligand and TfR1 receptor. We thus demonstrated that Sfp-catalyzed small molecule labeling of the PCP tag represents a practical and efficient tool for molecular imaging studies in living cells.

Publication types

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

MeSH terms

  • Bacterial Proteins / metabolism*
  • Blotting, Western
  • Catalysis
  • Cell Line
  • Cloning, Molecular
  • Endocytosis
  • Fluorescence Resonance Energy Transfer
  • Protein Transport
  • Receptors, Transferrin / metabolism*
  • Transferases (Other Substituted Phosphate Groups) / metabolism*
  • Transferrin / metabolism


  • Bacterial Proteins
  • Receptors, Transferrin
  • Transferrin
  • phosphopantetheinyl transferase
  • Transferases (Other Substituted Phosphate Groups)