A photoswitchable orange-to-far-red fluorescent protein, PSmOrange

Nat Methods. 2011 Jul 31;8(9):771-7. doi: 10.1038/nmeth.1664.


We report a photoswitchable monomeric Orange (PSmOrange) protein that is initially orange (excitation, 548 nm; emission, 565 nm) but becomes far-red (excitation, 636 nm; emission, 662 nm) after irradiation with blue-green light. Compared to its parental orange proteins, PSmOrange has greater brightness, faster maturation, higher photoconversion contrast and better photostability. The red-shifted spectra of both forms of PSmOrange enable its simultaneous use with cyan-to-green photoswitchable proteins to study four intracellular populations. Photoconverted PSmOrange has, to our knowledge, the most far-red excitation peak of all GFP-like fluorescent proteins, provides diffraction-limited and super-resolution imaging in the far-red light range, is optimally excited with common red lasers, and can be photoconverted subcutaneously in a mouse. PSmOrange photoswitching occurs via a two-step photo-oxidation process, which causes cleavage of the polypeptide backbone. The far-red fluorescence of photoconverted PSmOrange results from a new chromophore containing N-acylimine with a co-planar carbon-oxygen double bond.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • COS Cells
  • Chlorocebus aethiops
  • Color
  • Female
  • Fluorescence
  • Fluorescent Dyes / chemistry
  • Fluorescent Dyes / radiation effects
  • HEK293 Cells
  • HeLa Cells
  • Humans
  • Light
  • Luminescent Proteins / chemistry*
  • Luminescent Proteins / radiation effects
  • Mice
  • Molecular Sequence Data


  • Fluorescent Dyes
  • Luminescent Proteins
  • fluorescent protein 583

Associated data

  • GENBANK/JN376081