Cookies Disclaimer

OK Our site saves small pieces of text information (cookies) on your device in order to deliver better content and for statistical purposes. You can disable the usage of cookies by changing the settings of your browser. By browsing our website without changing the browser settings you grant us permission to store that information on your device.

Publication

Bibliographic Data

  • Authors: Brakemann, T.,Stiel, A.C.,Weber, G.,Andresen, M.,Testa, I.,Grotjohann, T.,Leutenegger, M.,Plessmann, U.,Urlaub, H.,Eggeling, C.,Wahl, M.C.,Hell, S.W. and Jakobs, S.
  • Title: A reversibly photoswitchable GFP-like protein with fluorescence excitation decoupled from switching
  • Journal: Nature Biotech.
  • Volume: 29
  • Issue: 10
  • Volume: 942-947
  • DOI: 10.1038/nbt.1952
  • Highlighted in Nature Biotechnoloy.

    Journal Cover Nature Biotechnology Oct. 2011.

    Access the recommendation on F1000Prime

Abstract

Photoswitchable fluorescent proteins have enabled new approaches for imaging cells, but their utility has been limited either because they cannot be switched repeatedly or because the wavelengths for switching and fluorescence imaging are strictly coupled. We report a bright, monomeric, reversibly photoswitchable variant of GFP, Dreiklang, whose fluorescence excitation spectrum is decoupled from that for optical switching. Reversible on-and-off switching in living cells is accomplished at illumination wavelengths of similar to 365 nm and similar to 405 nm, respectively, whereas fluorescence is elicited at similar to 515 nm. Mass spectrometry and high-resolution crystallographic analysis of the same protein crystal in the photoswitched on- and off-states demonstrate that switching is based on a reversible hydration/dehydration reaction that modifies the chromophore. The switching properties of Dreiklang enable far-field fluorescence nanoscopy in living mammalian cells using both a coordinate-targeted and a stochastic single molecule switching approach.