posted on 2018-08-08, 00:00authored byAlison
G. Tebo, Frederico M. Pimenta, Martha Zoumpoulaki, Carlos Kikuti, Helena Sirkia, Marie-Aude Plamont, Anne Houdusse, Arnaud Gautier
Fluorescent reporters
are essential components for the design of
optical biosensors that are able to image intracellular analytes in
living cells. Herein, we describe the development of circularly permuted
variants of Fluorescence-Activating and absorption-Shifting Tag (FAST)
and demonstrate their potential as reporting module in biosensors.
Circularly permutated FAST (cpFAST) variants allow one to condition
the binding and activation of a fluorogenic ligand (and thus fluorescence)
to analyte recognition by coupling them with analyte-binding domains.
We demonstrated their use for biosensor design by generating multicolor
plug-and-play fluorogenic biosensors for imaging the intracellular
levels of Ca2+ in living mammalian cells in real time.