We combine molecular chemistry, molecular biology, protein engineering and directed evolution techniques to create tools that allow biologists to explore life in new ways. These tools are composed of two parts: a protein module and a synthetic small molecule. The advantage of using a protein module is that instructions for its manufacture can be easily and specifically introduced into cells in the form of DNA. In addition, its properties can be adjusted using protein evolution techniques. The interest of using a small synthetic molecule is to be able to use molecular engineering to refine its properties, and thus benefit from the power of modern chemistry to explore biological processes.
This original approach allowed us to create FAST (Fluorescence-Activating and absorption-Shifting Tag), a new fluorescent chemogenetic reporter for biological imaging. FAST is a small genetically encoded protein tag of 14 kDa evolved from the Photoactive Yellow Protein (PYP). Using revolutionary directed evolution techniques, this protein has been redesigned to selectively and reversibly bind synthetic fluorogenic ligands (so-called fluorogens) from the hydroxybenzilidene rhodanine (HBR) family. These fluorogenic ligands strongly fluoresce only when bound to FAST, enabling to selectively image proteins fused to FAST without the need of washing the excess of fluorogenic ligand.
Half as large as the green fluorescent protein (GFP), FAST has a reduced genetic footprint and minimizes the risk of dysfunctional fusions. In addition, FAST fluorescence is instantaneous and does not need molecular oxygen, solving long standing issues of GFP-like fluorescent proteins. Combining the advantages of synthetic labels with the targeting specificity of genetically encoded tags, FAST allowed us to build and breed a collection of fluorescent reporters for e.g. the characterization of protein localization and dynamics, the detection of protein-protein interactions, the measure of analyte levels or the visualization of cellular events (e.g. cell cycle) with unprecedent spatial and temporal resolution in living systems.