title: A fluorescent sensor for GABA and synthetic GABA(B) receptor ligands.
abstract: While γ-aminobutyric acid (GABA) is the main inhibitory neurotransmitter, suitable tools to measure its concentration in living cells with high spatiotemporal resolution are missing. Herein, we describe the first ratiometric fluorescent sensor for GABA, dubbed GABA-Snifit, which senses GABA with high specificity and spatiotemporal resolution on the surface of living mammalian cells. GABA-Snifit is a semisynthetic fusion protein containing the GABA(B) receptor, SNAP- and CLIP-tag, a synthetic fluorophore and a fluorescent GABA(B) receptor antagonist. When assembled on cell surfaces, GABA-Snifit displays a GABA-dependent fluorescence emission spectrum in the range of 500-700 nm that permits sensing micromolar to millimolar GABA concentrations. The ratiometric change of the sensor on living cells is 1.8. Furthermore, GABA-Snifit can be utilized to quantify the relative binding affinities of GABA(B) receptor agonists, antagonists and the effect of allosteric modulators. These properties make GABA-Snifit a valuable tool to investigate the role of GABA and GABA(B) in biological systems.
authors.affiliation: Institute of Chemical Sciences and Engineering (ISIC), Institute of Bioengineering, NCCR in Chemical Biology, Ecole Polytechnique Fédérale de Lausanne (EPFL), 1015 Lausanne, Switzerland.
authors.forename: Anastasiya
authors.initials: A
authors.lastname: Masharina
authors.forename: Luc
authors.initials: L
authors.lastname: Reymond
authors.forename: Damien
authors.initials: D
authors.lastname: Maurel
authors.forename: Keitaro
authors.initials: K
authors.lastname: Umezawa
authors.forename: Kai
authors.initials: K
authors.lastname: Johnsson
mesh_terms: D000494:Allosteric Regulation; D004591:Electrophoresis, Polyacrylamide Gel; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry*; D057809:HEK293 Cells; D006801:Humans; D008024:Ligands; D018080:Receptors, GABA-B / Q000737:chemistry*; D005680:gamma-Aminobutyric Acid / Q000737:chemistry*
chemical_list: D005456:Fluorescent Dyes; D008024:Ligands; D018080:Receptors, GABA-B; D005680:gamma-Aminobutyric Acid