Novel Green Genetically Encoded Fluorescent Sensor for Sulfate for Probing Sulfate Transport Activity of Solute Carrier Family 26 Member a2 (Slc26a2) Protein

Abstract

Abstract Genetically encoded fluorescent biosensors became indispensable tools for biological research enabling real-time observation of physiological processes in live cells. Recent protein engineering effort has resulted in generation large variety of fluorescent biosensors for wide range of biologically relevant processes from small ions to enzymatic activity and signaling pathways. However, biosensors for imaging sulfate ion, the fourth most abundant physiological anion, in mammalian cells are still lacking. Here we report the development and characterization of green fluorescent biosensor for sulfate named Thyone. Thyone, derived through structure-guided design from bright green fluorescent protein mNeonGreen, exhibited large negative fluorescence response upon association with sulfate anion with affinity of 11 mM in mammalian cells. High anion selectivity and sensitivity of Thyone allowed for imaging of sulfate anion transients mediated by sulfate transporter heterologously expressed in cultured mammalian cells. We believe that Thyone will find a broad application for assaying the sulfate transport in mammalian cells via anion transporters and exchangers.