We have engineered a rhodopsin-based ratiometric fluorescent biosensor for chloride that is membrane-bound, operates across a physiological pH regime, and has the farthest red single-photon excitation and emission profile to date.
Chloride is a vital ion for all forms of life. Nature has evolved elegant supramolecular machines to facilitate chloride transport across cell membranes. Protein-based fluorescent biosensors can enable researchers to intercept and monitor chloride in living cells but remain underdeveloped. Here, we demonstrate how this can be achieved through the introduction of a single point mutation in an engineered, microbial rhodopsin from the archaebacterium Halorubrum sodomense resulting in ChloRED-1-CFP. This membrane-bound host is a far-red emitting, ratiometric sensor that provides a reversible readout of chloride in live bacteria at physiological pH, setting the stage to investigate the roles of chloride in a wide range of biological contexts.
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