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A genetically encoded single-wavelength sensor for imaging cytosolic and cell surface ATP

By Mark Lobas, Jun Nagai, Mira T. Kronschl├Ąger, Philip Borden, Jonathan S. Marvin, Loren L. Looger, Baljit S. Khakh

Posted 06 Aug 2018
bioRxiv DOI: 10.1101/385484 (published DOI: 10.1038/s41467-019-08441-5)

Adenosine triphosphate (ATP) is a universal intracellular energy source and an evolutionarily ancient extracellular signal. Here, we report the generation and characterization of single-wavelength genetically encoded fluorescent sensors (iATPSnFRs) for imaging extracellular and cytosolic ATP from insertion of circularly permuted superfolder GFP into the epsilon subunit of F0F1-ATPase from Bacillus PS3. On the cell surface and within the cytosol, iATPSnFR1.0 responded to relevant ATP concentrations (30 uM to 3 mM) with fast increases in fluorescence. iATPSnFRs can be genetically targeted to specific cell types and sub-cellular compartments, imaged with standard light microscopes, do not respond to other nucleotides and nucleosides, and when fused with a red fluorescent protein function as ratiometric indicators. iATPSnFRs represent promising new reagents for imaging ATP dynamics.

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