Ontology type: schema:ScholarlyArticle Open Access: True
2019-12
AUTHORSMark A. Lobas, Rongkun Tao, Jun Nagai, Mira T. Kronschläger, Philip M. Borden, Jonathan S. Marvin, Loren L. Looger, Baljit S. Khakh
ABSTRACTAdenosine 5' triphosphate (ATP) is a universal intracellular energy source and an evolutionarily ancient, ubiquitous extracellular signal in diverse species. 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 responds to relevant ATP concentrations (30 μM 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. After careful consideration of their modest pH sensitivity, iATPSnFRs represent promising reagents for imaging ATP in the extracellular space and within cells during a variety of settings, and for further application-specific refinements. More... »
PAGES711
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DOIhttp://dx.doi.org/10.1038/s41467-019-08441-5
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