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PLoS One. 2014 Apr 21;9(4):e95808. doi: 10.1371/journal.pone.0095808. eCollection 2014.

An engineered palette of metal ion quenchable fluorescent proteins.

Author information

1
Laboratory of Molecular Biophysics, National Heart Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
2
Laboratory of Molecular Biology, National Institute Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, United States of America.
3
Laboratory of Biochemistry, National Heart Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, United States of America.

Abstract

Many fluorescent proteins have been created to act as genetically encoded biosensors. With these sensors, changes in fluorescence report on chemical states in living cells. Transition metal ions such as copper, nickel, and zinc are crucial in many physiological and pathophysiological pathways. Here, we engineered a spectral series of optimized transition metal ion-binding fluorescent proteins that respond to metals with large changes in fluorescence intensity. These proteins can act as metal biosensors or imaging probes whose fluorescence can be tuned by metals. Each protein is uniquely modulated by four different metals (Cu2+, Ni2+, Co2+, and Zn2+). Crystallography revealed the geometry and location of metal binding to the engineered sites. When attached to the extracellular terminal of a membrane protein VAMP2, dimeric pairs of the sensors could be used in cells as ratiometric probes for transition metal ions. Thus, these engineered fluorescent proteins act as sensitive transition metal ion-responsive genetically encoded probes that span the visible spectrum.

PMID:
24752441
PMCID:
PMC3994163
DOI:
10.1371/journal.pone.0095808
[Indexed for MEDLINE]
Free PMC Article

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