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- W2023383439 abstract "Ca2+, the most ubiquitous signaling molecule in human body, regulates numerous biological functions by fluxing between the subcellular compartments. Quantitative and real-time detection of Ca2+ concentration fluctuation in specific cellular environments such as the endoplasmic reticulum (ER) is essential to explore the mechanism of Ca2+-dependent cellular signaling. Currently the reported Ca2+ sensors based on natural Ca2+ binding proteins are limited due to the perturbation of Ca2+ signaling. To overcome these limitations, we report the rational design of Ca2+ biosensors by engineering a Ca2+ binding site into a single enhanced green fluorescent protein (EGFP). These developed Ca2+ sensors exhibit a ratiometic fluorescent signal change after binding to Ca2+, with a Kd value optimal for the measurement of Ca2+ in the ER. Metal selectivity of the sensors for Ca2+ in comparison with Ln3+, and excessive biological metal ions such as Mg2+, K+, Na+ has also been examined. In addition, these developed sensors can be targeted to the ER, and exhibit high potential for living cell imaging. Further, their optical and conformational properties have been investigated using various spectroscopic methods. Moreover, pulsed-field-gradient nuclear magnetic resonance spectroscopy has been applied to probe their oligomeric state in solution and conformational changes of specific ligands due to Ca2+ binding were investigated using heteronuclear-labeled proteins with different 2D and 3D NMR techniques." @default.
- W2023383439 created "2016-06-24" @default.
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- W2023383439 date "2009-02-01" @default.
- W2023383439 modified "2023-09-27" @default.
- W2023383439 title "Rational Design of Ca2+ Biosensor" @default.
- W2023383439 doi "https://doi.org/10.1016/j.bpj.2008.12.2787" @default.
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