A new fluorescent biosensor for inositol trisphosphate |
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| Authors: | Morii Takashi Sugimoto Kenji Makino Keisuke Otsuka Masami Imoto Keiji Mori Yasuo |
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| Institution: | Institute of Advanced Energy, Kyoto University, PRESTO, Japan Science and Technology Corporation, Uji, Kyoto 611-0011, Japan. |
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| Abstract: | An intracellular second messenger d-myo-inositol-1,4,5-trisphosphate (IP3) is a key biological signaling molecule that controls the cellular Ca2+ concentration. We report the preparation and evaluation of a functionalized protein-based sensor for IP3 by exploring the selective IP3 binding properties of pleckstrin homology (PH) domain. Signal transduction is imparted to the protein by mutation of proximal residues to cysteine and then alkylation of the active site by various fluorophore derivatives. This creates functionalized proteins that show micromolar affinity for IP3, reasonably strong fluorescence emission, and wavelength changes in the fluorophore and selectivity higher than the original PH domain among different inositol phosphate derivatives. |
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