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  • Title: FRET-based in vivo Ca2+ imaging by a new calmodulin-GFP fusion molecule.
    Author: Truong K, Sawano A, Mizuno H, Hama H, Tong KI, Mal TK, Miyawaki A, Ikura M.
    Journal: Nat Struct Biol; 2001 Dec; 8(12):1069-73. PubMed ID: 11702071.
    Abstract:
    Intracellular Ca2+ acts as a second messenger that regulates numerous physiological cellular phenomena including development, differentiation and apoptosis. Cameleons, a class of fluorescent indicators for Ca2+ based on green fluorescent proteins (GFPs) and calmodulin (CaM), have proven to be a useful tool in measuring free Ca2+ concentrations in living cells. Traditional cameleons, however, have a small dynamic range of fluorescence resonance energy transfer (FRET), making subtle changes in Ca2+ concentrations difficult to detect and study in some cells and organelles. Using the NMR structure of CaM bound to the CaM binding peptide derived from CaM-dependent kinase kinase (CKKp), we have rationally designed a new cameleon that displays a two-fold increase in the FRET dynamic range within the physiologically significant range of cytoplasmic Ca2+ concentration of 0.05-1 microM.
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