FRET-based in vivo ca2+ a imaging by a new calmodulin-GFP fusion molecule

Kevin Truong, Asako Sawano, Hideaki Mizuno, Hiroshi Hama, Kit I. Tong, Tapas Kumar Mal, Atsushi Miyawaki, Mitsuhiko Ikura

Research output: Contribution to journalArticlepeer-review

180 Scopus citations

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 twofold increase in the FRET dynamic range within the physiologically significant range of cytoplasmic Ca2+ concentration of 0.05-1 μM.

Original languageEnglish (US)
Pages (from-to)1069-1073
Number of pages5
JournalNature Structural Biology
Volume8
Issue number12
DOIs
StatePublished - 2001

All Science Journal Classification (ASJC) codes

  • Structural Biology
  • Biochemistry
  • Genetics

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