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      Ca2+ indicators based on computationally redesigned calmodulin-peptide pairs.

      Chemistry & Biology
      Amino Acid Sequence, Calcium, chemistry, metabolism, Calmodulin, Cell Membrane, Cytosol, HeLa Cells, Humans, Models, Molecular, Molecular Sequence Data, Peptides, Protein Binding, Subcellular Fractions

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          Abstract

          The binding interface of calmodulin and a calmodulin binding peptide were reengineered by computationally designing complementary bumps and holes. This redesign led to the development of sensitive and specific pairs of mutant proteins used to sense Ca(2+) in a second generation of genetically encoded Ca(2+) indicators (cameleons). These cameleons are no longer perturbed by large excesses of native calmodulin, and they display Ca(2+) sensitivities tuned over a 100-fold range (0.6-160 microM). Incorporation of circularly permuted Venus in place of Citrine results in a 3- to 5-fold increase in the dynamic range. These redesigned cameleons show significant improvements over previous versions in the ability to monitor Ca(2+) in the cytoplasm as well as distinct subcellular localizations, such as the plasma membrane of neurons and the mitochondria.

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