Kamis, 01 Februari 2018

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Optimization of a GCaMP Calcium Indicator for Neural Activity ...
src: www.jneurosci.org

GCaMP is a genetically encoded calcium indicator, or GECI initially developed by Junichi Nakai. GCaMP is created from a fusion of green fluorescent protein (GFP), calmodulin, and M13, a peptide sequence from myosin light chain kinase. The advantage of GECI's is that they can be genetically specified for studies in living organisms. The first transgenic mouse expressing a GCaMP was reported in 2004 and GCaMP was subsequently improved to GCaMP2, which was stable at mammalian body temperatures and enabled the first in vivo mammalian recordings using a GECI. GCaMPs have been subsequently modified to progressively improve the range of the fluorescence signal, resulting in GCaMP3 through GCaMP8. Additionally, red fluorescence GECIs, termed "RCaMPs" have been developed to expand the spectral options for multi-lineage imaging.

GFP is circularly permutated so that the N- and C-termini are fused, creating a new terminus in the middle of the protein. Fused to the new terminus is calmodulin (CaM) and the M13 domain of a myosin light chain kinase. Calmodulin is a symmetrical, hinge-like protein that binds to four calcium ions via E-F motifs. When calcium is present, CaM undergoes a conformational change, and the hinge region is able to bind helical peptide chains on target proteins, such as M13. In the absence of calcium, the circularly permutated fluorescent proteins exist in a poorly fluorescent state due to a water pathway that enable protonation of the chromophore and poor absorbance at the excitation wavelengths. Ca2+ binding to the calmodulin moiety results in a structural shift that eliminates this solvent pathway, rapid de-protonation of the chromophore, and bright fluorescence.


Video GCaMP



See also

  • Optogenetics

Maps GCaMP



References

Source of the article : Wikipedia

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