6xu4
From Proteopedia
Crystal structure of the genetically-encoded FGCaMP calcium indicator in its calcium-bound state
Structural highlights
FunctionCALM_EMENI Calmodulin mediates the control of a large number of enzymes, ion channels and other proteins by Ca(2+). Among the enzymes to be stimulated by the calmodulin-Ca(2+) complex are a number of protein kinases and phosphatases. Publication Abstract from PubMedGenetically encoded calcium indicators (GECIs) have become a widespread tool for the visualization of neuronal activity. As compared to popular GCaMP GECIs, the FGCaMP indicator benefits from calmodulin and M13-peptide from the fungi Aspergillus niger and Aspergillus fumigatus, which prevent its interaction with the intracellular environment. However, FGCaMP exhibits a two-phase fluorescence behavior with the variation of calcium ion concentration, has moderate sensitivity in neurons (as compared to the GCaMP6s indicator), and has not been fully characterized in vitro and in vivo. To address these limitations, we developed an enhanced version of FGCaMP, called FGCaMP7. FGCaMP7 preserves the ratiometric phenotype of FGCaMP, with a 3.1-fold larger ratiometric dynamic range in vitro. FGCaMP7 demonstrates 2.7- and 8.7-fold greater photostability compared to mEGFP and mTagBFP2 fluorescent proteins in vitro, respectively. The ratiometric response of FGCaMP7 is 1.6- and 1.4-fold higher, compared to the intensiometric response of GCaMP6s, in non-stimulated and stimulated neuronal cultures, respectively. We reveal the inertness of FGCaMP7 to the intracellular environment of HeLa cells using its truncated version with a deleted M13-like peptide; in contrast to the similarly truncated variant of GCaMP6s. We characterize the crystal structure of the parental FGCaMP indicator. Finally, we test the in vivo performance of FGCaMP7 in mouse brain using a two-photon microscope and an NVista miniscope; and in zebrafish using two-color ratiometric confocal imaging. FGCaMP7, an Improved Version of Fungi-Based Ratiometric Calcium Indicator for In Vivo Visualization of Neuronal Activity.,Barykina NV, Sotskov VP, Gruzdeva AM, Wu YK, Portugues R, Subach OM, Chefanova ES, Plusnin VV, Ivashkina OI, Anokhin KV, Vlaskina AV, Korzhenevskiy DA, Nikolaeva AY, Boyko KM, Rakitina TV, Varizhuk AM, Pozmogova GE, Subach FV Int J Mol Sci. 2020 Apr 24;21(8). pii: ijms21083012. doi: 10.3390/ijms21083012. PMID:32344594[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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