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The kinetic mechanisms of fast-decay red-fluorescent genetically-encoded calcium indicators.

Kerruth, S; Coates, C; Dürst, CD; Oertner, TG; Török, K (2019) The kinetic mechanisms of fast-decay red-fluorescent genetically-encoded calcium indicators. J Biol Chem. ISSN 1083-351X
SGUL Authors: Torok, Katalin

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Genetically-encoded calcium indicators (GECIs) are useful reporters of cell signalling, neuronal and network activities. We have generated novel fast variants and investigated the kinetic mechanisms of two recently developed red-fluorescent GECIs (RGECIs), mApple-based jRGECO1a and mRuby-based jRCaMP1a. In the formation of fluorescent jRGECO1a and jRCaMP1a complexes calcium binding is followed by rate-limiting isomerisation. However fluorescence decay of calcium-bound jRGECO1a follows a different pathway form its formation: dissociation of calcium occurs first followed by the peptide, similarly to GCaMP-s. In contrast, fluorescence decay of calcium-bound jRCaMP1a occurs by the reversal of the on-pathway: peptide dissociation is followed by calcium. The mechanistic differences explain the generally slower off-kinetics of jRCaMP1a-type indicators compared to GCaMP-s and jRGECO1a-type GECI: the fluorescence decay rate of f-RCaMP1 was 21 s-1, compared to 109 s-1 for f-RGECO1 and f‑RGECO2 (37 °C). Thus, the CaM-peptide interface is an important determinant of the kinetic responses of GECIs, however the topology of the structural link to the fluorescent protein demonstrably affects the internal dynamics of the CaM-peptide complex. In the dendrites of hippocampal CA3 neurons, f-RGECO1 indicates calcium elevation in response to a 100 action potential train in a linear fashion, making the probe particularly useful for monitoring large amplitude, fast signals e.g. those in dendrites, muscle and immune cells.

Item Type: Article
Additional Information: This research was originally published in the Journal of Biological Chemistry.
Keywords: biosensor, calcium, calcium imaging, fluorescence, kinetics, 06 Biological Sciences, 11 Medical And Health Sciences, 03 Chemical Sciences, Biochemistry & Molecular Biology
SGUL Research Institute / Research Centre: Academic Structure > Molecular and Clinical Sciences Research Institute (MCS)
Journal or Publication Title: J Biol Chem
ISSN: 1083-351X
Language: eng
16 January 2019Published Online
16 January 2019Accepted
Publisher License: Publisher's own licence
Project IDFunderFunder ID
BB/M02556X/1Biotechnology and Biological Sciences Research Council
PubMed ID: 30651353
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