Deciphering the molecular mechanism responsible for GCaMP6m's Ca2+-dependent change in fluorescence

The goal of this work is to determine how GCaMP6m's fluorescence is altered in response to Ca2+-binding. Our detailed spectroscopic study reveals the simplest explanation for how GCaMP6m changes fluorescence in response to Ca2+ is with a four-state model, in which a Ca2+-dependent change of the...

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Bibliographic Details
Published in:PloS one 2017-02, Vol.12 (2), p.e0170934-e0170934
Main Authors: Barnett, Lauren M, Hughes, Thomas E, Drobizhev, Mikhail
Format: Article
Language:English
Subjects:
Biology
Biology and Life Sciences
Biosensors
Brain
Calcium
Calcium - metabolism
Calcium-binding protein
Calmodulin
Ecology and Environmental Sciences
Electrostatic properties
Fluorescence
Green Fluorescent Proteins - chemistry
Green Fluorescent Proteins - genetics
Green Fluorescent Proteins - metabolism
HEK293 Cells
Humans
Molecular Dynamics Simulation
Neurosciences
Physical Sciences
Protein Binding
Protein kinase A
Proteins
Protonation
Recombinant Proteins - chemistry
Recombinant Proteins - genetics
Recombinant Proteins - metabolism
Research and Analysis Methods
Sensors
Spectrum analysis
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Summary:The goal of this work is to determine how GCaMP6m's fluorescence is altered in response to Ca2+-binding. Our detailed spectroscopic study reveals the simplest explanation for how GCaMP6m changes fluorescence in response to Ca2+ is with a four-state model, in which a Ca2+-dependent change of the chromophore protonation state, due to a shift in pKa, is the predominant factor. The pKa shift is quantitatively explained by a change in electrostatic potential around the chromophore due to the conformational changes that occur in the protein when calmodulin binds Ca2+ and interacts with the M13 peptide. The absolute pKa values for the Ca2+-free and Ca2+-saturated states of GCaMP6m are critical to its high signal-to-noise ratio. This mechanism has important implications for further improvements to GCaMP6m and potentially for other similarly designed biosensors.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0170934