Open AccessDeciphering the molecular mechanism responsible for GCaMP6m's Ca2+-dependent change in fluorescence
Author(s) -
Lauren M. Barnett,
Thomas E. Hughes,
Mikhail Drobizhev
Publication year - 2017
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0170934
Subject(s) - calmodulin , chromophore , conformational change , protonation , fluorescence , biophysics , chemistry , biochemistry , biology , photochemistry , enzyme , physics , ion , organic chemistry , quantum mechanics
The goal of this work is to determine how GCaMP6m’s fluorescence is altered in response to Ca 2+ -binding. Our detailed spectroscopic study reveals the simplest explanation for how GCaMP6m changes fluorescence in response to Ca 2+ is with a four-state model, in which a Ca 2+ -dependent change of the chromophore protonation state, due to a shift in pK a , is the predominant factor. The pK a 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 Ca 2+ and interacts with the M13 peptide. The absolute pK a values for the Ca 2+ -free and Ca 2+ -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.