Hector Hernandez - Crystal Structures of the GCaMP Calcium Sensor Reveal the Mechanism of Fluorescence Signal Change and Aid Rational Design

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      Publication Details (including relevant citation   information):

      Akerboom, Jasper, Rivera, Jonathan D. Vélez, Guilbe, María M.   Rodríguez, Malavé, Elisa C. Alfaro, Hernandez, Hector H., Tian,   Lin, Hires, S. Andrew, Marvin, Jonathan S., Looger, Loren L.,   Schreiter, Eric R. Journal of Biological Chemistry  2009 284 (10 ) 6455 -6464

      Abstract: The genetically encoded calcium   indicator GCaMP2 shows promise for neural network activity   imaging, but is currently limited by low signal-to-noise ratio.   We describe x-ray crystal structures as well as solution   biophysical and spectroscopic characterization of GCaMP2 in the   calcium-free dark state, and in two calcium-bound bright states:   a monomeric form that dominates at intracellular concentrations   observed during imaging experiments and an unexpected   domain-swapped dimer with decreased fluorescence. This series of   structures provides insight into the mechanism of Ca2+-induced   fluorescence change. Upon calcium binding, the calmodulin (CaM)   domain wraps around the M13 peptide, creating a new domain   interface between CaM and the circularly permuted enhanced green   fluorescent protein domain. Residues from CaM alter the chemical   environment of the circularly permuted enhanced green fluorescent   protein chromophore and, together with flexible inter-domain   linkers, block solvent access to the chromophore. Guided by the   crystal structures, we engineered a series of GCaMP2 point   mutants to probe the mechanism of GCaMP2 function and   characterized one mutant with significantly improved   signal-to-noise. The mutation is located at a domain interface   and its effect on sensor function could not have been predicted   in the absence of structural data.

      Address (URL): http://www.jbc.org/content/284/10/6455.abstract