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Binding of calcium is sensed structurally and dynamically throughout the second calcium‐binding domain of the sodium/calcium exchanger
Author(s) -
Breukels Vincent,
Vuister Geerten W.
Publication year - 2010
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.22695
Subject(s) - calcium , chemistry , sodium calcium exchanger , biophysics , binding site , relaxation (psychology) , chemical shift , calcium binding protein , binding domain , calmodulin , in vivo , crystallography , biochemistry , biology , genetics , organic chemistry , neuroscience
We report the effects of Ca 2+ binding on the backbone relaxation rates and chemical shifts of the AD and BD splice variants of the second Ca 2+ ‐binding domain (CBD2) of the sodium–calcium exchanger. Analysis of the Ca 2+ ‐induced chemical shifts perturbations yields similar K D values of 16–24 μ M for the two CBD2‐AD Ca 2+ ‐binding sites, and significant effects are observed up to 20 Å away. To quantify the Ca 2+ ‐induced chemical shift changes, we performed a comparative analysis of eight Ca 2+ ‐binding proteins that revealed large differences between different protein folds. The CBD2 15 N relaxation data show the CBD2‐AD Ca 2+ coordinating loops to be more rigid in the Ca 2+ ‐bound state as well as to affect the FG‐loop located at the opposite site of the domain. The equivalent loops of the CBD2‐BD splice variant do not bind Ca 2+ and are much more dynamic relative to both the Ca 2+ ‐bound and apo forms of CBD2‐AD. A more structured FG‐loop in CBD2‐BD is suggested by increased S 2 order parameter values relative to both forms of CBD2‐AD. The chemical shift and relaxation data together indicate that, in spite of the small structural changes, the Ca 2+ ‐binding event is felt throughout the molecule. The data suggest that the FG‐loop plays an important role in connecting the Ca 2+ ‐binding event with the other cytosolic domains of the NCX, in line with in vivo and in vitro biochemical data as well as modeling results that connect the CBD2 FG‐loop with the first Ca 2+ ‐binding domain of NCX. Proteins 2010. © 2010 Wiley‐Liss, Inc.