Abstract
We investigated the impact of bound calmodulin (CaM)-target compound structure on the affinity of calcium (Ca(2+)) by integrating coarse-grained models and all-atomistic simulations with nonequilibrium physics. We focused on binding between CaM and two specific targets, Ca(2+)/CaM-dependent protein kinase II (CaMKII) and neurogranin (Ng), as they both regulate CaM-dependent Ca(2+) signaling pathways in neurons. It was shown experimentally that Ca(2+)/CaM (holoCaM) binds to the CaMKII peptide with overwhelmingly higher affinity than Ca(2+)-free CaM (apoCaM); the binding of CaMKII peptide to CaM in return increases the Ca(2+) affinity for CaM. However, this reciprocal relation was not observed in the Ng peptide (Ng(13-49)), which binds to apoCaM or holoCaM with binding affinities of the same order of magnitude. Unlike the holoCaM-CaMKII peptide, whose structure can be determined by crystallography, the structural description of the apoCaM-Ng(13-49) is unknown due to low binding affinity, therefore we computationally generated an ensemble of apoCaM-Ng(13-49) structures by matching the changes in the chemical shifts of CaM upon Ng(13-49) binding from nuclear magnetic resonance experiments. Next, we computed the changes in Ca(2+) affinity for CaM with and without binding targets in atomistic models using Jarzynski's equality. We discovered the molecular underpinnings of lowered affinity of Ca(2+) for CaM in the presence of Ng(13-49) by showing that the N-terminal acidic region of Ng peptide pries open the β-sheet structure between the Ca(2+) binding loops particularly at C-domain of CaM, enabling Ca(2+) release. In contrast, CaMKII peptide increases Ca(2+) affinity for the C-domain of CaM by stabilizing the two Ca(2+) binding loops. We speculate that the distinctive structural difference in the bound complexes of apoCaM-Ng(13-49) and holoCaM-CaMKII delineates the importance of CaM's progressive mechanism of target binding on its Ca(2+) binding affinities.