The Kinetics of Ca²⁺-Dependent Switching in a Calmodulin−IQ Domain Complex

Abstract

We have performed a kinetic analysis of Ca 2+ -dependent switching in the complex between calmodulin (CaM) and the IQ domain from neuromodulin, and have developed detailed kinetic models for this process. Our results indicate that the affinity of the C-ter Ca 2+ -binding sites in bound CaM is reduced due to a ~10-fold decrease in the Ca 2+ association rate, while the affinity of the Nter Ca 2+ -binding sites is increased due to a ~3-fold decrease in the Ca 2+ dissociation rate. Although the Ca 2+ -free and Ca 2+ -saturated forms of the CaM-IQ domain complex have identical affinities, CaM dissociates ~100 times faster in the presence of Ca 2+ . Furthermore, under these conditions CaM can be transferred to the CaM-binding domain from CaM kinase II via a ternary complex. These properties are consistent with the hypothesis that CaM bound to neuromodulin comprises a localized store that can be efficiently delivered to neuronal proteins in its Ca 2+ -bound form in response to a Ca 2+ signal.

Variations at the Semiconserved Glycine in the IQ Domain Consensus Sequence Have a Major Impact on Ca2+-Dependent Switching in Calmodulin−IQ Domain Complexes

“…As addressed in Results, due to energy coupling the N→C order does not play a significant role under equilibrium conditions and will not be discussed in detail here. However, as seen with the reference CaM–B IQ complex, this binding order may play a significant role under transient conditions, or in a ternary complex, where additional binding sites for the CaM lobes come into play (21). Investigations of the effects of amino acid variations at the semiconserved position under transient conditions are ongoing.…”

Calcium Input Frequency, Duration and Amplitude Differentially Modulate the Relative Activation of Calcineurin and CaMKII

“…Once calmodulin is in the R conformation, it can bind to target proteins, calcineurin and CaMKII in the model, and activate them. The transient dynamics of calcium association and dissociation with calmodulin was justified by stopped-flow fluorescence measurements [85] . This validation procedure was achieved before adding calcium pumps, buffer proteins, and other signaling molecules ( Figure 10 ).…”

“… Calcium ions released from calmodulin, normalized by the maximum capacity of release. Crosses, experimental data from Black [85] . Solid line, simulation result.…”

Designing proteins to combat disease: Cardiac troponin C as an example

“…Typically the binding of an EF-hand protein to its target protein enhances the overall Ca 2+ sensitivity of the system, often by orders of magnitude [15]. However, there are several examples where target interactions actually decrease the Ca 2+ sensitivity of the EF-hand protein, as can occur with CaM binding to IQ motif containing peptides [46]. Unlike traditional CaM binding motifs that require Ca 2+ -bound CaM to initiate an interaction, IQ motifs typically bind Ca 2+ -free CaM [47].…”