Biphasic Ca2+-Dependent Switching in a Calmodulin−IQ Domain Complex

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Abstract
The relationship between the free Ca2+ concentration and the apparent dissociation constant for the complex between calmodulin (CaM) and the neuromodulin IQ domain consists of two phases. In the first phase, Ca2+ bound to the C-ter EF hand pair in CaM increases the Kd for the complex from the Ca2+-free value of 2.3 ± 0.1 μM to a value of 14.4 ± 1.3 μM. In the second phase, Ca2+ bound to the N-ter EF hand pair reduces the Kd for the complex to a value of 2.5 ± 0.1 μM, reversing the effect of the first phase. Due to energy coupling effects associated with these phases, the mean dissociation constant for binding of Ca2+ to the C-ter EF hand pair is increased ∼3-fold, from 1.8 ± 0.1 to 5.1 ± 0.7 μM, and the mean dissociation constant for binding of Ca2+ to the N-ter EF hand pair is decreased by the same factor, from 11.2 ± 1.0 to 3.5 ± 0.6 μM. These characteristics produce a bell-shaped relationship between the apparent dissociation constant for the complex and the free Ca2+ concentration, with a distance of 5−6 μM between the midpoints of the rising and falling phases. Release of CaM from the neuromodulin IQ domain therefore appears to be promoted over a relatively narrow range of free Ca2+ concentrations. Our results demonstrate that CaM−IQ domain complexes can function as biphasic Ca2+ switches through opposing effects of Ca2+ bound sequentially to the two EF hand pairs in CaM.