Supplemental Information: Cooperative Binding Mitigates the High-Dose Hook Effect

In order to explore how the high-dose Hook effect is dependent on the parameters used in our model, we repeated the simulations done for the figure entitled “Reduced hook effect in cooperative (wt) calmodulin” with a range of parameter values. First, we varied all dissociation constants by factors of 10. The results (see figure 1) show that a high-dose Hook effect is observed in all the cases. Since the peak of CaM-Ca4 concentrations is highest at low Kd values (with tight binding), the decrease at increasing calmodulin concentrations is more severe in absolute terms. Next, we examined the impact of the allosteric equilibrium constant, L. High L values mean that the ratio of calmodulin molecules in the absence of Calcium is more highly skewed towards the T state (rather than the R state). L = 1 means that the T state and R state are equally populated in the absence of ligand. As seen in figure 2, at lower L values the R state is favoured, resulting in a higher overall peak (due to the higher affinity of the R state), but also a strong high-dose hook effect (because cooperativity between the T and R state is what mitigates the high-dose hook effect.)