Physiological calcium concentrations regulate calmodulin binding and catalysis of adenylyl cyclase exotoxins

Edema factor (EF) and CyaA are calmodulin (CaM)-activated adenylyl cyclase exotoxins involved in the pathogenesis of anthrax and whooping cough, respectively. Using spectroscopic, enzyme kinetic and surface plasmon resonance spectroscopy analyses, we show that low Ca²⁺ concentrations increase the affinity of CaM for EF and CyaA causing their activation, but higher Ca²⁺ concentrations directly inhibit catalysis. Both events occur in a physiologically relevant range of Ca²⁺ concentrations. Despite the similarity in Ca²⁺ sensitivity, EF and CyaA have substantial differences in CaM binding and activation. CyaA has 100-fold higher affinity for CaM than EF. CaM has N- and C-terminal globular domains, each binding two Ca²⁺ ions. CyaA can be fully activated by CaM mutants with one defective C-terminal Ca²⁺-binding site or by either terminal domain of CaM while EF cannot. EF consists of a catalytic core and a helical domain, and both are required for CaM activation of EF. Mutations that decrease the interaction of the helical domain with the catalytic core create an enzyme with higher sensitivity to Ca²⁺-CaM activation. However, CyaA is fully activated by CaM without the domain corresponding to the helical domain of EF.