title: Physiological calcium concentrations regulate calmodulin binding and catalysis of adenylyl cyclase exotoxins.
abstract: 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 Ca(2+) concentrations increase the affinity of CaM for EF and CyaA causing their activation, but higher Ca(2+) concentrations directly inhibit catalysis. Both events occur in a physiologically relevant range of Ca(2+) concentrations. Despite the similarity in Ca(2+) 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 Ca(2+) ions. CyaA can be fully activated by CaM mutants with one defective C-terminal Ca(2+)-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 Ca(2+)-CaM activation. However, CyaA is fully activated by CaM without the domain corresponding to the helical domain of EF.
authors.affiliation: Ben-May Institute for Cancer Research, The University of Chicago, Chicago, IL 60637, USA.
authors.forename: Yuequan
authors.initials: Y
authors.lastname: Shen
authors.forename: Young-Sam
authors.initials: YS
authors.lastname: Lee
authors.forename: Sandriyana
authors.initials: S
authors.lastname: Soelaiman
authors.forename: Pamela
authors.initials: P
authors.lastname: Bergson
authors.forename: Dan
authors.initials: D
authors.lastname: Lu
authors.forename: Alice
authors.initials: A
authors.lastname: Chen
authors.forename: Kathy
authors.initials: K
authors.lastname: Beckingham
authors.forename: Zenon
authors.initials: Z
authors.lastname: Grabarek
authors.forename: Milan
authors.initials: M
authors.lastname: Mrksich
authors.forename: Wei-Jen
authors.initials: WJ
authors.lastname: Tang
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism*; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002384:Catalysis; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004926:Escherichia coli / Q000378:metabolism; D005098:Exotoxins / Q000378:metabolism*; D008274:Magnesium / Q000494:pharmacology; D008958:Models, Molecular; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000378:metabolism; D020349:Surface Plasmon Resonance; D014757:Viper Venoms / Q000378:metabolism
chemical_list: D002147:Calmodulin; D005098:Exotoxins; D011994:Recombinant Proteins; D014757:Viper Venoms; C416405:edema factor; D000262:Adenylyl Cyclases; D008274:Magnesium; D002118:Calcium