title: Calmodulin-induced conformational and hydrodynamic changes in the catalytic domain of Bordetella pertussis adenylate cyclase toxin.
abstract: Bordetella pertussis, the causative agent of whooping cough, secretes among various toxins an adenylate cyclase (CyaA) that displays a unique mechanism of cell invasion, which involves a direct translocation of its N-terminal catalytic domain (AC, 400 residues) across the plasma membrane of the eukaryotic targeted cells. Once into the cytosol, AC is activated by endogenous calmodulin and produces toxic amounts of cAMP. The structure of AC in complex with the C-terminal part of calmodulin has recently been determined. However, as the structure of the catalytic domain in the absence of calmodulin is still lacking, the molecular basis of AC activation by calmodulin remains largely unknown. To characterize this activation mechanism, we investigated here the biophysical properties of the isolated catalytic domain in solution with or without calmodulin. We found that calmodulin triggered only minor modifications of the protein secondary and tertiary structure but had a pronounced effect on the hydrodynamic properties of AC. Indeed, while the isolated catalytic domain was spherical and hydrated, it underwent a significant elongation as well as compaction and dehydration upon calmodulin interaction. On the basis of these data, we propose a model for the structural transition between the calmodulin-free and calmodulin-bound AC.
authors.affiliation: Institut Pasteur, Unité de Biochimie des Interactions Macromoléculaires, CNRS URA 2185, Département de Biologie Structurale et Chimie, 25-28 rue du Dr. Roux, 75724 Paris Cedex 15, France.
authors.forename: Johanna C
authors.initials: JC
authors.lastname: Karst
authors.forename: Ana Cristina
authors.initials: AC
authors.lastname: Sotomayor Pérez
authors.forename: J Iñaki
authors.initials: JI
authors.lastname: Guijarro
authors.forename: Bertrand
authors.initials: B
authors.lastname: Raynal
authors.forename: Alexandre
authors.initials: A
authors.lastname: Chenal
authors.forename: Daniel
authors.initials: D
authors.lastname: Ladant
mesh_terms: D037361:Adenylate Cyclase Toxin / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001886:Bordetella pertussis / Q000201:enzymology* / Q000235:genetics; D002147:Calmodulin / Q000494:pharmacology*; D020134:Catalytic Domain; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D002942:Circular Dichroism; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D011487:Protein Conformation; D013053:Spectrophotometry
chemical_list: D037361:Adenylate Cyclase Toxin; D002147:Calmodulin