Modeling of Biomolecular Systems Interactions, Dynamics, and Allostery: Bridging Experiments and Computations - September 10-14, 2014, Istanbul, Turkey - page 118

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Modeling of Biomolecular Systems Interactions, Dynamics, and Allostery Poster Session II
57-POS Board 10
Activation of Bordetella Pertussis Adenyl Cyclase by Calmodulin
Therese Malliavin1, Edithe Selwa2,1, Marilyne Davi1, Alexandre Chenal1, Ana-Cristina
Sotomayor-Perez1, Elodie Laine2, Daniel Ladant1.
1CNRS/Institut Pasteur, Paris, France, 2Université Pierre et Marie Curie, Paris, France.
The adenyl cyclase (AC) is an essential toxin from Bordetella pertussis able to invade eukaryotic
cells where it is activated upon binding to calmodulin. Based on the crystal structure of the AC
catalytic domain in complex with the C-terminal half of calmodulin (C-CaM), we previously
carried out molecular dynamics simulations (Selwa et al, 2012) which suggested that three
residues, Arg338, Asn347 and Asp360, might be important for the stabilization of the AC/CaM
interaction. These residues belong to a loop-helix-loop motif, LHL, at the C-terminal end of AC,
located at the interface between CaM and the AC catalytic loop. In the present work, we have
characterized in silico and in vitro, three AC variants in which, one (Asn347; ACm1A), two
(Arg338 and Asp360; ACm2A), or three residues (Arg338, Asn 347 and Asp360; ACm3A) were
modified to Ala. Biochemical studies revealed that the affinity for CaM of ACm1A and ACm2A
was not significantly affected, while that of ACm3A was drastically reduced. To understand the
effect of the modifications, molecular dynamics simulations were performed on the modified
proteins. The MD trajectories recorded on the ACm3A/C-CaM complex revealed that the
calcium-binding loops of C-CaM display large fluctuations that could be related to the weakened
interaction of ACm3A with its activator. All together, our results suggest that the LHL motif at
the C-terminal end of AC is crucial to link CaM-binding to the stabilization of the AC catalytic
loop in an active configuration.
References:
Selwa et al. Model of the activation of Bordetella pertussis adenyl cyclase by calmodulin:
molecular dynamics and mutagenesis studies. In revision at J Biol Chem.
Selwa et al. Differential role of Calmodulin and Calcium ions in the stabilization of the catalytic
domain of adenyl cyclase CyaA from Bordetella pertussis. Proteins 2012;80:1028-40.