Secretion and cytotoxicity of the Bordetella pertussis effector BteA: Molecular insights from NMR

Adi Yahalom, Department of Chemistry, Bar Ilan University, Ramat Gan, Israel
Chen Guttman, Departments of Life Sciences and the National Institute for Biotechnology in the Negev (NIBN), Ben-Gurion University of the Negev, Be’er Sheva, Israel
Geula Davidov, Departments of Life Sciences and the National Institute for Biotechnology in the Negev (NIBN), Ben-Gurion University of the Negev, Be’er Sheva, Israel
Hadassa Shaked, Department of Chemistry, Bar Ilan University, Ramat Gan, Israel
Raz Zarivach, Departments of Life Sciences and the National Institute for Biotechnology in the Negev (NIBN), Ben-Gurion University of the Negev, Be’er Sheva, Israel
Jordan Chill, Department of Chemistry, Bar Ilan University, Ramat Gan, Israel

Bordetella pertussis, the etiological agent of “whooping cough” disease, utilizes the type III secretion system (T3SS) to deliver the cytotoxic effector BteA into host cells. As with other T3SS effectors, prior to its secretion BteA binds the class IA chaperone BtcA. The multifunctional BteA N-terminal domain acts as a docking platform for BtcA and targets lipid raft microdomains within the eukaryotic host cell. While this interaction in effector-chaperone pairs of other pathogens has been biochemically characterized, it has yet to be fully investigated in Bordetella; furthermore, a molecular understanding of BteA behavior in bacteria and host is conspicuously lacking.

Nuclear magnetic resonance (NMR) is a powerful structural method capable of visualizing biological events on the molecular level. Using a range of NMR experiments we have captured snapshots of a 131 amino-acid N-terminal domain of BteA as a free protein, in complex with its cognate chaperone, and in conditions mimicking membrane-targeting. These, in tandem with a recently solved crystal structure of this domain, provide unprecedented insight into BteA behavior on the atomic level. We bring evidence that BtcA binds its substrate effector through a dual-interface binding mechanism comprising of non-globular and globular interactions at moderate affinity, and that the chaperone triggers a partial unfolding of the effector which may be necessary for later injection. Furthermore, the BteA surface interacting with phosphatidylinositol(4,5)P₂ has been identified and compared to the chaperone interface. These findings represent an important step towards a molecular understanding of BteA secretion and cytotoxicity.