Departments of Life Sciences and the National Institute for Biotechnology in the Negev (NIBN), Ben-Gurion University of the Negev, Be'er Sheva, Israel.
PLoS One. 2013 Dec 2;8(12):e81557. doi: 10.1371/journal.pone.0081557. eCollection 2013.
Bordetella pertussis, the etiological agent of "whooping cough" disease, utilizes the type III secretion system (T3SS) to deliver a 69 kDa cytotoxic effector protein, BteA, directly into the host cells. As with other T3SS effectors, prior to its secretion BteA binds BtcA, a 13.9 kDa protein predicted to act as a T3SS class IA chaperone. While this interaction had been characterized for such effector-chaperone pairs in other pathogens, it has yet to be fully investigated in Bordetella. Here we provide the first biochemical proof that BtcA is indeed a class IA chaperone, responsible for the binding of BteA's N-terminal domain. We bring forth extensive evidence that BtcA binds its substrate effector through a dual-interface binding mechanism comprising of non-globular and bi-globular interactions at a moderate micromolar level binding affinity. We demonstrate that the non-globular interactions involve the first 31 N-terminal residues of BteA287 and their removal leads to destabilization of the effector-chaperone complex and lower binding affinities to BtcA. These findings represent an important first step towards a molecular understanding of BteA secretion and cell entry.
百日咳博德特氏菌是“百日咳”病的病原体，它利用 III 型分泌系统（T3SS）将一种 69 kDa 的细胞毒性效应蛋白 BteA 直接输送到宿主细胞中。与其他 T3SS 效应蛋白一样，BteA 在分泌之前与 BtcA 结合，BtcA 是一种预测作为 T3SS 类 IA 伴侣蛋白的 13.9 kDa 蛋白。虽然这种相互作用已经在其他病原体中的效应子-伴侣对中得到了描述，但在博德特氏菌中尚未得到充分研究。在这里，我们提供了第一个生化证据，证明 BtcA 确实是一种类 IA 伴侣蛋白，负责结合 BteA 的 N 端结构域。我们提出了广泛的证据，证明 BtcA 通过双接口结合机制与其底物效应子结合，该机制在中等微摩尔水平的结合亲和力下包含非球形和双球形相互作用。我们证明，非球形相互作用涉及 BteA287 的前 31 个 N 端残基，其去除导致效应子-伴侣复合物的不稳定性和与 BtcA 的结合亲和力降低。这些发现代表了对 BteA 分泌和细胞进入的分子理解的重要的第一步。
