Stability, structural and functional properties of a monomeric, calcium-loaded adenylate cyclase toxin, CyaA, from Bordetella pertussis
Stability, structural and functional properties of a monomeric, calcium-loaded adenylate cyclase toxin, CyaA, from Bordetella pertussis
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DOI：
10.1038/srep42065
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发表时间：
2017-02-10
影响因子：
4.6
通讯作者：
Chenal, Alexandre
中科院分区：
文献类型：
--
作者：
Cannella, Sara E.;Enguene, Veronique Yvette Ntsogo;Chenal, Alexandre
百日咳博德特氏菌是百日咳的病原体，分泌腺苷酸环化酶毒素CyaA，其侵入真核细胞并通过cAMP过量产生改变其生理学。钙是CyaA的一个重要辅因子，因为它是大多数重复毒素（RTX）家族成员的情况。我们发现，钙结合的，单体形式的CyaA，hCyaAm，保存其透化和溶血活性，即使在一个完全无钙的环境。相比之下，hCyaAm需要溶液中的亚毫摩尔钙用于细胞侵袭，表明溶液中的游离钙参与CyaA毒素易位过程。我们进一步报告的第一个在解决方案中的hCyaAm的结构表征，从SAXS推断，质谱和流体动力学研究。我们表明，hCyaAm采用紧凑和稳定的状态，可以暂时保存其构象，即使在完全无钙的环境。因此，我们的研究结果表明，在hCyaAm中，C-末端RTX-结构域通过N-末端结构域稳定在高亲和力钙结合状态，而相反，钙结合到C-末端RTX-结构域强烈稳定N-末端区域。因此，hCyaAm的不同区域看起来紧密连接，导致结构域之间的稳定效应。CyaA响应钙的滞后行为可能与其他RTX溶细胞素相同。
Bordetella pertussis, the causative agent of whooping cough, secretes an adenylate cyclase toxin, CyaA, which invades eukaryotic cells and alters their physiology by cAMP overproduction. Calcium is an essential cofactor of CyaA, as it is the case for most members of the Repeat-in-ToXins (RTX) family. We show that the calcium-bound, monomeric form of CyaA, hCyaAm, conserves its permeabilization and haemolytic activities, even in a fully calcium-free environment. In contrast, hCyaAm requires submillimolar calcium in solution for cell invasion, indicating that free calcium in solution is involved in the CyaA toxin translocation process. We further report the first in solution structural characterization of hCyaAm, as deduced from SAXS, mass spectrometry and hydrodynamic studies. We show that hCyaAm adopts a compact and stable state that can transiently conserve its conformation even in a fully calcium-free environment. Our results therefore suggest that in hCyaAm, the C-terminal RTX-domain is stabilized in a high-affinity calcium-binding state by the N-terminal domains while, conversely, calcium binding to the C-terminal RTX-domain strongly stabilizes the N-terminal regions. Hence, the different regions of hCyaAm appear tightly connected, leading to stabilization effects between domains. The hysteretic behaviour of CyaA in response to calcium is likely shared by other RTX cytolysins.