Influence of the passenger domain of a model autotransporter on the properties of its translocator domain
Influence of the passenger domain of a model autotransporter on the properties of its translocator domain
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DOI：
10.1080/09687680701771925
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发表时间：
2008-01-01
影响因子：
--
通讯作者：
Jacob-Dubuisson, Francoise
中科院分区：
文献类型：
--
作者：
De, Emmanuelle;Saint, Nathalie;Jacob-Dubuisson, Francoise
自转运蛋白是革兰氏阴性菌分泌的蛋白质超家族，包括许多毒力因子。它们是模块化蛋白质，由 N 端信号肽、表面暴露的携带蛋白质活性的“过客”结构域以及由螺旋接头区域和跨膜桶组成的 C 端“易位器”结构域组成。易位蛋白结构域对于跨过外膜的过客结构域的分泌起着重要作用；然而，人们对自动运输的机制仍然知之甚少。百日咳杆菌产生一种自转运丝氨酸蛋白酶 SphB1，它参与细菌表面粘附素的成熟。 SphB1 还介导其自身前体的蛋白水解成熟。我们使用 SphB1 作为模型自转运蛋白，并对分离的易位蛋白结构域的生化和生物物理特性与其自然乘客的 C 端部分之前的相同结构域的生化和生物物理特性进行了首次比较。通过使用交联和动态光散射，我们提供了证据表明乘客域促进了 SphB1 的自动关联，尽管这些相互作用看起来相当不稳定。电生理学研究表明，自转运蛋白的乘客结构域似乎将易位通道维持在低电导构象，很可能是通过稳定孔内的β螺旋。过客可能显着影响 AT 理化性质，这可能与 AT 蛋白的体内成熟和稳定性有关。
Autotransporters are a superfamily of proteins secreted by Gram-negative bacteria including many virulence factors. They are modular proteins composed of an N-terminal signal peptide, a surface-exposed 'passenger' domain carrying the activity of the protein, and a C-terminal 'translocator' domain composed of an -helical linker region and a transmembrane -barrel. The translocator domain plays an essential role for the secretion of the passenger domain across the outer membrane; however, the mechanism of autotransport remains poorly understood. The whooping cough agent Bordetella pertussis produces an autotransporter serine-protease, SphB1, which is involved in the maturation of an adhesin at the bacterial surface. SphB1 also mediates the proteolytic maturation of its own precursor. We used SphB1 as a model autotransporter and performed the first comparisons of the biochemical and biophysical properties of an isolated translocator domain with those of the same domain preceded by the C-terminal moiety of its natural passenger. By using cross-linking and dynamic light scattering, we provide evidence that the passenger domain promotes the auto-association of SphB1, although these interactions appear rather labile. Electrophysiological studies revealed that the passenger domain of the autotransporter appears to maintain the translocator channel in a low-conductance conformation, most likely by stabilizing the -helix inside the pore. That the passenger may significantly influence AT physicochemical properties is likely to be relevant for the in vivo maturation and stability of AT proteins.