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 alpha-helical linker region and a transmembrane beta-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 alpha-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.

中文翻译：

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模型自动运输车的乘员域对其移位器域的属性的影响。

自转运蛋白是革兰氏阴性细菌分泌的蛋白质的超家族，包括许多毒力因子。它们是由N端信号肽，携带该蛋白活性的表面暴露的“乘客”结构域以及由α-螺旋接头区域和跨膜β-结构组成的C端“转运蛋白”结构域组成的模块化蛋白。桶。转运蛋白结构域对于跨外膜分泌乘客结构域起着至关重要的作用。但是，自动运输的机制仍然知之甚少。百日咳药百日咳博德特氏菌产生一种自转运丝氨酸蛋白酶SphB1，该酶参与细菌表面黏附素的成熟。SphB1还介导其自身前体的蛋白水解成熟。我们将SphB1用作模型自动转运蛋白，并对分离的易位蛋白结构域与相同结构域的生化特性进行了首次比较，该相同结构域之前是其天然乘客的C末端部分。通过使用交联和动态光散射，我们提供了证据，即乘客域促进SphB1的自缔合，尽管这些相互作用看起来很不稳定。电生理研究表明，自转运蛋白的乘客结构域似乎以低电导构型维持易位通道，最可能是通过稳定孔内的α-螺旋。乘客可能显着影响AT的理化性质可能与AT蛋白的体内成熟和稳定性有关。