Laboratory of Intracellular Parasites, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA.
Cell Microbiol. 2011 Oct;13(10):1497-517. doi: 10.1111/j.1462-5822.2011.01635.x. Epub 2011 Jul 11.
Coiled-coil domains in eukaryotic and prokaryotic proteins contribute to diverse structural and regulatory functions. Here we have used in silico analysis to predict which proteins in the proteome of the enteric pathogen, Salmonella enterica serovar Typhimurium, harbour coiled-coil domains. We found that coiled-coil domains are especially prevalent in virulence-associated proteins, including type III effectors. Using SopB as a model coiled-coil domain type III effector, we have investigated the role of this motif in various aspects of effector function including chaperone binding, secretion and translocation, protein stability, localization and biological activity. Compared with wild-type SopB, SopB coiled-coil mutants were unstable, both inside bacteria and after translocation into host cells. In addition, the putative coiled-coil domain was required for the efficient membrane association of SopB in host cells. Since many other Salmonella effectors were predicted to contain coiled-coil domains, we also investigated the role of this motif in their intracellular targeting in mammalian cells. Mutation of the predicted coiled-coil domains in PipB2, SseJ and SopD2 also eliminated their membrane localization in mammalian cells. These findings suggest that coiled-coil domains represent a common membrane-targeting determinant for Salmonella type III effectors.
卷曲螺旋结构域在真核和原核蛋白质中发挥着多样化的结构和调节功能。在这里，我们通过计算机分析预测了肠道病原体鼠伤寒沙门氏菌的蛋白质组中哪些蛋白质含有卷曲螺旋结构域。我们发现卷曲螺旋结构域在与毒力相关的蛋白质中特别普遍，包括 III 型效应子。我们以 SopB 作为卷曲螺旋结构域 III 型效应子的模型，研究了该结构域在效应子功能的各个方面的作用，包括伴侣蛋白结合、分泌和易位、蛋白质稳定性、定位和生物活性。与野生型 SopB 相比，SopB 卷曲螺旋突变体在细菌内部和易位进入宿主细胞后都不稳定。此外，该卷曲螺旋结构域对于 SopB 在宿主细胞中的有效膜结合是必需的。由于预测许多其他沙门氏菌效应子含有卷曲螺旋结构域，我们还研究了该结构域在其在哺乳动物细胞中的细胞内靶向作用中的作用。在 PipB2、SseJ 和 SopD2 中突变预测的卷曲螺旋结构域也消除了它们在哺乳动物细胞中的膜定位。这些发现表明卷曲螺旋结构域是沙门氏菌 III 型效应子的一个共同的膜靶向决定因素。
