1] Research Institute of Molecular Pathology, Vienna, Austria. [2] Institute of Molecular Biotechnology, Austrian Academy of Sciences, Vienna, Austria. [3].
1] Research Institute of Molecular Pathology, Vienna, Austria. [2] Institute of Molecular Biotechnology, Austrian Academy of Sciences, Vienna, Austria. [3] Center for Structural Systems Biology, University Medical Center Hamburg-Eppendorf and Deutsches Elektronen-Synchrotron, Hamburg, Germany.
Nat Struct Mol Biol. 2014 Jan;21(1):82-7. doi: 10.1038/nsmb.2722. Epub 2013 Dec 8.
Type 3 secretion systems use 3.5-megadalton syringe-like, membrane-embedded 'injectisomes', each containing an ~800-Å-long needle complex to connect intracellular compartments of infectious bacteria and hosts. Here we identify requirements for substrate association with, transport through and exit from the injectisome of Salmonella enterica serovar Typhimurium. This guided the design of substrates that become trapped within the secretion path and enabled visualization of injectisomes in action in situ. We used cryo-EM to define the secretion path, providing a structural explanation as to why effector proteins must be unfolded during transport. Furthermore, trapping of a heterologous substrate in the needle prevents secretion of natural bacterial effectors. Together, the data reveal the path of protein secretion across multiple membranes and show that mechanisms rejecting unacceptable substrates can be undermined, and transport of bacterial effectors across an already assembled type 3 secretion system can be inhibited.
III 型分泌系统使用 3.5 兆道尔顿的注射器状、膜嵌入的“注入体”，每个注入体包含一个约 800Å 长的针复合物，用于连接感染细菌和宿主的细胞内隔室。在这里，我们确定了与沙门氏菌肠亚种 Typhimurium 注入体的底物结合、通过和退出的要求。这指导了设计成为分泌途径中捕获的底物，并能够在原位可视化注入体的作用。我们使用 cryo-EM 来定义分泌途径，为为什么效应蛋白在运输过程中必须展开提供了结构解释。此外，在针中捕获异源底物会阻止天然细菌效应蛋白的分泌。总之，这些数据揭示了跨多个膜的蛋白质分泌途径，并表明可以破坏拒绝不可接受底物的机制，并且可以抑制已经组装好的 III 型分泌系统中细菌效应蛋白的运输。
