Virulence-associated type III secretion systems serve the injection of bacterial effector proteins into eukaryotic host cells. These effector proteins modulate host cell biology in order to promote colonization and infection, hence type III secretion systems are often essential bacterial pathogenicity factors. The core of type III secretion systems is a cell envelope-spanning macromolecular machine called injectisome. It consists of almost twenty different components in a stoichiometry of one to more than one hundred. Assembly of this 6 MDa complex requires the coordinated integration of components from the cytoplasm, the inner membrane, the periplasm, the outer membrane and even the extracellular space of Gram-negative bacteria. Here, we review injectisome assembly with an emphasis on the techniques that were employed towards its investigation. In particular, we focus on in vivo photocrosslinking, a technique that exploits the encoding of the artificial UV-inducible crosslinking amino acid p-benzoyl-phenylalanine to identify protein-protein interactions and to delineate assembly pathways.

与毒性相关的III型分泌系统可将细菌效应蛋白注射到真核宿主细胞中。这些效应蛋白调节宿主细胞生物学以促进定植和感染，因此III型分泌系统通常是必不可少的细菌致病因素。III型分泌系统的核心是跨细胞包膜的大分子机器，称为注射体。它由化学计量比在一百到一百的几乎二十种不同成分组成。这6个MDa复合物的组装需要细胞质，内膜，周质，外膜乃至革兰氏阴性细菌的细胞外空间的组分的协调整合。在这里，我们将回顾注射体组装，重点是用于其研究的技术。体内光交联，一种利用人工UV诱导型交联氨基酸对苯甲酰苯丙氨酸的编码技术来识别蛋白质与蛋白质的相互作用并描绘装配途径。