Pathogenic species from the Mycobacterium genus are responsible for a number of adverse health conditions in humans and animals that threaten health security and the economy worldwide. Mycobacteria have up to five specialized secretion systems (ESX-1 to ESX-5) that transport virulence factors across their complex cell envelope to facilitate manipulation of their environment. In pathogenic species, these virulence factors influence the immune system’s response and are responsible for membrane disruption and contributing to cell death. While structural details of these secretion systems have been recently described, gaps still remain in the structural understanding of the secretion mechanisms of most substrates. Here, we describe the crystal structure of Mycobacterium tuberculosis ESX-1 secretion-associated substrate EspB bound to its chaperone EspK. We found that EspB interacts with the C-terminal domain of EspK through its helical tip. Furthermore, cryogenic electron microscopy, size exclusion chromatography analysis, and small-angle X-ray scattering experiments show that EspK keeps EspB in its secretion-competent monomeric form and prevents its oligomerization. The structure presented in this study suggests an additional secretion mechanism in ESX-1, analogous to the chaperoning of proline-glutamate (PE)–proline-proline-glutamate (PPE) proteins by EspG, where EspK facilitates the secretion of EspB in Mycobacterium species.

来自分枝杆菌属的致病菌种导致人类和动物的许多不良健康状况，威胁着全球的健康安全和经济。分枝杆菌有多达五个专门的分泌系统（ESX-1 到 ESX-5），可将毒力因子转运穿过其复杂的细胞包膜，以促进对环境的操纵。在致病物种中，这些毒力因子会影响免疫系统的反应，并导致膜破裂和细胞死亡。虽然最近描述了这些分泌系统的结构细节，但对大多数底物分泌机制的结构理解仍然存在差距。在这里，我们描述了结核分枝杆菌的晶体结构ESX-1 分泌相关底物 EspB 与其伴侣 EspK 结合。我们发现 EspB 通过其螺旋尖端与 EspK 的 C 末端结构域相互作用。此外，低温电子显微镜、尺寸排阻色谱分析和小角度 X 射线散射实验表明，EspK 使 EspB 保持其具有分泌能力的单体形式，并防止其寡聚化。本研究中呈现的结构表明 ESX-1 中存在一种额外的分泌机制，类似于 EspG 对脯氨酸-谷氨酸 (PE)–脯氨酸-脯氨酸-谷氨酸 (PPE) 蛋白的伴侣作用，其中 EspK 促进分枝杆菌中EspB 的分泌.