Structure of an active FlhA ring reveals allosteric coupling between substrate specificity and export engine activation

The bacterial flagellar type III secretion system transports structural subunits in a defined order to assemble the flagellum. FlhA is the core component of this nanomachine; its transmembrane domain functions as an ion-driven export engine and its cytoplasmic domain connected by a flexible linker forms a nonameric ring that controls substrate selection. The coupling mechanism of these activities has remained unclear because the transmembrane domain has resisted structural analysis. Here we identify linker deletions that autonomously activate the export gate and capture a functionally active, hook-type protein export state of the entire ring complex. Using cryo-electron microscopy, we determine the structure of the FlhA({Delta}331-347) ring at 2.73 [A] resolution. The structure shows linker-mediated docking of the cytoplasmic ring to the transmembrane ring to stabilize the entire ring while sterically excluding filament-type substrate binding. These findings reveal how the FlhA ring allosterically couples substrate specificity to ion-driven export during flagellar assembly.