Structural insights into the assembly mechanism of the flagellar MS-ring with three different symmetries

The flagellar basal body MS-ring, formed by 34 FliF subunits, is the core of the flagellar motor as well as the base for flagellar assembly. The MS-ring is also a housing for the flagellar protein export gate complex that is required for construction of the flagellum on the cell surface. A large periplasmic region of FliF contains three ring-building motifs named RBM1, RBM2, and RBM3. RBM3 forms the S-ring and β-collar with C34 symmetry. RBM2 forms the inner core ring of the M-ring with C23 symmetry surrounded by 11 cog-like structures formed by RBM1 and RBM2. However, it remains unknown how FliF assembles to generate these three different symmetries within the MS-ring. Here, we report the two cryoEM structures of the MS-ring formed by FliF co-expressed with FliG and transmembrane export gate proteins. Structural comparison of 33-mer and 34-mer MS-rings revealed that a subtle change in the conformation of RBM3 produces the different rotational symmetries. Combination of cryoEM structural and mutational analyses of the MS-ring with C33 symmetry provides evidence that the well-conserved DQxGxxL motif within a flexible loop connecting RBM2 and RBM3 allows RBM2 to take two different orientations relative to RBM3 to form not only 11 cog-like structures just outside the inner core ring with C22 symmetry but also an appropriately sized central pore in the inner core ring to accommodate the export gate complex.