A triangular loop of domain D1 of FlgE is essential for hook assembly but not for the mechanical function

Highlights: • The hook protein FlgE assembles into a curved tubular structure with the help of the FlgD cap. • The flagellar hook acts as a universal joint to smoothly transmit torque produced by the flagellar motor to the filament. • FlgE consists of four domains, D0, Dc, D1 and D2. • Genetic analysis suggests that a loop of domain D1 (D1-loop) is responsible for the interaction with FlgD. • A deletion of the D1-loop or domain D2 makes the hook straight but does not affect the bending flexibility of the hook. The bacterial flagellar hook is a short, curved tubular structure made of FlgE. The hook connects the basal body as a rotary motor and the filament as a helical propeller and functions as a universal joint to smoothly transmit torque produced by the motor to the filament. Salmonella FlgE consists of D0, Dc, D1 and D2 domains. Axial interactions between a triangular loop of domain D1 (D1-loop) and domain D2 are postulated to be responsible for hook supercoiling. In contrast, Bacillus FlgE lacks the D1-loop and domain D2. Here, to clarify the roles of the D1-loop and domain D2 in the mechanical function, we carried out deletion analysis of Salmonella FlgE. A deletion of the D1-loop conferred a loss-of-function phenotype whereas that of domain D2 did not. The D1-loop deletion inhibited hook polymerization. Suppressor mutations of the D1-loop deletion was located within FlgD, which acts as the hook cap to promote hook assembly. This suggests a possible interaction between the D1-loop of FlgE and FlgD. Suppressor mutant cells produced straight hooks, but retained the ability to form a flagellar bundle behind a cell body, suggesting that the loop deletion does not affect the bending flexibility of the Salmonella hook.