Genetic analysis of the bacterial hook-capping protein FlgD responsible for hook assembly

FlgD of Salmonella enterica is a 232 aa protein that acts as the hook cap to promote assembly of FlgE into the hook structure. The N-terminal 86 residues (FlgD(N)) complement flgD mutants, albeit to a small degree. However, little is known about the role of the C-terminal region of FlgD (FlgD(C)). Here we isolated pseudorevertants from Salmonella flgE mutants. About half of the extragenic mutations lay within FlgD(C) and only one in FlgD(N). These suppressor mutations prevented mutant FlgE subunits from leaking out to some degree. Two weakly motile flgD mutants encoding C-terminally truncated variants, FlgD₁₋₁₉₅ and FlgD((1-138f-s+4aa)), secreted larger amounts of FlgE into the culture medium than wild-type cells. Their hooks were shorter, and their length distributions were broader, with significant tailing towards smaller values. These results suggest that FlgD(C) contributes to efficient hook polymerization. Therefore, we propose that FlgD(N) attaches to the distal end of the hook to promote hook polymerization and that FlgD(C) blocks the exit of newly exported FlgE monomers into the culture medium, allowing FlgE to have more time to assemble into the hook.