The role of intrinsically disordered C-terminal region of FliK in substrate specificity switching of the bacterial flagellar type III export apparatus

Abstract

The bacterial flagellar export switching machinery consists of a ruler protein, FliK, and an export switch protein, FlhB and switches substrate specificity of the flagellar type III export apparatus upon completion of hook assembly. An interaction between the C-terminal domain of FliK (FliK_C ) and the C-terminal cytoplasmic domain of FlhB (FlhB_C ) is postulated to be responsible for this switch. FliK_C has a compactly folded domain termed FliK_T3S4 (residues 268-352) and an intrinsically disordered region composed of the last 53 residues, FliK_CT (residues 353-405). Residues 301-350 of FliK_T3S4 and the last five residues of FliK_CT are critical for the switching function of FliK. FliK_CT is postulated to regulate the interaction of FliK_T3S4 with FlhB_C , but it remains unknown how. Here we report the role of FliK_CT in the export switching mechanism. Systematic deletion analyses of FliK_CT revealed that residues of 351-370 are responsible for efficient switching of substrate specificity of the export apparatus. Suppressor mutant analyses showed that FliK_CT coordinates FliK_T3S4 action with the switching. Site-directed photo-cross-linking experiments showed that Val-302 and Ile-304 in the hydrophobic core of FliK_T3S4 bind to FlhB_C . We propose that FliK_CT may induce conformational rearrangements of FliK_T3S4 to bind to FlhB_C .