# Bacterial Flagellar Filament: A Supramolecular Multifunctional Nanostructure
## 5 Protein Synthesis, Export and Filament Elongation
### Chaperones
#### 5.2.2 FliT--Chaperone of FliD  
For the correct formation of the filament, a filament-capping protein FliD should be exported first during the filament assembly to form a penta- or hexameric cap that promotes self-assembly of FliC. In order to do this, FliD requires the assistance of its chaperon FliT [145]. FliT act as key flagellar chaperone in the assembly and operation of the flagellum because it binds to several flagellar proteins in the cytoplasm, such as the export apparatus components FliI, FliJ, and FliA, beyond interaction with its cognate FliD. As an example of its versatility, FliT also functions as a negative transcriptional regulator of flagellar genes by inhibiting the formation of a DNA complex with the master regulator FlhDC. Lately, several crystal structures of FliT alone and in complex with FliD or FliI have become available [130; 145]. Although in the crystal structure of _Salmonella_ FliT was present as a tetramer, the observation that FliT was in the monomer-dimer equilibrium under physiological conditions suggests that the tetrameric form was an artefact of crystal packing. The structure revealed, however, that FliT adopts an antiparallel four \(\alpha\)-helix bundle and uses a hydrophobic surface formed by the first three helices to recognize its substrate proteins. In the absence of a substrate protein, FliT adopts an auto-inhibited structure conformation in which both the binding site for the partner proteins (the hydrophobic surface formed by helices \(\alpha\)1-\(\alpha\)3) and the binding site for FlhA (helix \(\alpha\)4) are occluded. This auto-inhibited structure may serve to protect the substate-binding surface from aggregation in the absence of substrates and to mask the binding site for FlhA, adopting an uninhibited conformation only when the substrate protein is to be targeted to the export gate. The auto-inhibition/activation and targeting mechanism reported for FliT appears to be shared among other flagellar chaperones such as FliS, which also adopts an auto-inhibited structure, released upon FliC binding. The N-terminus of FliS serves as the FlhA binding site, and in these chaperones a highly conserved Tyr residue appears to be essential for efficient FlhA binding, suggesting that these chaperones may use similar strategies for substrate binding and activation of the complexes for binding to FlhA and thus for targeting to the export gate. A recent structure of the FlhA\(-\)FiliT\(-\)FiliD ternary complex revealed that there is no direct interaction between FliD and FlhA, as was observed with FlhA-FliS-FliC ternary complex (Figure 5) [140]. Although the structural architecture of FliT is similar to that of FliS, the arrangement of \(\alpha\)-helices is different. FliS forms a heterodimer with its cognate substrate FliC. The C-terminal region of FliC interacts with all three \(\alpha\)-helices of FliS in an extended conformation. Similarly, FliT forms a stable heterodimeric complex with FliD in solution through an interaction between the C-terminal half of FliT and the C-terminal region of FliD, similar to that of the FliS-FliC complex. However, only the highly conserved surface-exposed Lys79 in \(\alpha\)3, is critical for the interaction with FliD suggesting that the C-terminal region of FliD might interact with \(\alpha\)3 of FliT in a different manner from the FliS-FliC interaction. Database searches revealed that FliS is the most widely conserved and ancient flagellar chaperone and it has been proposed that other flagellar chaperones, such as FlgN and FliT, has evolved from FliS. Biochemical and structural analyses have provided insight into how the C-terminus of FliT regulates its interactions with the FlhDC complex, FliT ATPase, and Flij (subunits of the export apparatus that may function as a general chaperone), and the conformational change that is responsible for the switch between binding partners during flagellar protein export.