A Critical Region in the FlaA Flagellin Facilitates Filament Formation of the Vibrio cholerae Flagellum

Abstract

<i>Vibrio cholerae</i> is a Gram-negative bacterium with a monotrichous flagellum that causes the human disease cholera. Flagellum-mediated motility is an integral part of the bacterial life cycle inside the host and in the aquatic environment. The <i>V. cholerae</i> flagellar filament is composed of five flagellin subunits (FlaA, FlaB, FlaC, FlaD, and FlaE); however, only FlaA is necessary and sufficient for filament synthesis. <i>flaA</i> is transcribed from a class III flagellar promoter, whereas the other four flagellins are transcribed from class IV promoters. However, expressing <i>flaA</i> from a class IV promoter still facilitated motility in a strain that was otherwise lacking all five flagellins (Δ<i>flaA-E</i>). Furthermore, FlaA from <i>V. parahaemolyticus</i> (FlaA^VP; 77% identity) supported motility of the <i>V. cholerae</i> Δ<i>flaA-E</i> strain, whereas FlaA from <i>V. vulnificus</i> (FlaA^VV; 75% identity) did not, indicating that FlaA amino acid sequence is responsible for its critical role in flagellar synthesis. Chimeric proteins composed of different domains of FlaA^VC and FlaD or FlaA^VV revealed that the N-terminal D₁ domain (D_1N) contains an important region required for FlaA function. Further analyses of chimeric FlaA^VC-FlaD proteins identified a lysine residue present at position 145 of the other flagellins but absent from FlaA^VC that can prevent monofilament formation. Moreover, the D_1N region of amino acids 87 to 153 of FlaA^VV inserted into FlaA^VC allows monofilament formation but not motility, apparently due to the lack of filament curvature. These results identify residues within the D_1N domain that allow FlaA^VC to fold into a functional filament structure and suggest that FlaA^VC assists correct folding of the other flagellins.<b>IMPORTANCE</b><i>V. cholerae</i> causes the severe diarrheal disease cholera. Its ability to swim is mediated by rotation of a polar flagellum, and this motility is integral to its ability to cause disease and persist in the environment. The current studies illuminate how one specific flagellin (FlaA) within a multiflagellin structure mediates formation of the flagellar filament, thus allowing <i>V. cholerae</i> to swim. This knowledge can lead to safer vaccines and potential therapeutics to inhibit cholera.