Fig. 5: TLR5 targets a conserved region of bacterial flagellin within which we find little evidence of positive selection.

a Alignment of the domain of flagellin which TLR5 recognises in symbionts and pathogens. Red bars indicate residues predicted to be in the interface between flagellin and TLR5⁷⁴. Red residues have been identified as important for TLR5 binding by alanine scanning mutagenesis⁷⁴. As a member of the ε-proteobacteria, Helicobacter pylori has managed to escape TLR5 recognition and maintain motility by a serious of compensatory mutations⁷⁴. b Schematic of flagellin alignments for the 12 species tested. Numbers indicate the total number of sequences in the alignment (and the number of unique sequences). Red domains indicate the TLR5 binding region as shown in the above alignment, yellow domains are a second site that also interacts with TLR5 (a C-terminal region that also forms part of the D1 domain when the protein folds). Episodic positive selection was determined as any site with an LRT > 2 and p < 0.05 (calculated by MEME, and pervasive positive selection an ω > 1 and p < 0.05 calculated by FEL and are represented by ‘+’). Lines indicate pervasive negative selection at residues predicted by FEL to have a value of ω < 0.05. For C. freundii, E. cloacae and E. coli variable domains made aligning the full flagellin sequence inaccurate, therefore we focused only on the N-terminal D1 domain, which is the primary binding site for TLR5.