Distinctive Recognition of Flagellin by Human and Mouse Toll-Like Receptor 5

Abstract

Toll-like receptor 5 (TLR5) is a receptor of the innate immune system that recognizes flagellin from certain bacterial species and triggers an inflammatory response. The Salmonella dublin flagellin in complex with zebrafish TLR5 has been crystallized previously. In the present study, we extrapolate the structure of this complex using structure-guided mutagenesis to determine the recognition modes of human and mouse TLR5 receptors and demonstrate species-specific differences in flagellin recognition. In general, the recognition mode of the mouse receptor can be said to be more robust in comparison to that of the human receptor. All-atom molecular dynamics simulation showed differences between the two receptors within the primary binding region. Using a functional motility assay, we show that although the highly conserved area of the flagellin analyzed in this study encompasses key structural requirements for flagella formation, a direct correlation between immune recognition and structure on the level of amino acid residues is not observed.

Extension and refinement of the recognition motif for Toll-like receptor 5 activation by flagellin

“…The distribution of charged and polar residues at the convex site of LRR14‐16 differs between hTLR5 and mTLR5; therefore, this segment of the receptor may play a role in the selectivity for different flagellins. Stimulation of TLR5 mutants with flagellins of SaTy and SeMa resulted in a differential response, thereby indicating the contribution of these selected residues to species specificity . These effects also suggest direct binding of flagellin to the residues of the proposed extended secondary binding site.…”

“…Stimulation of TLR5 mutants with flagellins of SaTy and SeMa resulted in a differential response, thereby indicating the contribution of these selected residues to species specificity. 31 These effects also suggest direct binding of flagellin to the residues of the proposed extended secondary binding site. In general, SeMa flagellin was, with some exceptions, less sensitive to mutations than SaTy flagellin, which exhibits more potent agonistic activity.…”

“…In general, SeMa flagellin was, with some exceptions, less sensitive to mutations than SaTy flagellin, which exhibits more potent agonistic activity. 31 Mutations of mouse-to-human counterpart residues rendered mTLR5 more responsive to SaTy flagellin as compared to SeMa, additionally demonstrating that flagellin could interact with the convex solvent-exposed residues of LRR13-16 of TLR5.…”

“…Flagellin is a protein agonist, and as such the most complex among the TLR agonists characterized thus far. Species‐specific differences in the receptor‐ligand recognition were demonstrated both for flagellin from different bacteria as well as for TLR5 from different species . Activation of TLR5 with flagellin from β‐ and γ‐proteobacteria such as Salmonella typhimurium (SaTy) or Serratia marcescens (SeMa) triggers the MyD88‐dependent signaling pathway.…”

Duplicated TLR5 of zebrafish functions as a heterodimeric receptor

“…Flagellin binding and thus insertion of R90 in the hotspot pocket alters the conformation of the LRR9 loop (19). Mutation of R90 preserves binding of flagellin to hTLR5 and mouse TLR5 (mTLR5) but reduces receptor activation (43,44), suggesting that insertion of R90 into the hotspot pocket initiates a conformational change that is necessary for receptor activation (44). Although the hotspot pockets of mTLR5 and hTLR5 differ only by three residues (Fig.…”