Identification of Conserved Domains in Salmonella muenchen Flagellin That Are Essential for Its Ability to Activate TLR5 and to Induce an Inflammatory Response in Vitro

Kanneganti G.K. Murthy, Amitabha Deb, Sunali Goonesekera, Csaba Szabó, Andrew L. Salzman

    Research output: Contribution to journalArticle

    91 Scopus citations

    Abstract

    The bacterial surface protein flagellin is widely distributed and well conserved among distant bacterial species. We and other investigators have reported recently that purified flagellin from Salmonella dublin or recombinant flagellin of Salmonella muenchen origin binds to the eukaryotic toll receptor TLR5 and activates the nuclear translocation of NF-κB and mitogen-activated protein kinase, resulting in the release of a host of pro-inflammatory mediators in vitro and in vivo. The amino acid sequence alignment of flagellins from various Gram-negative bacteria shows that the C and N termini are well conserved. It is possible that sequences within the N and C termini or both may regulate the pro-inflammatory activity of flagellin. Here we set out to map more precisely the regions in both termini that are required for TLR5 activation and pro-inflammatory signaling. Systematic deletion of amino acids from either terminus progressively reduced eukaryotic pro-inflammatory activation. However, deletion of amino acids 95-108 (motif N) in the N terminus and 441-449 (motif C) in the C terminus abolished pro-inflammatory activity completely. Site-directed mutagenesis analysis provided further evidence for the importance of motifs N and C. We also present evidence for the functional role of motifs N and C with the TLR5 receptor using a reporter assay system. Taken together, our results demonstrate that the pro-inflammatory activity of flagellin results from the interaction of motif N with the TLR5 receptor on the cell surface.

    Original languageEnglish (US)
    Pages (from-to)5667-5675
    Number of pages9
    JournalJournal of Biological Chemistry
    Volume279
    Issue number7
    DOIs
    StatePublished - Feb 13 2004

    ASJC Scopus subject areas

    • Biochemistry
    • Molecular Biology
    • Cell Biology

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