Abstract
Spirochaetes are bacteria responsible for several serious diseases, including Lyme disease (Borrelia burgdorferi), syphilis (Treponema pallidum) and leptospirosis (Leptospira interrogans), and contribute to periodontal diseases (Treponema denticola)1. These spirochaetes employ an unusual form of flagella-based motility necessary for pathogenicity; indeed, spirochaete flagella (periplasmic flagella) reside and rotate within the periplasmic space2–11. The universal joint or hook that links the rotary motor to the filament is composed of ∼120–130 FlgE proteins, which in spirochaetes form an unusually stable, high-molecular-weight complex9,12–17. In other bacteria, the hook can be readily dissociated by treatments such as heat18. In contrast, spirochaete hooks are resistant to these treatments, and several lines of evidence indicate that the high-molecular-weight complex is the consequence of covalent crosslinking12,13,17. Here, we show that T. denticola FlgE self-catalyses an interpeptide crosslinking reaction between conserved lysine and cysteine, resulting in the formation of an unusual lysinoalanine adduct that polymerizes the hook subunits. Lysinoalanine crosslinks are not needed for flagellar assembly, but they are required for cell motility and hence infection. The self-catalytic nature of FlgE crosslinking has important implications for protein engineering, and its sensitivity to chemical inhibitors provides a new avenue for the development of antimicrobials targeting spirochaetes.
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Acknowledgements
Funding was provided by the National Institutes of Health (R01-DE023431, to N.C., M.M. and C.L.), R01 GM064664 (to B.C.), R01-AI087946 (to J.L.) and R01-DE023080 and R01-AI078958 (to C.L). J.L. was also supported by grant AU-1714 from the Welch Foundation. The authors thank B. Bachert, M. Barbier, R. Duda, R. Hendrix, D. McNitt, S. Norris, R. Silversmith and R. Sircar for suggestions, technical assistance and support. The findings and conclusions in this article are those of the authors and do not necessarily represent the official position of the Centers for Disease Control and Prevention or the National Institute for Occupational Safety and Health.
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N.W.C. and M.R.M. designed the project. B.R.C., N.W.C., C.L, J.L. and M.R.M. wrote the manuscript. P.S.C., N.W.C., B.R.C., J.M.H, M.E.J., C.L., J.L., K.A.M. and M.R.M. designed the experiments. J.B., A.C., J.L., K.A.M., M.R.M., M.E.J., M.L. and S.Z. carried out experiments.
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Supplementary information
Supplementary Figures 1–12, Supplementary Tables 1–6, Supplementary Video Legends 1–4 (PDF 3951 kb)
Supplementary Video 1
Cells WT T. denticola in 1% methylcelluose. (MOV 5395 kb)
Supplementary Video 2
Cells of mutant δflgE in 1% methylcellulose. (MOV 12170 kb)
Supplementary Video 3
Cells of substitution mutant TdC178A in 1% methylcellulose. (MOV 5836 kb)
Supplementary Video 4
Cells of substitution mutant TdK169A in 1% methylcellulose. (MOV 7137 kb)
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Miller, M., Miller, K., Bian, J. et al. Spirochaete flagella hook proteins self-catalyse a lysinoalanine covalent crosslink for motility. Nat Microbiol 1, 16134 (2016). https://doi.org/10.1038/nmicrobiol.2016.134
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DOI: https://doi.org/10.1038/nmicrobiol.2016.134
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