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Spirochaete flagella hook proteins self-catalyse a lysinoalanine covalent crosslink for motility

Nature Microbiology volume 1, Article number: 16134 (2016) | Download Citation


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,​3,​4,​5,​6,​7,​8,​9,​10,​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,​13,​14,​15,​16,​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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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.

Author information


  1. Department of Biochemistry, Robert C. Byrd Health Sciences Center, West Virginia University, Morgantown, West Virginia 26506, USA

    • Michael R. Miller
  2. Department of Microbiology, Immunology, and Cell Biology, Robert C. Byrd Health Sciences Center, West Virginia University, Morgantown, West Virginia 26506, USA

    • Kelly A. Miller
    • , Milinda E. James
    • , Andrew Cockburn
    •  & Nyles W. Charon
  3. Department of Oral Biology, State University of New York, Buffalo, New York 14214, USA

    • Jiang Bian
    •  & Chunhao Li
  4. Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853, USA

    • Sheng Zhang
    • , Michael J. Lynch
    •  & Brian R. Crane
  5. Department of Pharmaceutical Sciences, Robert C. Byrd Health Sciences Center, West Virginia University, Morgantown, West Virginia 26506, USA

    • Patrick S. Callery
  6. National Institute for Occupational Safety and Health, 1095 Willowdale Road, Morgantown West Virginia 26505, USA

    • Justin M. Hettick
  7. Department of Pathology and Laboratory Medicine, University of Texas Health Sciences Center, Houston, Texas 77030, USA

    • Jun Liu


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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.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Nyles W. Charon.

Supplementary information

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  1. 1.

    Supplementary information

    Supplementary Figures 1–12, Supplementary Tables 1–6, Supplementary Video Legends 1–4


  1. 1.

    Supplementary Video 1

    Cells WT T. denticola in 1% methylcelluose.

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    Supplementary Video 2

    Cells of mutant δflgE in 1% methylcellulose.

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    Supplementary Video 3

    Cells of substitution mutant TdC178A in 1% methylcellulose.

  4. 4.

    Supplementary Video 4

    Cells of substitution mutant TdK169A in 1% methylcellulose.

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