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Structural basis of cell surface receptor recognition by botulinum neurotoxin B


Botulinum neurotoxins (BoNTs) are potent bacterial toxins that cause paralysis at femtomolar concentrations1 by blocking neurotransmitter release. A ‘double receptor’ model has been proposed in which BoNTs recognize nerve terminals via interactions with both gangliosides and protein receptors that mediate their entry2. Of seven BoNTs (subtypes A–G), the putative receptors for BoNT/A3,4, BoNT/B5,6 and BoNT/G7 have been identified, but the molecular details that govern recognition remain undefined. Here we report the crystal structure of full-length BoNT/B in complex with the synaptotagmin II (Syt-II) recognition domain at 2.6 Å resolution. The structure of the complex reveals that Syt-II forms a short helix that binds to a hydrophobic groove within the binding domain of BoNT/B. In addition, mutagenesis of amino acid residues within this interface on Syt-II affects binding of BoNT/B. Structural and sequence analysis reveals that this hydrophobic groove is conserved in the BoNT/G and BoNT/B subtypes, but varies in other clostridial neurotoxins. Furthermore, molecular docking studies using the ganglioside GT1b indicate that its binding site is more extensive than previously proposed and might form contacts with both BoNT/B and synaptotagmin. The results provide structural insights into how BoNTs recognize protein receptors and reveal a promising target for blocking toxin–receptor recognition.

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Figure 1: The structure of BoNT/B–Syt-II recognition domain complex.
Figure 2: Interaction between the H CC domain of BoNT/B and the recognition domain of Syt-II.
Figure 3: Ganglioside-supported BoNT/B binding with synaptotagmin.
Figure 4: Model of BoNT/B binding to its receptors on a neuron.

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  1. Simpson, L. L. Identification of the characteristics that underlie botulinum toxin potency: implications for designing novel drugs. Biochimie 82, 943–953 (2000)

    Article  CAS  Google Scholar 

  2. Montecucco, C. How do tetanus and botulinum toxins bind to neuronal membranes?. Trends Biochem. Sci. 11, 315–317 (1986)

    Article  Google Scholar 

  3. Mahrhold, S., Rummel, A., Bigalke, H., Davletov, B. & Binz, T. The synaptic vesicle protein 2C mediates the uptake of botulinum neurotoxin A into phrenic nerves. FEBS Lett. 580, 2011–2014 (2006)

    Article  CAS  Google Scholar 

  4. Dong, M. et al. SV2 is the protein receptor for botulinum neurotoxin A. Science 312, 592–596 (2006)

    Article  CAS  ADS  Google Scholar 

  5. Nishiki, T. et al. The high-affinity binding of Clostridium botulinum type B neurotoxin to synaptotagmin II associated with gangliosides GT1b/GD1a. FEBS Lett. 378, 253–257 (1996)

    Article  CAS  Google Scholar 

  6. Dong, M. et al. Synaptotagmins I and II mediate entry of botulinum neurotoxin B into cells. J. Cell Biol. 162, 1293–1303 (2003)

    Article  CAS  Google Scholar 

  7. Rummel, A., Karnath, T., Henke, T., Bigalke, H. & Binz, T. Synaptotagmins I and II act as nerve cell receptors for botulinum neurotoxin G. J. Biol. Chem. 279, 30865–30870 (2004)

    Article  CAS  Google Scholar 

  8. Nishiki, T. et al. Binding of botulinum type B neurotoxin to Chinese hamster ovary cells transfected with rat synaptotagmin II cDNA. Neurosci. Lett. 208, 105–108 (1996)

    Article  CAS  Google Scholar 

  9. Swaminathan, S. & Eswaramoorthy, S. Structural analysis of the catalytic and binding sites of Clostridium botulinum neurotoxin B. Nature Struct. Biol. 7, 693–699 (2000)

    Article  CAS  Google Scholar 

  10. Breidenbach, M. A. & Brunger, A. T. Substrate recognition strategy for botulinum neurotoxin serotype A. Nature 432, 925–929 (2004)

    Article  CAS  ADS  Google Scholar 

  11. Simpson, L. L. Identification of the major steps in botulinum toxin action. Annu. Rev. Pharmacol. Toxicol. 44, 167–193 (2004)

    Article  CAS  Google Scholar 

  12. Jayaraman, S., Eswaramoorthy, S., Kumaran, D. & Swaminathan, S. Common binding site for disialyllactose and tri-peptide in C-fragment of tetanus neurotoxin. Proteins 61, 288–295 (2005)

    Article  CAS  Google Scholar 

  13. Lacy, D. B., Tepp, W., Cohen, A. C., DasGupta, B. R. & Stevens, R. C. Crystal structure of botulinum neurotoxin type A and implications for toxicity. Nature Struct. Biol. 5, 898–902 (1998)

    Article  CAS  Google Scholar 

  14. Schiavo, G., Matteoli, M. & Montecucco, C. Neurotoxins affecting neuroexocytosis. Physiol. Rev. 80, 717–766 (2000)

    Article  CAS  Google Scholar 

  15. Kitamura, M., Takamiya, K., Aizawa, S. & Furukawa, K. Gangliosides are the binding substances in neural cells for tetanus and botulinum toxins in mice. Biochim. Biophys. Acta 1441, 1–3 (1999)

    Article  CAS  Google Scholar 

  16. Kozaki, S., Kamata, Y., Watarai, S., Nishiki, T. & Mochida, S. Ganglioside GT1b as a complementary receptor component for Clostridium botulinum neurotoxins. Microbiol. Pathol. 25, 91–99 (1998)

    Article  CAS  Google Scholar 

  17. Rummel, A., Mahrhold, S., Bigalke, H. & Binz, T. The H-CC-domain of botulinum neurotoxins A and B exhibits a singular ganglioside binding site displaying serotype specific carbohydrate interaction. Mol. Microbiol. 51, 631–643 (2004)

    Article  CAS  Google Scholar 

  18. Ginalski, K., Venclovas, C., Lesyng, B. & Fidelis, K. Structure-based sequence alignment for the beta-trefoil subdomain of the clostridial neurotoxin family provides residue level information about the putative ganglioside binding site. FEBS Lett. 482, 119–124 (2000)

    Article  CAS  Google Scholar 

  19. Yowler, B. C., Kensinger, R. D. & Schengrund, C. L. Botulinum neurotoxin A activity is dependent upon the presence of specific gangliosides in neuroblastoma cells expressing synaptotagmin I. J. Biol. Chem. 277, 32815–32819 (2002)

    Article  CAS  Google Scholar 

  20. Rummel, A., Bade, S., Alves, J., Bigalke, H. & Binz, T. Two carbohydrate binding sites in the H-cc-domain of tetanus neurotoxin are required for toxicity. J. Mol. Biol. 326, 835–847 (2003)

    Article  CAS  Google Scholar 

  21. Fotinou, C. et al. The crystal structure of tetanus toxin Hc fragment complexed with a synthetic GT1b analogue suggests cross-linking between ganglioside receptors and the toxin. J. Biol. Chem. 276, 32274–32281 (2001)

    Article  CAS  Google Scholar 

  22. Weis, W. I. Cell-surface carbohydrate recognition by animal and viral lectins. Curr. Opin. Struct. Biol. 7, 624–630 (1997)

    Article  MathSciNet  CAS  Google Scholar 

  23. Eswaramoorthy, S., Kumaran, D., Keller, J. & Swaminathan, S. Role of metals in the biological activity of Clostridium botulinum neurotoxins. Biochemistry 43, 2209–2216 (2004)

    Article  CAS  Google Scholar 

  24. Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Meth. Enzymol. 276, 307–326 (1997)

    Article  CAS  Google Scholar 

  25. Vagin, A. & Teplyakov, A. MOLREP: an automated program for molecular replacement. J. Appl. Cryst. 30, 1022–1025 (1997)

    Article  CAS  Google Scholar 

  26. Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D 60, 2126–2132 (2004)

    Google Scholar 

  27. Collaborative Computational Project Number 4. The CCP4 suite: programs for protein crystallography. Acta Crystallogr. D 50, 760–763 (1994)

  28. Cheng, Y. et al. Crystallographic identification of Ca2+ and Sr2+ coordination sites in synaptotagmin I C2B domain. Protein Sci. 13, 2665–2672 (2004)

    Article  CAS  Google Scholar 

  29. Sutton, R. B., Ernst, J. A. & Brunger, A. T. Crystal structure of the cytosolic C2A–C2B domains of synaptotagmin III. Implications for Ca2+-independent SNARE complex interaction. J. Cell Biol. 147, 589–598 (1999)

    Article  CAS  Google Scholar 

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We thank E. Abola, K. Saikatendu and J. Ng for discussions and A. Walker for assistance with manuscript preparation. This work was supported by a grant from the Pacific Southwest Regional Center of Excellence (R.C.S. and E.A.J.), and grants from the NIH/NIAID (to E.R.C.). E.R.C. and E.A.J. acknowledge membership of and support from the Region V “Great Lakes” Regional Center of Excellence in Biodefense and Emerging Infectious Diseases. E.R.C. is an Investigator of the Howard Hughes Medical Institute. Portions of this research were carried out at the SSRL, a national user facility operated by Stanford University on behalf of the US Department of Energy, Office of Basic Energy Sciences. The SSRL Structural Molecular Biology Program is supported by the Department of Energy, Office of Biological and Environmental Research and by the National Institutes of Health, National Center for Research Resources, Biomedical Technology Program, and the National Institute of General Medical Sciences.

Author Contributions J.W.A., Q.C. (crystallography) and M.D. (molecular biology) contributed equally to this work and co-wrote the paper. All authors discussed the results and commented upon the manuscript.

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Correspondence to Edwin R. Chapman or Raymond C. Stevens.

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Atomic coordinates and experimental structure factors of the BoNT/B–Syt II ectodomain complex have been deposited in the RCSB Protein Data Bank (PDB) under accession code 2NP0. Reprints and permissions information is available at The authors declare no competing financial interests.

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This file contains Supplementary Methods, Supplementary Table 1 and 2 and Supplementary Figures 1-7 with legends.

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Chai, Q., Arndt, J., Dong, M. et al. Structural basis of cell surface receptor recognition by botulinum neurotoxin B . Nature 444, 1096–1100 (2006).

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