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Botulinum neurotoxin B recognizes its protein receptor with high affinity and specificity

Nature volume 444, pages 1092–1095 (2006)Cite this article

Abstract

Botulinum neurotoxins (BoNTs) are produced by Clostridium botulinum and cause the neuroparalytic syndrome of botulism. With a lethal dose of 1 ng kg-1, they pose a biological hazard to humans and a serious potential bioweapon threat1. BoNTs bind with high specificity at neuromuscular junctions and they impair exocytosis of synaptic vesicles containing acetylcholine through specific proteolysis of SNAREs (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptors), which constitute part of the synaptic vesicle fusion machinery2,3. The molecular details of the toxin–cell recognition have been elusive. Here we report the structure of a BoNT in complex with its protein receptor: the receptor-binding domain of botulinum neurotoxin serotype B (BoNT/B) bound to the luminal domain of synaptotagmin II, determined at 2.15 Å resolution. On binding, a helix is induced in the luminal domain which binds to a saddle-shaped crevice on a distal tip of BoNT/B. This crevice is adjacent to the non-overlapping ganglioside-binding site of BoNT/B. Synaptotagmin II interacts with BoNT/B with nanomolar affinity, at both neutral and acidic endosomal pH. Biochemical and neuronal ex vivo studies of structure-based mutations indicate high specificity and affinity of the interaction, and high selectivity of BoNT/B among synaptotagmin I and II isoforms. Synergistic binding of both synaptotagmin and ganglioside imposes geometric restrictions on the initiation of BoNT/B translocation after endocytosis. Our results provide the basis for the rational development of preventive vaccines or inhibitors against these neurotoxins.

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Figure 1: Structure of the H C B–Syt-II complex.
Figure 2: The H C B–Syt-II complex is stabilized by extensive intermolecular interactions involving two pronounced pockets on the H C B surface.
Figure 3: Site-directed mutagenesis analysis of the toxin-receptor binding site in BoNT/B and in the luminal domains of Syt-I and Syt-II.
Figure 4: Simultaneous binding with membrane-anchored Syt-II and ganglioside imposes geometric restrictions on how BoNT/B binds to the membrane surface.

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Acknowledgements

We thank P. Adams and J. Zhou for critical reading of the manuscript, and the staff of beamlines 9-1 at the Stanford Synchrotron Radiation Laboratory (SSRL) and beam line 8.2.2 at the Advanced Light Source (ALS) for help during data collection; H. Bigalke for providing the mouse phrenic nerve facility; and T. Henke and C. Knorr for technical assistance. The SSRL is 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. The ALS is supported by the Office of Energy Research (Office of Basic Energy Sciences, Material Sciences Division) of the US Department of Energy at Lawrence Berkeley National Laboratory. Support by the Department of Defense and Defense Threat Reduction Agency (to A.T.B.) and by a Deutsche Forschungsgemeinschaft grant (to T.B.) is acknowledged. The atomic coordinates and structure factors of the HCB–Syt-II complex are deposited in the Protein Data Bank under accession code 2NM1.

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Authors and Affiliations

  1. Howard Hughes Medical Institute,

    Rongsheng Jin & Axel T. Brunger

  2. Departments of Molecular and Cellular Physiology, Neurology and Neurological Science, Structural Biology, and Stanford Synchrotron Radiation Laboratory, Stanford University, California, 94305, Stanford, USA

    Rongsheng Jin & Axel T. Brunger

  3. Institut für Toxikologie,

    Andreas Rummel

  4. Institut für Biochemie, Medizinische Hochschule Hannover, Carl-Neuberg-Strasse 1, Hannover, 30625, Germany

    Thomas Binz

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  1. Rongsheng Jin
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Correspondence to Axel T. Brunger.

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The atomic coordinates and structure factors of the HCB–Syt-II complex are deposited in the Protein Data Bank under accession code 2NM1. Reprints and permissions information is available at www.nature.com/reprints. The authors declare no competing financial interests.

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Jin, R., Rummel, A., Binz, T. et al. Botulinum neurotoxin B recognizes its protein receptor with high affinity and specificity. Nature 444, 1092–1095 (2006). https://doi.org/10.1038/nature05387

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