Identification of the SV2-Protein Receptor Binding Site of Botulinum Neurotoxin Type E.

Biochemical Journal (Impact Factor: 4.4). 04/2013; 453(1). DOI: 10.1042/BJ20130391
Source: PubMed


The highly specific binding and uptake of botulinum neurotoxins (BoNT/A-G) into peripheral cholinergic motoneurons turns them into the most poisonous substances known. Interaction with gangliosides accumulates the neurotoxins on the plasma membrane and binding to a synaptic vesicle membrane protein leads to neurotoxin endocytosis. The synaptic vesicle (glyco-)protein 2 (SV2) mediates the uptake of BoNT/A and E, whereas synaptotagmin (Syt) is responsible for the endocytosis of BoNT/B and G. The Syt-binding site of the former was identified by co-crystallization and mutational analyses. Here we report the identification of the SV2‑binding interface of BoNT/E. Mutations interfering with SV2‑binding were located at a site that corresponds to the Syt‑binding site of BoNT/B and at an extended surface area located on the back side of the conserved ganglioside binding site, comprising the N- and C‑terminal half of the BoNT/E binding domain. Mutations impairing the affinity also reduced the neurotoxicity of full-length BoNT/E at mouse phrenic nerve hemidiaphragm preparations demonstrating the crucial role of the identified binding interface. Furthermore, we show that a monoclonal antibody neutralizes BoNT/E activity because it directly interferes with the BoNT/E-SV2 interaction. The data obtained suggest a novel mode of binding for BoNTs that exploit SV2 as cell surface receptor.

Full-text preview

Available from:
  • Source
    • "Similar to the serotype-specific BoNT binding to Syt discussed above, various BoNTs seem to develop different mechanisms recognizing three isoforms of SV2. For example, BoNT/A has the highest affinity to SV2C followed by SV2A and 2B (Dong et al., 2006); BoNT/D preferentially binds SV2B, and to a lesser extent, SV2A and 2C (Peng et al., 2011); BoNT/E exhibits higher binding affinity to SV2A over 2B but does not bind SV2C (Dong et al., 2008; Mahrhold et al., 2013; Peng et al., 2011). The binding-sites for BoNT/A and BoNT/E have been mapped to the large luminal domain 4 (LD4) of SV2, which is necessary and sufficient to mediate the entry of both toxins (Dong et al., 2008, 2006). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Botulinum neurotoxins (BoNTs) are among the most deadly toxins known. They act rapidly in a highly specific manner to block neurotransmitter release by cleaving the soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) complex at neuromuscular junctions. The extreme toxicity of BoNTs relies predominantly on their neurotropism that is accomplished by recognition of two host receptors, a polysialo-ganglioside and in the majority of cases a synaptic vesicle protein, through their receptor-binding domains. Two proteins, synaptotagmin and synaptic vesicles glycoprotein 2, have been identified as the receptors for various serotypes of BoNTs. Here, we review recent breakthroughs on the structural studies of BoNT-protein receptor recognitions that highlight a range of diverse mechanisms by which BoNTs manipulate host neuronal proteins for highly specific uptake at neuromuscular junctions. Copyright © 2015. Published by Elsevier Ltd.
    Progress in Biophysics and Molecular Biology 02/2015; 117(2-3). DOI:10.1016/j.pbiomolbio.2015.02.004 · 2.27 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The modular four domain structure of clostridial neurotoxins supports the idea to reassemble individual domains from tetanus and botulinum neurotoxins to generate novel molecules with altered pharmacological properties. To treat disorders of the central nervous system drug transporter molecules based on catalytically inactive clostridial neurotoxins circumventing the passage of the blood-brain-barrier are desired. Such molecules can be produced based on the highly effective botulinum neurotoxin serotype A incorporating the retrograde axonal sorting property of tetanus neurotoxin which is supposed to be encoded within its C-terminal cell binding domain HC. The corresponding exchange of the tetanus neurotoxin HC-fragment in botulinum neurotoxin A yielded the novel hybrid molecule AATT which displayed decreased potency at the neuromuscular junction like tetanus neurotoxin but exerted equal activity in cortical neurons compared to botulinum neurotoxin A wild-type. Minimizing the tetanus neurotoxin cell binding domain to its N- or C-terminal half drastically reduced the potencies of AATA and AAAT in cortical neurons indicating that the structural motif mediating sorting of tetanus neurotoxin is predominantly encoded within the entire HC-fragment. However, the reciprocal exchange resulted in TTAA which showed a similar potency as tetanus neurotoxin at the neuromuscular junction indicating that the tetanus neurotoxin portions prevents a high potency as observed for botulinum neurotoxins. In conclusion, clostridial neurotoxin based inactivated drug transporter for targeting central neurons should contain the cell binding domain of tetanus neurotoxin to exert its tropism for the central nervous system.
    Toxicon 06/2013; 75. DOI:10.1016/j.toxicon.2013.06.010 · 2.49 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Botulinum neurotoxins translocate their enzymatic domain across vesicular membranes. The molecular triggers of this process are unknown. Here, we tested the possibility that this is elicited by protonation of conserved surface carboxylates. Glutamate-48, glutamate-653 and aspartate-877 were identified as possible candidates and changed into amide. This triple mutant showed increased neurotoxicity due to faster cytosolic delivery of the enzymatic domain; membrane translocation could take place at less acidic pH. Thus, neutralisation of specific negative surface charges facilitates membrane contact permitting a faster initiation of the toxin membrane insertion.
    FEBS letters 10/2013; 587(23). DOI:10.1016/j.febslet.2013.10.010 · 3.17 Impact Factor
Show more

We use cookies to give you the best possible experience on ResearchGate. Read our cookies policy to learn more.