3ogg

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Crystal structure of the receptor binding domain of botulinum neurotoxin D

Structural highlights

3ogg is a 1 chain structure with sequence from Clostridium botulinum. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance.
Method:X-ray diffraction, Resolution 1.651Å
Resources:FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT

Function

BXD_CBDP Botulinum toxin causes flaccid paralysis by inhibiting neurotransmitter (acetylcholine) release from the presynaptic membranes of nerve terminals of the eukaryotic host skeletal and autonomic nervous system, with frequent heart or respiratory failure (PubMed:8175689, PubMed:16252491). Precursor of botulinum neurotoxin D for which a proteinaceous coreceptor is controversial. In double SV2A/SV2B knockout mice this toxin does not degrade its synaptobrevin target; introducing SV2A, SV2B or SV2C restores target cleavage (PubMed:21483489). Recognition of SV2 by this toxin does not occur via SV2 glycosylation or its large extracellular loop 4 (PubMed:21483489). Another group does not find a convincing interaction with SV2 (PubMed:21632541). Thus a protein receptor for this BoNT serotype has yet to be definitively proven. Recognizes at least 1 complex polysialylated ganglioside found on neural tissue. Electrical stimulation increases uptake of toxin in an ex vivo assay, presumably by transiently exposing a receptor usually found in eukaryotic target synaptic vesicles (PubMed:19650874, PubMed:21483489, PubMed:21632541). Upon synaptic vesicle recycling the toxin is taken up via the endocytic pathway; when the pH of the toxin-containing endosome drops a structural rearrangement occurs so that the N-terminus of the heavy chain (HC) forms pores that allows the light chain (LC) to translocate into the cytosol (By similarity). Once in the cytosol the disulfide bond linking the 2 subunits is reduced and LC cleaves its target protein on synaptic vesicles, preventing their fusion with the cytoplasmic membrane and thus neurotransmitter release (By similarity). Requires complex eukaryotic host polysialogangliosides for full neurotoxicity and for binding to neurons (PubMed:20704566, PubMed:21483489).[UniProtKB:P0DPI0][1] [2] [3] [4] [5] [6] Has proteolytic activity (PubMed:8175689, PubMed:8197120). After translocation into the eukaryotic host cytosol, inhibits neurotransmitter release by acting as a zinc endopeptidase that cleaves the '61-Lys-|-Leu-62' bond of synaptobrevin-1 (VAMP1), and the equivalent 'Lys-|-Leu' sites in VAMP2 and VAMP3 (PubMed:8175689). Cleaves the '49-Lys-|-Ile-50' bond of A.californica synaptobrevin (AC P35589) (PubMed:8197120). This chain probably has to be partially unfolded to translocate into the eukaryotic host cell cytosol (PubMed:15584922).[7] [8] [9] Responsible for host epithelial cell transcytosis, host nerve cell targeting and translocation of light chain (LC) into eukaryotic host cell cytosol. Composed of 3 subdomains; the translocation domain (TD), and N-terminus and C-terminus of the receptor-binding domain (RBD). The RBD is responsible for the adherence of the toxin to the eukaryotic target cell surface. The N-terminus of the TD wraps an extended belt around the perimeter of the LC, protecting Zn(2+) in the active site; it may also prevent premature LC dissociation from the translocation channel and protect toxin prior to translocation (PubMed:17907800). The TD inserts into synaptic vesicle membrane to allow translocation into the host cytosol (By similarity). The RBD binds eukaryotic host phosphatidylethanolamine, which may serve as toxin receptor (PubMed:16115873). Treatment of synaptosomes with proteinase K does not reduce HC binding, suggesting there is no protein receptor or it is protected from extracellular proteases (PubMed:16115873). HC significantly decreases uptake and toxicity of whole BoNT/D (PubMed:19650874, PubMed:21483489). HC also interferes with uptake of tetanus toxin (PubMed:19650874). Has 2 closely located carbohydrate-binding receptor sites and binds at least 1 GT1b ganglioside (PubMed:20704566). Bind gangliosides in the order GD2 > GT1b > GD1b (PubMed:21632541). Interacts with eukaryotic target protein SV2B (synaptic vesicle glycoprotein 2B) (PubMed:21483489). Expression of SV2A, SV2B or SV2C in mice knocked-out for the SV2 proteins restores entry of BoNT/D and cleavage of VAMP2, suggesting SV2 acts as its receptor (PubMed:21483489). Unlike BoNT/A and BoNT/E, toxin uptake is not mediated by large extracellular loop 4 of SV2 (PubMed:21483489). Another group finds very poor interaction with SV2 proteins, suggesting the possible protein receptor may not have been identified (PubMed:21632541).[UniProtKB:P0DPI0][10] [11] [12] [13] [14] [15]

Evolutionary Conservation

Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf.

Publication Abstract from PubMed

Botulinum neurotoxins (BoNTs) are the most toxic proteins known. The mechanism for entry into neuronal cells for serotypes A, B, E, F, and G involves a well understood dual receptor (protein and ganglioside) process, however, the mechanism of entry for serotypes C and D remains unclear. To provide structural insights into how BoNT/D enters neuronal cells, the crystal structure of the receptor binding domain (S863-E1276) for this serotype (BoNT/D-HCR) was determined at 1.65A resolution. While BoNT/D-HCR adopts an overall fold similar to that observed in other known BoNT HCRs, several major structural differences are present. These structural differences are located at, or near, putative receptor binding sites and may be responsible for BoNT/D host preferences. Two loops, S1195-I1204 and K1236-N1244, located on both sides of the putative protein receptor binding pocket, are displaced >10A relative to the corresponding residues in the crystal structures of BoNT/B and G. Obvious clashes were observed in the putative protein receptor binding site when the BoNT/B protein receptor synaptotagmin II was modeled into the BoNT/D-HCR structure. Although a ganglioside binding site has never been unambiguously identified in BoNT/D-HCR, a shallow cavity in an analogous location to the other BoNT serotypes HCR domains is observed in BoNT/D-HCR that has features compatible with membrane binding. A portion of a loop near the putative receptor binding site, K1236-N1244, is hydrophobic and solvent-exposed and may directly bind membrane lipids. Liposome-binding experiments with BoNT/D-HCR demonstrate that this membrane lipid may be phosphatidylethanolamine.

Structural analysis of the receptor binding domain of botulinum neurotoxin serotype D.,Zhang Y, Buchko GW, Qin L, Robinson H, Varnum SM Biochem Biophys Res Commun. 2010 Oct 29;401(4):498-503. Epub 2010 Sep 19. PMID:20858456[16]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

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Citations
3 reviews cite this structure
Botulinum neurotoxins: mechanism of action.
Tighe et al. (2013)
2 more
6 other articles
No citations found

See Also

References

  1. ↑ Takeda M, Tsukamoto K, Kohda T, Matsui M, Mukamoto M, Kozaki S. Characterization of the neurotoxin produced by isolates associated with avian botulism. Avian Dis. 2005 Sep;49(3):376-81. PMID:16252491 doi:10.1637/7347-022305R1.1
  2. ↑ Rummel A, Häfner K, Mahrhold S, Darashchonak N, Holt M, Jahn R, Beermann S, Karnath T, Bigalke H, Binz T. Botulinum neurotoxins C, E and F bind gangliosides via a conserved binding site prior to stimulation-dependent uptake with botulinum neurotoxin F utilising the three isoforms of SV2 as second receptor. J Neurochem. 2009 Sep;110(6):1942-54. PMID:19650874 doi:10.1111/j.1471-4159.2009.06298.x
  3. ↑ Strotmeier J, Lee K, Volker AK, Mahrhold S, Zong Y, Zeiser J, Zhou J, Pich A, Bigalke H, Binz T, Rummel A, Jin R. Botulinum neurotoxin serotype D attacks neurons via two carbohydrate binding sites in a ganglioside dependent manner. Biochem J. 2010 Aug 12. PMID:20704566 doi:10.1042/BJ20101042
  4. ↑ Peng L, Tepp WH, Johnson EA, Dong M. Botulinum neurotoxin D uses synaptic vesicle protein SV2 and gangliosides as receptors. PLoS Pathog. 2011 Mar;7(3):e1002008. PMID:21483489 doi:10.1371/journal.ppat.1002008
  5. ↑ Kroken AR, Karalewitz AP, Fu Z, Kim JJ, Barbieri JT. Novel ganglioside-mediated entry of botulinum neurotoxin serotype D into neurons. J Biol Chem. 2011 Jun 1. PMID:21632541 doi:10.1074/jbc.M111.254086
  6. ↑ Yamasaki S, Baumeister A, Binz T, Blasi J, Link E, Cornille F, Roques B, Fykse EM, Südhof TC, Jahn R, et al.. Cleavage of members of the synaptobrevin/VAMP family by types D and F botulinal neurotoxins and tetanus toxin. J Biol Chem. 1994 Apr 29;269(17):12764-72 PMID:8175689
  7. ↑ Yamasaki S, Baumeister A, Binz T, Blasi J, Link E, Cornille F, Roques B, Fykse EM, Südhof TC, Jahn R, et al.. Cleavage of members of the synaptobrevin/VAMP family by types D and F botulinal neurotoxins and tetanus toxin. J Biol Chem. 1994 Apr 29;269(17):12764-72 PMID:8175689
  8. ↑ Yamasaki S, Hu Y, Binz T, Kalkuhl A, Kurazono H, Tamura T, Jahn R, Kandel E, Niemann H. Synaptobrevin/vesicle-associated membrane protein (VAMP) of Aplysia californica: structure and proteolysis by tetanus toxin and botulinal neurotoxins type D and F. Proc Natl Acad Sci U S A. 1994 May 24;91(11):4688-92. PMID:8197120 doi:10.1073/pnas.91.11.4688
  9. ↑ Bade S, Rummel A, Reisinger C, Karnath T, Ahnert-Hilger G, Bigalke H, Binz T. Botulinum neurotoxin type D enables cytosolic delivery of enzymatically active cargo proteins to neurones via unfolded translocation intermediates. J Neurochem. 2004 Dec;91(6):1461-72. PMID:15584922 doi:10.1111/j.1471-4159.2004.02844.x
  10. ↑ Tsukamoto K, Kohda T, Mukamoto M, Takeuchi K, Ihara H, Saito M, Kozaki S. Binding of Clostridium botulinum type C and D neurotoxins to ganglioside and phospholipid. Novel insights into the receptor for clostridial neurotoxins. J Biol Chem. 2005 Oct 21;280(42):35164-71. PMID:16115873 doi:10.1074/jbc.M507596200
  11. ↑ Rummel A, Häfner K, Mahrhold S, Darashchonak N, Holt M, Jahn R, Beermann S, Karnath T, Bigalke H, Binz T. Botulinum neurotoxins C, E and F bind gangliosides via a conserved binding site prior to stimulation-dependent uptake with botulinum neurotoxin F utilising the three isoforms of SV2 as second receptor. J Neurochem. 2009 Sep;110(6):1942-54. PMID:19650874 doi:10.1111/j.1471-4159.2009.06298.x
  12. ↑ Strotmeier J, Lee K, Volker AK, Mahrhold S, Zong Y, Zeiser J, Zhou J, Pich A, Bigalke H, Binz T, Rummel A, Jin R. Botulinum neurotoxin serotype D attacks neurons via two carbohydrate binding sites in a ganglioside dependent manner. Biochem J. 2010 Aug 12. PMID:20704566 doi:10.1042/BJ20101042
  13. ↑ Peng L, Tepp WH, Johnson EA, Dong M. Botulinum neurotoxin D uses synaptic vesicle protein SV2 and gangliosides as receptors. PLoS Pathog. 2011 Mar;7(3):e1002008. PMID:21483489 doi:10.1371/journal.ppat.1002008
  14. ↑ Kroken AR, Karalewitz AP, Fu Z, Kim JJ, Barbieri JT. Novel ganglioside-mediated entry of botulinum neurotoxin serotype D into neurons. J Biol Chem. 2011 Jun 1. PMID:21632541 doi:10.1074/jbc.M111.254086
  15. ↑ Brunger AT, Breidenbach MA, Jin R, Fischer A, Santos JS, Montal M. Botulinum neurotoxin heavy chain belt as an intramolecular chaperone for the light chain. PLoS Pathog. 2007 Sep 7;3(9):1191-4. PMID:17907800 doi:10.1371/journal.ppat.0030113
  16. ↑ Zhang Y, Buchko GW, Qin L, Robinson H, Varnum SM. Structural analysis of the receptor binding domain of botulinum neurotoxin serotype D. Biochem Biophys Res Commun. 2010 Oct 29;401(4):498-503. Epub 2010 Sep 19. PMID:20858456 doi:10.1016/j.bbrc.2010.09.063

Contents


PDB ID 3ogg

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