Gangliosides interact with synaptotagmin to form the high-affinity receptor complex for botulinum neurotoxin B

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 116; no. 36; pp. 18098 - 18108
• Main Authors: Flores, Alessandra, Ramirez-Franco, Jorge, Desplantes, Richard, Debreux, Kévin, Ferracci, Géraldine, Wernert, Florian, Blanchard, Marie-Pierre, Maulet, Yves, Youssouf, Fahamoe, Sangiardi, Marion, Iborra, Cécile, Popoff, Michel Robert, Seagar, Michael, Fantini, Jacques, Lévêque, Christian, Far, Oussama El
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 03-09-2019
• Series: PNAS Plus
• Subjects: Affinity; Animals; Binding; Binding Sites; Biological Sciences; Botulinum toxin; Botulinum toxin type B; Botulinum Toxins, Type A - chemistry; Botulinum Toxins, Type A - metabolism; Cellular Biology; Ceramide; Circular dichroism; Dichroism; Domains; Fluorescence; Fluorescence spectroscopy; Gangliosides; Gangliosides - chemistry; Gangliosides - pharmacology; Life Sciences; Molecular modelling; Molecular structure; Monomolecular films; Mutation; Nerve endings; Neurotoxins; Oligosaccharides; Peptides; PNAS Plus; Point mutation; Potentiation; Protein Conformation, alpha-Helical; Protein Domains; Rats; Surface plasmon resonance; Synaptotagmin; Synaptotagmin I - chemistry; Synaptotagmin I - genetics; Synaptotagmin I - metabolism; Synaptotagmin II - chemistry; Synaptotagmin II - genetics; Synaptotagmin II - metabolism; Toxins; Transmembrane domains; Tryptophan; gangliosides; synaptotagmin; botulinum neurotoxin B receptor
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1908051116

Botulinum neurotoxin type B (BoNT/B) recognizes nerve terminals by binding to 2 receptor components: a polysialoganglioside, predominantly GT1b, and synaptotagmin 1/2. It is widely thought that BoNT/B initially binds to GT1b then diffuses in the plane of the membrane to interact with synaptotagmin. We have addressed the hypothesis that a GT1b–synaptotagmin cis complex forms the BoNT/B receptor. We identified a consensus glycosphingolipid-binding motif in the extracellular juxtamembrane domain of synaptotagmins 1/2 and confirmed by Langmuir monolayer, surface plasmon resonance, and circular dichroism that GT1b interacts with synaptotagmin peptides containing this sequence, inducing α-helical structure. Molecular modeling and tryptophan fluorescence spectroscopy were consistent with the intertwining of GT1b and synaptotagmin, involving cis interactions between the oligosaccharide and ceramide moieties of GT1b and the juxtamembrane and transmembrane domains of synaptotagmin, respectively. Furthermore, a point mutation on synaptotagmin, located outside of the BoNT/B-binding segment, inhibited GT1b binding and blocked GT1b-induced potentiation of BoNT/B binding to synaptotagmin-expressing cells. Our findings are consistent with amodel in which a preassembled GT1b–synaptotagmin complex constitutes the high-affinity BoNT/B receptor.