| 1. |
- Gregg, Brieana M., et al.
(author)
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Botulinum neurotoxin X lacks potency in mice and in human neurons
- 2024
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In: mBio. - 2161-2129 .- 2150-7511. ; 15:3
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Journal article (peer-reviewed)abstract
- Botulinum neurotoxins (BoNTs) are a class of toxins produced by Clostridium botulinum (C. botulinum) and other species of Clostridia. BoNT/X is a putative novel botulinum neurotoxin identified through genome sequencing and capable of SNARE cleavage, but its neurotoxic potential in humans and vertebrates remained unclear. The C. botulinum strain producing BoNT/X, Strain 111, encodes both a plasmid-borne bont/b2 as well as the chromosomal putative bont/x. This study utilized C. botulinum Strain 111 from Japan as well as recombinantly produced full-length BoNT/X to more fully analyze this putative pathogenic toxin. We confirmed production of full-length, catalytically active native BoNT/X by C. botulinum Strain 111, produced as a disulfide-bonded dichain polypeptide similar to other BoNTs. Both the purified native and the recombinant BoNT/X had high enzymatic activity in vitro but displayed very low potency in human-induced pluripotent stem cell-derived neuronal cells and in mice. Intraperitoneal injection of up to 50 µg of native BoNT/X in mice did not result in botulism; however, mild local paralysis was observed after injection of 2 μg into the gastrocnemius muscle. We further demonstrate that the lack of toxicity by BoNT/X is due to inefficient neuronal cell association and entry, which can be rescued by replacing the receptor binding domain of BoNT/X with that of BoNT/A. These data demonstrate that BoNT/X is not a potent vertebrate neurotoxin like the classical seven serotypes of BoNTs.
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| 2. |
- Peng, Lisheng, et al.
(author)
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Botulinum neurotoxin D-C uses synaptotagmin I and II as receptors, and human synaptotagmin II is not an effective receptor for type B, D-C and G toxins
- 2012
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In: Journal of Cell Science. - : The Company of Biologists. - 0021-9533 .- 1477-9137. ; 125:13, s. 3233-3242
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Journal article (peer-reviewed)abstract
- Botulinum neurotoxins (BoNTs) are classified into seven types (A-G), but multiple subtype and mosaic toxins exist. These subtype and mosaic toxins share a high sequence identity, and presumably the same receptors and substrates with their parental toxins. Here, we report that a mosaic toxin, type D-C (BoNT/D-C), uses different receptors from its parental toxin BoNT/C. BoNT/D-C, but not BoNT/C, binds directly to the luminal domains of synaptic vesicle proteins synaptotagmin (Syt) I and II, and requires expression of SytI/II to enter neurons. The SytII luminal fragment containing the toxin-binding site can block the entry of BoNT/D-C into neurons and reduce its toxicity in vivo in mice. We also found that gangliosides increase binding of BoNT/D-C to SytI/II and enhance the ability of the SytII luminal fragment to block BoNT/D-C entry into neurons. These data establish SytI/II, in conjunction with gangliosides, as the receptors for BoNT/D-C, and indicate that BoNT/D-C is functionally distinct from BoNT/C. We further found that BoNT/D-C recognizes the same binding site on SytI/II where BoNT/B and G also bind, but utilizes a receptor-binding interface that is distinct from BoNT/B and G. Finally, we also report that human and chimpanzee SytII has diminished binding and function as the receptor for BoNT/B, D-C and G owing to a single residue change from rodent SytII within the toxin binding site, potentially reducing the potency of these BoNTs in humans and chimpanzees.
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| 3. |
- Zhang, Sicai, et al.
(author)
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Structural basis for the unique ganglioside and cell membrane recognition mechanism of botulinum neurotoxin DC
- 2017
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In: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 8
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Journal article (peer-reviewed)abstract
- Botulinum neurotoxins (BoNTs), the most potent toxins known, are potential bioterrorism agents. It is well established that all seven serotypes of BoNTs (BoNT/A-G) require complex gangliosides as co-receptors. Here, we report that BoNT/DC, a presumed mosaic toxin between BoNT/D and BoNT/C1, binds and enters efficiently into neurons lacking complex gangliosides and shows no reduction in toxicity in mice deficient in complex gangliosides. The co-crystal structure of BoNT/DC with sialyl-Thomsen-Friedenreich antigen (Sialyl-T) suggests that BoNT/DC recognizes only the sialic acid, but not other moieties in gangliosides. Using liposome flotation assays, we demonstrate that an extended loop in BoNT/DC directly interacts with lipid membranes, and the co-occurring sialic acid binding and loop-membrane interactions mediate the recognition of gangliosides in membranes by BoNT/DC. These findings reveal a unique mechanism for cell membrane recognition and demonstrate that BoNT/DC can use a broad range of sialic acid-containing moieties as co-receptors.
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