| 1. |
- Contreras, Estefania, et al.
(författare)
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A neurotoxin that specifically targets Anopheles mosquitoes
- 2019
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 10
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Tidskriftsartikel (refereegranskat)abstract
- Clostridial neurotoxins, including tetanus and botulinum neurotoxins, generally target vertebrates. We show here that this family of toxins has a much broader host spectrum, by identifying PMP1, a clostridial-like neurotoxin that selectively targets anopheline mosquitoes. Isolation of PMP1 from Paraclostridium bifermentans strains collected in anopheline endemic areas on two continents indicates it is widely distributed. The toxin likely evolved from an ancestral form that targets the nervous system of similar organisms, using a common mechanism that disrupts SNARE-mediated exocytosis. It cleaves the mosquito syntaxin and employs a unique receptor recognition strategy. Our research has an important impact on the study of the evolution of clostridial neurotoxins and provides the basis for the use of P. bifermentans strains and PMP1 as innovative, environmentally friendly approaches to reduce malaria through anopheline control.
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| 2. |
- Davies, Jonathan R., et al.
(författare)
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Structural and Biochemical Characterization of Botulinum Neurotoxin Subtype B2 Binding to Its Receptors
- 2020
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Ingår i: Toxins. - : MDPI AG. - 2072-6651 .- 2072-6651. ; 12:9
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Tidskriftsartikel (refereegranskat)abstract
- Botulinum neurotoxins (BoNTs) can be used therapeutically to treat a wide range of neuromuscular and neurological conditions. A collection of natural BoNT variants exists which can be classified into serologically distinct serotypes (BoNT/B), and further divided into subtypes (BoNT/B1, B2, …). BoNT subtypes share a high degree of sequence identity within the same serotype yet can display large variation in toxicity. One such example is BoNT/B2, which was isolated from Clostridium botulinum strain 111 in a clinical case of botulism, and presents a 10-fold lower toxicity than BoNT/B1. In an effort to understand the molecular mechanisms behind this difference in potency, we here present the crystal structures of BoNT/B2 in complex with the ganglioside receptor GD1a, and with the human synaptotagmin I protein receptor. We show, using receptor-binding assays, that BoNT/B2 has a slightly higher affinity for GD1a than BoNT/B1, and confirm its considerably weaker affinity for its protein receptors. Although the overall receptor-binding mechanism is conserved for both receptors, structural analysis suggests the lower affinity of BoNT/B2 is the result of key substitutions, where hydrophobic interactions important for synaptotagmin-binding are replaced by polar residues. This study provides a template to drive the development of future BoNT therapeutic molecules centered on assessing the natural subtype variations in receptor-binding that appears to be one of the principal stages driving toxicity.
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| 3. |
- Dong, Min, et al.
(författare)
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Botulinum and tetanus neurotoxins
- 2019
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Ingår i: Annual Review of Biochemistry. - : Annual Reviews. - 0066-4154 .- 1545-4509. ; 88, s. 811-837
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Forskningsöversikt (refereegranskat)abstract
- Botulinum neurotoxins (BoNTs) and tetanus neurotoxin (TeNT) are the most potent toxins known and cause botulism and tetanus, respectively. BoNTs are also widely utilized as therapeutic toxins. They contain three functional domains responsible for receptor-binding, membrane translocation, and proteolytic cleavage of host proteins required for synaptic vesicle exocytosis. These toxins also have distinct features: BoNTs exist within a progenitor toxin complex (PTC), which protects the toxin and facilitates its absorption in the gastrointestinal tract, whereas TeNT is uniquely transported retrogradely within motor neurons. Our increasing knowledge of these toxins has allowed the development of engineered toxins for medical uses. The discovery of new BoNTs and BoNT-like proteins provides additional tools to understand the evolution of the toxins and to engineer toxin-based therapeutics. This review summarizes the progress on our understanding of BoNTs and TeNT, focusing on the PTC, receptor recognition, new BoNT-like toxins, and therapeutic toxin engineering.
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| 4. |
- Elliott, Mark, et al.
(författare)
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Engineered botulinum neurotoxin B with improved binding to human receptors has enhanced efficacy in preclinical models
- 2019
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Ingår i: Science Advances. - : American Association for the Advancement of Science (AAAS). - 2375-2548. ; 5:1
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Tidskriftsartikel (refereegranskat)abstract
- Although botulinum neurotoxin serotype A (BoNT/A) products are common treatments for various disorders, there is only one commercial BoNT/B product, whose low potency, likely stemming from low affinity toward its human receptor synaptotagmin 2 (hSyt2), has limited its therapeutic usefulness. We express and characterize two full-length recombinant BoNT/B1 proteins containing designed mutations E1191M/S1199Y (rBoNT/B1(MY)) and E1191Q/S1199W (rBoNT/B1(QW)) that enhance binding to hSyt2. In preclinical models including human-induced pluripotent stem cell neurons and a humanized transgenic mouse, this increased hSyt2 affinity results in high potency, comparable to that of BoNT/A. Last, we solve the cocrystal structure of rBoNT/B1(MY) in complex with peptides of hSyt2 and its homolog hSyt1. We demonstrate that neuronal surface receptor binding limits the clinical efficacy of unmodified BoNT/B and that modified BoNT/B proteins have promising clinical potential.
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| 5. |
- Gustafsson, Robert, et al.
(författare)
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Crystal Structure of Botulinum Neurotoxin A2 in Complex with the Human Protein Receptor SV2C Reveals Plasticity in Receptor Binding
- 2018
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Ingår i: Toxins. - : MDPI AG. - 2072-6651 .- 2072-6651. ; 10:4
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Tidskriftsartikel (refereegranskat)abstract
- Botulinum neurotoxins (BoNTs) are a family of highly dangerous bacterial toxins, with seven major serotypes (BoNT/A-G). Members of BoNTs, BoNT/A1 and BoNT/B1, have been utilized to treat an increasing number of medical conditions. The clinical trials are ongoing for BoNT/A2, another subtype of BoNT/A, which showed promising therapeutic properties. Both BoNT/A1 and BoNT/A2 utilize three isoforms of synaptic vesicle protein SV2 (SV2A, B, and C) as their protein receptors. We here present a high resolution (2.0 angstrom) co-crystal structure of the BoNT/A2 receptor-binding domain in complex with the human SV2C luminal domain. The structure is similar to previously reported BoNT/A-SV2C complexes, but a shift of the receptor-binding segment in BoNT/A2 rotates SV2C in two dimensions giving insight into the dynamic behavior of the interaction. Small differences in key residues at the binding interface may influence the binding to different SV2 isoforms, which may contribute to the differences between BoNT/A1 and BoNT/A2 observed in the clinic.
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| 6. |
- Hamark, Christoffer, et al.
(författare)
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Glycans Confer Specificity to the Recognition of Ganglioside Receptors by Botulinum Neurotoxin A
- 2017
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Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 0002-7863 .- 1520-5126. ; 139:1, s. 218-230
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Tidskriftsartikel (refereegranskat)abstract
- The highly poisonous botulinum neurotoxins, produced by the bacterium Clostridium botulinum, act on their hosts by a high-affinity association to two receptors on neuronal cell surfaces as the first step of invasion. The glycan motifs of gangliosides serve as initial coreceptors for these protein complexes, whereby a membrane protein receptor is bound. Herein we set out to characterize the carbohydrate minimal binding epitope of the botulinum neurotoxin serotype A. By means of ligand-based NMR spectroscopy, X-ray crystallography, computer simulations, and isothermal titration calorimetry, a screening of ganglioside analogues together with a detailed characterization of various carbohydrate ligand complexes with the toxin were accomplished. We show that the representation of the glycan epitope to the protein affects the details of binding. Notably, both branches of the oligosaccharide GD la can associate to botulinum neurotoxin serotype A when expressed as individual trisaccharides. It is, however, the terminal branch of GD1a as well as this trisaccharide motif alone, corresponding to the sialyl-Thomsen-Friedenreich antigen, that represents the active ligand epitope, and these compounds bind to the neurotoxin with a high degree of predisposition but with low affinities. This finding does not correlate with the oligosaccharide moieties having a strong contribution to the total affinity, which was expected to be the case. We here propose that the glycan part of the ganglioside receptors mainly provides abundance and specificity, whereas the interaction with the membrane itself and protein receptor brings about the strong total binding of the toxin to the neuronal membrane.
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| 7. |
- Kinsolving, Julia, et al.
(författare)
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Structural and functional insight into the interaction of Clostridioides difficile toxin B and FZD7
- 2024
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Ingår i: Cell Reports. - 2211-1247. ; 43:2
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Tidskriftsartikel (refereegranskat)abstract
- The G protein -coupled receptors of the Frizzled (FZD) family, in particular FZD1,2,7, are receptors that are exploited by Clostridioides difficile toxin B (TcdB), the major virulence factor responsible for pathogenesis associated with Clostridioides difficile infection. We employ a live -cell assay examining the affinity between full-length FZDs and TcdB. Moreover, we present cryoelectron microscopy structures of TcdB alone and in complex with full-length FZD7, which reveal that large structural rearrangements of the combined repetitive polypeptide domain are required for interaction with FZDs and other TcdB receptors, constituting a first step for receptor recognition. Furthermore, we show that bezlotoxumab, an FDA -approved monoclonal antibody to treat Clostridioides difficile infection, favors the apo-TcdB structure and thus disrupts binding with FZD7. The dynamic transition between the two conformations of TcdB also governs the stability of the pore -forming region. Thus, our work provides structural and functional insight into how conformational dynamics of TcdB determine receptor binding.
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| 8. |
- Kosenina, Sara, 1993-, et al.
(författare)
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Crystal structure of the catalytic domain of BoNT/X in complex with its substrate, VAMP2
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Botulinum neurotoxins (BoNTs) are the deadliest toxins known to man but are at the same time being used for the treatment of an increasing number of medical conditions. BoNTs are zinc proteases that act in presynaptic cholinergic motoneurons by cleaving SNARE proteins, hence inhibiting neurotransmission. Structural information on the BoNT catalytic domain is crucial for a detailed understanding of their substrate specificity and binding mechanism.Here we report the 1.85 Å crystal structure of the catalytic light chain of BoNT/X in complex with its substrate, VAMP2. The structure sheds light on the sites important for VAMP2 binding and will help in the engineering of novel therapeutic toxins with altered and improved substrate specificity.
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| 9. |
- Košenina, Sara, et al.
(författare)
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Crystal structure of the catalytic domain of the Weissella oryzae botulinum-like toxin
- 2019
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Ingår i: FEBS Letters. - : Wiley. - 0014-5793 .- 1873-3468. ; 593:12, s. 1403-1410
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Tidskriftsartikel (refereegranskat)abstract
- Botulinum neurotoxins (BoNTs) are the most potent toxins known. So far, eight serotypes have been identified that all act as zinc-dependent endopeptidases targeting SNARE proteins and inhibiting the release of neurotransmitters. Recently, the first botulinum toxin-like protein was identified outside the Clostridial genus, designated BoNT/Wo in the genome of Weissella oryzae. Here, we report the 1.6 angstrom X-ray crystal structure of the light chain of BoNT/Wo (LC/Wo). LC/Wo presents the core fold common to BoNTs but has an unusually wide, open and negatively charged catalytic pocket, with an additional Ca2+ ion besides the zinc ion and a unique ss-hairpin motif. The structural information will help establish the substrate profile of BoNT/Wo and help our understanding of how BoNT evolved.
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| 10. |
- Kosenina, Sara, et al.
(författare)
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Structural Analysis of Botulinum Neurotoxins Type B and E by Cryo-EM
- 2022
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Ingår i: Toxins. - : MDPI AG. - 2072-6651 .- 2072-6651. ; 14:1
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Tidskriftsartikel (refereegranskat)abstract
- Botulinum neurotoxins (BoNTs) are the causative agents of a potentially lethal paralytic disease targeting cholinergic nerve terminals. Multiple BoNT serotypes exist, with types A, B and E being the main cause of human botulism. Their extreme toxicity has been exploited for cosmetic and therapeutic uses to treat a wide range of neuromuscular disorders. Although naturally occurring BoNT types share a common end effect, their activity varies significantly based on the neuronal cell-surface receptors and intracellular SNARE substrates they target. These properties are the result of structural variations that have traditionally been studied using biophysical methods such as X-ray crystallography. Here, we determined the first structures of botulinum neurotoxins using single-particle cryogenic electron microscopy. The maps obtained at 3.6 and 3.7 Å for BoNT/B and /E, respectively, highlight the subtle structural dynamism between domains, and of the binding domain in particular. This study demonstrates how the recent advances made in the field of single-particle electron microscopy can be applied to bacterial toxins of clinical relevance and the botulinum neurotoxin family in particular.
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