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Query: EC:3.4.24.69 (botulinum neurotoxin)
 1,901 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Tetanus and botulinum neurotoxins are produced by several Clostridia and cause the paralytic syndromes of tetanus and botulism by blocking neurotransmitter release at central and peripheral synapses, respectively. They consist of two disulfide-linked polypeptides: H (100 kDa) is responsible for neurospecific binding and cell penetration of L (50 kDa), a zinc-endopeptidase specific for three protein subunits of the neuroexocytosis apparatus. Tetanus neurotoxin and botulinum neurotoxin serotypes B, D, F and G cleave at single sites, which differ for each neurotoxin, VAMP/synaptobrevin, a membrane protein of the synaptic vesicles. Botulinum A and E neurotoxins cleave SNAP-25, a protein of the presynaptic membrane, at two different carboxyl-terminal peptide bonds. Serotype C cleaves specifically syntaxin, another protein of the nerve plasmalemma. The target specificity of these metallo-proteinases relies on a double recognition of their substrates based on interactions with the cleavage site and with a non-contiguous segment that contains a structural motif common to VAMP, SNAP-25 and syntaxin.
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PMID:The metallo-proteinase activity of tetanus and botulism neurotoxins. 758 Dec 98

We investigated the effect of poisoning rat brain synaptosomes with botulinum neurotoxin A on the NSF-mediated disassembly of a complex consisting of syntaxin, SNAP-25 and synaptobrevin (fusion complex). Botulinum neurotoxin A specifically removes 9 amino acids from the C-terminus of SNAP-25 and efficiently blocks KCl-evoked glutamate release from synaptosomes. We report that truncated SNAP-25 is incorporated into the fusion complex of poisoned synaptosomes. The presence of truncated SNAP-25 does not interfere with the NSF-induced disassembly of the fusion complex. Also, the release of truncated SNAP-25 from the fusion complex is similar to that of the native SNAP-25. Since neither the formation of the complex nor its disassembly seems to be affected by the SNAP-25 fragment, this fragment is likely to block exocytosis by disrupting events between disassembly of the synaptosomal fusion complex and membrane fusion itself.
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PMID:Poisoning by botulinum neurotoxin A does not inhibit formation or disassembly of the synaptosomal fusion complex. 762 35

Western blotting of the insulin-secreting beta-cell lines HIT-15 and RINm5F with anti-SNAP-25 (synaptosomal associated protein of 25 kDa), anti-synaptobrevin, and anti-syntaxin 1 antibodies revealed the presence of proteins with the same electrophoretic mobility as found in neural tissue. Permeabilization of both of these insulinoma cell lines to botulinum neurotoxin A by electroporation resulted, after 3 days of culture, in the loss of approximately 90% of SNAP-25 immunoreactivity. A similar permeabilization of these cells with botulinum neurotoxin B resulted in the cleavage of approximately 90% of the synaptobrevin-like immunoreactivities. Botulinum neurotoxin F also cleaved approximately 90% of the synaptobrevin-like immunoreactivity in RINm5F cells. The permeabilization of both insulinoma cells to neurotoxin A resulted in a > 90% inhibition of potassium-stimulated, calcium-dependent insulin release. By contrast, permeabilization of the insulinoma cell lines to neurotoxin B resulted in only a approximately 60% inhibition of potassium-stimulated insulin release in HIT-15 cells, and neither neurotoxin B nor F caused inhibition in RINm5F cells. Thus HIT-15 and RINm5F cells contain the components of the putative exocytotic docking complex described in cells derived from the neural crest. In HIT-15 cells both SNAP-25 and synaptobrevin appear to be involved in calcium-dependent insulin secretion, whereas in RINm5F cells SNAP-25 but not synaptobrevin is involved.
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PMID:The effect of botulinum neurotoxins on the release of insulin from the insulinoma cell lines HIT-15 and RINm5F. 762 39

Botulinum neurotoxin serotype C (BoNT/C) is a 150-kDa protein produced by Clostridium botulinum, which causes animal botulism. In contrast to the other botulinum neurotoxins that contain one atom of zinc, highly purified preparations of BoNT/C bind two atoms of zinc per toxin molecule. BoNT/C is a zinc-endopeptidase that cleaves syntaxin 1A at the Lys253-Ala254 and syntaxin 1B at the Lys252-Ala253 peptide bonds, only when they are inserted into a lipid bilayer. The other Lys-Ala bond present within the carboxyl-terminal region is not hydrolyzed. Syntaxin isoforms 2 and 3 are also cleaved by BoNT/C, while syntaxin 4 is resistant. These data suggest that BoNT/C recognizes a specific spatial organization of syntaxin, adopted upon membrane insertion, which brings a selected Lys-Ala peptide bond of its carboxyl-terminal region to the active site of this novel metalloproteinase.
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PMID:Botulinum neurotoxin type C cleaves a single Lys-Ala bond within the carboxyl-terminal region of syntaxins. 773 92

Rat brain synaptosomes were used to study the effect of several clostridial neurotoxins on the neurotransmitter release. In this system the blockade of transmitter release correlated with the proteolytic activity of the toxins. Blockade of glutamate release was linked to selective proteolysis of one of the following synaptic proteins: synaptobrevin (BoNT/D, BoNT/F); SNAP-25 (BoNT/A, BoNT/E), or HPC-1/syntaxin (BoNT/C1). All the toxins used had an inhibitory effect on synaptosomes with the exception of BoNT/F. BoNT/F cleaved synaptobrevin in permeabilized synaptosomes but failed to produce the same effect on intact synaptosomes.
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PMID:Inhibition of neurotransmitter release by clostridial neurotoxins correlates with specific proteolysis of synaptosomal proteins. 787 84

The anaerobic bacterium Clostridium botulinum produces several related neurotoxins that block exocytosis of synaptic vesicles in nerve terminals and that are responsible for the clinical manifestations of botulism. Recently, it was reported that botulinum neurotoxin type B as well as tetanus toxin act as zinc-dependent proteases that specifically cleave synaptobrevin, a membrane protein of synaptic vesicles (Link et al., Biochem. Biophys. Res. Commun., 189, 1017-1023; Schiavo et al., Nature, 359, 832-835). Here we report that inhibition of neurotransmitter release by botulinum neurotoxin type C1 was associated with the proteolysis of HPC-1 (= syntaxin), a membrane protein present in axonal and synaptic membranes. Breakdown of HPC-1/syntaxin was selective since no other protein degradation was detectable. In vitro studies showed that the breakdown was due to a direct interaction between HPC-1/syntaxin and the toxin light chain which acts as a metallo-endoprotease. Toxin-induced cleavage resulted in the generation of a soluble fragment of HPC-1/syntaxin that is 2-4 kDa smaller than the native protein. When HPC-1/syntaxin was translated in vitro, cleavage occurred only when translation was performed in the presence of microsomes, although a full-length product was obtained in the absence of membranes. However, susceptibility to toxin cleavage was restored when the product of membrane-free translation was subsequently incorporated into artificial proteoliposomes. In addition, a translated form of HPC-1/syntaxin, which lacked the putative transmembrane domain at the C-terminus, was soluble and resistant to toxin action. We conclude that HPC-1/syntaxin is involved in exocytotic membrane fusion.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Botulinum neurotoxin C1 blocks neurotransmitter release by means of cleaving HPC-1/syntaxin. 790 Oct 2

The membrane proteins SNAP-25, syntaxin, and synaptobrevin (vesicle-associated membrane protein) have recently been implicated as central elements of an exocytotic membrane fusion complex in neurons. Here we report that SNAP-25 binds directly to both syntaxin and synaptobrevin. The SNAP-25-binding domain of syntaxin lies between residues 199 and 243, within the region previously shown to mediate synaptobrevin binding (Calakos, N., Bennett, M. K., Peterson, K. E., and Scheller, R. H. (1994) Science 263, 1146-1149). The syntaxin-binding domain of SNAP-25 encompasses most of the amino-terminal half of SNAP-25, including its putative palmitoylation sites. Truncation of the carboxyl-terminal 9 residues of SNAP-25, which yields a fragment corresponding to that generated by botulinum neurotoxin A, diminishes the interaction of SNAP-25 with synaptobrevin, but not with syntaxin. Sequence analysis revealed that the regions that mediate the interaction between SNAP-25 and syntaxin contain heptad repeats characteristic of certain classes of alpha-helices. Similar repeats are also present at the carboxyl terminus of SNAP-25 and in synaptobrevin. These domains have a moderate to high probability of forming coiled coils. We conclude that SNAP-25 can interact with both syntaxin and synaptobrevin and that binding may be mediated by alpha-helical domains that form intermolecular coiled-coil structures.
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PMID:SNAP-25, a t-SNARE which binds to both syntaxin and synaptobrevin via domains that may form coiled coils. 796 55

The clostridial neurotoxins responsible for tetanus and botulism are eight different proteins, composed of two disulfide-linked polypeptide chains. They bind specifically to the presynaptic membrane via the heavy chain, while the light chain enters the cytosol of the neurons, where it displays a zinc-endopeptidase activity directed to proteins of the neuroexocytosis apparatus. Tetanus neurotoxin and botulinum neurotoxin serotypes B, D, F and G cleave specifically and at single different peptide bonds VAMP/synaptobrevin, a component of small synaptic vesicles. In contrast, the other neurotoxins catalyze the hydrolysis of proteins of the presynaptic membrane. Serotypes A and E of botulinum neurotoxin cleave SNAP-25, at different sites located within the carboxyl-terminus, while the specific target of serotype C is syntaxin.
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PMID:Clostridial neurotoxins as tools to investigate the molecular events of neurotransmitter release. 799 6

Tetanus toxin (TeTx) and the various forms of botulinal neurotoxins (BoNT/A to BoNT/G) potently inhibit neurotransmission by means of their L chains which selectively proteolyze synaptic proteins such as synaptobrevin (TeTx, BoNT/B, BoNT/F), SNAP-25 (BoNT/A), and syntaxin (BoNT/C1). Here we show that BoNT/D cleaves rat synaptobrevin 1 and 2 in toxified synaptosomes and in isolated vesicles. In contrast, synaptobrevin 1, as generated by in vitro translation, is only a poor substrate for BoNT/D, whereas this species is cleaved by BoNT/F with similar potency. Cleavage by BoNT/D occurs at the peptide bond Lys59-Leu60 which is adjacent to the BoNT/F cleavage site (Gln58-Lys59) and again differs from the site hydrolyzed by TeTx and BoNT/B (Gln76-Phe77). Cellubrevin, a recently discovered isoform expressed outside the nervous system, is efficiently cleaved by all three toxins examined. For further characterization of the substrate requirements of BoNT/D, we tested amino- and carboxyl-terminal deletion mutants of synaptobrevin 2 as well as synthetic peptides. Shorter peptides containing up to 15 amino acids on either side of the cleavage site were not cleaved, and a peptide extending from Arg47 to Thr116 was a poor substrate for all three toxins tested. However, cleavability was restored when the peptide is further extended at the NH2 terminus (Thr27-Thr116) demonstrating that NH2 terminally located sequences of synaptobrevin which are distal from the respective cleavage sites are required for proteolysis. To further examine the isoform specificity, several mutants of rat synaptobrevin 2 were generated in which individual amino acids were replaced with those found in rat synaptobrevin 1. We show that a Met46 to Ile46 substitution drastically diminishes cleavability by BoNT/D and that the presence of Val76 instead of Gln76 dictates the reduced cleavability of synaptobrevin isoforms by TeTx.
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PMID:Cleavage of members of the synaptobrevin/VAMP family by types D and F botulinal neurotoxins and tetanus toxin. 817 89

Clostridial neurotoxins, tetanus toxin (TeTx) and the seven related but serologically distinct botulinal neurotoxins (BoNT/A to BoNT/G), are potent inhibitors of synaptic vesicle exocytosis in nerve endings. Recently it was reported that the light chains of clostridial neurotoxins act as zinc-dependent metalloproteases which specifically cleave synaptic target proteins such as synaptobrevin/VAMPs, HPC-1/syntaxin (BoNT/C1), and SNAP-25 (BoNT/A). We show here that BoNT/E, like BoNT/A, cleaves SNAP-25, as generated by in vitro translation or by expression in Escherichia coli. BoNT/E cleaves the Arg180-Ile181 bond. This site is different from that of BoNT/A, which cleaves SNAP-25 between the amino acid residues Gln197 and Arg198. These findings further support the view that clostridial neurotoxins have evolved from an ancestral protease recognizing the exocytotic fusion machinery of synaptic vesicles whereby individual toxins target different members of the membrane fusion complex.
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PMID:Proteolysis of SNAP-25 by types E and A botulinal neurotoxins. 829 7


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