Showing papers by "Thomas Binz published in 1994"
TL;DR: The data suggest that synaptobrevin, syntaxin and SNAP‐25 associate into a unique stable complex that functions in synaptic vesicle exocytosis, suggesting that membrane fusion involves intermolecular interactions via coiled‐coil structures.
Abstract: Clostridial neurotoxins inhibit neurotransmitter release by selective and specific intracellular proteolysis of synaptobrevin/VAMP, synaptosomal-associated protein of 25 kDa (SNAP-25) or syntaxin. Here we show that in binary reactions synaptobrevin binds weakly to both SNAP-25 and syntaxin, and SNAP-25 binds to syntaxin. In the presence of all three components, a dramatic increase in the interaction strengths occurs and a stable sodium dodecyl sulfate-resistant complex forms. Mapping of the interacting sequences reveals that complex formation correlates with the presence of predicted alpha-helical structures, suggesting that membrane fusion involves intermolecular interactions via coiled-coil structures. Most toxins only attack the free, and not the complexed, proteins, and proteolysis of the proteins by different clostridial neurotoxins has distinct inhibitory effects on the formation of synaptobrevin-syntaxin-SNAP-25 complexes. Our data suggest that synaptobrevin, syntaxin and SNAP-25 associate into a unique stable complex that functions in synaptic vesicle exocytosis.
800 citations
TL;DR: BoNT/E, like BoNT/A, cleaves SNAP-25, as generated by in vitro translation or by expression in Escherichia coli, and 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.
432 citations
TL;DR: It is shown that BoNT/D cleaves rat synaptobrevin 1 and 2 in toxified synaptosomes and in isolated vesicles and that the presence of Val76 instead of Gln76 dictates the reduced cleavability of synaptOBrevin isoforms by TeTx.
277 citations
TL;DR: The results indicate that cellubrevin plays an important role in the constitutive exocytosis of vesicles which recycle plasmalemma receptors, and the incomplete inhibition of transferrin release produced by the toxin suggests the existence of a cellUBrevin-independent exocyTotic mechanism, which may involve tetanus toxin-insensitive proteins of the synaptobrevin/VAMP family.
Abstract: Cellubrevin is a member of the synaptobrevin/VAMP family of SNAREs, which has a broad tissue distribution. In fibroblastic cells it is concentrated in the vesicles which recycle transferrin receptors but its role in membrane trafficking and fusion remains to be demonstrated. Cellubrevin, like the synaptic vesicle proteins synaptobrevins I and II, can be cleaved by tetanus toxin, a metallo-endoprotease which blocks neurotransmitter release. However, nonneuronal cells are unaffected by the toxin due to lack of cell surface receptors for its heavy chain. To determine whether cellubrevin cleavage impairs exocytosis of recycling vesicles, we tested the effect of tetanus toxin light chain on the release of preinternalized transferrin from streptolysin-O-perforated CHO cells. The release was found to be temperature and ATP dependent as well as NEM sensitive. Addition of tetanus toxin light chain, but not of a proteolytically inactive form of the toxin, resulted in a partial inhibition of transferrin release which correlated with the toxin-mediated cleavage of cellubrevin. The residual release of transferrin occurring after complete cellubrevin degradation was still ATP dependent. Our results indicate that cellubrevin plays an important role in the constitutive exocytosis of vesicles which recycle plasmalemma receptors. The incomplete inhibition of transferrin release produced by the toxin suggests the existence of a cellubrevin-independent exocytotic mechanism, which may involve tetanus toxin-insensitive proteins of the synaptobrevin/VAMP family.
218 citations
TL;DR: It is shown that botulinum toxin type G cleaves rat synaptobrevin 2 between Ala81 and Ala82, a peptide bond that differs from those attacked by tetanus toxin and the botulinal toxins types B, D, and F.
120 citations
TL;DR: The soluble and membrane-anchored forms of synaptobrevin are cleaved by the light chains of the botulinal toxins type D and F and by tetanus toxin involving the peptide bonds Lys49-Ile50, Gln48-Lys49, and Gln66-Phe67, respectively.
Abstract: Synaptobrevin/vesicle-associated membrane protein (VAMP) and syntaxin are potential vesicle donor and target membrane receptors of a docking complex that requires N-ethylmaleimide-sensitive factor (NSF) and soluble NSF-attachment proteins as soluble factors for vesicle fusion with target membranes. Members of this docking complex are the target of clostridial neurotoxins that act as zinc-dependent proteases. Molecular cloning of the Aplysia californica synaptobrevin cDNA revealed a 180-residue polypeptide (M(r), 19,745) with a central transmembrane region and an atypically large C-terminal intravesicular domain. This polypeptide integrates into membranes at both the co- and posttranslational level, as shown by modification of an artificially introduced N-glycosylation site. The soluble and membrane-anchored forms of synaptobrevin are cleaved by the light chains of the botulinal toxins type D and F and by tetanus toxin involving the peptide bonds Lys49-Ile50, Gln48-Lys49, and Gln66-Phe67, respectively. The active center of teh tetanus toxin light chain was identified by site-specific mutagenesis. His233, His237, Glu234, and Glu270/271 are essential to this proteolytic activity. Modification of histidine residues resulted in loss of zinc binding, whereas a replacement of Glu234 only slightly reduced the zinc content.
98 citations
TL;DR: The relatedness of Borna disease virus in various naturally infected animal species was determined by reverse transcriptase/PCR amplification and subsequent sequence determination of the p24 gene, indicative of a common ancestral virus pool and a remarkably low species barrier of BDV.
58 citations
TL;DR: Rat brain synaptosomes were used to study the effect of several clostridial neurotoxins on the neurotransmitter release and the blockade of transmitter release correlated with the proteolytic activity of the toxins.
Abstract: 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.
44 citations
TL;DR: The results indicate that zinc bound to the zinc binding motif constitutes the active site of the tetanus toxin light chain and suggest that cleavage of synaptobrevin by the neurotoxin causes the inhibition of exocytotic release of vasopressin from secretory granules.
42 citations
16 citations