Recombinant Clostridium botulinum Botulinum neurotoxin type E, partial

Botulinum toxin causes flaccid paralysis by inhibiting neurotransmitter (acetylcholine) release from the presynaptic membranes of nerve terminals of eukaryotic host skeletal and autonomic nervous system, with frequent heart or respiratory failure. Precursor of botulinum neurotoxin E which has 2 coreceptors; complex polysialylated gangliosides found on neural tissue and specific membrane-anchored proteins found in synaptic vesicles. Receptor proteins are exposed on host presynaptic cell membrane during neurotransmitter release, when the toxin heavy chain (HC) binds to them. Upon synaptic vesicle recycling the toxin is taken up via the endocytic pathway. When the pH of the toxin-containing endosome drops a structural rearrangement occurs so that the N-terminus of the HC forms pores that allows the light chain (LC) to translocate into the cytosol. Once in the cytosol the disulfide bond linking the 2 subunits is reduced and LC cleaves its target protein on synaptic vesicles, preventing their fusion with the cytoplasmic membrane and thus neurotransmitter release. Electrical stimulation increases uptake of toxin, probably by transiently exposing a receptor found in eukaryotic target synaptic vesicles. Uses the large lumenal domain of synaptic vesicle glycoproteins 2A and 2B (SV2A and SV2B) but not SV2C as receptor; an N-linked glycan of SV2 is essential for receptor function. Host cell gangliosides are also required for neurotoxin uptake and full toxicity. BoNT/E is a 'coincidence detector'; it requires simultaneous binding to coreceptor GT1b and low pH to transform into a membrane-bound, oligomeric channel. Requires trypsinization and reduction before it can be used in assays in vitro.; Has proteolytic activity. After translocation into the eukaryotic host cytosol, inhibits neurotransmitter release by acting as a zinc endopeptidase that catalyzes the hydrolysis of the '180-Arg-|-Ile-181' bond in SNAP25. Hydrolyzes the '185-Arg-|-Ile-186' bond of mouse SNAP23, but not in human which has a different sequence. Recognizes the '146-Met--Asp-186' region of SNAP25. The reaction mechanism probably has a nucleophilic water held in place by Glu-213. Reduction of the interchain disulfide bond occurs in the host cytosol and probably prevents retrotranslocation into the synaptic vesicle.; Responsible for host epithelial cell transcytosis, host nerve cell targeting and translocation of light chain (LC) into host cytosol. Composed of 3 subdomains; the translocation domain (TD), and N-terminus and C-terminus of the receptor-binding domain (RBD). The RBD is responsible for the adherence of the toxin to the cell surface. It probably simultaneously recognizes 2 coreceptors; polysialated gangliosides and either of the receptor proteins SV2A and SV2B in close proximity on host synaptic vesicles. The N-terminus of the TD wraps an extended belt around the perimeter of the light chain (LC), protecting Zn(2+) in the active site. The belt may also prevent premature LC dissociation from the translocation channel and protect toxin prior to translocation. The TD inserts into synaptic vesicle membrane to allow translocation into the host cytosol. Responsible for adherence of the toxin to the cell surface; HC alone prevents uptake of whole toxin by neural cells, and delays paralysis onset by 154%. Significantly decreases uptake and toxicity of whole BoNT/E, but also interferes with uptake of BoNT/C; binds GT1b in vitro. Binds to synaptic vesicle glycoproteins SV2A and SV2B which serve as coreceptors with gangliosides. Interaction with SV2 proteins requires SV2 glycosylation. HC alone significantly decreases uptake and toxicity of whole BoNT/E. HC is responsible for translocation of LC into the host cytosol; an intact disulfide bond between the 2 subunits is required for translocation, which is reduced upon contact with the host cytosol.