| IED ID | IndEnz0002006067 |
| Enzyme Type ID | protease006067 |
| Protein Name |
Botulinum neurotoxin type C BoNT/C Bontoxilysin-C1 BoNT/C1 Botulinum neurotoxin type C1 Cleaved into: Botulinum neurotoxin C light chain LC EC 3.4.24.69 ; Botulinum neurotoxin C heavy chain HC |
| Gene Name | |
| Organism | Clostridium botulinum C phage (Clostridium botulinum C bacteriophage) |
| Taxonomic Lineage | Viruses Duplodnaviria Heunggongvirae Uroviricota Caudoviricetes Caudovirales Siphoviridae (phages with long non-contractile tails) unclassified Siphoviridae Clostridium botulinum C phage (Clostridium botulinum C bacteriophage) |
| Enzyme Sequence | MPITINNFNYSDPVDNKNILYLDTHLNTLANEPEKAFRITGNIWVIPDRFSRNSNPNLNKPPRVTSPKSGYYDPNYLSTDSDKDTFLKEIIKLFKRINSREIGEELIYRLSTDIPFPGNNNTPINTFDFDVDFNSVDVKTRQGNNWVKTGSINPSVIITGPRENIIDPETSTFKLTNNTFAAQEGFGALSIISISPRFMLTYSNATNDVGEGRFSKSEFCMDPILILMHELNHAMHNLYGIAIPNDQTISSVTSNIFYSQYNVKLEYAEIYAFGGPTIDLIPKSARKYFEEKALDYYRSIAKRLNSITTANPSSFNKYIGEYKQKLIRKYRFVVESSGEVTVNRNKFVELYNELTQIFTEFNYAKIYNVQNRKIYLSNVYTPVTANILDDNVYDIQNGFNIPKSNLNVLFMGQNLSRNPALRKVNPENMLYLFTKFCHKAIDGRSLYNKTLDCRELLVKNTDLPFIGDISDVKTDIFLRKDINEETEVIYYPDNVSVDQVILSKNTSEHGQLDLLYPSIDSESEILPGENQVFYDNRTQNVDYLNSYYYLESQKLSDNVEDFTFTRSIEEALDNSAKVYTYFPTLANKVNAGVQGGLFLMWANDVVEDFTTNILRKDTLDKISDVSAIIPYIGPALNISNSVRRGNFTEAFAVTGVTILLEAFPEFTIPALGAFVIYSKVQERNEIIKTIDNCLEQRIKRWKDSYEWMMGTWLSRIITQFNNISYQMYDSLNYQAGAIKAKIDLEYKKYSGSDKENIKSQVENLKNSLDVKISEAMNNINKFIRECSVTYLFKNMLPKVIDELNEFDRNTKAKLINLIDSHNIILVGEVDKLKAKVNNSFQNTIPFNIFSYTNNSLLKDIINEYFNNINDSKILSLQNRKNTLVDTSGYNAEVSEEGDVQLNPIFPFDFKLGSSGEDRGKVIVTQNENIVYNSMYESFSISFWIRINKWVSNLPGYTIIDSVKNNSGWSIGIISNFLVFTLKQNEDSEQSINFSYDISNNAPGYNKWFFVTVTNNMMGNMKIYINGKLIDTIKVKELTGINFSKTITFEINKIPDTGLITSDSDNINMWIRDFYIFAKELDGKDINILFNSLQYTNVVKDYWGNDLRYNKEYYMVNIDYLNRYMYANSRQIVFNTRRNNNDFNEGYKIIIKRIRGNTNDTRVRGGDILYFDMTINNKAYNLFMKNETMYADNHSTEDIYAIGLREQTKDINDNIIFQIQPMNNTYYYASQIFKSNFNGENISGICSIGTYRFRLGGDWYRHNYLVPTVKQGNYASLLESTSTHWGFVPVSE |
| Enzyme Length | 1291 |
| Uniprot Accession Number | P18640 |
| Absorption | |
| Active Site | ACT_SITE 230; /evidence=ECO:0000255|PROSITE-ProRule:PRU10095 |
| Activity Regulation | ACTIVITY REGULATION: 1,10-phenanthroline, EDTA and partially captopril block cleavage of syntaxin in brain synaptosomes (PubMed:7901002, PubMed:7737992). Treatment of synaptosomes with a mild detergent also inhibits cleavage (PubMed:7737992). 1,10-phenanthroline partially antagonizes inhibitions of neurotransmitter release (PubMed:8611567). {ECO:0000269|PubMed:7737992, ECO:0000269|PubMed:7901002, ECO:0000269|PubMed:8611567}. |
| Binding Site | BINDING 1119; /note="Ganglioside GD1a; GBP2 binding site"; /evidence="ECO:0000305|PubMed:23027864"; BINDING 1146; /note="Ganglioside GD1b; Sia-1 binding site"; /evidence="ECO:0000305|PubMed:21542861, ECO:0007744|PDB:3R4S"; BINDING 1281; /note="Ganglioside GD1a; GBP2 binding site"; /evidence="ECO:0000305|PubMed:23027864" |
| Calcium Binding | |
| catalytic Activity | CATALYTIC ACTIVITY: Reaction=Limited hydrolysis of proteins of the neuroexocytosis apparatus, synaptobrevins, SNAP25 or syntaxin. No detected action on small molecule substrates.; EC=3.4.24.69; Evidence={ECO:0000269|PubMed:7737992, ECO:0000269|PubMed:8611567, ECO:0000269|PubMed:9886085}; |
| DNA Binding | |
| EC Number | 3.4.24.69 |
| Enzyme Function | FUNCTION: [Botulinum neurotoxin type C]: Botulinum toxin causes flaccid paralysis by inhibiting neurotransmitter (acetylcholine) release from the presynaptic membranes of nerve terminals of the eukaryotic host skeletal and autonomic nervous system, with frequent heart or respiratory failure (PubMed:16252491, PubMed:7901002, PubMed:8611567). Is unique among characterized BoNTs in having 2 substrates, syntaxin (STX) and SNAP25 (PubMed:7901002, PubMed:7737992, PubMed:8611567, PubMed:9886085, PubMed:17718519). Precursor of botulinum neurotoxin C which unlike most BoNTs seems not to have a proteinaceous coreceptor, and instead recognizes 2 different complex polysialylated gangliosides found on neural tissue probably found in synaptic vesicles (PubMed:21483489, PubMed:23027864). 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 heavy chain (HC) forms pores that allows the light chain (LC) to translocate into the cytosol (By similarity). 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 (By similarity). In vitro the whole toxin only has protease activity after reduction (PubMed:8611567). Electrical stimulation increases uptake of toxin, presumably by transiently exposing a receptor usually found in eukaryotic target synaptic vesicles (PubMed:19650874). Forms ion-conducting channels at around pH 6.1 (PubMed:2424493). Requires complex eukaryotic host polysialogangliosides for full neurotoxicity (PubMed:19650874, PubMed:21483489). Synaptic vesicle glycoproteins (SV2) do not seem to act as its receptor (PubMed:21483489). {ECO:0000250|UniProtKB:P0DPI0, ECO:0000269|PubMed:17718519, ECO:0000269|PubMed:19650874, ECO:0000269|PubMed:21483489, ECO:0000269|PubMed:2424493, ECO:0000269|PubMed:7737992, ECO:0000269|PubMed:7901002, ECO:0000269|PubMed:8611567, ECO:0000269|PubMed:9886085, ECO:0000305|PubMed:16252491, ECO:0000305|PubMed:23027864}.; FUNCTION: [Botulinum neurotoxin C light chain]: Has proteolytic activity. After translocation into the eukaryotic host cytosol, inhibits neurotransmitter release by acting as a zinc endopeptidase that cleaves syntaxin-1A/STX1A and syntaxin-1B/STX1B (PubMed:7901002, PubMed:7737992, PubMed:8611567). Cleaves the '253-Arg-|-Ala-254' bond of STX1 and the '252-Arg-|-Ala-253' bond of STX2; also acts on syntaxin 3 (STX3) but not 4 (STX4) (PubMed:7737992). Cleaves the '198-Arg-|-Ala-199' bond of SNAP25 (PubMed:8611567, PubMed:9886085, PubMed:17718519). Recognizes the '93-Asn--Met-202' region of SNAP25 (PubMed:9886085). {ECO:0000269|PubMed:17718519, ECO:0000269|PubMed:7737992, ECO:0000269|PubMed:7901002, ECO:0000269|PubMed:8611567, ECO:0000269|PubMed:9886085}.; FUNCTION: [Botulinum neurotoxin C heavy chain]: Responsible for host epithelial cell transcytosis, host nerve cell targeting and translocation of light chain (LC) into eukaryotic 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 eukaryotic target cell surface. It simultaneously recognizes 2 polysialated gangliosides coreceptors in close proximity on host synaptic vesicles (PubMed:23027864, PubMed:21542861). The N-terminus of the TD wraps an extended belt around the perimeter of the LC, protecting Zn(2+) in the active site; it may also prevent premature LC dissociation from the translocation channel and protect toxin prior to translocation (By similarity). The TD inserts into synaptic vesicle membrane to allow translocation into the host cytosol (Probable). The C-terminal half of the HC (residues 864-1291) binds neurons in a dose-dependent manner (PubMed:20731382). The C-terminal half of the HC (residues 863-1291) binds eukaryotic host gangliosides in the order GD1b > GT1b > GD1a > GM1a (PubMed:16115873, PubMed:20731382, PubMed:23027864, PubMed:19650874). Has 2 ganglioside binding sites; Sia-1 prefers a sia7 sialic acid and sugars within the ganglioside (GD1b > GT1b), whereas GBP2 recognizes a sia5 sialic acid (GT1b and GD1a) (PubMed:23027864, PubMed:21542861). Both sites are required for HC to enter neurons, acting via different gangliosides (PubMed:23027864). This suggests that 2 gangliosides serve as toxin receptors (PubMed:16115873, PubMed:20731382, PubMed:21542861, PubMed:23027864). Synaptic activity (depolarization with K(+)) increases uptake by neurons (PubMed:23027864). Treatment of synaptosomes with proteinase K does not reduce HC binding, suggesting there is no protein receptor or it is protected from extracellular proteases (PubMed:16115873). Decreases uptake and toxicity of whole BoNT/A, but also interferes with uptake of BoNT/E and BoNT/F (PubMed:19650874). HC also binds phosphoinositides, which might play a role in membrane-binding (PubMed:22120109). {ECO:0000250|UniProtKB:P0DPI0, ECO:0000269|PubMed:16115873, ECO:0000269|PubMed:19650874, ECO:0000269|PubMed:20731382, ECO:0000269|PubMed:21542861, ECO:0000269|PubMed:22120109, ECO:0000269|PubMed:23027864, ECO:0000305|PubMed:2424493}. |
| Temperature Dependency | |
| PH Dependency | |
| Pathway | |
| nucleotide Binding | |
| Features | Active site (1); Beta strand (50); Binding site (3); Chain (3); Disulfide bond (1); Helix (22); Initiator methionine (1); Metal binding (3); Motif (1); Mutagenesis (10); Region (7); Sequence conflict (1); Turn (5) |
| Keywords | 3D-structure;Direct protein sequencing;Disulfide bond;Hydrolase;Lipid-binding;Metal-binding;Metalloprotease;Neurotoxin;Pharmaceutical;Protease;Secreted;Toxin;Virulence;Zinc |
| Interact With | |
| Induction | |
| Subcellular Location | SUBCELLULAR LOCATION: [Botulinum neurotoxin type C]: Secreted {ECO:0000269|PubMed:16252491}.; SUBCELLULAR LOCATION: [Botulinum neurotoxin C light chain]: Secreted {ECO:0000269|PubMed:16252491, ECO:0000269|PubMed:7802661}. Note=In animals that have ingested BoNT/C the LC acts in the eukaryotic host cytosol (PubMed:7901002, PubMed:7737992, PubMed:8611567, PubMed:9886085, PubMed:17718519). {ECO:0000305|PubMed:17718519, ECO:0000305|PubMed:7737992, ECO:0000305|PubMed:7901002, ECO:0000305|PubMed:8611567, ECO:0000305|PubMed:9886085}.; SUBCELLULAR LOCATION: [Botulinum neurotoxin C heavy chain]: Secreted {ECO:0000269|PubMed:16252491, ECO:0000269|PubMed:7802661}. Note=Upon incubation with cultured neurons the HC is detected in a synaptophysin-positive intracellular compartment (probably host synaptic vesicles) (PubMed:23027864). It probably integrates into the eukaryotic host synaptic vesicle membrane (PubMed:2424493). {ECO:0000269|PubMed:23027864, ECO:0000305|PubMed:2424493}. |
| Modified Residue | |
| Post Translational Modification | |
| Signal Peptide | |
| Structure 3D | X-ray crystallography (5) |
| Cross Reference PDB | 2QN0; 3DEB; 3N7K; 3R4S; 3R4U; |
| Mapped Pubmed ID | - |
| Motif | MOTIF 1256..1258; /note="Host ganglioside-binding motif"; /evidence="ECO:0000250|UniProtKB:P10844, ECO:0000305|PubMed:19650874" |
| Gene Encoded By | |
| Mass | 148,870 |
| Kinetics | BIOPHYSICOCHEMICAL PROPERTIES: Kinetic parameters: KM=18.6 uM for purified SNAP25 with isolated botulinum neurotoxin C light chain {ECO:0000269|PubMed:17718519}; Note=kcat is 0.391 min(-1), for isolated botulinum neurotoxin C light chain. {ECO:0000269|PubMed:17718519}; |
| Metal Binding | METAL 229; /note="Zinc; via tele nitrogen; catalytic"; /evidence="ECO:0000255|PROSITE-ProRule:PRU10095, ECO:0000269|PubMed:17718519, ECO:0007744|PDB:2QN0"; METAL 233; /note="Zinc; via tele nitrogen; catalytic"; /evidence="ECO:0000255|PROSITE-ProRule:PRU10095, ECO:0000269|PubMed:17718519, ECO:0007744|PDB:2QN0"; METAL 269; /note="Zinc; catalytic"; /evidence="ECO:0000269|PubMed:17718519, ECO:0007744|PDB:2QN0" |
| Rhea ID | |
| Cross Reference Brenda | 3.4.24.69; |