| IED ID | IndEnz0002003439 |
| Enzyme Type ID | protease003439 |
| Protein Name |
Botulinum neurotoxin type X BoNT/X Bontoxilysin-X Cleaved into: Botulinum neurotoxin X light chain LC EC 3.4.24.69 ; Botulinum neurotoxin X heavy chain HC |
| Gene Name | CBB2_0680 |
| Organism | Clostridium botulinum |
| Taxonomic Lineage | cellular organisms Bacteria Terrabacteria group Firmicutes Clostridia Eubacteriales Clostridiaceae Clostridium Clostridium botulinum |
| Enzyme Sequence | MKLEINKFNYNDPIDGINVITMRPPRHSDKINKGKGPFKAFQVIKNIWIVPERYNFTNNTNDLNIPSEPIMEADAIYNPNYLNTPSEKDEFLQGVIKVLERIKSKPEGEKLLELISSSIPLPLVSNGALTLSDNETIAYQENNNIVSNLQANLVIYGPGPDIANNATYGLYSTPISNGEGTLSEVSFSPFYLKPFDESYGNYRSLVNIVNKFVKREFAPDPASTLMHELVHVTHNLYGISNRNFYYNFDTGKIETSRQQNSLIFEELLTFGGIDSKAISSLIIKKIIETAKNNYTTLISERLNTVTVENDLLKYIKNKIPVQGRLGNFKLDTAEFEKKLNTILFVLNESNLAQRFSILVRKHYLKERPIDPIYVNILDDNSYSTLEGFNISSQGSNDFQGQLLESSYFEKIESNALRAFIKICPRNGLLYNAIYRNSKNYLNNIDLEDKKTTSKTNVSYPCSLLNGCIEVENKDLFLISNKDSLNDINLSEEKIKPETTVFFKDKLPPQDITLSNYDFTEANSIPSISQQNILERNEELYEPIRNSLFEIKTIYVDKLTTFHFLEAQNIDESIDSSKIRVELTDSVDEALSNPNKVYSPFKNMSNTINSIETGITSTYIFYQWLRSIVKDFSDETGKIDVIDKSSDTLAIVPYIGPLLNIGNDIRHGDFVGAIELAGITALLEYVPEFTIPILVGLEVIGGELAREQVEAIVNNALDKRDQKWAEVYNITKAQWWGTIHLQINTRLAHTYKALSRQANAIKMNMEFQLANYKGNIDDKAKIKNAISETEILLNKSVEQAMKNTEKFMIKLSNSYLTKEMIPKVQDNLKNFDLETKKTLDKFIKEKEDILGTNLSSSLRRKVSIRLNKNIAFDINDIPFSEFDDLINQYKNEIEDYEVLNLGAEDGKIKDLSGTTSDINIGSDIELADGRENKAIKIKGSENSTIKIAMNKYLRFSATDNFSISFWIKHPKPTNLLNNGIEYTLVENFNQRGWKISIQDSKLIWYLRDHNNSIKIVTPDYIAFNGWNLITITNNRSKGSIVYVNGSKIEEKDISSIWNTEVDDPIIFRLKNNRDTQAFTLLDQFSIYRKELNQNEVVKLYNYYFNSNYIRDIWGNPLQYNKKYYLQTQDKPGKGLIREYWSSFGYDYVILSDSKTITFPNNIRYGALYNGSKVLIKNSKKLDGLVRNKDFIQLEIDGYNMGISADRFNEDTNYIGTTYGTTHDLTTDFEIIQRQEKYRNYCQLKTPYNIFHKSGLMSTETSKPTFHDYRDWVYSSAWYFQNYENLNLRKHTKTNWYFIPKDEGWDED |
| Enzyme Length | 1306 |
| Uniprot Accession Number | P0DPK1 |
| Absorption | |
| Active Site | ACT_SITE 228; /evidence="ECO:0000250|UniProtKB:P0DPI0, ECO:0000255|PROSITE-ProRule:PRU10095" |
| Activity Regulation | ACTIVITY REGULATION: VAMP2 cleavage inhibited by EDTA. {ECO:0000269|PubMed:28770820}. |
| Binding Site | |
| 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:28770820}; |
| DNA Binding | |
| EC Number | 3.4.24.69 |
| Enzyme Function | FUNCTION: [Botulinum neurotoxin type X]: 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 X 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 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 (By similarity). Artificially assembled BoNT/X cleaves synaptobrevin-2/VAMP2 and VAMP4 in cultured rat neurons and causes flaccid paralysis in mice (PubMed:28770820). {ECO:0000250|UniProtKB:P0DPI0, ECO:0000269|PubMed:28770820}.; FUNCTION: [Botulinum neurotoxin X light chain]: Has proteolytic activity. After translocation into the eukaryotic host cytosol, LC hydrolyzes the '66-Arg-|-Ala-67' bond in synaptobrevin-2/VAMP2, and the equivalent bonds in 'Arg-|-Ala' bonds in VAMP1 and VAMP3, thus blocking neurotransmitter release (PubMed:28770820). Has a wider target range than most BoNTs, as it also cleaves the '87-Arg-|-Ser-89' bond in VAMP4, the '40-Arg-|-Ser-41' bond in VAMP5 and the '173-Lys-|-Ser-174' bond in YKT6; whether these are physiologically relevant substrates is unknown (PubMed:28770820). BoNT/X is 10-fold more efficient than BoNT/B and 40-fold more efficient than TeTX on an artificial human VAMP1 substrate (PubMed:29540745). {ECO:0000269|PubMed:28770820, ECO:0000269|PubMed:29540745}.; FUNCTION: [Botulinum neurotoxin X heavy chain]: Responsible for 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 simultaneously recognizes 2 coreceptors; polysialated gangliosides and an unknown receptor protein in close proximity on host synaptic vesicles. The N-terminus of the TD wraps an extended belt around the perimeter of the LC, protecting Zn(2+) in the active site (By similarity). The TD inserts into synaptic vesicle membrane to allow translocation into the host cytosol (By similarity). Protein ligation of the RBD to the rest of the toxin (creates an artificial whole toxin) greatly increases VAMP2 degradation, and thus neuron uptake (PubMed:28770820). {ECO:0000250|UniProtKB:P10844, ECO:0000269|PubMed:28770820}. |
| Temperature Dependency | |
| PH Dependency | |
| Pathway | |
| nucleotide Binding | |
| Features | Active site (1); Beta strand (22); Chain (3); Disulfide bond (1); Helix (13); Metal binding (3); Motif (1); Mutagenesis (4); Region (4); Turn (8) |
| Keywords | 3D-structure;Disulfide bond;Host cell junction;Host cell membrane;Host cytoplasm;Host cytoplasmic vesicle;Host membrane;Host synapse;Hydrolase;Membrane;Metal-binding;Metalloprotease;Neurotoxin;Protease;Secreted;Toxin;Virulence;Zinc |
| Interact With | |
| Induction | |
| Subcellular Location | SUBCELLULAR LOCATION: [Botulinum neurotoxin type X]: Secreted {ECO:0000250|UniProtKB:P0DPI0}.; SUBCELLULAR LOCATION: [Botulinum neurotoxin X light chain]: Secreted {ECO:0000250|UniProtKB:P0DPI0}. Host cytoplasm, host cytosol {ECO:0000250|UniProtKB:P0DPI0}.; SUBCELLULAR LOCATION: [Botulinum neurotoxin X heavy chain]: Secreted {ECO:0000250|UniProtKB:P0DPI0}. Host cell junction, host synapse, host presynaptic cell membrane {ECO:0000250|UniProtKB:P0DPI0}. Host cytoplasmic vesicle, host secretory vesicle, host synaptic vesicle membrane {ECO:0000250|UniProtKB:P0DPI0}. |
| Modified Residue | |
| Post Translational Modification | PTM: An interchain disulfide bond is required for toxin stability in an artificial construct with the light chain and translocation domain; which of Cys-461 or Cys-467 forms the disulfide bond with Cys-423 in vivo is unknown. {ECO:0000305|PubMed:28770820}. |
| Signal Peptide | |
| Structure 3D | X-ray crystallography (5) |
| Cross Reference PDB | 6F47; 6F4E; 6G8U; 6G8V; 7KZ7; |
| Mapped Pubmed ID | 33602850; |
| Motif | MOTIF 1274..1277; /note="Host ganglioside-binding motif"; /evidence="ECO:0000250|UniProtKB:P0DPI0, ECO:0000305|PubMed:28770820" |
| Gene Encoded By | |
| Mass | 150,285 |
| Kinetics | BIOPHYSICOCHEMICAL PROPERTIES: Kinetic parameters: KM=4.3 uM for a human VAMP1 fragment (residues 34-87) construct {ECO:0000269|PubMed:29540745}; Note=Apparent rate is 271 min(-1). {ECO:0000269|PubMed:29540745}; |
| Metal Binding | METAL 227; /note="Zinc; via tele nitrogen; catalytic"; /evidence="ECO:0000255|PROSITE-ProRule:PRU10095, ECO:0000269|PubMed:29540745, ECO:0007744|PDB:6F47"; METAL 231; /note="Zinc; via tele nitrogen; catalytic"; /evidence="ECO:0000255|PROSITE-ProRule:PRU10095, ECO:0000269|PubMed:29540745, ECO:0007744|PDB:6F47"; METAL 266; /note="Zinc; catalytic"; /evidence="ECO:0000269|PubMed:29540745, ECO:0007744|PDB:6F47" |
| Rhea ID | |
| Cross Reference Brenda | 3.4.24.69; |