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- PDB-6qns: Crystal structure of the binding domain of Botulinum Neurotoxin t... -

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Entry
Database: PDB / ID: 6qns
TitleCrystal structure of the binding domain of Botulinum Neurotoxin type B mutant I1248W/V1249W in complex with human synaptotagmin 1 and GD1a receptors
Components
  • Botulinum neurotoxin type B
  • Synaptotagmin-1
KeywordsTOXIN / botulinum toxin
Function / homology
Function and homology information


clathrin-sculpted acetylcholine transport vesicle membrane / Toxicity of botulinum toxin type G (botG) / clathrin-sculpted glutamate transport vesicle membrane / synchronous neurotransmitter secretion / fast, calcium ion-dependent exocytosis of neurotransmitter / positive regulation of calcium ion-dependent exocytosis of neurotransmitter / syntaxin-3 binding / regulation of regulated secretory pathway / spontaneous neurotransmitter secretion / Toxicity of botulinum toxin type B (botB) ...clathrin-sculpted acetylcholine transport vesicle membrane / Toxicity of botulinum toxin type G (botG) / clathrin-sculpted glutamate transport vesicle membrane / synchronous neurotransmitter secretion / fast, calcium ion-dependent exocytosis of neurotransmitter / positive regulation of calcium ion-dependent exocytosis of neurotransmitter / syntaxin-3 binding / regulation of regulated secretory pathway / spontaneous neurotransmitter secretion / Toxicity of botulinum toxin type B (botB) / clathrin-sculpted gamma-aminobutyric acid transport vesicle membrane / calcium-dependent activation of synaptic vesicle fusion / chromaffin granule membrane / dense core granule / GABA synthesis, release, reuptake and degradation / Acetylcholine Neurotransmitter Release Cycle / clathrin-sculpted monoamine transport vesicle membrane / regulation of calcium ion-dependent exocytosis / calcium ion sensor activity / Serotonin Neurotransmitter Release Cycle / vesicle docking / exocytic vesicle / regulation of exocytosis / Dopamine Neurotransmitter Release Cycle / Norepinephrine Neurotransmitter Release Cycle / vesicle organization / protein heterooligomerization / vesicle fusion / positive regulation of dopamine secretion / Glutamate Neurotransmitter Release Cycle / bontoxilysin / positive regulation of dendrite extension / neurotransmitter secretion / host cell presynaptic membrane / calcium-dependent phospholipid binding / host cell cytoplasmic vesicle / neuron projection terminus / membraneless organelle assembly / syntaxin-1 binding / Neurexins and neuroligins / host cell cytosol / clathrin binding / low-density lipoprotein particle receptor binding / regulation of synaptic vesicle exocytosis / phosphatidylserine binding / presynaptic active zone / postsynaptic cytosol / excitatory synapse / protein transmembrane transporter activity / detection of calcium ion / positive regulation of synaptic transmission / presynaptic cytosol / regulation of synaptic transmission, glutamatergic / vesicle-mediated transport / phosphatidylinositol-4,5-bisphosphate binding / cellular response to calcium ion / SNARE binding / hippocampal mossy fiber to CA3 synapse / clathrin-coated endocytic vesicle membrane / molecular condensate scaffold activity / metalloendopeptidase activity / synaptic vesicle membrane / calcium-dependent protein binding / synaptic vesicle / Cargo recognition for clathrin-mediated endocytosis / Clathrin-mediated endocytosis / presynaptic membrane / toxin activity / chemical synaptic transmission / postsynaptic membrane / cell differentiation / calmodulin binding / neuron projection / postsynaptic density / protein heterodimerization activity / axon / lipid binding / calcium ion binding / host cell plasma membrane / glutamatergic synapse / Golgi apparatus / proteolysis / extracellular region / zinc ion binding / identical protein binding / membrane / plasma membrane / cytoplasm
Similarity search - Function
Synaptotagmin / Clostridium neurotoxin, translocation / Clostridium neurotoxin, Translocation domain / Clostridium neurotoxin, translocation domain / Clostridial neurotoxin zinc protease / Botulinum/Tetanus toxin, catalytic chain / Clostridium neurotoxin, receptor-binding C-terminal / Clostridium neurotoxin, receptor binding N-terminal / Clostridium neurotoxin, C-terminal receptor binding / Clostridium neurotoxin, N-terminal receptor binding ...Synaptotagmin / Clostridium neurotoxin, translocation / Clostridium neurotoxin, Translocation domain / Clostridium neurotoxin, translocation domain / Clostridial neurotoxin zinc protease / Botulinum/Tetanus toxin, catalytic chain / Clostridium neurotoxin, receptor-binding C-terminal / Clostridium neurotoxin, receptor binding N-terminal / Clostridium neurotoxin, C-terminal receptor binding / Clostridium neurotoxin, N-terminal receptor binding / Kunitz inhibitor STI-like superfamily / Protein kinase C conserved region 2 (CalB) / C2 domain / C2 domain / C2 domain profile. / Trefoil (Acidic Fibroblast Growth Factor, subunit A) - #50 / Trefoil (Acidic Fibroblast Growth Factor, subunit A) / Trefoil / Jelly Rolls - #200 / C2 domain superfamily / Neutral zinc metallopeptidases, zinc-binding region signature. / Concanavalin A-like lectin/glucanase domain superfamily / Jelly Rolls / Sandwich / Mainly Beta
Similarity search - Domain/homology
Botulinum neurotoxin type B / Synaptotagmin-1
Similarity search - Component
Biological speciesClostridium botulinum (bacteria)
Homo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.4 Å
AuthorsMasuyer, G. / Yin, L. / Zhang, S. / Miyashita, S.I. / Dong, M. / Stenmark, P.
Funding support Sweden, 1items
OrganizationGrant numberCountry
Swedish Research Council Sweden
CitationJournal: Plos Biol. / Year: 2020
Title: Characterization of a membrane binding loop leads to engineering botulinum neurotoxin B with improved therapeutic efficacy.
Authors: Yin, L. / Masuyer, G. / Zhang, S. / Zhang, J. / Miyashita, S.I. / Burgin, D. / Lovelock, L. / Coker, S.F. / Fu, T.M. / Stenmark, P. / Dong, M.
History
DepositionFeb 12, 2019Deposition site: PDBE / Processing site: PDBE
Revision 1.0Feb 26, 2020Provider: repository / Type: Initial release
Revision 1.1Mar 25, 2020Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 2.0Jul 29, 2020Group: Advisory / Atomic model ...Advisory / Atomic model / Data collection / Derived calculations / Structure summary
Category: atom_site / chem_comp ...atom_site / chem_comp / entity / pdbx_branch_scheme / pdbx_chem_comp_identifier / pdbx_entity_branch / pdbx_entity_branch_descriptor / pdbx_entity_branch_link / pdbx_entity_branch_list / pdbx_entity_nonpoly / pdbx_nonpoly_scheme / pdbx_struct_assembly_gen / pdbx_validate_close_contact / struct_asym / struct_conn / struct_site / struct_site_gen
Item: _atom_site.auth_asym_id / _atom_site.auth_seq_id ..._atom_site.auth_asym_id / _atom_site.auth_seq_id / _atom_site.label_asym_id / _atom_site.label_entity_id / _chem_comp.name / _pdbx_struct_assembly_gen.asym_id_list / _pdbx_validate_close_contact.auth_asym_id_1 / _pdbx_validate_close_contact.auth_asym_id_2 / _pdbx_validate_close_contact.auth_seq_id_1 / _pdbx_validate_close_contact.auth_seq_id_2 / _struct_conn.pdbx_dist_value / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id
Description: Carbohydrate remediation / Provider: repository / Type: Remediation
Revision 2.1Jan 24, 2024Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description / Structure summary
Category: chem_comp / chem_comp_atom ...chem_comp / chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_conn
Item: _chem_comp.pdbx_synonyms / _database_2.pdbx_DOI ..._chem_comp.pdbx_synonyms / _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_conn.pdbx_leaving_atom_flag

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Botulinum neurotoxin type B
S: Synaptotagmin-1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)56,9493
Polymers55,6592
Non-polymers1,2901
Water1,53185
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area3110 Å2
ΔGint8 kcal/mol
Surface area22200 Å2
MethodPISA
Unit cell
Length a, b, c (Å)50.344, 78.491, 149.103
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number19
Space group name H-MP212121

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Components

#1: Protein Botulinum neurotoxin type B / BoNT/B / Bontoxilysin-B


Mass: 53186.824 Da / Num. of mol.: 1 / Mutation: I1248W, V1249W
Source method: isolated from a genetically manipulated source
Details: The N- and C-termini were not visible. Please keep sequence numbering of the full botulinum toxin, as per the uploaded PDB, so that it starts at N862 and terminates at E1291. Thank you.
Source: (gene. exp.) Clostridium botulinum (bacteria) / Gene: botB / Production host: Escherichia coli (E. coli) / Strain (production host): BL21 DE3 / References: UniProt: P10844, bontoxilysin
#2: Protein/peptide Synaptotagmin-1 / Synaptotagmin I / SytI / p65


Mass: 2471.825 Da / Num. of mol.: 1 / Source method: obtained synthetically
Details: Synthetic peptide corresponding to human synaptotagmin 1 residues 33-53
Source: (synth.) Homo sapiens (human) / References: UniProt: P21579
#3: Polysaccharide N-acetyl-alpha-neuraminic acid-(2-3)-beta-D-galactopyranose-(1-3)-2-acetamido-2-deoxy-beta-D- ...N-acetyl-alpha-neuraminic acid-(2-3)-beta-D-galactopyranose-(1-3)-2-acetamido-2-deoxy-beta-D-galactopyranose-(1-4)-[N-acetyl-alpha-neuraminic acid-(2-3)]beta-D-galactopyranose-(1-4)-beta-D-glucopyranose


Type: oligosaccharide / Mass: 1290.140 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DNeup5Aca2-3DGalpb1-3DGalpNAcb1-4[DNeup5Aca2-3]DGalpb1-4DGlcpb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/4,6,5/[a2122h-1b_1-5][a2112h-1b_1-5][Aad21122h-2a_2-6_5*NCC/3=O][a2112h-1b_1-5_2*NCC/3=O]/1-2-3-4-2-3/a4-b1_b3-c2_b4-d1_d3-e1_e3-f2WURCSPDB2Glycan 1.1.0
[][b-D-Glcp]{[(4+1)][b-D-Galp]{[(3+2)][a-D-Neup5Ac]{}[(4+1)][b-D-GalpNAc]{[(3+1)][b-D-Galp]{[(3+2)][a-D-Neup5Ac]{}}}}}LINUCSPDB-CARE
#4: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 85 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.6 Å3/Da / Density % sol: 53.9 %
Crystal growTemperature: 294.15 K / Method: vapor diffusion, sitting drop
Details: 20% v/v PEG6000, 0.1 M MES pH 6.0, 0.2 M sodium chloride

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: Diamond / Beamline: I04 / Wavelength: 0.979 Å
DetectorType: DECTRIS PILATUS3 S 6M / Detector: PIXEL / Date: Jul 8, 2018
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.979 Å / Relative weight: 1
ReflectionResolution: 2.4→74.6 Å / Num. obs: 23939 / % possible obs: 100 % / Redundancy: 10.4 % / CC1/2: 0.998 / Rmerge(I) obs: 0.117 / Rpim(I) all: 0.054 / Rrim(I) all: 0.129 / Net I/σ(I): 9.9
Reflection shellResolution: 2.4→2.46 Å / Redundancy: 10.5 % / Rmerge(I) obs: 1.469 / Mean I/σ(I) obs: 1.2 / Num. unique obs: 1728 / CC1/2: 0.907 / Rpim(I) all: 0.676 / Rrim(I) all: 1.62 / % possible all: 100

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Processing

Software
NameVersionClassification
REFMAC5.8.0238refinement
DIALSdata reduction
Aimlessdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 4KBB

4kbb


Resolution: 2.4→74.55 Å / Cor.coef. Fo:Fc: 0.956 / Cor.coef. Fo:Fc free: 0.943 / SU B: 19.277 / SU ML: 0.206 / Cross valid method: THROUGHOUT / ESU R: 0.375 / ESU R Free: 0.234
RfactorNum. reflection% reflectionSelection details
Rfree0.22872 1203 5 %RANDOM
Rwork0.19945 ---
obs0.20098 22656 99.76 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å
Displacement parametersBiso mean: 65.14 Å2
Baniso -1Baniso -2Baniso -3
1-4.38 Å20 Å20 Å2
2--2.92 Å20 Å2
3----7.3 Å2
Refinement stepCycle: 1 / Resolution: 2.4→74.55 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3813 0 88 85 3986
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0080.0134000
X-RAY DIFFRACTIONr_bond_other_d0.0010.0183559
X-RAY DIFFRACTIONr_angle_refined_deg1.5431.6655401
X-RAY DIFFRACTIONr_angle_other_deg1.2011.5998305
X-RAY DIFFRACTIONr_dihedral_angle_1_deg9.9215444
X-RAY DIFFRACTIONr_dihedral_angle_2_deg31.45523.598239
X-RAY DIFFRACTIONr_dihedral_angle_3_deg12.93515724
X-RAY DIFFRACTIONr_dihedral_angle_4_deg14.0041518
X-RAY DIFFRACTIONr_chiral_restr0.0660.2524
X-RAY DIFFRACTIONr_gen_planes_refined0.0070.024362
X-RAY DIFFRACTIONr_gen_planes_other0.0010.02885
X-RAY DIFFRACTIONr_nbd_refined
X-RAY DIFFRACTIONr_nbd_other
X-RAY DIFFRACTIONr_nbtor_refined
X-RAY DIFFRACTIONr_nbtor_other
X-RAY DIFFRACTIONr_xyhbond_nbd_refined
X-RAY DIFFRACTIONr_xyhbond_nbd_other
X-RAY DIFFRACTIONr_metal_ion_refined
X-RAY DIFFRACTIONr_metal_ion_other
X-RAY DIFFRACTIONr_symmetry_vdw_refined
X-RAY DIFFRACTIONr_symmetry_vdw_other
X-RAY DIFFRACTIONr_symmetry_hbond_refined
X-RAY DIFFRACTIONr_symmetry_hbond_other
X-RAY DIFFRACTIONr_symmetry_metal_ion_refined
X-RAY DIFFRACTIONr_symmetry_metal_ion_other
X-RAY DIFFRACTIONr_mcbond_it3.7595.8691782
X-RAY DIFFRACTIONr_mcbond_other3.7555.8681781
X-RAY DIFFRACTIONr_mcangle_it6.1038.82224
X-RAY DIFFRACTIONr_mcangle_other6.1028.8012225
X-RAY DIFFRACTIONr_scbond_it4.0316.4082218
X-RAY DIFFRACTIONr_scbond_other4.0316.4122219
X-RAY DIFFRACTIONr_scangle_it
X-RAY DIFFRACTIONr_scangle_other6.6769.4313178
X-RAY DIFFRACTIONr_long_range_B_refined9.49564.9134376
X-RAY DIFFRACTIONr_long_range_B_other9.49764.934373
X-RAY DIFFRACTIONr_rigid_bond_restr
X-RAY DIFFRACTIONr_sphericity_free
X-RAY DIFFRACTIONr_sphericity_bonded
LS refinement shellResolution: 2.398→2.46 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.378 74 -
Rwork0.358 1620 -
obs--98.2 %
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
10.06580.1902-0.06940.5693-0.16540.7659-0.01830.01080.0169-0.0445-0.01660.05820.0748-0.1050.03490.05260.025-0.00790.1158-0.02320.0124-10.347310.9489-13.6855
21.77792.4197-0.88496.66153.46917.0466-0.0739-0.05290.0219-0.3334-0.04660.0294-0.17050.14630.12050.20490.0573-0.00780.12620.10380.10421.717242.8483-31.2027
Refinement TLS group
IDRefine-IDRefine TLS-IDAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1A862 - 1291
2X-RAY DIFFRACTION2S37 - 52

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