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- PDB-3ddb: Crystal structure of the catalytic domain of Botulinum neurotoxin... -

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Basic information

Entry
Database: PDB / ID: 3ddb
TitleCrystal structure of the catalytic domain of Botulinum neurotoxin serotype a with a substrate analog peptide
Components
  • Botulinum neurotoxin A light chain
  • Synaptosomal-associated protein 25SNAP25
KeywordsHYDROLASE / BOTULINUM NEUROTOXIN TYPE A / BOTOX / CATALYTIC DOMAIN / ENDOPEPTIDASE / SYNTAXIN / BIO-WARFARE AGENT / METAL-BINDING / METALLOPROTEASE / PROTEASE / SECRETED / TRANSMEMBRANE / ZINC / ENZYME-SUBSTRATE COMPLEX / Pharmaceutical
Function / homology
Function and homology information


Toxicity of botulinum toxin type C (botC) / neurotransmitter uptake / Toxicity of botulinum toxin type E (botE) / host cell junction / exocytic insertion of neurotransmitter receptor to postsynaptic membrane / Toxicity of botulinum toxin type A (botA) / Acetylcholine Neurotransmitter Release Cycle / extrinsic component of presynaptic membrane / synaptic vesicle fusion to presynaptic active zone membrane / synaptobrevin 2-SNAP-25-syntaxin-1a-complexin II complex ...Toxicity of botulinum toxin type C (botC) / neurotransmitter uptake / Toxicity of botulinum toxin type E (botE) / host cell junction / exocytic insertion of neurotransmitter receptor to postsynaptic membrane / Toxicity of botulinum toxin type A (botA) / Acetylcholine Neurotransmitter Release Cycle / extrinsic component of presynaptic membrane / synaptic vesicle fusion to presynaptic active zone membrane / synaptobrevin 2-SNAP-25-syntaxin-1a-complexin II complex / synaptobrevin 2-SNAP-25-syntaxin-1a complex / synaptobrevin 2-SNAP-25-syntaxin-1a-complexin I complex / Serotonin Neurotransmitter Release Cycle / GABA synthesis, release, reuptake and degradation / presynaptic dense core vesicle exocytosis / ribbon synapse / synaptic vesicle docking / Dopamine Neurotransmitter Release Cycle / Norepinephrine Neurotransmitter Release Cycle / SNARE complex / negative regulation of neurotransmitter secretion / SNAP receptor activity / Glutamate Neurotransmitter Release Cycle / bontoxilysin / host cell presynaptic membrane / neurotransmitter receptor internalization / host cell cytoplasmic vesicle / Sensory processing of sound by inner hair cells of the cochlea / syntaxin-1 binding / SNARE complex assembly / synaptic vesicle priming / host cell cytosol / regulation of synapse assembly / endosomal transport / Other interleukin signaling / myosin binding / regulation of neuron projection development / exocytosis / synaptic vesicle exocytosis / voltage-gated potassium channel activity / tertiary granule membrane / associative learning / regulation of insulin secretion / protein transmembrane transporter activity / long-term memory / specific granule membrane / voltage-gated potassium channel complex / axonal growth cone / presynaptic active zone membrane / photoreceptor inner segment / axonogenesis / locomotory behavior / filopodium / long-term synaptic potentiation / Regulation of insulin secretion / trans-Golgi network / positive regulation of insulin secretion / metalloendopeptidase activity / calcium-dependent protein binding / synaptic vesicle / actin cytoskeleton / presynaptic membrane / lamellipodium / cell cortex / growth cone / toxin activity / chemical synaptic transmission / postsynapse / transmembrane transporter binding / membrane => GO:0016020 / cytoskeleton / endosome / neuron projection / protein domain specific binding / neuronal cell body / glutamatergic synapse / lipid binding / Neutrophil degranulation / host cell plasma membrane / perinuclear region of cytoplasm / proteolysis / zinc ion binding / extracellular region / membrane / plasma membrane / cytosol / cytoplasm
Similarity search - Function
Synaptosomal-associated protein 25 / SNAP-25 domain / SNAP-25 family / Zincin-like / Metalloproteases ("zincins"), catalytic domain like / Helical region found in SNAREs / t-SNARE coiled-coil homology domain profile. / Target SNARE coiled-coil homology domain / Clostridium neurotoxin, translocation / Clostridium neurotoxin, Translocation domain ...Synaptosomal-associated protein 25 / SNAP-25 domain / SNAP-25 family / Zincin-like / Metalloproteases ("zincins"), catalytic domain like / Helical region found in SNAREs / t-SNARE coiled-coil homology domain profile. / Target SNARE coiled-coil homology domain / Clostridium neurotoxin, translocation / Clostridium neurotoxin, Translocation domain / Clostridium neurotoxin, translocation domain / Clostridial neurotoxin zinc protease / Botulinum/Tetanus toxin, catalytic chain / Clostridium neurotoxin, receptor binding N-terminal / Clostridium neurotoxin, receptor-binding C-terminal / Clostridium neurotoxin, C-terminal receptor binding / Clostridium neurotoxin, N-terminal receptor binding / Kunitz inhibitor STI-like superfamily / Neutral zinc metallopeptidases, zinc-binding region signature. / Concanavalin A-like lectin/glucanase domain superfamily / Alpha-Beta Complex / Alpha Beta
Similarity search - Domain/homology
Botulinum neurotoxin type A / Botulinum neurotoxin type A / Synaptosomal-associated protein 25
Similarity search - Component
Biological speciesClostridium botulinum (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / FOURIER SYNTHESIS / Resolution: 1.6 Å
AuthorsKumaran, D. / Swaminathan, S.
CitationJournal: Plos Pathog. / Year: 2008
Title: Substrate binding mode and its implication on drug design for botulinum neurotoxin A
Authors: Kumaran, D. / Rawat, R. / Ahmed, S.A. / Swaminathan, S.
History
DepositionJun 5, 2008Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 16, 2008Provider: repository / Type: Initial release
Revision 1.1Jul 13, 2011Group: Version format compliance
Revision 1.2Aug 30, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_conn / struct_conn_type / struct_ref_seq / struct_ref_seq_dif / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_conn.conn_type_id / _struct_conn.id / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_leaving_atom_flag / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_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 / _struct_conn.ptnr2_label_seq_id / _struct_conn_type.id / _struct_ref_seq.db_align_beg / _struct_ref_seq.db_align_end / _struct_ref_seq_dif.details / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Botulinum neurotoxin A light chain
B: Synaptosomal-associated protein 25
hetero molecules


Theoretical massNumber of molelcules
Total (without water)50,4365
Polymers50,1792
Non-polymers2583
Water6,756375
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area1960 Å2
ΔGint-65 kcal/mol
Surface area18900 Å2
MethodPISA
Unit cell
Length a, b, c (Å)50.871, 66.581, 65.062
Angle α, β, γ (deg.)90.00, 98.34, 90.00
Int Tables number4
Space group name H-MP1211

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Components

#1: Protein Botulinum neurotoxin A light chain / Botulinum neurotoxin type A / BoNT/A / Bontoxilysin-A / BOTOX


Mass: 49415.727 Da / Num. of mol.: 1 / Fragment: residues 1-424
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Clostridium botulinum (bacteria) / Strain: Hall / Gene: botA / Plasmid: PET28B / Production host: Escherichia coli (E. coli) / Strain (production host): BL21-DE3-RIL
References: UniProt: A5HZZ9, UniProt: P0DPI1*PLUS, bontoxilysin
#2: Protein/peptide Synaptosomal-associated protein 25 / SNAP25 / SNAP-25 / Synaptosomal-associated 25 kDa protein / Super protein / SUP


Mass: 762.968 Da / Num. of mol.: 1 / Source method: obtained synthetically
Details: The SNAP-25 peptide is naturally found in Homo sapiens.
References: UniProt: P60880*PLUS
#3: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Zn
#4: Chemical ChemComp-SO4 / SULFATE ION / Sulfate


Mass: 96.063 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: SO4
#5: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 375 / 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: 2.17 Å3/Da / Density % sol: 43.38 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / pH: 6.5
Details: 20% PEG8000, 0.2M Ammoium Sulfate, 0.1M Sodium Cacodylate, 5% Ethylene Glycol, pH 6.5, VAPOR DIFFUSION, SITTING DROP, temperature 293K

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: NSLS / Beamline: X29A / Wavelength: 0.979 Å
DetectorType: ADSC QUANTUM 315 / Detector: CCD / Date: Mar 28, 2008 / Details: mirrors
RadiationMonochromator: SI III / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.979 Å / Relative weight: 1
ReflectionResolution: 1.6→50 Å / Num. all: 53272 / Num. obs: 53272 / % possible obs: 95.4 % / Observed criterion σ(F): 0 / Observed criterion σ(I): 0 / Redundancy: 12.1 % / Biso Wilson estimate: 15.8 Å2 / Rmerge(I) obs: 0.059 / Net I/σ(I): 23
Reflection shellResolution: 1.6→1.65 Å / Redundancy: 7.7 % / Rmerge(I) obs: 0.147 / Mean I/σ(I) obs: 2.5 / Num. unique all: 3514 / % possible all: 76.3

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Processing

Software
NameVersionClassification
CNS1.1refinement
CBASSdata collection
HKL-2000data reduction
HKL-2000data scaling
CNSphasing
RefinementMethod to determine structure: FOURIER SYNTHESIS
Starting model: PDB entry 3BWI

3bwi


Resolution: 1.6→32.42 Å / Rfactor Rfree error: 0.004 / Data cutoff high absF: 152506 / Data cutoff low absF: 0 / Isotropic thermal model: RESTRAINED / Cross valid method: THROUGHOUT / σ(F): 0 / Stereochemistry target values: Engh & Huber
RfactorNum. reflection% reflectionSelection details
Rfree0.218 2672 5.1 %RANDOM
Rwork0.2 ---
obs0.2 52650 92.7 %-
all-52650 --
Solvent computationSolvent model: FLAT MODEL / Bsol: 45.3482 Å2 / ksol: 0.346887 e/Å3
Displacement parametersBiso mean: 23 Å2
Baniso -1Baniso -2Baniso -3
1-14.2 Å20 Å20.87 Å2
2---6.47 Å20 Å2
3----7.73 Å2
Refine analyze
FreeObs
Luzzati coordinate error0.21 Å0.18 Å
Luzzati d res low-5 Å
Luzzati sigma a0.13 Å0.1 Å
Refinement stepCycle: LAST / Resolution: 1.6→32.42 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3474 0 12 375 3861
Refine LS restraints
Refine-IDTypeDev ideal
X-RAY DIFFRACTIONc_bond_d0.005
X-RAY DIFFRACTIONc_angle_deg1.2
X-RAY DIFFRACTIONc_dihedral_angle_d22.7
X-RAY DIFFRACTIONc_improper_angle_d0.74
LS refinement shellResolution: 1.6→1.7 Å / Rfactor Rfree error: 0.013 / Total num. of bins used: 6
RfactorNum. reflection% reflection
Rfree0.25 352 5 %
Rwork0.217 6669 -
obs--74.3 %

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