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- PDB-5vl3: CD22 d1-d3 in complex with therapeutic Fab Epratuzumab -

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Basic information

Entry
Database: PDB / ID: 5vl3
TitleCD22 d1-d3 in complex with therapeutic Fab Epratuzumab
Components
  • (Epratuzumab Fab ...) x 2
  • B-cell receptor CD22
KeywordsIMMUNE SYSTEM / therapeutic antibody / Siglec / B cell / Fab
Function / homology
Function and homology information


regulation of B cell proliferation / IgM binding / negative regulation of immunoglobulin production / negative regulation of B cell receptor signaling pathway / sialic acid binding / CD4 receptor binding / negative regulation of calcium-mediated signaling / CD22 mediated BCR regulation / neuronal cell body membrane / B cell activation ...regulation of B cell proliferation / IgM binding / negative regulation of immunoglobulin production / negative regulation of B cell receptor signaling pathway / sialic acid binding / CD4 receptor binding / negative regulation of calcium-mediated signaling / CD22 mediated BCR regulation / neuronal cell body membrane / B cell activation / immunoglobulin complex / regulation of endocytosis / regulation of immune response / Antigen activates B Cell Receptor (BCR) leading to generation of second messengers / recycling endosome / Immunoregulatory interactions between a Lymphoid and a non-Lymphoid cell / carbohydrate binding / protein phosphatase binding / adaptive immune response / early endosome / cell adhesion / immune response / external side of plasma membrane / signaling receptor binding / cell surface / extracellular space / extracellular exosome / membrane / metal ion binding / plasma membrane / cytoplasm
Similarity search - Function
CD80-like, immunoglobulin C2-set / CD80-like C2-set immunoglobulin domain / Immunoglobulin domain / : / Immunoglobulin domain / : / Immunoglobulin subtype 2 / Immunoglobulin C-2 Type / Immunoglobulin V-Type / Immunoglobulin V-set domain ...CD80-like, immunoglobulin C2-set / CD80-like C2-set immunoglobulin domain / Immunoglobulin domain / : / Immunoglobulin domain / : / Immunoglobulin subtype 2 / Immunoglobulin C-2 Type / Immunoglobulin V-Type / Immunoglobulin V-set domain / Immunoglobulin V-set domain / Immunoglobulin subtype / Immunoglobulin / Immunoglobulin/major histocompatibility complex, conserved site / Immunoglobulins and major histocompatibility complex proteins signature. / Immunoglobulin C-Type / Immunoglobulin C1-set / Immunoglobulin C1-set domain / Ig-like domain profile. / Immunoglobulin-like domain / Immunoglobulin-like domain superfamily / Immunoglobulins / Immunoglobulin-like fold / Immunoglobulin-like / Sandwich / Mainly Beta
Similarity search - Domain/homology
B-cell receptor CD22 / Ig-like domain-containing protein / IgG H chain
Similarity search - Component
Biological speciesHomo sapiens (human)
Mus musculus (house mouse)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.1 Å
AuthorsSicard, T. / Ereno-Orbea, J. / Julien, J.P.
Funding support Canada, 3items
OrganizationGrant numberCountry
Canadian Institutes of Health Research (CIHR)PJT-148811 Canada
Canadian Institutes of Health Research (CIHR)BPF-144483 Canada
Canadian Institutes of Health Research (CIHR)Canada Graduate Scholarship Masters Award Canada
CitationJournal: Nat Commun / Year: 2017
Title: Molecular basis of human CD22 function and therapeutic targeting.
Authors: June Ereño-Orbea / Taylor Sicard / Hong Cui / Mohammad T Mazhab-Jafari / Samir Benlekbir / Alba Guarné / John L Rubinstein / Jean-Philippe Julien /
Abstract: CD22 maintains a baseline level of B-cell inhibition to keep humoral immunity in check. As a B-cell-restricted antigen, CD22 is targeted in therapies against dysregulated B cells that cause ...CD22 maintains a baseline level of B-cell inhibition to keep humoral immunity in check. As a B-cell-restricted antigen, CD22 is targeted in therapies against dysregulated B cells that cause autoimmune diseases and blood cancers. Here we report the crystal structure of human CD22 at 2.1 Å resolution, which reveals that specificity for α2-6 sialic acid ligands is dictated by a pre-formed β-hairpin as a unique mode of recognition across sialic acid-binding immunoglobulin-type lectins. The CD22 ectodomain adopts an extended conformation that facilitates concomitant CD22 nanocluster formation on B cells and binding to trans ligands to avert autoimmunity in mammals. We structurally delineate the CD22 site targeted by the therapeutic antibody epratuzumab at 3.1 Å resolution and determine a critical role for CD22 N-linked glycosylation in antibody engagement. Our studies provide molecular insights into mechanisms governing B-cell inhibition and valuable clues for the design of immune modulators in B-cell dysfunction.The B-cell-specific co-receptor CD22 is a therapeutic target for depleting dysregulated B cells. Here the authors structurally characterize the ectodomain of CD22 and present its crystal structure with the bound therapeutic antibody epratuzumab, which gives insights into the mechanism of inhibition of B-cell activation.
History
DepositionApr 24, 2017Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 11, 2017Provider: repository / Type: Initial release
Revision 1.1Dec 6, 2017Group: Database references / Category: pdbx_database_related / Item: _pdbx_database_related.content_type
Revision 1.2Nov 27, 2019Group: Database references / Category: pdbx_database_related / Item: _pdbx_database_related.content_type
Revision 1.3Jan 8, 2020Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 2.0Jul 29, 2020Group: Atomic model / Data collection ...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 / struct_asym / struct_conn / struct_site / struct_site_gen
Item: _atom_site.B_iso_or_equiv / _atom_site.Cartn_x ..._atom_site.B_iso_or_equiv / _atom_site.Cartn_x / _atom_site.Cartn_y / _atom_site.Cartn_z / _atom_site.auth_asym_id / _atom_site.auth_atom_id / _atom_site.auth_comp_id / _atom_site.auth_seq_id / _atom_site.label_asym_id / _atom_site.label_atom_id / _atom_site.label_comp_id / _atom_site.label_entity_id / _atom_site.type_symbol / _chem_comp.name / _chem_comp.type / _pdbx_struct_assembly_gen.asym_id_list / _struct_conn.pdbx_role / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id
Description: Carbohydrate remediation / Provider: repository / Type: Remediation

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
Q: B-cell receptor CD22
R: B-cell receptor CD22
S: B-cell receptor CD22
T: B-cell receptor CD22
A: Epratuzumab Fab Heavy Chain
C: Epratuzumab Fab Heavy Chain
E: Epratuzumab Fab Heavy Chain
H: Epratuzumab Fab Heavy Chain
B: Epratuzumab Fab Light Chain
D: Epratuzumab Fab Light Chain
F: Epratuzumab Fab Light Chain
L: Epratuzumab Fab Light Chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)345,47321
Polymers340,68812
Non-polymers4,7849
Water00
1
Q: B-cell receptor CD22
H: Epratuzumab Fab Heavy Chain
L: Epratuzumab Fab Light Chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)86,5075
Polymers85,1723
Non-polymers1,3352
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area7950 Å2
ΔGint-7 kcal/mol
Surface area35190 Å2
MethodPISA
2
R: B-cell receptor CD22
A: Epratuzumab Fab Heavy Chain
B: Epratuzumab Fab Light Chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)86,3455
Polymers85,1723
Non-polymers1,1732
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area7670 Å2
ΔGint-10 kcal/mol
Surface area34560 Å2
MethodPISA
3
S: B-cell receptor CD22
C: Epratuzumab Fab Heavy Chain
D: Epratuzumab Fab Light Chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)86,2756
Polymers85,1723
Non-polymers1,1033
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area7530 Å2
ΔGint-16 kcal/mol
Surface area35020 Å2
MethodPISA
4
T: B-cell receptor CD22
E: Epratuzumab Fab Heavy Chain
F: Epratuzumab Fab Light Chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)86,3455
Polymers85,1723
Non-polymers1,1732
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area7780 Å2
ΔGint-10 kcal/mol
Surface area34640 Å2
MethodPISA
Unit cell
Length a, b, c (Å)87.110, 90.220, 136.632
Angle α, β, γ (deg.)70.83, 80.81, 80.99
Int Tables number1
Space group name H-MP1

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Components

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Antibody , 2 types, 8 molecules ACEHBDFL

#2: Antibody
Epratuzumab Fab Heavy Chain


Mass: 24028.684 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse), (gene. exp.) Homo sapiens (human)
Production host: Homo sapiens (human) / References: UniProt: S6B291
#3: Antibody
Epratuzumab Fab Light Chain


Mass: 24340.979 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse), (gene. exp.) Homo sapiens (human)
Production host: Homo sapiens (human) / References: UniProt: Q8TCD0

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Protein / Non-polymers , 2 types, 5 molecules QRST

#1: Protein
B-cell receptor CD22 / B-lymphocyte cell adhesion molecule / BL-CAM / Sialic acid-binding Ig-like lectin 2 / Siglec-2 / T- ...B-lymphocyte cell adhesion molecule / BL-CAM / Sialic acid-binding Ig-like lectin 2 / Siglec-2 / T-cell surface antigen Leu-14


Mass: 36802.406 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CD22, SIGLEC2 / Production host: Homo sapiens (human) / References: UniProt: P20273
#8: Chemical ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL


Mass: 92.094 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C3H8O3

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Sugars , 4 types, 8 molecules

#4: Polysaccharide
2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}}LINUCSPDB-CARE
#5: Polysaccharide alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2- ...alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 910.823 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpa1-3[DManpa1-6]DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/3,5,4/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5][a1122h-1a_1-5]/1-1-2-3-3/a4-b1_b4-c1_c3-d1_c6-e1WURCSPDB2Glycan 1.1.0
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(3+1)][a-D-Manp]{}[(6+1)][a-D-Manp]{}}}}}LINUCSPDB-CARE
#6: Polysaccharide alpha-D-mannopyranose-(1-3)-beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1- ...alpha-D-mannopyranose-(1-3)-beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 748.682 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpa1-3DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/3,4,3/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5][a1122h-1a_1-5]/1-1-2-3/a4-b1_b4-c1_c3-d1WURCSPDB2Glycan 1.1.0
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(3+1)][a-D-Manp]{}}}}}LINUCSPDB-CARE
#7: Polysaccharide beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta- ...beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 586.542 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/2,3,2/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5]/1-1-2/a4-b1_b4-c1WURCSPDB2Glycan 1.1.0
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{}}}}LINUCSPDB-CARE

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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: 2.99 Å3/Da / Density % sol: 58.84 %
Crystal growTemperature: 293 K / Method: vapor diffusion / Details: 20 mM Tris pH 8.0 and 150 mM NaCl, 5mg/mL

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: CLSI / Beamline: 08ID-1 / Wavelength: 0.97949 Å
DetectorType: RAYONIX MX-300 / Detector: CCD / Date: Dec 1, 2016
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97949 Å / Relative weight: 1
ReflectionResolution: 3.1→39.345 Å / Num. obs: 65863 / % possible obs: 94.2 % / Redundancy: 1.9 % / Biso Wilson estimate: 65.68 Å2 / CC1/2: 0.987 / Rmerge(I) obs: 0.09 / Rpim(I) all: 0.09 / Net I/σ(I): 8.2
Reflection shellResolution: 3.1→3.2 Å / Redundancy: 1.9 % / Rmerge(I) obs: 0.39 / Mean I/σ(I) obs: 1.9 / CC1/2: 0.74 / Rpim(I) all: 0.39 / % possible all: 96.7

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Processing

Software
NameVersionClassification
PHENIX(1.12_2829: ???)refinement
XPREP2015/1data scaling
PDB_EXTRACT3.22data extraction
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 3.1→39.345 Å / SU ML: 0.43 / Cross valid method: FREE R-VALUE / σ(F): 1.96 / Phase error: 32.03 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.2953 2000 3.04 %
Rwork0.2801 --
obs0.2806 65847 94.2 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 3.1→39.345 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms22594 0 318 0 22912
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00423512
X-RAY DIFFRACTIONf_angle_d0.85332007
X-RAY DIFFRACTIONf_dihedral_angle_d14.83414083
X-RAY DIFFRACTIONf_chiral_restr0.0493634
X-RAY DIFFRACTIONf_plane_restr0.0054019
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.1-3.17750.33851460.35784671X-RAY DIFFRACTION97
3.1775-3.26340.36621470.35034692X-RAY DIFFRACTION97
3.2634-3.35930.29831460.3344660X-RAY DIFFRACTION96
3.3593-3.46770.34141460.32314644X-RAY DIFFRACTION96
3.4677-3.59160.33031440.31844631X-RAY DIFFRACTION96
3.5916-3.73530.29991440.31364586X-RAY DIFFRACTION95
3.7353-3.90510.3581440.30374612X-RAY DIFFRACTION95
3.9051-4.11080.32531420.28844512X-RAY DIFFRACTION93
4.1108-4.3680.32681400.26354498X-RAY DIFFRACTION93
4.368-4.70480.25181400.25264440X-RAY DIFFRACTION92
4.7048-5.17730.27341400.2444480X-RAY DIFFRACTION92
5.1773-5.92430.25521400.25384478X-RAY DIFFRACTION93
5.9243-7.45570.30571430.27614526X-RAY DIFFRACTION93
7.4557-39.34830.22871380.23794417X-RAY DIFFRACTION91

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