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- PDB-7te4: Crystal structure of Fab2 anti-GluN2B antibody -

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Basic information

Entry
Database: PDB / ID: 7te4
TitleCrystal structure of Fab2 anti-GluN2B antibody
Components
  • Fab anti-GluN2B antibody, heavy chain
  • Fab2 anti-GluN2B antibody, light chain
KeywordsIMMUNE SYSTEM / Fab fragment
Biological speciesMus musculus (house mouse)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.456 Å
AuthorsTajima, N. / Furukawa, H.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Mental Health (NIH/NIMH)MH085926 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS111745 United States
CitationJournal: Nat Commun / Year: 2022
Title: Development and characterization of functional antibodies targeting NMDA receptors.
Authors: Nami Tajima / Noriko Simorowski / Remy A Yovanno / Michael C Regan / Kevin Michalski / Ricardo Gómez / Albert Y Lau / Hiro Furukawa /
Abstract: N-methyl-D-aspartate receptors (NMDARs) are critically involved in basic brain functions and neurodegeneration as well as tumor invasiveness. Targeting specific subtypes of NMDARs with distinct ...N-methyl-D-aspartate receptors (NMDARs) are critically involved in basic brain functions and neurodegeneration as well as tumor invasiveness. Targeting specific subtypes of NMDARs with distinct activities has been considered an effective therapeutic strategy for neurological disorders and diseases. However, complete elimination of off-target effects of small chemical compounds has been challenging and thus, there is a need to explore alternative strategies for targeting NMDAR subtypes. Here we report identification of a functional antibody that specifically targets the GluN1-GluN2B NMDAR subtype and allosterically down-regulates ion channel activity as assessed by electrophysiology. Through biochemical analysis, x-ray crystallography, single-particle electron cryomicroscopy, and molecular dynamics simulations, we show that this inhibitory antibody recognizes the amino terminal domain of the GluN2B subunit and increases the population of the non-active conformational state. The current study demonstrates that antibodies may serve as specific reagents to regulate NMDAR functions for basic research and therapeutic objectives.
History
DepositionJan 4, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 2, 2022Provider: repository / Type: Initial release
Revision 1.1Oct 18, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model / struct_ncs_dom_lim
Item: _struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id ..._struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id / _struct_ncs_dom_lim.beg_label_comp_id / _struct_ncs_dom_lim.beg_label_seq_id / _struct_ncs_dom_lim.end_auth_comp_id / _struct_ncs_dom_lim.end_label_asym_id / _struct_ncs_dom_lim.end_label_comp_id / _struct_ncs_dom_lim.end_label_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
L: Fab2 anti-GluN2B antibody, light chain
H: Fab anti-GluN2B antibody, heavy chain
A: Fab2 anti-GluN2B antibody, light chain
B: Fab anti-GluN2B antibody, heavy chain


Theoretical massNumber of molelcules
Total (without water)94,9744
Polymers94,9744
Non-polymers00
Water1086
1
L: Fab2 anti-GluN2B antibody, light chain
H: Fab anti-GluN2B antibody, heavy chain


Theoretical massNumber of molelcules
Total (without water)47,4872
Polymers47,4872
Non-polymers00
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area3340 Å2
ΔGint-28 kcal/mol
Surface area18170 Å2
MethodPISA
2
A: Fab2 anti-GluN2B antibody, light chain
B: Fab anti-GluN2B antibody, heavy chain


Theoretical massNumber of molelcules
Total (without water)47,4872
Polymers47,4872
Non-polymers00
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area3200 Å2
ΔGint-26 kcal/mol
Surface area17970 Å2
MethodPISA
Unit cell
Length a, b, c (Å)54.744, 66.901, 240.417
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
11(chain B and (resid 2 through 4 or (resid 5...
21(chain H and (resid 2 through 11 or (resid 12...
12(chain A and (resid 1 through 8 or (resid 9...
22(chain L and (resid 1 through 38 or (resid 39...

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg auth comp-IDBeg label comp-IDEnd auth comp-IDEnd label comp-IDSelection detailsAuth asym-IDLabel asym-IDAuth seq-IDLabel seq-ID
111VALVALLEULEU(chain B and (resid 2 through 4 or (resid 5...BD2 - 42 - 4
121GLNGLNGLNGLN(chain B and (resid 2 through 4 or (resid 5...BD55
131VALVALPROPRO(chain B and (resid 2 through 4 or (resid 5...BD2 - 2212 - 220
211VALVALLEULEU(chain H and (resid 2 through 11 or (resid 12...HB2 - 112 - 11
221VALVALGLNGLN(chain H and (resid 2 through 11 or (resid 12...HB12 - 1312 - 13
231VALVALARGARG(chain H and (resid 2 through 11 or (resid 12...HB2 - 2222 - 221
241VALVALARGARG(chain H and (resid 2 through 11 or (resid 12...HB2 - 2222 - 221
251VALVALARGARG(chain H and (resid 2 through 11 or (resid 12...HB2 - 2222 - 221
261VALVALARGARG(chain H and (resid 2 through 11 or (resid 12...HB2 - 2222 - 221
112ASPASPPROPRO(chain A and (resid 1 through 8 or (resid 9...AC1 - 81 - 8
122SERSERSERSER(chain A and (resid 1 through 8 or (resid 9...AC99
132ASPASPASNASN(chain A and (resid 1 through 8 or (resid 9...AC1 - 2111 - 211
142ASPASPASNASN(chain A and (resid 1 through 8 or (resid 9...AC1 - 2111 - 211
152ASPASPASNASN(chain A and (resid 1 through 8 or (resid 9...AC1 - 2111 - 211
162ASPASPASNASN(chain A and (resid 1 through 8 or (resid 9...AC1 - 2111 - 211
212ASPASPHISHIS(chain L and (resid 1 through 38 or (resid 39...LA1 - 381 - 38
222LYSLYSLYSLYS(chain L and (resid 1 through 38 or (resid 39...LA3939
232ASPASPASNASN(chain L and (resid 1 through 38 or (resid 39...LA1 - 2111 - 211

NCS ensembles :
ID
1
2

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Components

#1: Antibody Fab2 anti-GluN2B antibody, light chain


Mass: 23572.031 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)
#2: Antibody Fab anti-GluN2B antibody, heavy chain


Mass: 23914.781 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)
#3: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 6 / 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: 2.31 Å3/Da / Density % sol: 46.83 %
Crystal growTemperature: 290 K / Method: vapor diffusion, hanging drop / Details: 20% w/w PEG3350, 8% Tacsimate

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 23-ID-B / Wavelength: 1.1 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Feb 17, 2016
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.1 Å / Relative weight: 1
ReflectionResolution: 2.456→30 Å / Num. obs: 33229 / % possible obs: 99.9 % / Redundancy: 7.8 % / Biso Wilson estimate: 49.33 Å2 / Rmerge(I) obs: 0.182 / Net I/σ(I): 10.3
Reflection shellResolution: 2.456→2.499 Å / Rmerge(I) obs: 1.168 / Num. unique obs: 2877

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Processing

Software
NameVersionClassification
HKL-2000data scaling
PHENIX1.13_2998refinement
PDB_EXTRACT3.27data extraction
HKL-2000data reduction
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB entry 5B3J

5b3j


Resolution: 2.456→29.229 Å / SU ML: 0.4 / Cross valid method: THROUGHOUT / σ(F): 1.34 / Phase error: 33.94 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.2966 1663 5.02 %
Rwork0.2609 31490 -
obs0.2627 33153 99.89 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 59.25 Å2 / Biso mean: 46.4238 Å2 / Biso min: 36.59 Å2
Refinement stepCycle: final / Resolution: 2.456→29.229 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms6017 0 0 6 6023
Biso mean---41.51 -
Num. residues----824
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDNumberRefine-IDRmsType
11B1735X-RAY DIFFRACTION12.808TORSIONAL
12H1735X-RAY DIFFRACTION12.808TORSIONAL
21A1578X-RAY DIFFRACTION12.808TORSIONAL
22L1578X-RAY DIFFRACTION12.808TORSIONAL
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / % reflection obs: 100 %

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection Rwork
2.4563-2.52860.38841420.36482551
2.5286-2.61010.40251180.3452620
2.6101-2.70330.33781520.3352562
2.7033-2.81150.38681150.32272586
2.8115-2.93930.3411170.32292605
2.9393-3.09420.37591300.31442607
3.0942-3.28780.33071520.29992593
3.2878-3.54120.3191510.27312597
3.5412-3.89690.31781460.25222639
3.8969-4.4590.24681370.21612636
4.459-5.61140.20821520.21072680
5.6114-29.2290.3031510.23222814

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