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- EMDB-34871: Cryo-EM structure of biparatopic antibody Bp109-92 in complex wit... -

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Entry
Database: EMDB / ID: EMD-34871
TitleCryo-EM structure of biparatopic antibody Bp109-92 in complex with TNFR2
Map data
Sample
  • Complex: Cryo-EM structure of biparatopic antibody Bp109-92 in complex with TNFR2
    • Protein or peptide: Tumor necrosis factor receptor superfamily member 1B,Maltose/maltodextrin-binding periplasmic protein
    • Protein or peptide: TR109 heavy chain
    • Protein or peptide: TR109 light chain
    • Protein or peptide: TR92 heavy chain
    • Protein or peptide: TR92 light chain
KeywordsTNFR2 / biparatopic antibody / antagonist / IMMUNE SYSTEM
Function / homology
Function and homology information


glial cell-neuron signaling / regulation of cytokine production involved in immune response / tumor necrosis factor receptor superfamily complex / pulmonary valve development / RNA destabilization / aortic valve development / tumor necrosis factor receptor activity / negative regulation of extracellular matrix constituent secretion / positive regulation of apoptotic process involved in morphogenesis / regulation of T cell cytokine production ...glial cell-neuron signaling / regulation of cytokine production involved in immune response / tumor necrosis factor receptor superfamily complex / pulmonary valve development / RNA destabilization / aortic valve development / tumor necrosis factor receptor activity / negative regulation of extracellular matrix constituent secretion / positive regulation of apoptotic process involved in morphogenesis / regulation of T cell cytokine production / varicosity / negative regulation of neuroinflammatory response / tumor necrosis factor binding / TNFs bind their physiological receptors / negative regulation of cardiac muscle hypertrophy / positive regulation of myelination / regulation of neuroinflammatory response / positive regulation of membrane protein ectodomain proteolysis / regulation of myelination / positive regulation of oligodendrocyte differentiation / detection of maltose stimulus / maltose binding / maltose transport complex / regulation of T cell proliferation / maltose transport / maltodextrin transmembrane transport / Interleukin-10 signaling / carbohydrate transmembrane transporter activity / ATP-binding cassette (ABC) transporter complex, substrate-binding subunit-containing / carbohydrate transport / specific granule membrane / extrinsic apoptotic signaling pathway / tumor necrosis factor-mediated signaling pathway / ATP-binding cassette (ABC) transporter complex / cell chemotaxis / TNFR2 non-canonical NF-kB pathway / cellular response to growth factor stimulus / intrinsic apoptotic signaling pathway in response to DNA damage / outer membrane-bounded periplasmic space / Interleukin-4 and Interleukin-13 signaling / cellular response to lipopolysaccharide / periplasmic space / immune response / inflammatory response / membrane raft / neuronal cell body / DNA damage response / ubiquitin protein ligase binding / Neutrophil degranulation / perinuclear region of cytoplasm / extracellular region / membrane / plasma membrane
Similarity search - Function
Tumour necrosis factor receptor 1B / Tumor necrosis factor receptor 1B, N-terminal / TNFR/NGFR family cysteine-rich region domain profile. / TNFR/NGFR cysteine-rich region / TNFR/NGFR family cysteine-rich region signature. / Tumor necrosis factor receptor / nerve growth factor receptor repeats. / TNFR/NGFR cysteine-rich region / Maltose/Cyclodextrin ABC transporter, substrate-binding protein / Solute-binding family 1, conserved site / Bacterial extracellular solute-binding proteins, family 1 signature. ...Tumour necrosis factor receptor 1B / Tumor necrosis factor receptor 1B, N-terminal / TNFR/NGFR family cysteine-rich region domain profile. / TNFR/NGFR cysteine-rich region / TNFR/NGFR family cysteine-rich region signature. / Tumor necrosis factor receptor / nerve growth factor receptor repeats. / TNFR/NGFR cysteine-rich region / Maltose/Cyclodextrin ABC transporter, substrate-binding protein / Solute-binding family 1, conserved site / Bacterial extracellular solute-binding proteins, family 1 signature. / Bacterial extracellular solute-binding protein / Bacterial extracellular solute-binding protein
Similarity search - Domain/homology
Maltose/maltodextrin-binding periplasmic protein / Tumor necrosis factor receptor superfamily member 1B
Similarity search - Component
Biological speciesHomo sapiens (human) / Escherichia coli (strain K12) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.63 Å
AuthorsAkiba H / Fujita J / Ise T / Nishiyama K / Miyata T / Kato T / Namba K / Ohno H / Kamada H / Nagata S / Tsumoto K
Funding support Japan, 10 items
OrganizationGrant numberCountry
Japan Agency for Medical Research and Development (AMED)JP22ak0101099 Japan
Japan Agency for Medical Research and Development (AMED)JP21am0101117 Japan
Japan Agency for Medical Research and Development (AMED)JP22ama121003 Japan
Japan Agency for Medical Research and Development (AMED)JP17pc0101020 Japan
Japan Science and TechnologyJPMJOP1861 Japan
Japan Society for the Promotion of Science (JSPS)JP21K06453 Japan
Japan Society for the Promotion of Science (JSPS)JP20K22630
Kyoto University Foundation Japan
Takeda Science Foundation Japan
JEOL YOKOGUSHI Research Alliance Laboratories of Osaka University Japan
CitationJournal: Commun Biol / Year: 2023
Title: Development of a 1:1-binding biparatopic anti-TNFR2 antagonist by reducing signaling activity through epitope selection.
Authors: Hiroki Akiba / Junso Fujita / Tomoko Ise / Kentaro Nishiyama / Tomoko Miyata / Takayuki Kato / Keiichi Namba / Hiroaki Ohno / Haruhiko Kamada / Satoshi Nagata / Kouhei Tsumoto /
Abstract: Conventional bivalent antibodies against cell surface receptors often initiate unwanted signal transduction by crosslinking two antigen molecules. Biparatopic antibodies (BpAbs) bind to two different ...Conventional bivalent antibodies against cell surface receptors often initiate unwanted signal transduction by crosslinking two antigen molecules. Biparatopic antibodies (BpAbs) bind to two different epitopes on the same antigen, thus altering crosslinking ability. In this study, we develop BpAbs against tumor necrosis factor receptor 2 (TNFR2), which is an attractive immune checkpoint target. Using different pairs of antibody variable regions specific to topographically distinct TNFR2 epitopes, we successfully regulate the size of BpAb-TNFR2 immunocomplexes to result in controlled agonistic activities. Our series of results indicate that the relative positions of the two epitopes recognized by the BpAb are critical for controlling its signaling activity. One particular antagonist, Bp109-92, binds TNFR2 in a 1:1 manner without unwanted signal transduction, and its structural basis is determined using cryo-electron microscopy. This antagonist suppresses the proliferation of regulatory T cells expressing TNFR2. Therefore, the BpAb format would be useful in designing specific and distinct antibody functions.
History
DepositionNov 29, 2022-
Header (metadata) releaseOct 4, 2023-
Map releaseOct 4, 2023-
UpdateOct 4, 2023-
Current statusOct 4, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_34871.png

Downloads & links

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Map

FileDownload / File: emd_34871.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.0875 Å
Density
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-1.6517265 - 2.37173
Average (Standard dev.)0.00004395386 (±0.03385866)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 278.4 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_34871_msk_1.map
Projections & Slices
AxesZYX

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Half map: #2

Fileemd_34871_half_map_1.map
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Half map: #1

Fileemd_34871_half_map_2.map
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Sample components

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Entire : Cryo-EM structure of biparatopic antibody Bp109-92 in complex wit...

EntireName: Cryo-EM structure of biparatopic antibody Bp109-92 in complex with TNFR2
Components
  • Complex: Cryo-EM structure of biparatopic antibody Bp109-92 in complex with TNFR2
    • Protein or peptide: Tumor necrosis factor receptor superfamily member 1B,Maltose/maltodextrin-binding periplasmic protein
    • Protein or peptide: TR109 heavy chain
    • Protein or peptide: TR109 light chain
    • Protein or peptide: TR92 heavy chain
    • Protein or peptide: TR92 light chain

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Supramolecule #1: Cryo-EM structure of biparatopic antibody Bp109-92 in complex wit...

SupramoleculeName: Cryo-EM structure of biparatopic antibody Bp109-92 in complex with TNFR2
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Tumor necrosis factor receptor superfamily member 1B,Maltose/malt...

MacromoleculeName: Tumor necrosis factor receptor superfamily member 1B,Maltose/maltodextrin-binding periplasmic protein
type: protein_or_peptide / ID: 1
Details: fusion protein of TNFR2 from human fused with MalE from E.Coli K-12
Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (strain K12) (bacteria)
Molecular weightTheoretical: 60.447305 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: LPAQVAFTPY APEPGSTCRL REYYDQTAQM CCSKCSPGQH AKVFCTKTSD TVCDSCEDST YTQLWNWVPE CLSCGSRCSS DQVETQACT REQNRICTCR PGWYCALSKQ EGCRLCAPLR KCRPGFGVAR PGTETSDVVC KPCAPGTFSN TTSSTDICRP H QICNVVAI ...String:
LPAQVAFTPY APEPGSTCRL REYYDQTAQM CCSKCSPGQH AKVFCTKTSD TVCDSCEDST YTQLWNWVPE CLSCGSRCSS DQVETQACT REQNRICTCR PGWYCALSKQ EGCRLCAPLR KCRPGFGVAR PGTETSDVVC KPCAPGTFSN TTSSTDICRP H QICNVVAI PGNASMDAVC KIEEGKLVIW INGDKGYNGL AEVGKKFEKD TGIKVTVEHP DKLEEKFPQV AATGDGPDII FW AHDRFGG YAQSGLLAEI TPDKAFQDKL YPFTWDAVRY NGKLIAYPIA VEALSLIYNK DLLPNPPKTW EEIPALDKEL KAK GKSALM FNLQEPYFTW PLIAADGGYA FKYENGKYDI KDVGVDNAGA KAGLTFLVDL IKNKHMNADT DYSIAEAAFN KGET AMTIN GPWAWSNIDT SKVNYGVTVL PTFKGQPSKP FVGVLSAGIN AASPNKELAK EFLENYLLTD EGLEAVNKDK PLGAV ALKS YEEELVKDPR IAATMENAQK GEIMPNIPQM SAFWYAVRTA VINAASGRQT VDEALKDAQG HHHHHH

UniProtKB: Tumor necrosis factor receptor superfamily member 1B, Maltose/maltodextrin-binding periplasmic protein

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Macromolecule #2: TR109 heavy chain

MacromoleculeName: TR109 heavy chain / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 25.080273 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: QVQLKESGPG LVAPSQSLSI TCTVSGFSLT VYGVNWVRQP PGKGLEWLGM IWGDGSTAYN SALKSRLTIT KDNSKTQVFL KMNSLQTDD TARYYCARDG RRYALDYWGQ GTSVTVSSAS TKGPSVFPLA PSSKSTSGGT AALGCLVKDY FPEPVTVSWN S GALTSGVH ...String:
QVQLKESGPG LVAPSQSLSI TCTVSGFSLT VYGVNWVRQP PGKGLEWLGM IWGDGSTAYN SALKSRLTIT KDNSKTQVFL KMNSLQTDD TARYYCARDG RRYALDYWGQ GTSVTVSSAS TKGPSVFPLA PSSKSTSGGT AALGCLVKDY FPEPVTVSWN S GALTSGVH TFPAVLQSSG LYSLSSVVTV PSSSLGTQTY ICNVNHKPSN TKVDKKVEPK SCFNTHTCPP CPAPELLG

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Macromolecule #3: TR109 light chain

MacromoleculeName: TR109 light chain / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 23.852311 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: DIVLTQSPTS LAVSLGQRAT ISCRASESVD SYGDSFLHWY QQKPGQPPIL LIYRASNLDS GIPARFSGSG SRTDFTLTIN PVEADDVAT YYCQQSNEDP YTFGGGTKLE IKRTVAAPSV FIFPPSDEQL KSGTASVVCL LNNFYPREAK VQWKVDNALQ S GNSQESVT ...String:
DIVLTQSPTS LAVSLGQRAT ISCRASESVD SYGDSFLHWY QQKPGQPPIL LIYRASNLDS GIPARFSGSG SRTDFTLTIN PVEADDVAT YYCQQSNEDP YTFGGGTKLE IKRTVAAPSV FIFPPSDEQL KSGTASVVCL LNNFYPREAK VQWKVDNALQ S GNSQESVT EQDSKDSTYS LSSTLTLSKA DYEKHKVYAC EVTHQGLSSP VTKSFNRGEC

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Macromolecule #4: TR92 heavy chain

MacromoleculeName: TR92 heavy chain / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 25.543641 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: KVQLQQSGAE LVKPGASVKL SCKASGYTFT ESIIHWVKQR SGQGLEWIGW FYPGSDNINY NEKFKDKATL TADKSSSTVY MELTRLTSE DSAVYFCASH EGPYVYFDYW GQGTTLTVSS ASTKGPSVFP LAPSSKSTSG GTAALGCLVK DYFPEPVTVS W NSGALTSG ...String:
KVQLQQSGAE LVKPGASVKL SCKASGYTFT ESIIHWVKQR SGQGLEWIGW FYPGSDNINY NEKFKDKATL TADKSSSTVY MELTRLTSE DSAVYFCASH EGPYVYFDYW GQGTTLTVSS ASTKGPSVFP LAPSSKSTSG GTAALGCLVK DYFPEPVTVS W NSGALTSG VHTFPAVLQS SGLYSLSSVV TVPSSSLGTQ TYICNVNHKP SNTKVDKKVE PKSCDKTHTC PPCPAPELLG

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Macromolecule #5: TR92 light chain

MacromoleculeName: TR92 light chain / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 23.590156 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: DIVMTQSHKF MSTSVGDRVS ITCKASQDVS TAVAWYQQKP GQSPKLLIYW TSTRHTGVPD RFTGSGSGTD YTLTISSVQA EDLALYYCQ HHYSTPYTFG GGTKLEIQRT VAAPSVFIFP PSDEQLKSGT ASVVCLLNNF YPREAKVQWK VDNALQSGNS Q ESVTEQDS ...String:
DIVMTQSHKF MSTSVGDRVS ITCKASQDVS TAVAWYQQKP GQSPKLLIYW TSTRHTGVPD RFTGSGSGTD YTLTISSVQA EDLALYYCQ HHYSTPYTFG GGTKLEIQRT VAAPSVFIFP PSDEQLKSGT ASVVCLLNNF YPREAKVQWK VDNALQSGNS Q ESVTEQDS KDSTYSLSST LTLSKADYEK HKVYACEVTH QGLSSPVTKS FNRGEC

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.2 mg/mL
BufferpH: 7.4 / Details: 1x PBS
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 20 sec. / Pretreatment - Atmosphere: AIR / Details: 20 mA
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeJEOL CRYO ARM 300
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 60000
Specialist opticsEnergy filter - Name: In-column Omega Filter / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: JEOL CRYOSPECPORTER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Average exposure time: 4.9 sec. / Average electron dose: 60.0 e/Å2

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Image processing

Particle selectionNumber selected: 2218071
Startup modelType of model: NONE
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.2)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.63 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.2) / Number images used: 100391
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

3alq


Chain - Chain ID: A / Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL
Output model

PDB-8hlb:
Cryo-EM structure of biparatopic antibody Bp109-92 in complex with TNFR2

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