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- EMDB-63050: Cryo-EM structure of linker-extended biparatopic antibody BA1-GP4... -
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Open data
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Basic information
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Title | Cryo-EM structure of linker-extended biparatopic antibody BA1-GP4 in complex with TNFR2 | |||||||||||||||
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![]() | Antibody / biparatopic antibody / antagonist / IMMUNE SYSTEM | |||||||||||||||
Function / homology | ![]() 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 / negative regulation of neuroinflammatory response / TNFs bind their physiological receptors / tumor necrosis factor binding / negative regulation of cardiac muscle hypertrophy / regulation of neuroinflammatory response / positive regulation of myelination / positive regulation of membrane protein ectodomain proteolysis / regulation of myelination / Interleukin-10 signaling / regulation of T cell proliferation / positive regulation of oligodendrocyte differentiation / specific granule membrane / extrinsic apoptotic signaling pathway / TNFR2 non-canonical NF-kB pathway / tumor necrosis factor-mediated signaling pathway / intrinsic apoptotic signaling pathway in response to DNA damage / cellular response to lipopolysaccharide / Interleukin-4 and Interleukin-13 signaling / inflammatory response / membrane raft / ubiquitin protein ligase binding / Neutrophil degranulation / extracellular region / membrane / plasma membrane Similarity search - Function | |||||||||||||||
Biological species | ![]() ![]() ![]() | |||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.73 Å | |||||||||||||||
![]() | Otsuki T / Matsumoto S / Fujita J / Miyata T / Namba K / Kanada R / Okuno Y / Kamada H / Ohno H / Akiba H | |||||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Conversion of an agonistic anti-TNFR2 biparatopic antibody into an antagonist by insertion of peptide linkers into the hinge region. Authors: Takuya Otsuki / Shigeyuki Matsumoto / Junso Fujita / Tomoko Miyata / Keiichi Namba / Ryo Kanada / Yasushi Okuno / Haruhiko Kamada / Hiroaki Ohno / Hiroki Akiba / ![]() Abstract: Biparatopic antibodies (BpAbs) bind two different antigen epitopes to form characteristic immunocomplexes. Many BpAbs have been developed for enhanced cross-linking to induce signal transduction or ...Biparatopic antibodies (BpAbs) bind two different antigen epitopes to form characteristic immunocomplexes. Many BpAbs have been developed for enhanced cross-linking to induce signal transduction or cell internalization, whereas few were reported with smaller immunocomplexes to suppress unwanted signaling. Here, we developed a strategy to induce 1:1 immunocomplex formation to maximize antagonistic function. Various peptide linkers were introduced into the hinge regions of IgG-like agonist BpAbs against tumor necrosis factor receptor 2. Loss of crosslinking activity was observed for one BpAb, allowing the conversion of its function from an agonist to an antagonist. However, cross-linking activity was retained for another agonist BpAb, which binds to a different epitope pair. In a combined analysis of cryo-electron microscopy and coarse-grained molecular dynamics simulations, effect of epitope combination on the stability of 1:1 complexes was observed. These results lead to an understanding of the mechanism and design of BpAbs to adopt a 1:1-binding mode. | |||||||||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 51.7 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 24.7 KB 24.7 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 9.9 KB | Display | ![]() |
Images | ![]() | 47 KB | ||
Masks | ![]() | 103 MB | ![]() | |
Filedesc metadata | ![]() | 7.2 KB | ||
Others | ![]() ![]() | 95.5 MB 95.5 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 851.7 KB | Display | ![]() |
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Full document | ![]() | 851.3 KB | Display | |
Data in XML | ![]() | 18.2 KB | Display | |
Data in CIF | ![]() | 23.5 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 9lflMC M: atomic model generated by this map C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.048 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
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Density Histograms |
-Half map: #2
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Projections & Slices |
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Density Histograms |
-Half map: #1
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Density Histograms |
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Sample components
-Entire : Linker-extended biparatopic antibody BA1-GP4 in complex with TNFR2
Entire | Name: Linker-extended biparatopic antibody BA1-GP4 in complex with TNFR2 |
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Components |
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-Supramolecule #1: Linker-extended biparatopic antibody BA1-GP4 in complex with TNFR2
Supramolecule | Name: Linker-extended biparatopic antibody BA1-GP4 in complex with TNFR2 type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 200 KDa |
-Macromolecule #1: Tumor necrosis factor receptor superfamily member 1B
Macromolecule | Name: Tumor necrosis factor receptor superfamily member 1B / type: protein_or_peptide / ID: 1 Details: 1-178, Tumor necrosis factor receptor 1B, Homo sapiens; 179-543, Maltose/maltodextrin-binding periplasmic protein - Escherichia coli K-12; 545-550, hexahistidine tag Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 16.747895 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: TPYAPEPGST CRLREYYDQT AQMCCSKCSP GQHAKVFCTK TSDTVCDSCE DSTYTQLWNW VPECLSCGSR CSSDQVETQA CTREQNRIC TCRPGWYCAL SKQEGCRLCA PLRKCRPGFG VARPGTETSD VVCKPCAPGT FSNTTSSTDI CRP UniProtKB: Tumor necrosis factor receptor superfamily member 1B |
-Macromolecule #2: TR92 heavy chain
Macromolecule | Name: TR92 heavy chain / type: protein_or_peptide / ID: 2 / Details: TR92 antibody heavy chain / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 23.777564 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: 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 PKS |
-Macromolecule #3: TR92 light chain
Macromolecule | Name: TR92 light chain / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 23.242812 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: IVMTQSHKFM STSVGDRVSI TCKASQDVST AVAWYQQKPG QSPKLLIYWT STRHTGVPDR FTGSGSGTDY TLTISSVQAE DLALYYCQH HYSTPYTFGG GTKLEIQRTV AAPSVFIFPP SDEQLKSGTA SVVCLLNNFY PREAKVQWKV DNALQSGNSQ E SVTEQDSK ...String: IVMTQSHKFM STSVGDRVSI TCKASQDVST AVAWYQQKPG QSPKLLIYWT STRHTGVPDR FTGSGSGTDY TLTISSVQAE DLALYYCQH HYSTPYTFGG GTKLEIQRTV AAPSVFIFPP SDEQLKSGTA SVVCLLNNFY PREAKVQWKV DNALQSGNSQ E SVTEQDSK DSTYSLSSTL TLSKADYEKH KVYACEVTHQ GLSSPVTKSF NRG |
-Macromolecule #4: TR96 heavy chain
Macromolecule | Name: TR96 heavy chain / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 24.093518 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: EVQLQQSGAE LVKPGASVKL SCTPSGFNIK DTYMHWVKQR PEQGLEWIGR IDPANGYTEY DPKFQDKATI TADTSSNTAY LQLSSLTSE DTAVYYCADT QLYYWGQGTT LTVSSASVAA PSVFIFPPSD EQLKSGTASV VCLLNNFYPR EAKVQWKVDN A LQSGNSQE ...String: EVQLQQSGAE LVKPGASVKL SCTPSGFNIK DTYMHWVKQR PEQGLEWIGR IDPANGYTEY DPKFQDKATI TADTSSNTAY LQLSSLTSE DTAVYYCADT QLYYWGQGTT LTVSSASVAA PSVFIFPPSD EQLKSGTASV VCLLNNFYPR EAKVQWKVDN A LQSGNSQE SVTEQDSKDS TYSLSSTLTL SKADYEKHKV YACEVTHQGL SSPVTKSFNR G |
-Macromolecule #5: TR96 light chain
Macromolecule | Name: TR96 light chain / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 22.055656 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: QIVLTQSPAI MSASLGERVT MTCTASSSVS STYLHWYQQK PGSSPKLWIY STSNLASGVP ARFSGSGSGT SYSLTISNME AEDAATYYC HQYHRSPLTF GAGTKLELKS SASTKGPSVF PLAPSSKSTS GGTAALGCLV KDYFPEPVTV SWNSGALTSG V HTFPAVLQ ...String: QIVLTQSPAI MSASLGERVT MTCTASSSVS STYLHWYQQK PGSSPKLWIY STSNLASGVP ARFSGSGSGT SYSLTISNME AEDAATYYC HQYHRSPLTF GAGTKLELKS SASTKGPSVF PLAPSSKSTS GGTAALGCLV KDYFPEPVTV SWNSGALTSG V HTFPAVLQ SSGLYSLSSV VTVPSSSLGT QTYICNVNHK PSNTKVDKKV EP |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.71 mg/mL |
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Buffer | pH: 7.5 |
Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 20 sec. |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV |
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Electron microscopy
Microscope | JEOL CRYO ARM 300 |
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Specialist optics | Energy filter - Name: In-column Omega Filter / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4098 pixel / Number grids imaged: 1 / Number real images: 5508 / Average exposure time: 4.87 sec. / Average electron dose: 80.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 60000 |
Sample stage | Specimen holder model: JEOL CRYOSPECPORTER / Cooling holder cryogen: NITROGEN |