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- EMDB-60766: HL-type bispecific diabody Ex3 composed of 528 and OKT3 Fvs in te... -

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Basic information

Entry
Database: EMDB / ID: EMD-60766
TitleHL-type bispecific diabody Ex3 composed of 528 and OKT3 Fvs in ternary complex with sEGFR and CD3gamma-epsilon (middle conformation)
Map datasharpened map
Sample
  • Complex: HL-type bispecific antibody Ex3 composed of 528 and OKT3 Fvs in ternary complex with sEGFR and CD3gamma-epsilon
    • Complex: Soluble Epidermal Growth Factor Receptor (sEGFR)
      • Protein or peptide: Epidermal growth factor receptor
    • Complex: HL-type bispecific diabody Ex3
      • Protein or peptide: HL-type bispecific diabody Ex3
    • Complex: CD3 gamma-epsilon
      • Protein or peptide: T-cell surface glycoprotein CD3 gamma chain,T-cell surface glycoprotein CD3 epsilon chain
Keywordsbispecific antibody / diabody / EGFR / CD3 / ternary complex / HL / Ex3 / 528 / OKT3 / ANTITUMOR PROTEIN / ANTITUMOR PROTEIN-IMMUNE SYSTEM complex
Function / homology
Function and homology information


regulation of lymphocyte apoptotic process / gamma-delta T cell receptor complex / T cell anergy / positive regulation of cell-cell adhesion mediated by integrin / positive regulation of T cell anergy / gamma-delta T cell activation / CD4-positive, alpha-beta T cell proliferation / negative thymic T cell selection / positive regulation of CD4-positive, alpha-beta T cell proliferation / alpha-beta T cell receptor complex ...regulation of lymphocyte apoptotic process / gamma-delta T cell receptor complex / T cell anergy / positive regulation of cell-cell adhesion mediated by integrin / positive regulation of T cell anergy / gamma-delta T cell activation / CD4-positive, alpha-beta T cell proliferation / negative thymic T cell selection / positive regulation of CD4-positive, alpha-beta T cell proliferation / alpha-beta T cell receptor complex / positive thymic T cell selection / signal complex assembly / positive regulation of protein kinase C activity / multivesicular body, internal vesicle lumen / positive regulation of prolactin secretion / negative regulation of cardiocyte differentiation / response to hydroxyisoflavone / diterpenoid metabolic process / Shc-EGFR complex / Inhibition of Signaling by Overexpressed EGFR / ovulation cycle / EGFR interacts with phospholipase C-gamma / positive regulation of mucus secretion / epidermal growth factor binding / regulation of peptidyl-tyrosine phosphorylation / response to UV-A / tongue development / positive regulation of cell-matrix adhesion / PLCG1 events in ERBB2 signaling / midgut development / T cell receptor complex / ERBB2-EGFR signaling pathway / digestive tract morphogenesis / hydrogen peroxide metabolic process / morphogenesis of an epithelial fold / PTK6 promotes HIF1A stabilization / smoothened signaling pathway / ERBB2 Activates PTK6 Signaling / Signaling by EGFR / intracellular vesicle / establishment or maintenance of cell polarity / response to cobalamin / Translocation of ZAP-70 to Immunological synapse / Phosphorylation of CD3 and TCR zeta chains / positive regulation of interleukin-4 production / negative regulation of epidermal growth factor receptor signaling pathway / eyelid development in camera-type eye / dendrite development / cerebral cortex cell migration / protein insertion into membrane / ERBB2 Regulates Cell Motility / protein tyrosine kinase activator activity / Signaling by ERBB4 / Respiratory syncytial virus (RSV) attachment and entry / alpha-beta T cell activation / PI3K events in ERBB2 signaling / Generation of second messenger molecules / negative regulation of mitotic cell cycle / FCGR activation / immunological synapse / MAP kinase kinase kinase activity / Co-inhibition by PD-1 / hair follicle development / Estrogen-dependent nuclear events downstream of ESR-membrane signaling / positive regulation of G1/S transition of mitotic cell cycle / GAB1 signalosome / Role of phospholipids in phagocytosis / positive regulation of bone resorption / embryonic placenta development / T cell receptor binding / positive regulation of phosphorylation / salivary gland morphogenesis / peptidyl-tyrosine autophosphorylation / positive regulation of T cell proliferation / positive regulation of peptidyl-serine phosphorylation / positive regulation of glial cell proliferation / positive regulation of vasoconstriction / T cell costimulation / Signaling by ERBB2 / transmembrane receptor protein tyrosine kinase activity / GRB2 events in EGFR signaling / TFAP2 (AP-2) family regulates transcription of growth factors and their receptors / SHC1 events in EGFR signaling / EGFR Transactivation by Gastrin / GRB2 events in ERBB2 signaling / positive regulation of interleukin-2 production / cerebellum development / SHC1 events in ERBB2 signaling / cellular response to epidermal growth factor stimulus / cellular response to dexamethasone stimulus / positive regulation of synaptic transmission, glutamatergic / FCGR3A-mediated IL10 synthesis / positive regulation of calcium-mediated signaling / ossification / positive regulation of DNA repair / T cell activation / cell surface receptor protein tyrosine kinase signaling pathway / neuron projection morphogenesis / positive regulation of superoxide anion generation / positive regulation of epithelial cell proliferation
Similarity search - Function
CD3 gamma/delta subunit, Ig-like domain / T-cell surface glycoprotein CD3 delta chain / Ig-like domain on T-cell surface glycoprotein CD3 epsilon chain / CD3 protein, epsilon/gamma/delta subunit / Immunoreceptor tyrosine-based activation motif / Phosphorylated immunoreceptor signalling ITAM / ITAM motif mammalian type profile. / Immunoreceptor tyrosine-based activation motif / : / Epidermal growth factor receptor transmembrane-juxtamembrane segment ...CD3 gamma/delta subunit, Ig-like domain / T-cell surface glycoprotein CD3 delta chain / Ig-like domain on T-cell surface glycoprotein CD3 epsilon chain / CD3 protein, epsilon/gamma/delta subunit / Immunoreceptor tyrosine-based activation motif / Phosphorylated immunoreceptor signalling ITAM / ITAM motif mammalian type profile. / Immunoreceptor tyrosine-based activation motif / : / Epidermal growth factor receptor transmembrane-juxtamembrane segment / Tyrosine protein kinase, EGF/ERB/XmrK receptor / Growth factor receptor domain 4 / Growth factor receptor domain IV / Receptor L-domain / Furin-like cysteine-rich domain / Receptor L-domain superfamily / Furin-like cysteine rich region / Receptor L domain / Furin-like repeat / Furin-like repeats / Growth factor receptor cysteine-rich domain superfamily / : / Immunoglobulin subtype 2 / Immunoglobulin C-2 Type / Tyrosine-protein kinase, catalytic domain / Tyrosine kinase, catalytic domain / Tyrosine protein kinases specific active-site signature. / Tyrosine-protein kinase, active site / Serine-threonine/tyrosine-protein kinase, catalytic domain / Protein tyrosine and serine/threonine kinase / Immunoglobulin-like domain superfamily / Protein kinase, ATP binding site / Protein kinases ATP-binding region signature. / Immunoglobulin-like fold / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
Epidermal growth factor receptor / T-cell surface glycoprotein CD3 epsilon chain / T-cell surface glycoprotein CD3 gamma chain
Similarity search - Component
Biological speciessynthetic construct (others) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.64 Å
AuthorsSato K / Uehara S / Tsugita A / Matsui T / Asano R / Makabe K / Yokoyama T / Tanaka Y
Funding support Japan, 1 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS) Japan
CitationJournal: Cell Rep / Year: 2025
Title: Bispecific antibody-antigen complex structures reveal activity enhancement by domain rearrangement
Authors: Sato K / Uehara S / Tsugita A / Ishii M / Ishiyama S / Maejima A / Nakahara I / Nazuka M / Matsui T / Christos G / Yokoyama T / Kumagai I / Makabe K / Asano R / Tanaka Y
History
DepositionJul 10, 2024-
Header (metadata) releaseMay 28, 2025-
Map releaseMay 28, 2025-
UpdateJun 18, 2025-
Current statusJun 18, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_60766.png

Downloads & links

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Map

FileDownload / File: emd_60766.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationsharpened map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.79 Å/pix.
x 384 pix.
= 302.592 Å		0.79 Å/pix.
x 384 pix.
= 302.592 Å		0.79 Å/pix.
x 384 pix.
= 302.592 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.788 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0165
Minimum - Maximum-0.09545655 - 0.16656104
Average (Standard dev.)0.00009967814 (±0.0026073325)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 302.59198 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: half map B

Fileemd_60766_half_map_1.map
Annotationhalf map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map A

Fileemd_60766_half_map_2.map
Annotationhalf map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : HL-type bispecific antibody Ex3 composed of 528 and OKT3 Fvs in t...

EntireName: HL-type bispecific antibody Ex3 composed of 528 and OKT3 Fvs in ternary complex with sEGFR and CD3gamma-epsilon
Components
  • Complex: HL-type bispecific antibody Ex3 composed of 528 and OKT3 Fvs in ternary complex with sEGFR and CD3gamma-epsilon
    • Complex: Soluble Epidermal Growth Factor Receptor (sEGFR)
      • Protein or peptide: Epidermal growth factor receptor
    • Complex: HL-type bispecific diabody Ex3
      • Protein or peptide: HL-type bispecific diabody Ex3
    • Complex: CD3 gamma-epsilon
      • Protein or peptide: T-cell surface glycoprotein CD3 gamma chain,T-cell surface glycoprotein CD3 epsilon chain

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Supramolecule #1: HL-type bispecific antibody Ex3 composed of 528 and OKT3 Fvs in t...

SupramoleculeName: HL-type bispecific antibody Ex3 composed of 528 and OKT3 Fvs in ternary complex with sEGFR and CD3gamma-epsilon
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all

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Supramolecule #2: Soluble Epidermal Growth Factor Receptor (sEGFR)

SupramoleculeName: Soluble Epidermal Growth Factor Receptor (sEGFR) / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: synthetic construct (others)

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Supramolecule #3: HL-type bispecific diabody Ex3

SupramoleculeName: HL-type bispecific diabody Ex3 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #4: CD3 gamma-epsilon

SupramoleculeName: CD3 gamma-epsilon / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #3
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Epidermal growth factor receptor

MacromoleculeName: Epidermal growth factor receptor / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: receptor protein-tyrosine kinase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 69.496062 KDa
Recombinant expressionOrganism: Cricetulus griseus (Chinese hamster)
SequenceString: LEEKKVCQGT SNKLTQLGTF EDHFLSLQRM FNNCEVVLGN LEITYVQRNY DLSFLKTIQE VAGYVLIALN TVERIPLENL QIIRGNMYY ENSYALAVLS NYDANKTGLK ELPMRNLQEI LHGAVRFSNN PALCNVESIQ WRDIVSSDFL SNMSMDFQNH L GSCQKCDP ...String:
LEEKKVCQGT SNKLTQLGTF EDHFLSLQRM FNNCEVVLGN LEITYVQRNY DLSFLKTIQE VAGYVLIALN TVERIPLENL QIIRGNMYY ENSYALAVLS NYDANKTGLK ELPMRNLQEI LHGAVRFSNN PALCNVESIQ WRDIVSSDFL SNMSMDFQNH L GSCQKCDP SCPNGSCWGA GEENCQKLTK IICAQQCSGR CRGKSPSDCC HNQCAAGCTG PRESDCLVCR KFRDEATCKD TC PPLMLYN PTTYQMDVNP EGKYSFGATC VKKCPRNYVV TDHGSCVRAC GADSYEMEED GVRKCKKCEG PCRKVCNGIG IGE FKDSLS INATNIKHFK NCTSISGDLH ILPVAFRGDS FTHTPPLDPQ ELDILKTVKE ITGFLLIQAW PENRTDLHAF ENLE IIRGR TKQHGQFSLA VVSLNITSLG LRSLKEISDG DVIISGNKNL CYANTINWKK LFGTSGQKTK IISNRGENSC KATGQ VCHA LCSPEGCWGP EPRDCVSCRN VSRGRECVDK CNLLEGEPRE FVENSECIQC HPECLPQAMN ITCTGRGPDN CIQCAH YID GPHCVKTCPA GVMGENNTLV WKYADAGHVC HLCHPNCTYG CTGPGLEGCP TNGPKIPSHH HHHH

UniProtKB: Epidermal growth factor receptor

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Macromolecule #2: HL-type bispecific diabody Ex3

MacromoleculeName: HL-type bispecific diabody Ex3 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 56.781754 KDa
Recombinant expressionOrganism: Brevibacillus choshinensis (bacteria)
SequenceString: MAFAAQVQLV QSGGGVVQPG RSLRLSCKAS GYTFTRYTMH WVRQAPGKGL EWIGYINPSR GYTNYNQKVK DRFTISRDNS KNTAFLQMD SLRPEDTGVY FCARYYDDHY SLDYWGQGTP VTVSSAGGGG SDIVMTQSPL SLPVTPGEPA SISCRSSQNI V HNNGITYL ...String:
MAFAAQVQLV QSGGGVVQPG RSLRLSCKAS GYTFTRYTMH WVRQAPGKGL EWIGYINPSR GYTNYNQKVK DRFTISRDNS KNTAFLQMD SLRPEDTGVY FCARYYDDHY SLDYWGQGTP VTVSSAGGGG SDIVMTQSPL SLPVTPGEPA SISCRSSQNI V HNNGITYL EWYLQKPGQS PQLLIYKVSD RFSGVPDRFS GSGSGTDFTL KISRVEAEDV GVYYCFQGSH IPPTFGQGTK VE IKRAAAA GGGGSGGGGS GGGGSGGGGS QVQLVQSGAE VKKPGASVKV SCKASGYTFT SYWMHWVRQA PGQGLEWMGN IWP GSGGTN YAEKFKNRVT MTRDTSISTA YMELSRLRSD DTAVYYCARS GGPYFFDYWG QGTLVTVSSA GGGGSDIQMT QSPS SLSAS VGDRVTITCS ASSSVSYMNW YQQTPGKAPK RWIYDTSKLA SGVPSRFSGS GSGTDYTFTI SSLQPEDIAT YYCQQ WSSN PFTFGQGTKL QITRAAAAEQ KLISEEDLNL GGGMRGSHHH HHH

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Macromolecule #3: T-cell surface glycoprotein CD3 gamma chain,T-cell surface glycop...

MacromoleculeName: T-cell surface glycoprotein CD3 gamma chain,T-cell surface glycoprotein CD3 epsilon chain
type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 22.940545 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MQSIKGNHLV KVYDYQEDGS VLLTCDAEAK NITWFKDGKM IGFLTEDKKK WNLGSNAKDP RGMYQCKGSQ NKSKPLQVYY RMGSADDAK KDAAKKDDAK KDDAKKDGSD GNEEMGGITQ TPYKVSISGT TVILTCPQYP GSEILWQHND KNIGGDEDDK N IGSDEDHL ...String:
MQSIKGNHLV KVYDYQEDGS VLLTCDAEAK NITWFKDGKM IGFLTEDKKK WNLGSNAKDP RGMYQCKGSQ NKSKPLQVYY RMGSADDAK KDAAKKDDAK KDDAKKDGSD GNEEMGGITQ TPYKVSISGT TVILTCPQYP GSEILWQHND KNIGGDEDDK N IGSDEDHL SLKEFSELEQ SGYYVCYPRG SKPEDANFYL YLRARV

UniProtKB: T-cell surface glycoprotein CD3 gamma chain, T-cell surface glycoprotein CD3 epsilon chain

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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

BufferpH: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeJEOL CRYO ARM 300
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.4 µm / Nominal defocus min: 0.9 µm

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.64 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 89363
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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