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- EMDB-20055: Cryo-EM structure of Her2 extracellular domain-Trastuzumab Fab-Pe... -

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Entry
Database: EMDB / ID: EMD-20055
TitleCryo-EM structure of Her2 extracellular domain-Trastuzumab Fab-Pertuzumab Fab complex
Map data
SampleHer2 extracellular domain-Trastuzumab Fab-Pertuzumab Fab complex:
Human HER2 extracellular domain / Pertuzumab Fab / Trastuzumab Fab / Receptor tyrosine-protein kinase erbB-2 / (Pertuzumab FAB ...) x 2 / (Trastuzumab FAB ...) x 2 / (ligand) x 3
Function / homology
Function and homology information


Classical antibody-mediated complement activation / Role of phospholipids in phagocytosis / Regulation of actin dynamics for phagocytic cup formation / FCGR activation / Cell surface interactions at the vascular wall / Immunoregulatory interactions between a Lymphoid and a non-Lymphoid cell / PI3K events in ERBB2 signaling / GRB2 events in ERBB2 signaling / Constitutive Signaling by Aberrant PI3K in Cancer / Initial triggering of complement ...Classical antibody-mediated complement activation / Role of phospholipids in phagocytosis / Regulation of actin dynamics for phagocytic cup formation / FCGR activation / Cell surface interactions at the vascular wall / Immunoregulatory interactions between a Lymphoid and a non-Lymphoid cell / PI3K events in ERBB2 signaling / GRB2 events in ERBB2 signaling / Constitutive Signaling by Aberrant PI3K in Cancer / Initial triggering of complement / Downregulation of ERBB2:ERBB3 signaling / GRB7 events in ERBB2 signaling / PIP3 activates AKT signaling / PLCG1 events in ERBB2 signaling / SHC1 events in ERBB2 signaling / Scavenging of heme from plasma / Signaling by ERBB2 / CD22 mediated BCR regulation / ERBB2 Regulates Cell Motility / PI5P, PP2A and IER3 Regulate PI3K/AKT Signaling / ERBB2 Activates PTK6 Signaling / Downregulation of ERBB2 signaling / Sema4D induced cell migration and growth-cone collapse / TFAP2 (AP-2) family regulates transcription of growth factors and their receptors / Regulation of Complement cascade / Antigen activates B Cell Receptor (BCR) leading to generation of second messengers / RAF/MAP kinase cascade / FCERI mediated NF-kB activation / FCERI mediated Ca+2 mobilization / FCERI mediated MAPK activation / Role of LAT2/NTAL/LAB on calcium mobilization / Fc epsilon receptor (FCERI) signaling / negative regulation of immature T cell proliferation in thymus / RNA polymerase I core binding / regulation of microtubule-based process / positive regulation of transcription by RNA polymerase I / ErbB-3 class receptor binding / peripheral nervous system development / oligodendrocyte differentiation / motor neuron axon guidance / myelination / neuromuscular junction development / enzyme linked receptor protein signaling pathway / negative regulation of ERBB signaling pathway / positive regulation of cell adhesion / phosphatidylinositol 3-kinase signaling / regulation of angiogenesis / regulation of cell motility / positive regulation of protein targeting to membrane / basal plasma membrane / complement activation / cellular response to growth factor stimulus / regulation of ERK1 and ERK2 cascade / immunoglobulin complex, circulating / immunoglobulin receptor binding / phagocytosis, recognition / positive regulation of B cell activation / cellular response to epidermal growth factor stimulus / positive regulation of translation / positive regulation of epithelial cell proliferation / ERBB2 signaling pathway / neuron differentiation / phagocytosis, engulfment / positive regulation of GTPase activity / complement activation, classical pathway / regulation of complement activation / antigen binding / receptor protein-tyrosine kinase / transmembrane receptor protein tyrosine kinase activity / Ras guanyl-nucleotide exchange factor activity / transmembrane signaling receptor activity / Fc-epsilon receptor signaling pathway / receptor-mediated endocytosis / positive regulation of MAP kinase activity / retina homeostasis / B cell receptor signaling pathway / wound healing / phosphatidylinositol-4,5-bisphosphate 3-kinase activity / Fc-gamma receptor signaling pathway involved in phagocytosis / leukocyte migration / transmembrane receptor protein tyrosine kinase signaling pathway / adaptive immune response / heart development / apical plasma membrane / regulation of immune response / positive regulation of MAPK cascade / myelin sheath / positive regulation of cell growth / basolateral plasma membrane / protein phosphatase binding / receptor complex / protein C-terminus binding / cell surface receptor signaling pathway / endosome membrane / blood microparticle / protein tyrosine kinase activity / MAPK cascade / immune response / protein dimerization activity / intracellular signal transduction
Immunoglobulin C1-set domain / Furin-like cysteine-rich domain / Immunoglobulin subtype / Immunoglobulin C1-set / Immunoglobulin/major histocompatibility complex, conserved site / Serine-threonine/tyrosine-protein kinase, catalytic domain / Protein kinase domain / Receptor L-domain / Protein tyrosine kinase / Growth factor receptor domain IV ...Immunoglobulin C1-set domain / Furin-like cysteine-rich domain / Immunoglobulin subtype / Immunoglobulin C1-set / Immunoglobulin/major histocompatibility complex, conserved site / Serine-threonine/tyrosine-protein kinase, catalytic domain / Protein kinase domain / Receptor L-domain / Protein tyrosine kinase / Growth factor receptor domain IV / Protein kinases ATP-binding region signature. / Immunoglobulin V-set domain / Tyrosine protein kinases specific active-site signature. / Immunoglobulins and major histocompatibility complex proteins signature. / Protein kinase domain profile. / Ig-like domain profile. / Furin-like repeat / Growth factor receptor domain 4 / Tyrosine-protein kinase, catalytic domain / Immunoglobulin-like domain superfamily / Receptor L-domain superfamily / Furin-like cysteine rich region / Receptor L domain / Immunoglobulin-like domain / Protein kinase, ATP binding site / Tyrosine protein kinase, EGF/ERB/XmrK receptor / Immunoglobulin-like fold / Immunoglobulin V-set domain / Protein kinase-like domain superfamily / Growth factor receptor cysteine-rich domain superfamily / Tyrosine-protein kinase, active site
Immunoglobulin kappa constant / Receptor tyrosine-protein kinase erbB-2 / Immunoglobulin gamma-1 heavy chain / IGH@ protein
SourceHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.36 Å
AuthorsHao Y / Yu X / Bai Y / Huang X
CitationJournal: PLoS ONE / Year: 2019
Title: Cryo-EM Structure of HER2-trastuzumab-pertuzumab complex.
Authors: Yue Hao / Xinchao Yu / Yonghong Bai / Helen J McBride / Xin Huang /
Abstract: Trastuzumab and pertuzumab are monoclonal antibodies that bind to distinct subdomains of the extracellular domain of human epidermal growth factor receptor 2 (HER2). Adding these monoclonal ...Trastuzumab and pertuzumab are monoclonal antibodies that bind to distinct subdomains of the extracellular domain of human epidermal growth factor receptor 2 (HER2). Adding these monoclonal antibodies to the treatment regimen of HER2-positive breast cancer has changed the paradigm for treatment in that form of cancer. Synergistic activity has been observed with the combination of these two antibodies leading to hypotheses regarding the mechanism(s) and to the development of bispecific antibodies to maximize the clinical effect further. Although the individual crystal structures of HER2-trastuzumab and HER2-pertuzumab revealed the distinct binding sites and provided the structural basis for their anti-tumor activities, detailed structural information on the HER2-trastuzumab-pertuzumab complex has been elusive. Here we present the cryo-EM structure of HER2-trastuzumab-pertuzumab at 4.36 Å resolution. Comparison with the binary complexes reveals no cooperative interaction between trastuzumab and pertuzumab, and provides key insights into the design of novel, high-avidity bispecific molecules with potentially greater clinical efficacy.
Validation ReportPDB-ID: 6oge

SummaryFull reportAbout validation report
DateDeposition: Apr 2, 2019 / Header (metadata) release: May 1, 2019 / Map release: May 15, 2019 / Update: May 15, 2019

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 4.2
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 4.2
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6oge
  • Surface level: 4.2
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_20055.map.gz / Format: CCP4 / Size: 34.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
1.06 Å/pix.
x 208 pix.
= 220.272 Å
1.06 Å/pix.
x 208 pix.
= 220.272 Å
1.06 Å/pix.
x 208 pix.
= 220.272 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.059 Å
Density
Contour LevelBy AUTHOR: 4.2 / Movie #1: 4.2
Minimum - Maximum-10.745524 - 18.676493000000001
Average (Standard dev.)-0.000000000007725 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions208208208
Spacing208208208
CellA=B=C: 220.272 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0591.0591.059
M x/y/z208208208
origin x/y/z0.0000.0000.000
length x/y/z220.272220.272220.272
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ208208208
MAP C/R/S321
start NC/NR/NS000
NC/NR/NS208208208
D min/max/mean-10.74618.676-0.000

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Supplemental data

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Sample components

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Entire Her2 extracellular domain-Trastuzumab Fab-Pertuzumab Fab complex

EntireName: Her2 extracellular domain-Trastuzumab Fab-Pertuzumab Fab complex
Number of components: 12

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Component #1: protein, Her2 extracellular domain-Trastuzumab Fab-Pertuzumab Fab...

ProteinName: Her2 extracellular domain-Trastuzumab Fab-Pertuzumab Fab complex
Recombinant expression: No
SourceSpecies: Homo sapiens (human)

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Component #2: protein, Human HER2 extracellular domain

ProteinName: Human HER2 extracellular domain / Recombinant expression: No
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

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Component #3: protein, Pertuzumab Fab

ProteinName: Pertuzumab Fab / Recombinant expression: No
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Cricetulus griseus (Chinese hamster)

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Component #4: protein, Trastuzumab Fab

ProteinName: Trastuzumab Fab / Recombinant expression: No
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Cricetulus griseus (Chinese hamster)

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Component #5: protein, Receptor tyrosine-protein kinase erbB-2

ProteinName: Receptor tyrosine-protein kinase erbB-2 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 68.536844 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

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Component #6: protein, Pertuzumab FAB LIGHT CHAIN

ProteinName: Pertuzumab FAB LIGHT CHAIN / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 23.548152 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Cricetulus griseus (Chinese hamster)

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Component #7: protein, Pertuzumab FAB HEAVY CHAIN

ProteinName: Pertuzumab FAB HEAVY CHAIN / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 23.674486 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Cricetulus griseus (Chinese hamster)

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Component #8: protein, Trastuzumab FAB LIGHT CHAIN

ProteinName: Trastuzumab FAB LIGHT CHAIN / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 23.466031 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Cricetulus griseus (Chinese hamster)

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Component #9: protein, Trastuzumab FAB HEAVY CHAIN

ProteinName: Trastuzumab FAB HEAVY CHAIN / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 23.42518 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Cricetulus griseus (Chinese hamster)

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Component #10: ligand, N-ACETYL-D-GLUCOSAMINE

LigandName: N-ACETYL-D-GLUCOSAMINEN-Acetylglucosamine / Number of Copies: 7 / Recombinant expression: No
MassTheoretical: 0.221208 kDa

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Component #11: ligand, BETA-D-MANNOSE

LigandName: BETA-D-MANNOSE / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 0.180156 kDa

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Component #12: ligand, ALPHA-D-MANNOSE

LigandName: ALPHA-D-MANNOSE / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 0.180156 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 2.4 mg/mL / pH: 7.5
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 45 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 130000.0 X (nominal) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: -3500.0 - -1500.0 nm
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 398409
3D reconstructionSoftware: cisTEM / Resolution: 4.36 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Refinement protocol: flexible / Refinement space: REAL
Output model

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