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- PDB-7t6x: Cryo-EM structure of full-length hepatitis C virus E1E2 glycoprot... -

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Basic information

Entry
Database: PDB / ID: 7t6x
TitleCryo-EM structure of full-length hepatitis C virus E1E2 glycoprotein in complex with AR4A, AT12009, and IGH505 Fabs
Components
  • (Envelope glycoprotein ...) x 2
  • AR4A Fab, heavy chain
  • AR4A, kappa light chain
  • AT12009 CrossMab, heavy chain
  • AT12009 crossMab, light kappa chain
  • IGH505, heavy chain
  • IGH505, lambda light chain
KeywordsVIRAL PROTEIN/IMMUNE SYSTEM / hepatitis C virus / E1E2 / viral protein-immune system complex / antibody-antigen complex
Function / homology
Function and homology information


host cell mitochondrial membrane / host cell lipid droplet / virus-mediated perturbation of host defense response / lipid droplet / clathrin-dependent endocytosis of virus by host cell / viral nucleocapsid / host cell endoplasmic reticulum membrane / ribonucleoprotein complex / viral envelope / virion membrane ...host cell mitochondrial membrane / host cell lipid droplet / virus-mediated perturbation of host defense response / lipid droplet / clathrin-dependent endocytosis of virus by host cell / viral nucleocapsid / host cell endoplasmic reticulum membrane / ribonucleoprotein complex / viral envelope / virion membrane / structural molecule activity / membrane / cytoplasm
Similarity search - Function
Hepatitis C virus, Core protein, C-terminal / Hepatitis C virus core protein / Hepatitis C virus, Non-structural protein E2/NS1 / Hepatitis C virus non-structural protein E2/NS1 / Hepatitis C virus, Envelope glycoprotein E1 / Hepatitis C virus envelope glycoprotein E1
Similarity search - Domain/homology
Biological speciesHepacivirus C
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.83 Å
AuthorsTorrents de la Pena, A. / Ward, A.B. / Eshun-Wilson, L. / Lander, G.C.
Funding support Netherlands, 1items
OrganizationGrant numberCountry
Netherlands Organisation for Scientific Research (NWO) Netherlands
CitationJournal: Science / Year: 2022
Title: Structure of the hepatitis C virus E1E2 glycoprotein complex.
Authors: Alba Torrents de la Peña / Kwinten Sliepen / Lisa Eshun-Wilson / Maddy L Newby / Joel D Allen / Ian Zon / Sylvie Koekkoek / Ana Chumbe / Max Crispin / Janke Schinkel / Gabriel C Lander / ...Authors: Alba Torrents de la Peña / Kwinten Sliepen / Lisa Eshun-Wilson / Maddy L Newby / Joel D Allen / Ian Zon / Sylvie Koekkoek / Ana Chumbe / Max Crispin / Janke Schinkel / Gabriel C Lander / Rogier W Sanders / Andrew B Ward /
Abstract: Hepatitis C virus (HCV) infection is a leading cause of chronic liver disease, cirrhosis, and hepatocellular carcinoma in humans and afflicts more than 58 million people worldwide. The HCV envelope ...Hepatitis C virus (HCV) infection is a leading cause of chronic liver disease, cirrhosis, and hepatocellular carcinoma in humans and afflicts more than 58 million people worldwide. The HCV envelope E1 and E2 glycoproteins are essential for viral entry and comprise the primary antigenic target for neutralizing antibody responses. The molecular mechanisms of E1E2 assembly, as well as how the E1E2 heterodimer binds broadly neutralizing antibodies, remain elusive. Here, we present the cryo-electron microscopy structure of the membrane-extracted full-length E1E2 heterodimer in complex with three broadly neutralizing antibodies-AR4A, AT1209, and IGH505-at ~3.5-angstrom resolution. We resolve the interface between the E1 and E2 ectodomains and deliver a blueprint for the rational design of vaccine immunogens and antiviral drugs.
History
DepositionDec 14, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 2, 2022Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
U: Envelope glycoprotein E2
H: AR4A Fab, heavy chain
L: AR4A, kappa light chain
A: IGH505, heavy chain
B: IGH505, lambda light chain
D: AT12009 crossMab, light kappa chain
C: AT12009 CrossMab, heavy chain
E: Envelope glycoprotein E1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)253,88423
Polymers246,6298
Non-polymers7,25515
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Envelope glycoprotein ... , 2 types, 2 molecules UE

#1: Protein Envelope glycoprotein E2


Mass: 40156.699 Da / Num. of mol.: 1 / Fragment: UNP residues 216-578
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Hepacivirus C / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: A0A159UHX2
#8: Protein Envelope glycoprotein E1


Mass: 20956.279 Da / Num. of mol.: 1 / Fragment: UNP residues 24-215
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Hepacivirus C / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: A0A159UHX2

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Antibody , 6 types, 6 molecules HLABDC

#2: Antibody AR4A Fab, heavy chain


Mass: 29808.500 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK293F / Production host: Homo sapiens (human)
#3: Antibody AR4A, kappa light chain


Mass: 25290.209 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK293F / Production host: Homo sapiens (human)
#4: Antibody IGH505, heavy chain


Mass: 52277.027 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK293F / Production host: Homo sapiens (human)
#5: Antibody IGH505, lambda light chain


Mass: 24974.764 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK293F / Production host: Homo sapiens (human)
#6: Antibody AT12009 crossMab, light kappa chain


Mass: 24014.084 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK293F / Production host: Homo sapiens (human)
#7: Antibody AT12009 CrossMab, heavy chain


Mass: 29151.678 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK293F / Production host: Homo sapiens (human)

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Sugars , 5 types, 15 molecules

#9: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#10: Polysaccharide
beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta- ...beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 586.542 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/2,3,2/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5]/1-1-2/a4-b1_b4-c1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{}}}LINUCSPDB-CARE
#11: Polysaccharide alpha-D-mannopyranose-(1-6)-alpha-D-mannopyranose-(1-6)-beta-D-mannopyranose-(1-4)-2-acetamido-2- ...alpha-D-mannopyranose-(1-6)-alpha-D-mannopyranose-(1-6)-beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 910.823 Da / Num. of mol.: 1 / Source method: obtained synthetically
DescriptorTypeProgram
DManpa1-6DManpa1-6DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/3,5,4/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5][a1122h-1a_1-5]/1-1-2-3-3/a4-b1_b4-c1_c6-d1_d6-e1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(6+1)][a-D-Manp]{[(6+1)][a-D-Manp]{}}}}}LINUCSPDB-CARE
#12: Polysaccharide alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2- ...alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 910.823 Da / Num. of mol.: 2 / Source method: obtained synthetically
DescriptorTypeProgram
DManpa1-3[DManpa1-6]DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/3,5,4/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5][a1122h-1a_1-5]/1-1-2-3-3/a4-b1_b4-c1_c3-d1_c6-e1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(3+1)][a-D-Manp]{}[(6+1)][a-D-Manp]{}}}}LINUCSPDB-CARE
#13: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE / N-Acetylglucosamine


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C8H15NO6 / Feature type: SUBJECT OF INVESTIGATION
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Details

Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: cryoEM structure of full-length hepatitis C virus E1E2 glycoprotein in complex with AR4A, AT12009 and IGH505 Fabs
Type: COMPLEX / Entity ID: #1-#8 / Source: MULTIPLE SOURCES
Source (natural)Organism: Hepacivirus C / Strain: AMS0232
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK 293F / Plasmid: pPPI4
Buffer solutionpH: 7.4 / Details: TBS buffer, pH 7.4
Buffer component
IDConc.NameFormulaBuffer-ID
125 mMTRIS hydrochloride1
2150 mMsodium ChlorideNaClSodium chloride1
SpecimenConc.: 0.8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Full-length hepatitis C virus E1E2 glycoprotein complexed with AR4A, AT12009, and IGH505 Fabs and embedded in peptidiscs is diluted in TBS to a final concentration of 0.8 mg/ml.
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 40 K
Details: Blotting time: 5 s; Blotting force: 0; Waiting time: 7 s

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 36000 X / Calibrated magnification: 43478 X / Nominal defocus max: 1120 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: OTHER
Image recordingAverage exposure time: 9 sec. / Electron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 2 / Num. of real images: 5121
Details: images were collected at 250 ms per frame. 36 frames were collected.

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Processing

SoftwareName: PHENIX / Version: 1.17.1_3660: / Classification: refinement
EM software
IDNameVersionCategory
2Leginonimage acquisition
4Gctf1.06CTF correction
10cryoSPARCv3.2.0initial Euler assignment
11cryoSPARCv3.2.0final Euler assignment
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 2017845
3D reconstructionResolution: 3.83 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 48546 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0219462
ELECTRON MICROSCOPYf_angle_d1.96112902
ELECTRON MICROSCOPYf_dihedral_angle_d10.1553453
ELECTRON MICROSCOPYf_chiral_restr0.1271505
ELECTRON MICROSCOPYf_plane_restr0.0191604

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