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- PDB-6pzy: CryoEM derived model of NA-73 Fab in complex with N9 Shanghai2 -

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Basic information

Entry
Database: PDB / ID: 6pzy
TitleCryoEM derived model of NA-73 Fab in complex with N9 Shanghai2
Components
  • (NA-73 fragment antibody ...) x 2
  • Neuraminidase
KeywordsVIRAL PROTEIN/IMMUNE SYSTEM / Neuraminidase / NA / Sh2 / N9Shanghai / N9Sh2 / antibody / Fab / NA-73 / Influenza / VIRAL PROTEIN-IMMUNE SYSTEM complex / H7N9
Function / homology
Function and homology information


exo-alpha-(2->6)-sialidase activity / exo-alpha-(2->8)-sialidase activity / exo-alpha-sialidase / exo-alpha-(2->3)-sialidase activity / viral budding from plasma membrane / carbohydrate metabolic process / host cell plasma membrane / virion membrane / integral component of membrane / metal ion binding
Glycoside hydrolase, family 34 / Neuraminidase / Sialidase superfamily / Sialidase, Influenza viruses A/B / Neuraminidase - #10 / 6 Propeller / Neuraminidase / Mainly Beta
Neuraminidase / polysac:dmanpa1-2dmanpa1-2dmanpa1-3[dmanpa1-3[dmanpa1-6]dmanpa1-6]dmanpb1-4dglcpnacb1-4dglcpnacb1-: / polysac:dglcpnacb1-4dglcpnacb1-:
Biological speciesInfluenza A virus
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.17 Å
AuthorsWard, A.B. / Turner, H.L. / Zhu, X.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)U19 AI117905 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)HHSN 272201400024C United States
CitationJournal: Cell Host Microbe / Year: 2019
Title: Structural Basis of Protection against H7N9 Influenza Virus by Human Anti-N9 Neuraminidase Antibodies.
Authors: Xueyong Zhu / Hannah L Turner / Shanshan Lang / Ryan McBride / Sandhya Bangaru / Iuliia M Gilchuk / Wenli Yu / James C Paulson / James E Crowe / Andrew B Ward / Ian A Wilson /
Abstract: Influenza virus neuraminidase (NA) is a major target for small-molecule antiviral drugs. Antibodies targeting the NA surface antigen could also inhibit virus entry and egress to provide host ...Influenza virus neuraminidase (NA) is a major target for small-molecule antiviral drugs. Antibodies targeting the NA surface antigen could also inhibit virus entry and egress to provide host protection. However, our understanding of the nature and range of target epitopes is limited because of a lack of human antibody structures with influenza neuraminidase. Here, we describe crystal and cryogenic electron microscopy (cryo-EM) structures of NAs from human-infecting avian H7N9 viruses in complex with five human anti-N9 antibodies, systematically defining several antigenic sites and antibody epitope footprints. These antibodies either fully or partially block the NA active site or bind to epitopes distant from the active site while still showing neuraminidase inhibition. The inhibition of antibodies to NAs was further analyzed by glycan array and solution-based NA activity assays. Together, these structural studies provide insights into protection by anti-NA antibodies and templates for the development of NA-based influenza virus vaccines and therapeutics.
Validation Report
SummaryFull reportAbout validation report
History
DepositionAug 1, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 4, 2019Provider: repository / Type: Initial release
Revision 1.1Dec 25, 2019Group: Database references / Category: citation / Item: _citation.journal_volume / _citation.page_first
Revision 2.0Jul 29, 2020Group: Atomic model / Data collection ...Atomic model / Data collection / Derived calculations / Structure summary
Category: atom_site / chem_comp ...atom_site / chem_comp / em_entity_assembly / entity / pdbx_branch_scheme / pdbx_chem_comp_identifier / pdbx_entity_branch / pdbx_entity_branch_descriptor / pdbx_entity_branch_link / pdbx_entity_branch_list / pdbx_entity_nonpoly / pdbx_nonpoly_scheme / pdbx_struct_assembly_gen / struct_asym / struct_conn / struct_site / struct_site_gen
Item: _atom_site.B_iso_or_equiv / _atom_site.Cartn_x ..._atom_site.B_iso_or_equiv / _atom_site.Cartn_x / _atom_site.Cartn_y / _atom_site.Cartn_z / _atom_site.auth_asym_id / _atom_site.auth_atom_id / _atom_site.auth_comp_id / _atom_site.auth_seq_id / _atom_site.label_asym_id / _atom_site.label_atom_id / _atom_site.label_comp_id / _atom_site.label_entity_id / _atom_site.type_symbol / _chem_comp.name / _chem_comp.type / _em_entity_assembly.entity_id_list / _entity.formula_weight / _entity.pdbx_description / _entity.pdbx_number_of_molecules / _entity.type / _pdbx_struct_assembly_gen.asym_id_list / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_role / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id
Description: Carbohydrate remediation / Provider: repository / Type: Remediation

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Structure visualization

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Assembly

Deposited unit
A: Neuraminidase
C: NA-73 fragment antibody heavy chain
D: NA-73 fragment antibody light chain
B: Neuraminidase
G: NA-73 fragment antibody heavy chain
J: NA-73 fragment antibody light chain
E: Neuraminidase
H: NA-73 fragment antibody heavy chain
K: NA-73 fragment antibody light chain
F: Neuraminidase
I: NA-73 fragment antibody heavy chain
L: NA-73 fragment antibody light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)392,13824
Polymers383,31812
Non-polymers8,82012
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Buried area37230 Å2
ΔGint92 kcal/mol
Surface area88060 Å2

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Components

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Protein , 1 types, 4 molecules ABEF

#1: Protein
Neuraminidase /


Mass: 48214.508 Da / Num. of mol.: 4 / Fragment: UNP residues 37-465
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Influenza A virus (A/environment/Shanghai/S1439/2013(H7N9))
Strain: A/environment/Shanghai/S1439/2013(H7N9) / Gene: NA / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: S5MF06, exo-alpha-sialidase

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Antibody , 2 types, 8 molecules CGHIDJKL

#2: Antibody
NA-73 fragment antibody heavy chain


Mass: 24822.797 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): ExpiCHO / Production host: Cricetinae gen. sp. (mammal)
#3: Antibody
NA-73 fragment antibody light chain


Mass: 22792.084 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): ExpiCHO / Production host: Cricetinae gen. sp. (mammal)

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Sugars , 3 types, 12 molecules

#4: Polysaccharide
alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-3)-[alpha-D- ...alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-3)-[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: 1559.386 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpa1-2DManpa1-2DManpa1-3[DManpa1-3[DManpa1-6]DManpa1-6]DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/3,9,8/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5][a1122h-1a_1-5]/1-1-2-3-3-3-3-3-3/a4-b1_b4-c1_c3-d1_c6-g1_d2-e1_e2-f1_g3-h1_g6-i1WURCSPDB2Glycan 1.1.0
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(3+1)][a-D-Manp]{[(2+1)][a-D-Manp]{[(2+1)][a-D-Manp]{}}}[(6+1)][a-D-Manp]{[(3+1)][a-D-Manp]{}[(6+1)][a-D-Manp]{}}}}}}LINUCSPDB-CARE
#5: 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.: 4
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-Glycam 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
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}}LINUCSPDB-CARE
#6: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-Acetylglucosamine


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Formula: C8H15NO6
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 interestN

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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

Component
IDNameTypeEntity IDParent-IDSource
1NA-73 Fab in complex with N9 Shanghai2COMPLEX#1-#30MULTIPLE SOURCES
2N9 Shanghai2COMPLEX#11RECOMBINANT
3NA-73 FabCOMPLEX#2-#31RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Influenza A virus (A/environment/Shanghai/S1439/2013(H7N9))1347105
23Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
12Spodoptera frugiperda (fall armyworm)7108
23Cricetinae gen. sp. (mammal)36483
Buffer solutionpH: 7.4
SpecimenConc.: 0.14 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: unspecified
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 29000 X / Nominal defocus max: 1500 nm / Nominal defocus min: 500 nm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

EM software
IDNameVersionCategoryDetails
2Leginonimage acquisition
4GctfCTF correction
7UCSF Chimeramodel fitting
9cryoSPARC2initial Euler assignment
10cryoSPARC2final Euler assignment
11cryoSPARC2classification
12cryoSPARC23D reconstruction
13Cootmodel refinement
14Rosetta3.1model refinementrelax
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C4 (4 fold cyclic)
3D reconstructionResolution: 3.17 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 157751 / Symmetry type: POINT
Atomic model buildingPDB-ID: 5L14

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