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- PDB-6pzw: CryoEM derived model of NA-22 Fab in complex with N9 Shanghai2 -

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Basic information

Entry
Database: PDB / ID: 6pzw
TitleCryoEM derived model of NA-22 Fab in complex with N9 Shanghai2
Components
  • (NA-22 fragment antigen binding ...) x 2
  • Neuraminidase
KeywordsVIRAL PROTEIN/IMMUNE SYSTEM / Neuraminidase / NA / Sh2 / N9Shanghai / N9Sh2 / antibody / Fab / NA-22 / Influenza / VIRAL PROTEIN-IMMUNE SYSTEM complex / H7N9
Function / homology
Function and homology information


exo-alpha-(2->3)-sialidase activity / exo-alpha-sialidase / exo-alpha-(2->6)-sialidase activity / exo-alpha-(2->8)-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 / Sialidase, Influenza viruses A/B / Sialidase superfamily / Neuraminidase
Neuraminidase
Biological speciesInfluenza A virus
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3 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

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Structure visualization

Movie
  • Deposited structure unit
  • Imaged by Jmol
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  • Superimposition on EM map
  • EMDB-20538
  • Imaged by UCSF Chimera
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Structure viewerMolecule:
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Assembly

Deposited unit
A: Neuraminidase
I: NA-22 fragment antigen binding light chain
K: NA-22 fragment antigen binding heavy chain
D: Neuraminidase
E: NA-22 fragment antigen binding light chain
F: NA-22 fragment antigen binding heavy chain
G: NA-22 fragment antigen binding light chain
J: NA-22 fragment antigen binding heavy chain
C: Neuraminidase
B: Neuraminidase
L: NA-22 fragment antigen binding light chain
H: NA-22 fragment antigen binding heavy chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)390,69959
Polymers381,45212
Non-polymers9,24747
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Buried area38630 Å2
ΔGint59 kcal/mol
Surface area83290 Å2

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Components

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Protein/peptide , 1 types, 4 molecules ADCB

#1: Protein/peptide
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): SF6 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: S5MF06, exo-alpha-sialidase

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NA-22 fragment antigen binding ... , 2 types, 8 molecules IEGLKFJH

#2: Protein/peptide
NA-22 fragment antigen binding light chain


Mass: 22774.986 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: Protein/peptide
NA-22 fragment antigen binding heavy chain


Mass: 24373.496 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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Non-polymers , 3 types, 47 molecules

#4: Chemical
ChemComp-NAG / N-ACETYL-D-GLUCOSAMINE


Mass: 221.208 Da / Num. of mol.: 19
Source method: isolated from a genetically manipulated source
Formula: C8H15NO6 / N-Acetylglucosamine
#5: Chemical
ChemComp-BMA / BETA-D-MANNOSE


Mass: 180.156 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Formula: C6H12O6 / Mannose
#6: Chemical...
ChemComp-MAN / ALPHA-D-MANNOSE


Mass: 180.156 Da / Num. of mol.: 24
Source method: isolated from a genetically manipulated source
Formula: C6H12O6 / Mannose

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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

Type: COMPLEX

IDNameEntity IDParent-IDSource
1NA-22 Fab in complex with N9 Shanghai21, 2, 30MULTIPLE SOURCES
2N9 Shanghai211RECOMBINANT
3NA-22 Fab2, 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 f7108
23Cricetinae gen. sp. (mammal)36483
Buffer solutionpH: 7.4
SpecimenConc.: 0.478 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: unspecified
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen 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: 2000 nm / Nominal defocus min: 1200 nm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE
Specimen holderModel: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 8.25 sec. / Electron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
Image scansMovie frames/image: 33

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Processing

EM software
IDNameVersionCategoryDetails
2Leginonimage acquisition
4Gctf1.06CTF correction
7UCSF Chimeramodel fitting
9Cootmodel refinement
10Rosetta3.1model refinementrelax
11cryoSPARC2initial Euler assignment
12cryoSPARC2final Euler assignment
13cryoSPARC2classification
14cryoSPARC23D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C4 (4 fold cyclic)
3D reconstructionResolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 20922 / Symmetry type: POINT
Atomic model buildingSpace: REAL
Atomic model buildingPDB-ID: 5L14

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