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- EMDB-21174: De novo designed icosahedral nanoparticle I53_dn5 -

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Basic information

Entry
Database: EMDB / ID: EMD-21174
TitleDe novo designed icosahedral nanoparticle I53_dn5
Map dataI53_dn5 Nanoparticle Map
Sample
  • Complex: De novo designed nanoparticle of icosahedral symmetry I53_dn5_NP
    • Protein or peptide: I53_dn5B
    • Protein or peptide: I53_dn5A
KeywordsDe novo / Nanoparticle / DE NOVO PROTEIN
Biological speciessynthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 5.35 Å
AuthorsAntanasijevic A / Ward AB
Funding support United States, 1 items
OrganizationGrant numberCountry
Bill & Melinda Gates FoundationOPP1115782 United States
CitationJournal: Elife / Year: 2020
Title: Tailored design of protein nanoparticle scaffolds for multivalent presentation of viral glycoprotein antigens.
Authors: George Ueda / Aleksandar Antanasijevic / Jorge A Fallas / William Sheffler / Jeffrey Copps / Daniel Ellis / Geoffrey B Hutchinson / Adam Moyer / Anila Yasmeen / Yaroslav Tsybovsky / Young- ...Authors: George Ueda / Aleksandar Antanasijevic / Jorge A Fallas / William Sheffler / Jeffrey Copps / Daniel Ellis / Geoffrey B Hutchinson / Adam Moyer / Anila Yasmeen / Yaroslav Tsybovsky / Young-Jun Park / Matthew J Bick / Banumathi Sankaran / Rebecca A Gillespie / Philip Jm Brouwer / Peter H Zwart / David Veesler / Masaru Kanekiyo / Barney S Graham / Rogier W Sanders / John P Moore / Per Johan Klasse / Andrew B Ward / Neil P King / David Baker /
Abstract: Multivalent presentation of viral glycoproteins can substantially increase the elicitation of antigen-specific antibodies. To enable a new generation of anti-viral vaccines, we designed self- ...Multivalent presentation of viral glycoproteins can substantially increase the elicitation of antigen-specific antibodies. To enable a new generation of anti-viral vaccines, we designed self-assembling protein nanoparticles with geometries tailored to present the ectodomains of influenza, HIV, and RSV viral glycoprotein trimers. We first designed trimers tailored for antigen fusion, featuring N-terminal helices positioned to match the C termini of the viral glycoproteins. Trimers that experimentally adopted their designed configurations were incorporated as components of tetrahedral, octahedral, and icosahedral nanoparticles, which were characterized by cryo-electron microscopy and assessed for their ability to present viral glycoproteins. Electron microscopy and antibody binding experiments demonstrated that the designed nanoparticles presented antigenically intact prefusion HIV-1 Env, influenza hemagglutinin, and RSV F trimers in the predicted geometries. This work demonstrates that antigen-displaying protein nanoparticles can be designed from scratch, and provides a systematic way to investigate the influence of antigen presentation geometry on the immune response to vaccination.
History
DepositionJan 5, 2020-
Header (metadata) releaseJan 29, 2020-
Map releaseAug 12, 2020-
UpdateMar 6, 2024-
Current statusMar 6, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.015
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6vfj
  • Surface level: 0.015
  • Imaged by UCSF Chimera
  • Download
  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6vfj
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21174.png

Downloads & links

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Map

FileDownload / File: emd_21174.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationI53_dn5 Nanoparticle Map
Voxel sizeX=Y=Z: 1.03 Å
Density
Contour LevelBy AUTHOR: 0.015 / Movie #1: 0.015
Minimum - Maximum-0.014618222 - 0.04184899
Average (Standard dev.)0.00016296933 (±0.0028055245)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 412.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.031.031.03
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z412.000412.000412.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-205-205-205
NX/NY/NZ411411411
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-0.0150.0420.000

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Supplemental data

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Mask #1

Fileemd_21174_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: I53 dn5 Nanoparticle Half-Map 1

Fileemd_21174_half_map_1.map
AnnotationI53_dn5 Nanoparticle Half-Map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: I53 dn5 Nanoparticle Half-Map 2

Fileemd_21174_half_map_2.map
AnnotationI53_dn5 Nanoparticle Half-Map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : De novo designed nanoparticle of icosahedral symmetry I53_dn5_NP

EntireName: De novo designed nanoparticle of icosahedral symmetry I53_dn5_NP
Components
  • Complex: De novo designed nanoparticle of icosahedral symmetry I53_dn5_NP
    • Protein or peptide: I53_dn5B
    • Protein or peptide: I53_dn5A

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Supramolecule #1: De novo designed nanoparticle of icosahedral symmetry I53_dn5_NP

SupramoleculeName: De novo designed nanoparticle of icosahedral symmetry I53_dn5_NP
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Self-assembling nanoparticle of icosahedral symmetry
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 1.96 MDa

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Macromolecule #1: I53_dn5B

MacromoleculeName: I53_dn5B / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 15.356009 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MEEAELAYLL GELAYKLGEY RIAIRAYRIA LKRDPNNAEA WYNLGNAYYK QGRYREAIEY YQKALELDPN NAEAWYNLGN AYYERGEYE EAIEYYRKAL RLDPNNADAM QNLLNAKMRE ELEHHHHHH

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Macromolecule #2: I53_dn5A

MacromoleculeName: I53_dn5A / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 17.215148 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MGKYDGSKLR IGILHARWNA EIILALVLGA LKRLQEFGVK RENIIIETVP GSFELPYGSK LFVEKQKRLG KPLDAIIPIG VLIKGSTMH FEYICDSTTH QLMKLNFELG IPVIFGVLTC LTDEQAEARA GLIEGKMHNH GEDWGAAAVE MATKFN

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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

Concentration1.9 mg/mL
BufferpH: 7.4
Component:
ConcentrationNameFormula
25.0 mMTris-HClTris
150.0 mMSodium ChlorideNaClSodium chloride

Details: Fresh TBS buffer, 0.2um filtered, 0.06mM DDM detergent added immediately before freezing
GridSupport film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 11 / Details: unspecified
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV
Details: 0.06mM DDM detergent (from an 8X stock) added immediately before freezing.
DetailsNanoparticle was generated by co-expression of two components (A and B) in E coli. Assembled particles purified using a combination of Ni-affinity chromatography and gel filtration chromatography.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.6 µm / Nominal magnification: 29000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
TemperatureMin: 70.0 K / Max: 90.0 K
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 1548 / Average electron dose: 50.4 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 207651
Startup modelType of model: OTHER / Details: Ab Initio generated model from cryoSPARC
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final 3D classificationNumber classes: 10 / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: I (icosahedral) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 5.35 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 31353

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Atomic model buiding 1

DetailsIcosahedral nanoparticle I53_dn5 model was docked into a reconstructed map using UCSF Chimera and Rosetta relaxed refinement was applied to relax the model into the map.
RefinementSpace: REAL / Protocol: OTHER
Output model

PDB-6vfj:
De novo designed icosahedral nanoparticle I53_dn5

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