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- EMDB-21170: Tetrahedral nanoparticle T33_dn10 presenting BG505-SOSIP -

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Basic information

Entry
Database: EMDB / ID: EMD-21170
TitleTetrahedral nanoparticle T33_dn10 presenting BG505-SOSIP
Map dataBG505-SOSIP-T33_dn10-NP, Negative Stain EM map
Sample
  • Complex: Tetrahedral nanoparticle, T33_dn10, presenting BG505-SOSIP
    • Other: BG505-SOSIP-T33_dn10A
    • Protein or peptide: T33_dn10B
Biological speciessynthetic construct (others)
Methodsingle particle reconstruction / negative staining / Resolution: 25.67 Å
AuthorsWard AB / Antanasijevic A
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-
UpdateAug 19, 2020-
Current statusAug 19, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0055
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.0055
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21170.png

Downloads & links

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Map

FileDownload / File: emd_21170.map.gz / Format: CCP4 / Size: 91.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationBG505-SOSIP-T33_dn10-NP, Negative Stain EM map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.05 Å/pix.
x 288 pix.
= 590.4 Å		2.05 Å/pix.
x 288 pix.
= 590.4 Å		2.05 Å/pix.
x 288 pix.
= 590.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.05 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0055 / Movie #1: 0.0055
Minimum - Maximum-0.048289906 - 0.060376342
Average (Standard dev.)-0.00022629763 (±0.0027946713)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions288288288
Spacing288288288
CellA=B=C: 590.39996 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.052.052.05
M x/y/z288288288
origin x/y/z0.0000.0000.000
length x/y/z590.400590.400590.400
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/NS288288288
D min/max/mean-0.0480.060-0.000

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Supplemental data

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Sample components

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Entire : Tetrahedral nanoparticle, T33_dn10, presenting BG505-SOSIP

EntireName: Tetrahedral nanoparticle, T33_dn10, presenting BG505-SOSIP
Components
  • Complex: Tetrahedral nanoparticle, T33_dn10, presenting BG505-SOSIP
    • Other: BG505-SOSIP-T33_dn10A
    • Protein or peptide: T33_dn10B

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Supramolecule #1: Tetrahedral nanoparticle, T33_dn10, presenting BG505-SOSIP

SupramoleculeName: Tetrahedral nanoparticle, T33_dn10, presenting BG505-SOSIP
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all / Details: 4 copies of BG505-SOSIP presented
Source (natural)Organism: synthetic construct (others)
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant strain: HEK293F / Recombinant plasmid: pPPI4

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Macromolecule #1: BG505-SOSIP-T33_dn10A

MacromoleculeName: BG505-SOSIP-T33_dn10A / type: other / ID: 1 / Classification: other
Source (natural)Organism: synthetic construct (others)
SequenceString: MKRGLCCVLL LCGAVFVSPS QEIHARFRRG ARAENLWVTV YYGVPVWKDA ETTLFCASDA KAYETKKHNV WATHCCVPTD PNPQEIHLEN VTEEFNMWKN NMVEQMHTDI ISLWDQSLKP CVKLTPLCVT LQCTNVTNNI TDDMRGELKN CSFNMTTELR DKKQKVYSLF ...String:
MKRGLCCVLL LCGAVFVSPS QEIHARFRRG ARAENLWVTV YYGVPVWKDA ETTLFCASDA KAYETKKHNV WATHCCVPTD PNPQEIHLEN VTEEFNMWKN NMVEQMHTDI ISLWDQSLKP CVKLTPLCVT LQCTNVTNNI TDDMRGELKN CSFNMTTELR DKKQKVYSLF YRLDVVQINE NQGNRSNNSN KEYRLINCNT SAITQACPKV SFEPIPIHYC APAGFAILKC KDKKFNGTGP CTNVSTVQCT HGIKPVVSTQ LLLNGSLAEE EVIIRSENIT NNAKNILVQL NESVQINCTR PNNNTVKSIR IGPGQWFYYT GDIIGDIRQA HCNVSKATWN ETLGKVVKQL RKHFGNNTII RFANSSGGDL EVTTHSFNCG GEFFYCNTSG LFNSTWISNT SVQGSNSTGS NDSITLPCRI KQIINMWQRI GQAMYAPPIQ GVIRCVSNIT GLILTRDGGS TNSTTETFRP GGGDMRDNWR SELYKYKVVK IEPLGVAPTR CKRRVVGRRR RRRAVGIGAV SLGFLGAAGS TMGAASMTLT VQARNLLSGI VQQQSNLLRA PECQQHLLKD THWGIKQLQA RVLAVEHYLR DQQLLGIWGC SGKLICCTNV PWNSSWSNRN LSEIWDNMTW LQWDKEISNY TQIIYGLLEE SQNQQEKNEQ GSGSGSGSGG EEAELAYLLG ELAYKLGEYR IAIRAYRIAL KRDPNNAEAW YNLGNAYYKQ GDYDEAIEYY QKALELDPNN AEAWYNLGNA YYKQGDYDEA IEYYEKALEL DPENLEALQN LLNAMDKQG
Recombinant expressionOrganism: Homo sapiens (human)

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Macromolecule #2: T33_dn10B

MacromoleculeName: T33_dn10B / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MIEEVVAEMI DILAESSKKS IEELARAADN KTTEKAVAEA IEEIARLATA AIQLIEALAK NLASEEFMAR AISAIAELAK KAIEAIYRLA DNHTTDTFMA RAIAAIANLA VTAILAIAAL ASNHTTEEFM ARAISAIAEL AKKAIEAIYR LADNHTTDKF MAAAIEAIAL ...String:
MIEEVVAEMI DILAESSKKS IEELARAADN KTTEKAVAEA IEEIARLATA AIQLIEALAK NLASEEFMAR AISAIAELAK KAIEAIYRLA DNHTTDTFMA RAIAAIANLA VTAILAIAAL ASNHTTEEFM ARAISAIAEL AKKAIEAIYR LADNHTTDKF MAAAIEAIAL LATLAILAIA LLASNHTTEK FMARAIMAIA ILAAKAIEAI YRLADNHTSP TYIEKAIEAI EKIARKAIKA IEMLAKNITT EEYKEKAKKI IDIIRKLAKM AIKKLEDNRT LEHHHHHH

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Experimental details

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Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.05 mg/mL
BufferpH: 7.4
Component:
ConcentrationNameFormula
25.0 mMTris-HCl
150.0 mMsodium chlorideNaCl

Details: TBS buffer, pH 7.4
StainingType: NEGATIVE / Material: Uranyl Formate
Details: Sample diluted to 0.05 mg/mL. 3 uL was applied onto the grid, blotted off, and then stained with 2% uranyl formate for 60 seconds.
GridMaterial: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS
DetailsBG505-SOSIP-T33_dn10 nanoparticle was purified by SEC, diluted to 0.05mg/ml and loaded onto a grid.

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Electron microscopy

MicroscopeFEI TECNAI SPIRIT
Image recordingFilm or detector model: TVIPS TEMCAM-F416 (4k x 4k) / Average electron dose: 25.0 e/Å2
Electron beamAcceleration voltage: 120 kV / Electron source: LAB6
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 1.5 µm
Experimental equipment
Model: Tecnai Spirit / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Details: Negative stain EM map of the non-liganded T33_dn10 nanoparticle
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: T (tetrahedral) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 25.67 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 375
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final 3D classificationSoftware - Name: RELION (ver. 3.0)

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