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- EMDB-25107: Lassa virus glycoprotein construct(Josiah GPCysR4) recovered from... -

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Basic information

Entry
Database: EMDB / ID: EMD-25107
TitleLassa virus glycoprotein construct(Josiah GPCysR4) recovered from GPC-I53-50 nanoparticle by localized reconstruction
Map dataLassa virus glycoprotein (GPCysR4) map reconstructed from GPCysR4-I53-50 nanoparticle by localized reconstruction - Main Map
Sample
  • Complex: Lassa virus glycoprotein construct(Josiah GPCysR4) recovered from GPC-I53-50 nanoparticle
    • Protein or peptide: Josiah GPCysR4 I53-50A
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
Function / homology
Function and homology information


host cell Golgi membrane / receptor-mediated endocytosis of virus by host cell / host cell endoplasmic reticulum membrane / fusion of virus membrane with host endosome membrane / viral envelope / virion attachment to host cell / host cell plasma membrane / virion membrane / membrane / metal ion binding
Similarity search - Function
Arenavirus glycoprotein, zinc binding domain / Arenavirus glycoprotein / Arenavirus glycoprotein
Similarity search - Domain/homology
Pre-glycoprotein polyprotein GP complex
Similarity search - Component
Biological speciesLassa mammarenavirus
Methodsingle particle reconstruction / cryo EM / Resolution: 3.97 Å
AuthorsAntanasijevic A / Brouwer PJM / Ward AB
Funding support United States, 1 items
OrganizationGrant numberCountry
Bill & Melinda Gates FoundationOPP1170236 United States
CitationJournal: Cell Host Microbe / Year: 2022
Title: Lassa virus glycoprotein nanoparticles elicit neutralizing antibody responses and protection.
Authors: Philip J M Brouwer / Aleksandar Antanasijevic / Adam J Ronk / Helena Müller-Kräuter / Yasunori Watanabe / Mathieu Claireaux / Hailee R Perrett / Tom P L Bijl / Marloes Grobben / Jeffrey C ...Authors: Philip J M Brouwer / Aleksandar Antanasijevic / Adam J Ronk / Helena Müller-Kräuter / Yasunori Watanabe / Mathieu Claireaux / Hailee R Perrett / Tom P L Bijl / Marloes Grobben / Jeffrey C Umotoy / Angela I Schriek / Judith A Burger / Khadija Tejjani / Nicole M Lloyd / Thijs H Steijaert / Marlies M van Haaren / Kwinten Sliepen / Steven W de Taeye / Marit J van Gils / Max Crispin / Thomas Strecker / Alexander Bukreyev / Andrew B Ward / Rogier W Sanders /
Abstract: The Lassa virus is endemic in parts of West Africa, and it causes hemorrhagic fever with high mortality. The development of a recombinant protein vaccine has been hampered by the instability of ...The Lassa virus is endemic in parts of West Africa, and it causes hemorrhagic fever with high mortality. The development of a recombinant protein vaccine has been hampered by the instability of soluble Lassa virus glycoprotein complex (GPC) trimers, which disassemble into monomeric subunits after expression. Here, we use two-component protein nanoparticles consisting of trimeric and pentameric subunits to stabilize GPC in a trimeric conformation. These GPC nanoparticles present twenty prefusion GPC trimers on the surface of an icosahedral particle. Cryo-EM studies of GPC nanoparticles demonstrated a well-ordered structure and yielded a high-resolution structure of an unliganded GPC. These nanoparticles induced potent humoral immune responses in rabbits and protective immunity against the lethal Lassa virus challenge in guinea pigs. Additionally, we isolated a neutralizing antibody that mapped to the putative receptor-binding site, revealing a previously undefined site of vulnerability. Collectively, these findings offer potential approaches to vaccine and therapeutic design for the Lassa virus.
History
DepositionOct 5, 2021-
Header (metadata) releaseOct 12, 2022-
Map releaseOct 12, 2022-
UpdateDec 28, 2022-
Current statusDec 28, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_25107.png

Downloads & links

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Map

FileDownload / File: emd_25107.map.gz / Format: CCP4 / Size: 40.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationLassa virus glycoprotein (GPCysR4) map reconstructed from GPCysR4-I53-50 nanoparticle by localized reconstruction - Main Map
Voxel sizeX=Y=Z: 1.03 Å
Density
Contour LevelBy AUTHOR: 0.02
Minimum - Maximum-0.031668592 - 0.086114384
Average (Standard dev.)-1.2201465e-05 (±0.0033173314)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions220220220
Spacing220220220
CellA=B=C: 226.59999 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_25107_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Lassa virus glycoprotein (GPCysR4) map reconstructed from GPCysR4-I53-50...

Fileemd_25107_half_map_1.map
AnnotationLassa virus glycoprotein (GPCysR4) map reconstructed from GPCysR4-I53-50 nanoparticle by localized reconstruction - Half Map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Lassa virus glycoprotein (GPCysR4) map reconstructed from GPCysR4-I53-50...

Fileemd_25107_half_map_2.map
AnnotationLassa virus glycoprotein (GPCysR4) map reconstructed from GPCysR4-I53-50 nanoparticle by localized reconstruction - 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 : Lassa virus glycoprotein construct(Josiah GPCysR4) recovered from...

EntireName: Lassa virus glycoprotein construct(Josiah GPCysR4) recovered from GPC-I53-50 nanoparticle
Components
  • Complex: Lassa virus glycoprotein construct(Josiah GPCysR4) recovered from GPC-I53-50 nanoparticle
    • Protein or peptide: Josiah GPCysR4 I53-50A
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: Lassa virus glycoprotein construct(Josiah GPCysR4) recovered from...

SupramoleculeName: Lassa virus glycoprotein construct(Josiah GPCysR4) recovered from GPC-I53-50 nanoparticle
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Josiah GPCysR4 is a trimeric complex consisting of 3 copies of the head domain and 3 copies of the stem domain. Nanoparticle was assembled by combining equimolar amounts of GPC-I53-50A and I53-50B.
Source (natural)Organism: Lassa mammarenavirus

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Macromolecule #1: Josiah GPCysR4 I53-50A

MacromoleculeName: Josiah GPCysR4 I53-50A / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Lassa mammarenavirus
Molecular weightTheoretical: 73.84557 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGQIVTFFQE VPHVIEEVMN IVLIALSVLA VLKGLYNFAT CGLVGLVTFL LLCGRSCTTS LYKGVYELQT LELNMETLNM TMPLSCTKN NSHHYIMVGN ETGLELTLTN TSIINHKFCN LSDAHKKNLY DHALMSIIST FHLSIPNFNQ YEAMSCDFNG G KISVQYNL ...String:
MGQIVTFFQE VPHVIEEVMN IVLIALSVLA VLKGLYNFAT CGLVGLVTFL LLCGRSCTTS LYKGVYELQT LELNMETLNM TMPLSCTKN NSHHYIMVGN ETGLELTLTN TSIINHKFCN LSDAHKKNLY DHALMSIIST FHLSIPNFNQ YEAMSCDFNG G KISVQYNL SHSYAGDAAN HCGTVANGVL QTFMRMAWGG SYIALDSGCG NWDCIMTSYQ YLIIQNTTWE DHCQFSRPSP IG YLGLLSQ RTRDIYISRR RRGTFTWTLS DSEGKDTPGG YCLTRWMLIE AELKCFGNTA VAKCNEKHDE EFCDMLRLFD FNK QAIQRL KAPAQMSIQL INKAVNALIN DQLIMKNHLR DIMCIPYCNY SKYWYLNHTT TGRTSLPKCW LVSNGSYLNE THFS DDIEQ QADNMITEML QKEYMERQGG SGGSGGSGGS GGSEKAAKAE EAARKMEELF KKHKIVAVLR ANSVEEAIEK AVAVF AGGV HLIEITFTVP DADTVIKALS VLKEKGAIIG AGTVTSVEQC RKAVESGAEF IVSPHLDEEI SQFCKEKGVF YMPGVM TPT ELVKAMKLGH DILKLFPGEV VGPEFVKAMK GPFPNVKFVP TGGVDLDNVC EWFDAGVLAV GVGDALVEGD PDEVREK AK EFVEKIRGCT EGSLEWSHPQ FEK

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Macromolecule #6: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 6 / Number of copies: 6 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose / N-Acetylglucosamine

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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

Concentration2 mg/mL
BufferpH: 7
Component:
ConcentrationFormulaName
10.0 mMTris-HClTrisTris
150.0 mMNaClSodium chlorideSodium chloride

Details: TBS
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 7 sec. / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV / Details: Blot time 4s, Wait time 10s, Blot force 0.
DetailsLassa virus glycoprotein construct(Josiah GPCysR4) recovered from GPC-I53-50 nanoparticle by localized reconstruction. Nanoparticle was assembled by combining equimolar amounts of GPC-I53-50A and I53-50B.

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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: 1.6 µm / Nominal defocus min: 0.6 µm / Nominal magnification: 29000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 2 / Average exposure time: 10.5 sec. / 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: 1728229
Details: This is the number of GPCysR4 trimer subparticles extracted from the GPC-I53-50 nanoparticle by localized reconstruction: 86,411 nanoparticles X 20 = 1728229.
Startup modelType of model: OTHER / Details: Map reconstructed from negative stain EM
Initial angle assignmentType: OTHER / Software - Name: RELION (ver. 3.0) / Details: Regularized likelihood
Final 3D classificationSoftware - Name: RELION (ver. 3.0)
Final angle assignmentType: OTHER / Software - Name: RELION (ver. 3.0) / Details: Regularized likelihood
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C3 (3 fold cyclic) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.97 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0)
Details: C3 symmetry used for refinement. Solvent mask applied.
Number images used: 124891
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

5vk2

RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-7sgd:
Lassa virus glycoprotein construct(Josiah GPCysR4) recovered from GPC-I53-50 nanoparticle by localized reconstruction

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