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- EMDB-23625: HMPV F v3-B in complex with MPE33 Fab -

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Basic information

Entry
Database: EMDB / ID: EMD-23625
TitleHMPV F v3-B in complex with MPE33 Fab
Map data
Sample
  • Complex: HMPV F v3-B in complex with MPE33
    • Complex: HMPV F v3-B
      • Protein or peptide: HMPV F v3-B
    • Complex: MPE33
      • Protein or peptide: MPE33 Heavy chain
      • Protein or peptide: MPE33 Light chain
Function / homologyPrecursor fusion glycoprotein F0, Paramyxoviridae / Fusion glycoprotein F0 / membrane => GO:0016020 / fusion of virus membrane with host plasma membrane / host cell plasma membrane / virion membrane / plasma membrane / Fusion glycoprotein F0
Function and homology information
Biological speciesHuman metapneumovirus / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.79 Å
AuthorsGorman J / Kwong PD
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM103310 United States
Other privateSimons Foundation (SF349247) United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2021
Title: Interprotomer disulfide-stabilized variants of the human metapneumovirus fusion glycoprotein induce high titer-neutralizing responses.
Authors: Guillaume B E Stewart-Jones / Jason Gorman / Li Ou / Baoshan Zhang / M Gordon Joyce / Lijuan Yang / Cheng Cheng / Gwo-Yu Chuang / Kathryn E Foulds / Wing-Pui Kong / Adam S Olia / Mallika ...Authors: Guillaume B E Stewart-Jones / Jason Gorman / Li Ou / Baoshan Zhang / M Gordon Joyce / Lijuan Yang / Cheng Cheng / Gwo-Yu Chuang / Kathryn E Foulds / Wing-Pui Kong / Adam S Olia / Mallika Sastry / Chen-Hsiang Shen / John-Paul Todd / Yaroslav Tsybovsky / Raffaello Verardi / Yongping Yang / Peter L Collins / Davide Corti / Antonio Lanzavecchia / Diana G Scorpio / John R Mascola / Ursula J Buchholz / Peter D Kwong /
Abstract: Human metapneumovirus (HMPV) is a major cause of respiratory disease worldwide, particularly among children and the elderly. Although there is no licensed HMPV vaccine, promising candidates have been ...Human metapneumovirus (HMPV) is a major cause of respiratory disease worldwide, particularly among children and the elderly. Although there is no licensed HMPV vaccine, promising candidates have been identified for related pneumoviruses based on the structure-based stabilization of the fusion (F) glycoprotein trimer, with prefusion-stabilized F glycoprotein trimers eliciting significantly higher neutralizing responses than their postfusion F counterparts. However, immunization with HMPV F trimers in either prefusion or postfusion conformations has been reported to elicit equivalent neutralization responses. Here we investigate the impact of stabilizing disulfides, especially interprotomer disulfides (IP-DSs) linking protomers of the F trimer, on the elicitation of HMPV-neutralizing responses. We designed F trimer disulfides, screened for their expression, and used electron microscopy (EM) to confirm their formation, including that of an unexpected postfusion variant. In mice, IP-DS-stabilized prefusion and postfusion HMPV F elicited significantly higher neutralizing responses than non-IP-DS-stabilized HMPV Fs. In macaques, the impact of IP-DS stabilization was more measured, although IP-DS-stabilized variants of either prefusion or postfusion HMPV F induced neutralizing responses many times the average titers observed in a healthy human cohort. Serological and absorption-based analyses of macaque responses revealed elicited HMPV-neutralizing responses to be absorbed differently by IP-DS-containing and by non-IP-DS-containing postfusion Fs, suggesting IP-DS stabilization to alter not only the immunogenicity of select epitopes but their antigenicity as well. We speculate the observed increase in immunogenicity by IP-DS trimers to be related to reduced interprotomer flexibility within the HMPV F trimer.
History
DepositionMar 15, 2021-
Header (metadata) releaseAug 25, 2021-
Map releaseAug 25, 2021-
UpdateOct 6, 2021-
Current statusOct 6, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.55
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.55
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23625.png

Downloads & links

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Map

FileDownload / File: emd_23625.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.0961 Å
Density
Contour LevelBy AUTHOR: 0.55 / Movie #1: 0.55
Minimum - Maximum-0.34403914 - 1.9057205
Average (Standard dev.)4.0516108e-05 (±0.04565462)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 438.44 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.09611.09611.0961
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z438.440438.440438.440
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ450450450
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-0.3441.9060.000

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Supplemental data

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Mask #1

Fileemd_23625_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: unsharpened map

Fileemd_23625_additional_1.map
Annotationunsharpened map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: density modified map with ResolveCryoEM

Fileemd_23625_additional_2.map
Annotationdensity modified map with ResolveCryoEM
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map B

Fileemd_23625_half_map_1.map
Annotationhalf map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map A

Fileemd_23625_half_map_2.map
Annotationhalf map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : HMPV F v3-B in complex with MPE33

EntireName: HMPV F v3-B in complex with MPE33
Components
  • Complex: HMPV F v3-B in complex with MPE33
    • Complex: HMPV F v3-B
      • Protein or peptide: HMPV F v3-B
    • Complex: MPE33
      • Protein or peptide: MPE33 Heavy chain
      • Protein or peptide: MPE33 Light chain

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Supramolecule #1: HMPV F v3-B in complex with MPE33

SupramoleculeName: HMPV F v3-B in complex with MPE33 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all

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Supramolecule #2: HMPV F v3-B

SupramoleculeName: HMPV F v3-B / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Human metapneumovirus
Recombinant expressionOrganism: Homo sapiens (human)

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Supramolecule #3: MPE33

SupramoleculeName: MPE33 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2-#3
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)

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Macromolecule #1: HMPV F v3-B

MacromoleculeName: HMPV F v3-B / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Human metapneumovirus
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: TPQHGLKESY LEESCSTITE GYLSVLRTGW YTNVFTLEVG DVENLTCTDG PSLIKTELDL TKSALRELKT CSADQLAREE QIEGGGGGGF VLGAIALGVA TAAAVTAGIA IAKTIRLESE VNAIKGCLKT TNECVSTLGN GVRVLATAVR ELKEFVSKNL TSAINKNKCD ...String:
TPQHGLKESY LEESCSTITE GYLSVLRTGW YTNVFTLEVG DVENLTCTDG PSLIKTELDL TKSALRELKT CSADQLAREE QIEGGGGGGF VLGAIALGVA TAAAVTAGIA IAKTIRLESE VNAIKGCLKT TNECVSTLGN GVRVLATAVR ELKEFVSKNL TSAINKNKCD IPDLKMAVSF SQFNRRFLNV VRQFSDNAGI TPAISLDLMT DAELARAVSY MPTSAGQIKL MLENRCMVRR KGFGILIGVY GSSVIYMVQL PIFGVIDTPC WIIKAAPSCS EKDGNYACLL REDQGWYCKN AGSTVYYPND KDCETRGDHV FCDTAAGINV AEQSRECNIN ISTTNYPCKV STGRHPISMV ALSPLGALVA CYKGVSCSIG SNRVGIIKQL PKGCSYITNQ DADTVTIDNT VYQLSKVEGE QHVIKGRPVS SSFDPIKFPE DQFNVALDQV FESIENSQAL VDQSNKILNS AESAIGGYIP EAPRDGQAYV RKDGEWVLLS TFLGGLVPR

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Macromolecule #2: MPE33 Heavy chain

MacromoleculeName: MPE33 Heavy chain / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: tgvhsqvqlq esgpglvkps qtlsltctvs gdsisngsyc wnwvrqpagk glewigriyp igntnynpsl ksrvtisldt sknqfslkla svtaadtavy ycarearife gyyyyyygld vwgqgttvtv ssastkgpsv fplapsskst sggtaalgcl vkdyfpepvt ...String:
tgvhsqvqlq esgpglvkps qtlsltctvs gdsisngsyc wnwvrqpagk glewigriyp igntnynpsl ksrvtisldt sknqfslkla svtaadtavy ycarearife gyyyyyygld vwgqgttvtv ssastkgpsv fplapsskst sggtaalgcl vkdyfpepvt vswnsgalts gvhtfpavlq ssglyslssv vtvpssslgt qtyicnvnhk psntkvdkkv epkscdkglv pr

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Macromolecule #3: MPE33 Light chain

MacromoleculeName: MPE33 Light chain / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
EIVLTQSPAT LSLSPGERAT LSCRASQTIS SYLAWYQQKP GQAPRLLISD VSKRATGIPA RFSGSGSGTD FTLTISSLEP EDFAVYYCHQ RSNWDTFGQG TKLEIK

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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

Concentration1.0 mg/mL
BufferpH: 7.4 / Component - Formula: PBS
GridModel: C-flat-1.2/1.3 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Average exposure time: 10.0 sec. / Average electron dose: 63.78 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: CTFFIND (ver. 4)
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 4.79 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 2.15) / Number images used: 33058
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final 3D classificationSoftware - Name: cryoSPARC (ver. 2.15)

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Atomic model buiding 1

Initial modelPDB ID:

5wb0

RefinementSpace: REAL

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