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- EMDB-17170: Local refinement of the SARS-CoV-2 Spike RBD bound to TMEM106B -

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Basic information

Entry
Database: EMDB / ID: EMD-17170
TitleLocal refinement of the SARS-CoV-2 Spike RBD bound to TMEM106B
Map data
Sample
  • Complex: Trimeric SARS-CoV-2 Spike bound to one molecule of TMEM106B
    • Complex: SARS-CoV-2 Spike ectodomain (Belgium/GHB-03021) stabilized in trimeric pre-fusion state
      • Protein or peptide: SARS-CoV-2 Spike ectodomain (Belgium/GHB-03021) stabilized in trimeric pre-fusion state
    • Complex: TMEM106B lumenall domain
      • Protein or peptide: TMEM106B lumenall domain
KeywordsSARS-CoV-2 / Spike / ectodomain / TMEM106B / luminal domain / VIRAL PROTEIN
Biological speciesSevere acute respiratory syndrome coronavirus 2 / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.83 Å
AuthorsCHEREPANOV P
Funding support United Kingdom, 4 items
OrganizationGrant numberCountry
Wellcome TrustFC001061 United Kingdom
Medical Research Council (MRC, United Kingdom)FC001061 United Kingdom
Cancer Research UKFC001061 United Kingdom
Wellcome Trust206175/Z/17/Z United Kingdom
CitationJournal: Cell / Year: 2023
Title: TMEM106B is a receptor mediating ACE2-independent SARS-CoV-2 cell entry.
Authors: Jim Baggen / Maarten Jacquemyn / Leentje Persoons / Els Vanstreels / Valerie E Pye / Antoni G Wrobel / Valeria Calvaresi / Stephen R Martin / Chloë Roustan / Nora B Cronin / Eamonn Reading ...Authors: Jim Baggen / Maarten Jacquemyn / Leentje Persoons / Els Vanstreels / Valerie E Pye / Antoni G Wrobel / Valeria Calvaresi / Stephen R Martin / Chloë Roustan / Nora B Cronin / Eamonn Reading / Hendrik Jan Thibaut / Thomas Vercruysse / Piet Maes / Frederik De Smet / Angie Yee / Toey Nivitchanyong / Marina Roell / Natalia Franco-Hernandez / Herve Rhinn / Alusha Andre Mamchak / Maxime Ah Young-Chapon / Eric Brown / Peter Cherepanov / Dirk Daelemans /
Abstract: SARS-CoV-2 is associated with broad tissue tropism, a characteristic often determined by the availability of entry receptors on host cells. Here, we show that TMEM106B, a lysosomal transmembrane ...SARS-CoV-2 is associated with broad tissue tropism, a characteristic often determined by the availability of entry receptors on host cells. Here, we show that TMEM106B, a lysosomal transmembrane protein, can serve as an alternative receptor for SARS-CoV-2 entry into angiotensin-converting enzyme 2 (ACE2)-negative cells. Spike substitution E484D increased TMEM106B binding, thereby enhancing TMEM106B-mediated entry. TMEM106B-specific monoclonal antibodies blocked SARS-CoV-2 infection, demonstrating a role of TMEM106B in viral entry. Using X-ray crystallography, cryogenic electron microscopy (cryo-EM), and hydrogen-deuterium exchange mass spectrometry (HDX-MS), we show that the luminal domain (LD) of TMEM106B engages the receptor-binding motif of SARS-CoV-2 spike. Finally, we show that TMEM106B promotes spike-mediated syncytium formation, suggesting a role of TMEM106B in viral fusion. Together, our findings identify an ACE2-independent SARS-CoV-2 infection mechanism that involves cooperative interactions with the receptors heparan sulfate and TMEM106B.
History
DepositionApr 20, 2023-
Header (metadata) releaseJul 19, 2023-
Map releaseJul 19, 2023-
UpdateAug 16, 2023-
Current statusAug 16, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_17170.png

Downloads & links

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Map

FileDownload / File: emd_17170.map.gz / Format: CCP4 / Size: 282.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.85 Å/pix.
x 420 pix.
= 357. Å		0.85 Å/pix.
x 420 pix.
= 357. Å		0.85 Å/pix.
x 420 pix.
= 357. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.85 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.3
Minimum - Maximum-1.0913188 - 1.5507456
Average (Standard dev.)0.000018003651 (±0.010572615)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions420420420
Spacing420420420
CellA=B=C: 357.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_17170_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_17170_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Trimeric SARS-CoV-2 Spike bound to one molecule of TMEM106B

EntireName: Trimeric SARS-CoV-2 Spike bound to one molecule of TMEM106B
Components
  • Complex: Trimeric SARS-CoV-2 Spike bound to one molecule of TMEM106B
    • Complex: SARS-CoV-2 Spike ectodomain (Belgium/GHB-03021) stabilized in trimeric pre-fusion state
      • Protein or peptide: SARS-CoV-2 Spike ectodomain (Belgium/GHB-03021) stabilized in trimeric pre-fusion state
    • Complex: TMEM106B lumenall domain
      • Protein or peptide: TMEM106B lumenall domain

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Supramolecule #1: Trimeric SARS-CoV-2 Spike bound to one molecule of TMEM106B

SupramoleculeName: Trimeric SARS-CoV-2 Spike bound to one molecule of TMEM106B
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Molecular weightTheoretical: 450 KDa

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Supramolecule #2: SARS-CoV-2 Spike ectodomain (Belgium/GHB-03021) stabilized in tri...

SupramoleculeName: SARS-CoV-2 Spike ectodomain (Belgium/GHB-03021) stabilized in trimeric pre-fusion state
type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2

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Supramolecule #3: TMEM106B lumenall domain

SupramoleculeName: TMEM106B lumenall domain / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: SARS-CoV-2 Spike ectodomain (Belgium/GHB-03021) stabilized in tri...

MacromoleculeName: SARS-CoV-2 Spike ectodomain (Belgium/GHB-03021) stabilized in trimeric pre-fusion state
type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2 / Strain: Belgium/GHB-03021
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: mfvflvllpl vssqcvnltt rtqlppaytn sftrgvyypd kvfrssvlhs tqdlflpffs nvtwfhakrf dnpvlpfndg vyfasteksn iirgwifgtt ldsktqslli vnnatnvvik vcefqfcndp flgvyyhknn kswmesefrv yssannctfe yvsqpflmdl ...String:
mfvflvllpl vssqcvnltt rtqlppaytn sftrgvyypd kvfrssvlhs tqdlflpffs nvtwfhakrf dnpvlpfndg vyfasteksn iirgwifgtt ldsktqslli vnnatnvvik vcefqfcndp flgvyyhknn kswmesefrv yssannctfe yvsqpflmdl egkqgnfknl refvfknidg yfkiyskhtp inlvrdlpqg fsaleplvdl piginitrfq tllalhrsyl tpgdsssgwt agaaayyvgy lqprtfllky nengtitdav dcaldplset kctlksftve kgiyqtsnfr vqptesivrf pnitnlcpfg evfnatrfas vyawnrkris ncvadysvly nsasfstfkc ygvsptklnd lcftnvyads fvirgdevrq iapgqtgkia dynyklpddf tgcviawnsn nldskvggny nylyrlfrks nlkpferdis teiyqagstp cngvdgfncy fplqsygfqp tngvgyqpyr vvvlsfellh apatvcgpkk stnlvknkcv nfnfngltgt gvltesnkkf lpfqqfgrdi adttdavrdp qtleilditp csfggvsvit pgtntsnqva vlyqdvncte vpvaihadql tptwrvystg snvfqtragc ligaehvnns yecdipigag icasyqprra rsvasqsiia ytmslgaens vaysnnsiai ptnftisvtt eilpvsmtkt svdctmyicg dstecsnlll qygsfctqln raltgiaveq dkntqevfaq vkqiyktppi kdfggfnfsq ilpdpskpik rspiedllfn kvtladagfi kqygdclgdi aardlicaqk fngltvlppl ltdemiaqyt sallagtits gwtfgagpal qipfpmqmay rfngigvtqn vlyenqklia nqfnsaigki qdslsstpsa lgklqdvvnq naqalntlvk qlssnfgais svlndilsrl dppeaevqid rlitgrlqsl qtyvtqqlir aaeirasanl aatkmsecvl gqskrvdfcg kgyhlmsfpq saphgvvflh vtyvpaqekn fttapaichd gkahfpregv fvsngthwfv tqrnfyepqi ittdntfvsg ncdvvigivn ntvydplqpe ldsfkeeldk yfknhtspdv dlgdisgina svvniqkeid rlnevaknln eslidlqelg kyeqgsgyip eaprdgqayv rkdgewvlls tflgrslevl fqgpgsawsh pqfekgggsg gggsggsaws hpqfek

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Macromolecule #2: TMEM106B lumenall domain

MacromoleculeName: TMEM106B lumenall domain / type: protein_or_peptide / ID: 2
Details: Lumenal domain of human TMEM106B with an N-terminal signal peptide and a C-terminal His6 tag.
Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: metdtlllwv lllwvpgstg daaqprsidv kyigvksayv sydvqkrtiy lnitntlnit nnnyysveve nitaqvqfsk tvigkarlnn itiigpldmk qidytvptvi aeemsymydf ctlisikvhn ivlmmqvtvt ttyfghseqi sqeryqyvdc grnttyqlgq ...String:
metdtlllwv lllwvpgstg daaqprsidv kyigvksayv sydvqkrtiy lnitntlnit nnnyysveve nitaqvqfsk tvigkarlnn itiigpldmk qidytvptvi aeemsymydf ctlisikvhn ivlmmqvtvt ttyfghseqi sqeryqyvdc grnttyqlgq seylnvlqpq qGSGENLYFQ SAGHHHHHH

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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 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
150.0 mMNaClsodium chloride
1.0 mMEDTA
25.0 mMTris
0.1 %n-octyl glucoside

Details: 150 mM NaCl, 1 mM EDTA, and 25 mM Tris-HCl, pH 8.0
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 2 / Number real images: 37079 / Average electron dose: 30.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.1 µm / Nominal defocus min: 0.5 µm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1799554
Startup modelType of model: OTHER
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.83 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.1)
Details: Local refinement, focussed on TMEM106B and the associated RBD, as implemented in cryoSPARC
Number images used: 25781
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.1)
Final 3D classificationNumber classes: 4 / Avg.num./class: 50000 / Software - Name: RELION (ver. 4.0)
FSC plot (resolution estimation)

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