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- EMDB-37632: Cryo-EM structure of bat RsSHC014 spike glycoprotein -

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Basic information

Entry
Database: EMDB / ID: EMD-37632
TitleCryo-EM structure of bat RsSHC014 spike glycoprotein
Map data
Sample
  • Complex: RsSHC014 glycoprotein
    • Protein or peptide: Spike glycoprotein,Fibritin
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
Keywordsspike / VIRAL PROTEIN
Function / homology
Function and homology information


endocytosis involved in viral entry into host cell / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / receptor-mediated virion attachment to host cell / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / host cell plasma membrane / virion membrane / membrane
Similarity search - Function
Fibritin C-terminal / Fibritin C-terminal region / Spike (S) protein S1 subunit, N-terminal domain, SARS-CoV-like / Spike glycoprotein, betacoronavirus / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike S1 subunit, receptor binding domain superfamily, betacoronavirus / Betacoronavirus spike glycoprotein S1, receptor binding ...Fibritin C-terminal / Fibritin C-terminal region / Spike (S) protein S1 subunit, N-terminal domain, SARS-CoV-like / Spike glycoprotein, betacoronavirus / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike S1 subunit, receptor binding domain superfamily, betacoronavirus / Betacoronavirus spike glycoprotein S1, receptor binding / Spike glycoprotein S1, N-terminal domain, betacoronavirus-like / Betacoronavirus-like spike glycoprotein S1, N-terminal / Spike glycoprotein S2, coronavirus, heptad repeat 1 / Spike glycoprotein S2, coronavirus, heptad repeat 2 / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 1 (HR1) region profile. / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 2 (HR2) region profile. / Spike glycoprotein S2 superfamily, coronavirus / Spike glycoprotein S2, coronavirus / Coronavirus spike glycoprotein S2
Similarity search - Domain/homology
Fibritin / Spike glycoprotein
Similarity search - Component
Biological speciesBat SARS-like coronavirus RsSHC014 / Enterobacteria phage T6 (virus)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.87 Å
AuthorsWang X / Qiao S
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: J Virol / Year: 2024
Title: Structural determinants of spike infectivity in bat SARS-like coronaviruses RsSHC014 and WIV1.
Authors: Shuyuan Qiao / Xinquan Wang /
Abstract: The recurrent spillovers of coronaviruses (CoVs) have posed severe threats to public health and the global economy. Bat severe acute respiratory syndrome (SARS)-like CoVs RsSHC014 and WIV1, currently ...The recurrent spillovers of coronaviruses (CoVs) have posed severe threats to public health and the global economy. Bat severe acute respiratory syndrome (SARS)-like CoVs RsSHC014 and WIV1, currently circulating in bat populations, are poised for human emergence. The trimeric spike (S) glycoprotein, responsible for receptor recognition and membrane fusion, plays a critical role in cross-species transmission and infection. Here, we determined the cryo-electron microscopy (EM) structures of the RsSHC014 S protein in the closed state at 2.9 Å, the WIV1 S protein in the closed state at 2.8 Å, and the intermediate state at 4.0 Å. In the intermediate state, one receptor-binding domain (RBD) is in the "down" position, while the other two RBDs exhibit poor density. We also resolved the complex structure of the WIV1 S protein bound to human ACE2 (hACE2) at 4.5 Å, which provides structural basis for the future emergence of WIV1 in humans. Through biochemical experiments, we found that despite strong binding affinities between the RBDs and both human and civet ACE2, the pseudoviruses of RsSHC014, but not WIV1, failed to infect 293T cells overexpressing either human or civet ACE2. Mutagenesis analysis revealed that the Y623H substitution, located in the SD2 region, significantly improved the cell entry efficiency of RsSHC014 pseudoviruses, which is likely accomplished by promoting the open conformation of spike glycoproteins. Our findings emphasize the necessity of both efficient RBD lifting and tight RBD-hACE2 binding for viral infection and underscore the significance of the 623 site of the spike glycoprotein for the infectivity of bat SARS-like CoVs.
IMPORTANCE: The bat SARS-like CoVs RsSHC014 and WIV1 can use hACE2 for cell entry without further adaptation, indicating their potential risk of emergence in human populations. The S glycoprotein, ...IMPORTANCE: The bat SARS-like CoVs RsSHC014 and WIV1 can use hACE2 for cell entry without further adaptation, indicating their potential risk of emergence in human populations. The S glycoprotein, responsible for receptor recognition and membrane fusion, plays a crucial role in cross-species transmission and infection. In this study, we determined the cryo-EM structures of the S glycoproteins of RsSHC014 and WIV1. Detailed comparisons revealed dynamic structural variations within spike proteins. We also elucidated the complex structure of WIV1 S-hACE2, providing structural evidence for the potential emergence of WIV1 in humans. Although RsSHC014 and WIV1 had similar hACE2-binding affinities, they exhibited distinct pseudovirus cell entry behavior. Through mutagenesis and cryo-EM analysis, we revealed that besides the structural variations, the 623 site in the SD2 region is another important structural determinant of spike infectivity.
History
DepositionOct 1, 2023-
Header (metadata) releaseJul 17, 2024-
Map releaseJul 17, 2024-
UpdateNov 6, 2024-
Current statusNov 6, 2024Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_37632.png

Downloads & links

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Map

FileDownload / File: emd_37632.map.gz / Format: CCP4 / Size: 98.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 296 pix.
= 324.978 Å		1.1 Å/pix.
x 296 pix.
= 324.978 Å		1.1 Å/pix.
x 296 pix.
= 324.978 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.0979 Å
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Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.00821
Minimum - Maximum-0.06319178 - 0.12838806
Average (Standard dev.)0.00022473121 (±0.0031713962)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions296296296
Spacing296296296
CellA=B=C: 324.97842 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_37632_half_map_1.map
Projections & Slices
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Slices (1/2)
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Half map: #2

Fileemd_37632_half_map_2.map
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Sample components

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Entire : RsSHC014 glycoprotein

EntireName: RsSHC014 glycoprotein
Components
  • Complex: RsSHC014 glycoprotein
    • Protein or peptide: Spike glycoprotein,Fibritin
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: RsSHC014 glycoprotein

SupramoleculeName: RsSHC014 glycoprotein / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Bat SARS-like coronavirus RsSHC014

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Macromolecule #1: Spike glycoprotein,Fibritin

MacromoleculeName: Spike glycoprotein,Fibritin / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Enterobacteria phage T6 (virus)
Molecular weightTheoretical: 140.530312 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MKLLVLVFAT LVSSYTIEKC LDFDDRTPPA NTQFLSSHRG VYYPDDIFRS NVLHLVQDHF LPFDSNVTRF ITFGLNFDNP IIPFRDGIY FAATEKSNVI RGWVFGSTMN NKSQSVIIMN NSTNLVIRAC NFELCDNPFF VVLKSNNTQI PSYIFNNAFN C TFEYVSKD ...String:
MKLLVLVFAT LVSSYTIEKC LDFDDRTPPA NTQFLSSHRG VYYPDDIFRS NVLHLVQDHF LPFDSNVTRF ITFGLNFDNP IIPFRDGIY FAATEKSNVI RGWVFGSTMN NKSQSVIIMN NSTNLVIRAC NFELCDNPFF VVLKSNNTQI PSYIFNNAFN C TFEYVSKD FNLDLGEKPG NFKDLREFVF RNKDGFLHVY SGYQPISAAS GLPTGFNALK PIFKLPLGIN ITNFRTLLTA FP PRPDYWG TSAAAYFVGY LKPTTFMLKY DENGTITDAV DCSQNPLAEL KCSVKSFEID KGIYQTSNFR VAPSKEVVRF PNI TNLCPF GEVFNATTFP SVYAWERKRI SNCVADYSVL YNSTSFSTFK CYGVSATKLN DLCFSNVYAD SFVVKGDDVR QIAP GQTGV IADYNYKLPD DFLGCVLAWN TNSKDSSTSG NYNYLYRWVR RSKLNPYERD LSNDIYSPGG QSCSAVGPNC YNPLR PYGF FTTAGVGHQP YRVVVLSFEL LNAPATVCGP KLSTDLIKNQ CVNFNFNGLT GTGVLTPSSK RFQPFQQFGR DVSDFT DSV RDPKTSEILD ISPCSFGGVS VITPGTNTSS EVAVLYQDVN CTDVPVAIHA DQLTPSWRVY STGNNVFQTQ AGCLIGA EH VDTSYECDIP IGAGICASYH TVSSLRSTSQ KSIVAYTMSL GADSSIAYSN NTIAIPTNFS ISITTEVMPV SMAKTSVD C NMYICGDSTE CANLLLQYGS FCTQLNRALS GIAVEQDRNT REVFAQVKQM YKTPTLKDFG GFNFSQILPD PLKPTKRSF IEDLLFNKVT LADAGFMKQY GECLGDINAR DLICAQKFNG LTVLPPLLTD DMIAAYTAAL VSGTATAGWT FGAGAALQIP FAMQMAYRF NGIGVTQNVL YENQKQIANQ FNKAISQIQE SLTTTSTALG KLQDVVNQNA QALNTLVKQL SSNFGAISSV L NDILSRLD PPEAEVQIDR LITGRLQSLQ TYVTQQLIRA AEIRASANLA ATKMSECVLG QSKRVDFCGK GYHLMSFPQA AP HGVVFLH VTYVPSQERN FTTAPAICHE GKAYFPREGV FVFNGTSWFI TQRNFFSPQI ITTDNTFVSG SCDVVIGIIN NTV YDPLQP ELDSFKEELD KYFKNHTSPD VDLGDISGIN ASVVNIQKEI DRLNEVAKNL NESLIDLQEL GKYEQGSGYI PEAP RDGQA YVRKDGEWVL LSTFLGRSLE VLFQGPGHHH HHHHHSAWSH PQFEKGGGSG GGGSGGSAWS HPQFEK

UniProtKB: Spike glycoprotein, Fibritin

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Macromolecule #4: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 4 / Number of copies: 6 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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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

Concentration0.3 mg/mL
BufferpH: 7.2
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.8 µm / Nominal defocus min: 1.5 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

5x58

Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.87 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 250439
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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