+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-23970 | |||||||||||||||
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Title | Full length SARS-CoV-2 Nsp2 | |||||||||||||||
Map data | SARS-CoV-2 Nsp2 main map | |||||||||||||||
Sample |
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Function / homology | Function and homology information protein guanylyltransferase activity / RNA endonuclease activity, producing 3'-phosphomonoesters / mRNA guanylyltransferase activity / 5'-3' RNA helicase activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of TBK1 activity / Lyases; Phosphorus-oxygen lyases / Assembly of the SARS-CoV-2 Replication-Transcription Complex (RTC) / Maturation of replicase proteins / ISG15-specific peptidase activity / Transcription of SARS-CoV-2 sgRNAs ...protein guanylyltransferase activity / RNA endonuclease activity, producing 3'-phosphomonoesters / mRNA guanylyltransferase activity / 5'-3' RNA helicase activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of TBK1 activity / Lyases; Phosphorus-oxygen lyases / Assembly of the SARS-CoV-2 Replication-Transcription Complex (RTC) / Maturation of replicase proteins / ISG15-specific peptidase activity / Transcription of SARS-CoV-2 sgRNAs / Translation of Replicase and Assembly of the Replication Transcription Complex / TRAF3-dependent IRF activation pathway / snRNP Assembly / Replication of the SARS-CoV-2 genome / double membrane vesicle viral factory outer membrane / Hydrolases; Acting on ester bonds; Exoribonucleases producing 5'-phosphomonoesters / SARS coronavirus main proteinase / host cell endosome / 3'-5'-RNA exonuclease activity / : / host cell endoplasmic reticulum-Golgi intermediate compartment / symbiont-mediated suppression of host toll-like receptor signaling pathway / symbiont-mediated degradation of host mRNA / mRNA guanylyltransferase / symbiont-mediated suppression of host ISG15-protein conjugation / G-quadruplex RNA binding / SARS-CoV-2 modulates host translation machinery / omega peptidase activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of IRF3 activity / mRNA (guanine-N7)-methyltransferase / methyltransferase cap1 / host cell Golgi apparatus / symbiont-mediated perturbation of host ubiquitin-like protein modification / mRNA (nucleoside-2'-O-)-methyltransferase activity / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / DNA helicase / ubiquitinyl hydrolase 1 / cysteine-type deubiquitinase activity / Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases / single-stranded RNA binding / host cell endoplasmic reticulum membrane / host cell perinuclear region of cytoplasm / viral protein processing / lyase activity / RNA helicase / induction by virus of host autophagy / RNA-directed RNA polymerase / copper ion binding / symbiont-mediated suppression of host gene expression / viral RNA genome replication / cysteine-type endopeptidase activity / RNA-dependent RNA polymerase activity / DNA-templated transcription / lipid binding / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / host cell nucleus / SARS-CoV-2 activates/modulates innate and adaptive immune responses / ATP hydrolysis activity / proteolysis / RNA binding / zinc ion binding / ATP binding / membrane Similarity search - Function | |||||||||||||||
Biological species | Severe acute respiratory syndrome coronavirus 2 | |||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.76 Å | |||||||||||||||
Authors | QCRG Structural Biology Consortium | |||||||||||||||
Funding support | United States, 4 items
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Citation | Journal: bioRxiv / Year: 2021 Title: CryoEM and AI reveal a structure of SARS-CoV-2 Nsp2, a multifunctional protein involved in key host processes. Authors: Meghna Gupta / Caleigh M Azumaya / Michelle Moritz / Sergei Pourmal / Amy Diallo / Gregory E Merz / Gwendolyn Jang / Mehdi Bouhaddou / Andrea Fossati / Axel F Brilot / Devan Diwanji / Evelyn ...Authors: Meghna Gupta / Caleigh M Azumaya / Michelle Moritz / Sergei Pourmal / Amy Diallo / Gregory E Merz / Gwendolyn Jang / Mehdi Bouhaddou / Andrea Fossati / Axel F Brilot / Devan Diwanji / Evelyn Hernandez / Nadia Herrera / Huong T Kratochvil / Victor L Lam / Fei Li / Yang Li / Henry C Nguyen / Carlos Nowotny / Tristan W Owens / Jessica K Peters / Alexandrea N Rizo / Ursula Schulze-Gahmen / Amber M Smith / Iris D Young / Zanlin Yu / Daniel Asarnow / Christian Billesbølle / Melody G Campbell / Jen Chen / Kuei-Ho Chen / Un Seng Chio / Miles Sasha Dickinson / Loan Doan / Mingliang Jin / Kate Kim / Junrui Li / Yen-Li Li / Edmond Linossi / Yanxin Liu / Megan Lo / Jocelyne Lopez / Kyle E Lopez / Adamo Mancino / Frank R Moss / Michael D Paul / Komal Ishwar Pawar / Adrian Pelin / Thomas H Pospiech / Cristina Puchades / Soumya Govinda Remesh / Maliheh Safari / Kaitlin Schaefer / Ming Sun / Mariano C Tabios / Aye C Thwin / Erron W Titus / Raphael Trenker / Eric Tse / Tsz Kin Martin Tsui / Feng Wang / Kaihua Zhang / Yang Zhang / Jianhua Zhao / Fengbo Zhou / Yuan Zhou / Lorena Zuliani-Alvarez / / David A Agard / Yifan Cheng / James S Fraser / Natalia Jura / Tanja Kortemme / Aashish Manglik / Daniel R Southworth / Robert M Stroud / Danielle L Swaney / Nevan J Krogan / Adam Frost / Oren S Rosenberg / Kliment A Verba / Abstract: The SARS-CoV-2 protein Nsp2 has been implicated in a wide range of viral processes, but its exact functions, and the structural basis of those functions, remain unknown. Here, we report an atomic ...The SARS-CoV-2 protein Nsp2 has been implicated in a wide range of viral processes, but its exact functions, and the structural basis of those functions, remain unknown. Here, we report an atomic model for full-length Nsp2 obtained by combining cryo-electron microscopy with deep learning-based structure prediction from AlphaFold2. The resulting structure reveals a highly-conserved zinc ion-binding site, suggesting a role for Nsp2 in RNA binding. Mapping emerging mutations from variants of SARS-CoV-2 on the resulting structure shows potential host-Nsp2 interaction regions. Using structural analysis together with affinity tagged purification mass spectrometry experiments, we identify Nsp2 mutants that are unable to interact with the actin-nucleation-promoting WASH protein complex or with GIGYF2, an inhibitor of translation initiation and modulator of ribosome-associated quality control. Our work suggests a potential role of Nsp2 in linking viral transcription within the viral replication-transcription complexes (RTC) to the translation initiation of the viral message. Collectively, the structure reported here, combined with mutant interaction mapping, provides a foundation for functional studies of this evolutionary conserved coronavirus protein and may assist future drug design. | |||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_23970.map.gz | 78.3 MB | EMDB map data format | |
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Header (meta data) | emd-23970-v30.xml emd-23970.xml | 17.7 KB 17.7 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_23970_fsc.xml | 10 KB | Display | FSC data file |
Images | emd_23970.png | 41.7 KB | ||
Masks | emd_23970_msk_1.map | 83.7 MB | Mask map | |
Others | emd_23970_half_map_1.map.gz emd_23970_half_map_2.map.gz | 65.6 MB 65.4 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-23970 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-23970 | HTTPS FTP |
-Related structure data
Related structure data | 7mswMC 7msxC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_23970.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | SARS-CoV-2 Nsp2 main map | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.834 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_23970_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: SARS-CoV-2 Nsp2 half map - Odd
File | emd_23970_half_map_1.map | ||||||||||||
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Annotation | SARS-CoV-2 Nsp2 half map - Odd | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: SARS-CoV-2 Nsp2 half map - Even
File | emd_23970_half_map_2.map | ||||||||||||
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Annotation | SARS-CoV-2 Nsp2 half map - Even | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : SARS-CoV-2 Nsp2
Entire | Name: SARS-CoV-2 Nsp2 |
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Components |
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-Supramolecule #1: SARS-CoV-2 Nsp2
Supramolecule | Name: SARS-CoV-2 Nsp2 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Molecular weight | Experimental: 71 KDa |
-Macromolecule #1: Non-structural protein 2
Macromolecule | Name: Non-structural protein 2 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 |
Molecular weight | Theoretical: 70.590219 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: AYTRYVDNNF CGPDGYPLEC IKDLLARAGK ASCTLSEQLD FIDTKRGVYC CREHEHEIAW YTERSEKSYE LQTPFEIKLA KKFDTFNGE CPNFVFPLNS IIKTIQPRVE KKKLDGFMGR IRSVYPVASP NECNQMCLST LMKCDHCGET SWQTGDFVKA T CEFCGTEN ...String: AYTRYVDNNF CGPDGYPLEC IKDLLARAGK ASCTLSEQLD FIDTKRGVYC CREHEHEIAW YTERSEKSYE LQTPFEIKLA KKFDTFNGE CPNFVFPLNS IIKTIQPRVE KKKLDGFMGR IRSVYPVASP NECNQMCLST LMKCDHCGET SWQTGDFVKA T CEFCGTEN LTKEGATTCG YLPQNAVVKI YCPACHNSEV GPEHSLAEYH NESGLKTILR KGGRTIAFGG CVFSYVGCHN KC AYWVPRA SANIGCNHTG VVGEGSEGLN DNLLEILQKE KVNINIVGDF KLNEEIAIIL ASFSASTSAF VETVKGLDYK AFK QIVESC GNFKVTKGKA KKGAWNIGEQ KSILSPLYAF ASEAARVVRS IFSRTLETAQ NSVRVLQKAA ITILDGISQY SLRL IDAMM FTSDLATNNL VVMAYITGGV VQLTSQWLTN IFGTVYEKLK PVLDWLEEKF KEGVEFLRDG WEIVKFISTC ACEIV GGQI VTCAKEIKES VQTFFKLVNK FLALCADSII IGGAKLKALN LGETFVTHSK GLYRKCVKSR EETGLLMPLK APKEII FLE GETLPTEVLT EEVVLKTGDL QPLEQPTSEA VEAPLVGTPV CINGLMLLEI KDTEKYCALA PNMMVTNNTF TLKGG |
-Macromolecule #2: ZINC ION
Macromolecule | Name: ZINC ION / type: ligand / ID: 2 / Number of copies: 3 / Formula: ZN |
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Molecular weight | Theoretical: 65.409 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.42 mg/mL | ||||||||||||
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Buffer | pH: 8 Component:
Details: Filtered and degassed before running FPLC | ||||||||||||
Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE | ||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV Details: Blot for 5 seconds before plunging into liquid ethane. | ||||||||||||
Details | This sample was monodisperse |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 105000 |
Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 66.0 e/Å2 Details: Two datasets comprising of 804 and 1116 118-frame super-resolution movies each were collected with a 3x3 image shift at a magnification of 105,000x with physical pixel size of 0.834 ang/pix ...Details: Two datasets comprising of 804 and 1116 118-frame super-resolution movies each were collected with a 3x3 image shift at a magnification of 105,000x with physical pixel size of 0.834 ang/pix on a Titan Krios (ThermoFisher) equipped with a K3 camera and a Bioquantum energy filter (Gatan) set to a slit width of 20 eV. Collection dose rate was 8 e-/pixel/second for a total dose of 66 e-/ang^2. Defocus range was 0.8 to 2.4 micron. Each collection was performed with semi-automated scripts in SerialEM. |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |