+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-22610 | |||||||||
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Title | Nucleotide bound SARS-CoV-2 Nsp15 | |||||||||
Map data | Nucleotide bound SARS-CoV-2 Nsp15 | |||||||||
Sample |
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Keywords | Ribonuclease / VIRAL PROTEIN | |||||||||
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 / snRNP Assembly / TRAF3-dependent IRF activation pathway / Replication of the SARS-CoV-2 genome / Hydrolases; Acting on ester bonds; Exoribonucleases producing 5'-phosphomonoesters / : / double membrane vesicle viral factory outer membrane / 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: 3.3 Å | |||||||||
Authors | Pillon MC / Stanley RE | |||||||||
Funding support | United States, 1 items
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Citation | Journal: Nat Commun / Year: 2021 Title: Cryo-EM structures of the SARS-CoV-2 endoribonuclease Nsp15 reveal insight into nuclease specificity and dynamics. Authors: Monica C Pillon / Meredith N Frazier / Lucas B Dillard / Jason G Williams / Seda Kocaman / Juno M Krahn / Lalith Perera / Cassandra K Hayne / Jacob Gordon / Zachary D Stewart / Mack Sobhany ...Authors: Monica C Pillon / Meredith N Frazier / Lucas B Dillard / Jason G Williams / Seda Kocaman / Juno M Krahn / Lalith Perera / Cassandra K Hayne / Jacob Gordon / Zachary D Stewart / Mack Sobhany / Leesa J Deterding / Allen L Hsu / Venkata P Dandey / Mario J Borgnia / Robin E Stanley / Abstract: Nsp15, a uridine specific endoribonuclease conserved across coronaviruses, processes viral RNA to evade detection by host defense systems. Crystal structures of Nsp15 from different coronaviruses ...Nsp15, a uridine specific endoribonuclease conserved across coronaviruses, processes viral RNA to evade detection by host defense systems. Crystal structures of Nsp15 from different coronaviruses have shown a common hexameric assembly, yet how the enzyme recognizes and processes RNA remains poorly understood. Here we report a series of cryo-EM reconstructions of SARS-CoV-2 Nsp15, in both apo and UTP-bound states. The cryo-EM reconstructions, combined with biochemistry, mass spectrometry, and molecular dynamics, expose molecular details of how critical active site residues recognize uridine and facilitate catalysis of the phosphodiester bond. Mass spectrometry revealed the accumulation of cyclic phosphate cleavage products, while analysis of the apo and UTP-bound datasets revealed conformational dynamics not observed by crystal structures that are likely important to facilitate substrate recognition and regulate nuclease activity. Collectively, these findings advance understanding of how Nsp15 processes viral RNA and provide a structural framework for the development of new therapeutics. | |||||||||
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_22610.map.gz | 43.4 MB | EMDB map data format | |
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Header (meta data) | emd-22610-v30.xml emd-22610.xml | 10.4 KB 10.4 KB | Display Display | EMDB header |
Images | emd_22610.png | 76.9 KB | ||
Filedesc metadata | emd-22610.cif.gz | 5.3 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-22610 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-22610 | HTTPS FTP |
-Related structure data
Related structure data | 7k0rMC 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_22610.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Nucleotide bound SARS-CoV-2 Nsp15 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.145 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Nucleotide bound complex of SARS-CoV-2 Nsp15
Entire | Name: Nucleotide bound complex of SARS-CoV-2 Nsp15 |
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Components |
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-Supramolecule #1: Nucleotide bound complex of SARS-CoV-2 Nsp15
Supramolecule | Name: Nucleotide bound complex of SARS-CoV-2 Nsp15 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 |
-Macromolecule #1: Uridylate-specific endoribonuclease
Macromolecule | Name: Uridylate-specific endoribonuclease / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO / EC number: Hydrolases; Acting on ester bonds |
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Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 |
Molecular weight | Theoretical: 40.764336 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: GSHMLEENLY FQSLEMSLEN VAFNVVNKGH FDGQQGEVPV SIINNTVYTK VDGVDVELFE NKTTLPVNVA FELWAKRNIK PVPEVKILN NLGVDIAANT VIWDYKRDAP AHISTIGVCS MTDIAKKPTE TICAPLTVFF DGRVDGQVDL FRNARNGVLI T EGSVKGLQ ...String: GSHMLEENLY FQSLEMSLEN VAFNVVNKGH FDGQQGEVPV SIINNTVYTK VDGVDVELFE NKTTLPVNVA FELWAKRNIK PVPEVKILN NLGVDIAANT VIWDYKRDAP AHISTIGVCS MTDIAKKPTE TICAPLTVFF DGRVDGQVDL FRNARNGVLI T EGSVKGLQ PSVGPKQASL NGVTLIGEAV KTQFNYYKKV DGVVQQLPET YFTQSRNLQE FKPRSQMEID FLELAMDEFI ER YKLEGYA FEHIVYGDFS HSQLGGLHLL IGLAKRFKES PFELEDFIPM DSTVKNYFIT DAQTGSSKCV CSVIDLLLDD FVE IIKSQD LSVVSKVVKV TIDYTEISFM LWCKDGHVET FYPKLQ UniProtKB: Replicase polyprotein 1ab |
-Macromolecule #2: URIDINE-5'-MONOPHOSPHATE
Macromolecule | Name: URIDINE-5'-MONOPHOSPHATE / type: ligand / ID: 2 / Number of copies: 6 / Formula: U5P |
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Molecular weight | Theoretical: 324.181 Da |
Chemical component information | ChemComp-U: |
-Macromolecule #3: PHOSPHATE ION
Macromolecule | Name: PHOSPHATE ION / type: ligand / ID: 3 / Number of copies: 6 / Formula: PO4 |
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Molecular weight | Theoretical: 94.971 Da |
Chemical component information | ChemComp-PO4: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TALOS ARCTICA |
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Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 54.0 e/Å2 |
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: NONE |
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Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 65393 |