+
Open data
-
Basic information
Entry | ![]() | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Title | SARS-CoV-2 endoribonuclease Nsp15 bound to dsRNA | ||||||||||||
![]() | SARS-CoV-2 endoribonuclease Nsp15 bound to dsRNA | ||||||||||||
![]() |
| ||||||||||||
![]() | endoribonuclease / VIRAL PROTEIN / VIRAL PROTEIN-RNA complex | ||||||||||||
Function / homology | ![]() protein guanylyltransferase activity / RNA endonuclease activity, producing 3'-phosphomonoesters / mRNA guanylyltransferase activity / 5'-3' RNA helicase activity / Lyases; Phosphorus-oxygen lyases / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of TBK1 activity / 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 / Lyases; Phosphorus-oxygen lyases / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of TBK1 activity / 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 / Replication of the SARS-CoV-2 genome / snRNP Assembly / double membrane vesicle viral factory outer membrane / Hydrolases; Acting on ester bonds; Exoribonucleases producing 5'-phosphomonoesters / 5'-3' DNA helicase activity / SARS coronavirus main proteinase / host cell endosome / host cell endoplasmic reticulum-Golgi intermediate compartment / 3'-5'-RNA exonuclease activity / symbiont-mediated degradation of host mRNA / symbiont-mediated suppression of host toll-like receptor signaling pathway / 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 / host cell Golgi apparatus / methyltransferase cap1 / symbiont-mediated perturbation of host ubiquitin-like protein modification / mRNA (nucleoside-2'-O-)-methyltransferase activity / DNA helicase / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / ubiquitinyl hydrolase 1 / cysteine-type deubiquitinase activity / Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases / single-stranded RNA binding / host cell perinuclear region of cytoplasm / host cell endoplasmic reticulum membrane / viral protein processing / lyase activity / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / RNA helicase / induction by virus of host autophagy / copper ion binding / RNA-directed RNA polymerase / viral RNA genome replication / cysteine-type endopeptidase activity / RNA-dependent RNA polymerase activity / virus-mediated perturbation of host defense response / DNA-templated transcription / lipid binding / 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 | ![]() ![]() | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.43 Å | ||||||||||||
![]() | Frazier MN / Krahn JM | ||||||||||||
Funding support | ![]()
| ||||||||||||
![]() | Journal: bioRxiv / Year: 2022 Title: Flipped Over U: Structural Basis for dsRNA Cleavage by the SARS-CoV-2 Endoribonuclease. Authors: Meredith N Frazier / Isha M Wilson / Juno M Krahn / Kevin John Butay / Lucas B Dillard / Mario J Borgnia / Robin E Stanley / ![]() Abstract: Coronaviruses generate double-stranded (ds) RNA intermediates during viral replication that can activate host immune sensors. To evade activation of the host pattern recognition receptor MDA5, ...Coronaviruses generate double-stranded (ds) RNA intermediates during viral replication that can activate host immune sensors. To evade activation of the host pattern recognition receptor MDA5, coronaviruses employ Nsp15, which is uridine-specific endoribonuclease. Nsp15 is proposed to associate with the coronavirus replication-transcription complex within double-membrane vesicles to cleave these dsRNA intermediates. How Nsp15 recognizes and processes dsRNA is poorly understood because previous structural studies of Nsp15 have been limited to small single-stranded (ss) RNA substrates. Here we present cryo-EM structures of SARS-CoV-2 Nsp15 bound to a 52nt dsRNA. We observed that the Nsp15 hexamer forms a platform for engaging dsRNA across multiple protomers. The structures, along with site-directed mutagenesis and RNA cleavage assays revealed critical insight into dsRNA recognition and processing. To process dsRNA Nsp15 utilizes a base-flipping mechanism to properly orient the uridine within the active site for cleavage. Our findings show that Nsp15 is a distinctive endoribonuclease that can cleave both ss- and dsRNA effectively. | ||||||||||||
History |
|
-
Structure visualization
Supplemental images |
---|
-
Downloads & links
-EMDB archive
Map data | ![]() | 46.7 MB | ![]() | |
---|---|---|---|---|
Header (meta data) | ![]() ![]() | 13 KB 13 KB | Display Display | ![]() |
Images | ![]() | 61.5 KB | ||
Filedesc metadata | ![]() | 5.7 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 512.3 KB | Display | ![]() |
---|---|---|---|---|
Full document | ![]() | 511.9 KB | Display | |
Data in XML | ![]() | 6.3 KB | Display | |
Data in CIF | ![]() | 7.1 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 7tqvMC ![]() 7tj2C C: citing same article ( M: atomic model generated by this map |
---|---|
Similar structure data | Similarity search - Function & homology ![]() |
-
Links
EMDB pages | ![]() ![]() |
---|---|
Related items in Molecule of the Month |
-
Map
File | ![]() | ||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | SARS-CoV-2 endoribonuclease Nsp15 bound to dsRNA | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.90396 Å | ||||||||||||||||||||
Density |
| ||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
|
-Supplemental data
-
Sample components
-Entire : Nsp15/dsRNA
Entire | Name: Nsp15/dsRNA |
---|---|
Components |
|
-Supramolecule #1: Nsp15/dsRNA
Supramolecule | Name: Nsp15/dsRNA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
---|---|
Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 250 KDa |
-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 |
---|---|
Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 40.69727 KDa |
Recombinant expression | Organism: ![]() ![]() |
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 FEAIVYGDFS HSQLGGLHLL IGLAKRFKES PFELEDFIPM DSTVKNYFIT DAQTGSSKCV CSVIDLLLDD FVE IIKSQD LSVVSKVVKV TIDYTEISFM LWCKDGHVET FYPKLQ UniProtKB: Replicase polyprotein 1ab |
-Macromolecule #2: RNA (33-MER)
Macromolecule | Name: RNA (33-MER) / type: rna / ID: 2 / Number of copies: 1 |
---|---|
Source (natural) | Organism: unidentified (others) |
Molecular weight | Theoretical: 16.781023 KDa |
Sequence | String: GGAGGUAGUA GGUUGUAUAG UAGUAAGACC AGACCCUAGA CCAAUUCAUG CC |
-Macromolecule #3: RNA (33-MER)
Macromolecule | Name: RNA (33-MER) / type: rna / ID: 3 / Number of copies: 1 |
---|---|
Source (natural) | Organism: unidentified (others) |
Molecular weight | Theoretical: 16.528768 KDa |
Sequence | String: GGCAUGAAUU GGUCUAGGGU CUGGUCUUAC UACUAUACAA CCUACUACCU CC |
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
![]() | single particle reconstruction |
Aggregation state | particle |
-
Sample preparation
Buffer | pH: 7.5 |
---|---|
Vitrification | Cryogen name: ETHANE |
-
Electron microscopy
Microscope | FEI TALOS ARCTICA |
---|---|
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 54.0 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: -0.8 µm / Nominal defocus min: 1.8 µm |
Experimental equipment | ![]() Model: Talos Arctica / Image courtesy: FEI Company |
-
Image processing
Startup model | Type of model: NONE |
---|---|
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.43 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 398516 |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |