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- PDB-7krn: Structure of SARS-CoV-2 backtracked complex bound to nsp13 helica... -
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Basic information
Entry | Database: PDB / ID: 7krn | |||||||||||||||||||||||||||||||||||||||||||||
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Title | Structure of SARS-CoV-2 backtracked complex bound to nsp13 helicase - nsp13(1)-BTC | |||||||||||||||||||||||||||||||||||||||||||||
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![]() | TRANSFERASE/HYDROLASE/RNA / RNA-dependent RNA polymerase / viral replication-transcription complex / transcription / viral proteins / TRANSFERASE-HYDROLASE-RNA complex | |||||||||||||||||||||||||||||||||||||||||||||
Function / homology | ![]() protein guanylyltransferase activity / RNA endonuclease activity producing 3'-phosphomonoesters, hydrolytic mechanism / mRNA guanylyltransferase activity / 5'-3' RNA helicase activity / Lyases; Phosphorus-oxygen lyases / Assembly of the SARS-CoV-2 Replication-Transcription Complex (RTC) / Maturation of replicase proteins / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of TBK1 activity / ISG15-specific peptidase activity / TRAF3-dependent IRF activation pathway ...protein guanylyltransferase activity / RNA endonuclease activity producing 3'-phosphomonoesters, hydrolytic mechanism / mRNA guanylyltransferase activity / 5'-3' RNA helicase activity / Lyases; Phosphorus-oxygen lyases / Assembly of the SARS-CoV-2 Replication-Transcription Complex (RTC) / Maturation of replicase proteins / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of TBK1 activity / ISG15-specific peptidase activity / TRAF3-dependent IRF activation pathway / Transcription of SARS-CoV-2 sgRNAs / Translation of Replicase and Assembly of the Replication Transcription Complex / 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 / host cell endoplasmic reticulum-Golgi intermediate compartment / SARS coronavirus main proteinase / 3'-5'-RNA exonuclease activity / 5'-3' DNA helicase activity / host cell endosome / symbiont-mediated degradation of host mRNA / mRNA guanylyltransferase / symbiont-mediated suppression of host ISG15-protein conjugation / G-quadruplex RNA binding / symbiont-mediated suppression of host toll-like receptor signaling pathway / omega peptidase activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of IRF3 activity / SARS-CoV-2 modulates host translation machinery / mRNA (guanine-N7)-methyltransferase / host cell Golgi apparatus / methyltransferase cap1 / symbiont-mediated perturbation of host ubiquitin-like protein modification / symbiont-mediated suppression of host NF-kappaB cascade / DNA helicase / methyltransferase cap1 activity / ubiquitinyl hydrolase 1 / cysteine-type deubiquitinase activity / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / forked DNA-dependent helicase activity / single-stranded 3'-5' DNA helicase activity / four-way junction helicase activity / double-stranded DNA helicase activity / Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases / single-stranded RNA binding / regulation of autophagy / host cell perinuclear region of cytoplasm / viral protein processing / lyase activity / host cell endoplasmic reticulum membrane / RNA helicase / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / symbiont-mediated suppression of host gene expression / copper ion binding / viral translational frameshifting / symbiont-mediated activation of host autophagy / RNA-directed RNA polymerase / cysteine-type endopeptidase activity / viral RNA genome replication / RNA-directed RNA polymerase activity / 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 | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å | |||||||||||||||||||||||||||||||||||||||||||||
![]() | Chen, J. / Malone, B. / Campbell, E.A. / Darst, S.A. | |||||||||||||||||||||||||||||||||||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural basis for backtracking by the SARS-CoV-2 replication-transcription complex. Authors: Brandon Malone / James Chen / Qi Wang / Eliza Llewellyn / Young Joo Choi / Paul Dominic B Olinares / Xinyun Cao / Carolina Hernandez / Edward T Eng / Brian T Chait / David E Shaw / Robert ...Authors: Brandon Malone / James Chen / Qi Wang / Eliza Llewellyn / Young Joo Choi / Paul Dominic B Olinares / Xinyun Cao / Carolina Hernandez / Edward T Eng / Brian T Chait / David E Shaw / Robert Landick / Seth A Darst / Elizabeth A Campbell / ![]() Abstract: Backtracking, the reverse motion of the transcriptase enzyme on the nucleic acid template, is a universal regulatory feature of transcription in cellular organisms but its role in viruses is not ...Backtracking, the reverse motion of the transcriptase enzyme on the nucleic acid template, is a universal regulatory feature of transcription in cellular organisms but its role in viruses is not established. Here we present evidence that backtracking extends into the viral realm, where backtracking by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA-dependent RNA polymerase (RdRp) may aid viral transcription and replication. Structures of SARS-CoV-2 RdRp bound to the essential nsp13 helicase and RNA suggested the helicase facilitates backtracking. We use cryo-electron microscopy, RNA-protein cross-linking, and unbiased molecular dynamics simulations to characterize SARS-CoV-2 RdRp backtracking. The results establish that the single-stranded 3' segment of the product RNA generated by backtracking extrudes through the RdRp nucleoside triphosphate (NTP) entry tunnel, that a mismatched nucleotide at the product RNA 3' end frays and enters the NTP entry tunnel to initiate backtracking, and that nsp13 stimulates RdRp backtracking. Backtracking may aid proofreading, a crucial process for SARS-CoV-2 resistance against antivirals. | |||||||||||||||||||||||||||||||||||||||||||||
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Structure visualization
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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PDBx/mmCIF format | ![]() | 400.6 KB | Display | ![]() |
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PDB format | ![]() | 315.1 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.1 MB | Display | ![]() |
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Full document | ![]() | 1.2 MB | Display | |
Data in XML | ![]() | 65.4 KB | Display | |
Data in CIF | ![]() | 101.5 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 23007MC ![]() 7kroC ![]() 7krpC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
-Protein , 2 types, 2 molecules AE
#1: Protein | Mass: 106780.977 Da / Num. of mol.: 1 / Fragment: UNP residues 4393-5324 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: rep, 1a-1b / Production host: ![]() ![]() |
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#4: Protein | Mass: 67354.039 Da / Num. of mol.: 1 / Fragment: UNP residues 5325-5925 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: rep, 1a-1b / Production host: ![]() ![]() |
-Non-structural protein ... , 2 types, 3 molecules BDC
#2: Protein | Mass: 22034.242 Da / Num. of mol.: 2 / Fragment: UNP residues 3943-4140 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: rep, 1a-1b / Production host: ![]() ![]() #3: Protein | | Mass: 9748.385 Da / Num. of mol.: 1 / Fragment: UNP residues 3860-3942 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: rep, 1a-1b / Production host: ![]() ![]() |
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-RNA chain , 2 types, 2 molecules PT
#5: RNA chain | Mass: 12667.554 Da / Num. of mol.: 1 / Source method: obtained synthetically Source: (synth.) ![]() ![]() |
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#6: RNA chain | Mass: 17573.391 Da / Num. of mol.: 1 / Source method: obtained synthetically Source: (synth.) ![]() ![]() |
-Non-polymers , 5 types, 13 molecules 








#7: Chemical | ChemComp-ZN / #8: Chemical | #9: Chemical | #10: Chemical | #11: Chemical | ChemComp-AF3 / | |
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-Details
Has ligand of interest | N |
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Has protein modification | N |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: SARS-CoV-2 backtracked complex bound to nsp13 helicase - nsp13(1)-BTC Type: COMPLEX / Entity ID: #1-#6 / Source: RECOMBINANT |
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Source (natural) | Organism: ![]() ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 8 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Electron dose: 66 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
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Processing
Software | Name: PHENIX / Version: 1.18.2_3874: / Classification: refinement | ||||||||||||||||||||||||
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EM software | Name: PHENIX / Category: model refinement | ||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 404706 / Symmetry type: POINT | ||||||||||||||||||||||||
Refine LS restraints |
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