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- PDB-8ey2: Cryo-EM structure of SARS-CoV-2 Main protease C145S in complex wi... -
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Basic information
Entry | Database: PDB / ID: 8ey2 | ||||||
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Title | Cryo-EM structure of SARS-CoV-2 Main protease C145S in complex with N-terminal peptide | ||||||
![]() | 3C-like proteinase | ||||||
![]() | VIRAL PROTEIN / covid19 / mpro / main protease / cryo-Em / 3cl / sars-cov-2 | ||||||
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 / 3'-5'-RNA exonuclease activity / host cell endoplasmic reticulum-Golgi intermediate compartment / host cell endosome / 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 / 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 / DNA helicase / mRNA (nucleoside-2'-O-)-methyltransferase activity / 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 / cysteine-type endopeptidase activity / RNA-directed RNA polymerase / viral RNA genome replication / virus-mediated perturbation of host defense response / RNA-dependent 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: 3.5 Å | ||||||
![]() | Noske, G.D. / Song, Y. / Fernandes, R.S. / Oliva, G. / Godoy, A.S. | ||||||
Funding support | ![]()
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![]() | ![]() Title: An in-solution snapshot of SARS-COV-2 main protease maturation process and inhibition. Authors: Gabriela Dias Noske / Yun Song / Rafaela Sachetto Fernandes / Rod Chalk / Haitem Elmassoudi / Lizbé Koekemoer / C David Owen / Tarick J El-Baba / Carol V Robinson / / Glaucius Oliva / Andre Schutzer Godoy / ![]() ![]() Abstract: The main protease from SARS-CoV-2 (M) is responsible for cleavage of the viral polyprotein. M self-processing is called maturation, and it is crucial for enzyme dimerization and activity. Here we use ...The main protease from SARS-CoV-2 (M) is responsible for cleavage of the viral polyprotein. M self-processing is called maturation, and it is crucial for enzyme dimerization and activity. Here we use C145S M to study the structure and dynamics of N-terminal cleavage in solution. Native mass spectroscopy analysis shows that mixed oligomeric states are composed of cleaved and uncleaved particles, indicating that N-terminal processing is not critical for dimerization. A 3.5 Å cryo-EM structure provides details of M N-terminal cleavage outside the constrains of crystal environment. We show that different classes of inhibitors shift the balance between oligomeric states. While non-covalent inhibitor MAT-POS-e194df51-1 prevents dimerization, the covalent inhibitor nirmatrelvir induces the conversion of monomers into dimers, even with intact N-termini. Our data indicates that the M dimerization is triggered by induced fit due to covalent linkage during substrate processing rather than the N-terminal processing. | ||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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PDBx/mmCIF format | ![]() | 126.4 KB | Display | ![]() |
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PDB format | ![]() | 93.6 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 28666MC ![]() 8eyjC M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 34567.371 Da / Num. of mol.: 4 / Fragment: UNP residues 3259-3569 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: rep, 1a-1b / Production host: ![]() ![]() References: UniProt: P0DTD1, SARS coronavirus main proteinase |
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-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: Cryo-EM structure of SARS-CoV-2 Main protease C145S mutant Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Value: 68 kDa/nm / Experimental value: YES |
Source (natural) | Organism: ![]() ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 7.8 Details: 20 mM Tris pH 7.8, 150 mM NaCl, 1 mM EDTA, 1 mM DTT |
Specimen | Conc.: 0.25 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE / Humidity: 100 % |
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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 / Nominal defocus max: 2500 nm / Nominal defocus min: 500 nm |
Image recording | Electron dose: 43.38557582 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of real images: 13770 |
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Processing
Software | Name: PHENIX / Version: 1.19.2_4158: / Classification: refinement | ||||||||||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: NONE | ||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 1956496 | ||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 227533 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 218 / Protocol: RIGID BODY FIT / Space: REAL / Target criteria: CC | ||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 7N6N Accession code: 7N6N / Source name: PDB / Type: experimental model | ||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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