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Yorodumi- EMDB-43144: MicroED structure of SARS-CoV-2 main protease (MPro/3CLPro) with ... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-43144 | ||||||||||||
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Title | MicroED structure of SARS-CoV-2 main protease (MPro/3CLPro) with missing cone eliminated by suspended drop | ||||||||||||
Map data | 1.5 sigma | ||||||||||||
Sample |
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Keywords | Cysteine Protease / Viral Protease / Viral Assembly / 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 / 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 / SARS coronavirus main proteinase / host cell endoplasmic reticulum-Golgi intermediate compartment / 3'-5'-RNA exonuclease activity / symbiont-mediated suppression of host NF-kappaB cascade / 5'-3' DNA helicase activity / 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 / mRNA (guanine-N7)-methyltransferase / omega peptidase activity / methyltransferase cap1 / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of IRF3 activity / SARS-CoV-2 modulates host translation machinery / host cell Golgi apparatus / 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 / host cell perinuclear region of cytoplasm / single-stranded RNA binding / host cell endoplasmic reticulum membrane / viral protein processing / lyase activity / RNA helicase / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / induction by virus of host autophagy / viral translational frameshifting / RNA-directed RNA polymerase / copper ion binding / viral RNA genome replication / cysteine-type endopeptidase activity / RNA-dependent RNA polymerase activity / virus-mediated perturbation of host defense response / lipid binding / DNA-templated transcription / 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 | electron crystallography / cryo EM / Resolution: 2.15 Å | ||||||||||||
Authors | Bu G / Gillman C / Danelius E / Hattne J / Nannenga BL / Gonen T | ||||||||||||
Funding support | United States, 3 items
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Citation | Journal: J Struct Biol X / Year: 2024 Title: Eliminating the missing cone challenge through innovative approaches. Authors: Cody Gillman / Guanhong Bu / Emma Danelius / Johan Hattne / Brent L Nannenga / Tamir Gonen / Abstract: Microcrystal electron diffraction (MicroED) has emerged as a powerful technique for unraveling molecular structures from microcrystals too small for X-ray diffraction. However, a significant hurdle ...Microcrystal electron diffraction (MicroED) has emerged as a powerful technique for unraveling molecular structures from microcrystals too small for X-ray diffraction. However, a significant hurdle arises with plate-like crystals that consistently orient themselves flat on the electron microscopy grid. If the normal of the plate correlates with the axes of the crystal lattice, the crystal orientations accessible for measurement are restricted because the crystal cannot be arbitrarily rotated. This limits the information that can be acquired, resulting in a missing cone of information. We recently introduced a novel crystallization strategy called suspended drop crystallization and proposed that crystals in a suspended drop could effectively address the challenge of preferred crystal orientation. Here we demonstrate the success of the suspended drop approach in eliminating the missing cone in two samples that crystallize as thin plates: bovine liver catalase and the SARS‑CoV‑2 main protease (Mpro). This innovative solution proves indispensable for crystals exhibiting systematic preferred orientations, unlocking new possibilities for structure determination by MicroED. | ||||||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_43144.map.gz | 1.8 MB | EMDB map data format | |
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Header (meta data) | emd-43144-v30.xml emd-43144.xml | 14.7 KB 14.7 KB | Display Display | EMDB header |
Images | emd_43144.png | 126.1 KB | ||
Filedesc metadata | emd-43144.cif.gz | 5.7 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-43144 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-43144 | HTTPS FTP |
-Validation report
Summary document | emd_43144_validation.pdf.gz | 555.2 KB | Display | EMDB validaton report |
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Full document | emd_43144_full_validation.pdf.gz | 554.8 KB | Display | |
Data in XML | emd_43144_validation.xml.gz | 4.3 KB | Display | |
Data in CIF | emd_43144_validation.cif.gz | 4.7 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-43144 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-43144 | HTTPS FTP |
-Related structure data
Related structure data | 8vd7MC 9bdjC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_43144.map.gz / Format: CCP4 / Size: 3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Annotation | 1.5 sigma | ||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. generated in cubic-lattice coordinate | ||||||||||||||||||||||||||||||||||||
Voxel size | X: 0.713 Å / Y: 0.694 Å / Z: 0.709 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 5 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Sample components
-Entire : SARS-CoV-2 main protease (Mpro)
Entire | Name: SARS-CoV-2 main protease (Mpro) |
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Components |
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-Supramolecule #1: SARS-CoV-2 main protease (Mpro)
Supramolecule | Name: SARS-CoV-2 main protease (Mpro) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 |
-Macromolecule #1: 3C-like proteinase nsp5
Macromolecule | Name: 3C-like proteinase nsp5 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: SARS coronavirus main proteinase |
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Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 |
Molecular weight | Theoretical: 33.825547 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: SGFRKMAFPS GKVEGCMVQV TCGTTTLNGL WLDDVVYCPR HVICTSEDML NPNYEDLLIR KSNHNFLVQA GNVQLRVIGH SMQNCVLKL KVDTANPKTP KYKFVRIQPG QTFSVLACYN GSPSGVYQCA MRPNFTIKGS FLNGSCGSVG FNIDYDCVSF C YMHHMELP ...String: SGFRKMAFPS GKVEGCMVQV TCGTTTLNGL WLDDVVYCPR HVICTSEDML NPNYEDLLIR KSNHNFLVQA GNVQLRVIGH SMQNCVLKL KVDTANPKTP KYKFVRIQPG QTFSVLACYN GSPSGVYQCA MRPNFTIKGS FLNGSCGSVG FNIDYDCVSF C YMHHMELP TGVHAGTDLE GNFYGPFVDR QTAQAAGTDT TITVNVLAWL YAAVINGDRW FLNRFTTTLN DFNLVAMKYN YE PLTQDHV DILGPLSAQT GIAVLDMCAS LKELLQNGMN GRTILGSALL EDEFTPFDVV RQCSGVTFQ UniProtKB: Replicase polyprotein 1ab |
-Macromolecule #2: CHLORIDE ION
Macromolecule | Name: CHLORIDE ION / type: ligand / ID: 2 / Number of copies: 1 / Formula: CL |
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Molecular weight | Theoretical: 35.453 Da |
-Macromolecule #3: water
Macromolecule | Name: water / type: ligand / ID: 3 / Number of copies: 6 / Formula: HOH |
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Molecular weight | Theoretical: 18.015 Da |
Chemical component information | ChemComp-HOH: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | electron crystallography |
Aggregation state | 3D array |
-Sample preparation
Buffer | pH: 6.5 / Details: 0.1 M MES pH 6.5, 20% PEG 3350, 5% DMSO. |
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Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 293 K |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Temperature | Min: 75.0 K / Max: 85.0 K |
Image recording | Film or detector model: FEI FALCON IV (4k x 4k) / Number diffraction images: 420 / Average exposure time: 1.0 sec. / Average electron dose: 0.0025 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: DIFFRACTION / Nominal defocus max: 0.0 µm / Nominal defocus min: 0.0 µm / Camera length: 2480 mm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN / Tilt angle: -40.0, 70.0 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 2.15 Å / Resolution method: DIFFRACTION PATTERN/LAYERLINES | ||||||||||||||||||||||||||||||||||||||||||
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Crystallography statistics | Number intensities measured: 123734 / Number structure factors: 14825 / Fourier space coverage: 95.5 / R merge: 25.6 / Overall phase residual: 0 / Phase error rejection criteria: none / High resolution: 2.15 Å Shell:
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-Atomic model buiding 1
Refinement | Protocol: OTHER / Target criteria: Rwork/Rfree gap of less than 5% |
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Output model | PDB-8vd7: |