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- PDB-8spj: Crystal Structure of SARS-CoV-2 Main Protease (Mpro) N28T Mutant -

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Entry
Database: PDB / ID: 8spj
TitleCrystal Structure of SARS-CoV-2 Main Protease (Mpro) N28T Mutant
Components3C-like proteinase nsp5
KeywordsVIRAL PROTEIN / HYDROLASE / Protease / SARS-CoV-2 / Mpro / Apo / COVID-19
Function / homology
Function and homology information


viral genome replication / methyltransferase 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 / Replication of the SARS-CoV-2 genome / double membrane vesicle viral factory outer membrane / SARS coronavirus main proteinase ...viral genome replication / methyltransferase 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 / Replication of the SARS-CoV-2 genome / double membrane vesicle viral factory outer membrane / SARS coronavirus main proteinase / host cell endosome / 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 / host cell Golgi apparatus / symbiont-mediated perturbation of host ubiquitin-like protein modification / endonuclease activity / ubiquitinyl hydrolase 1 / methylation / cysteine-type deubiquitinase activity / Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases / single-stranded RNA binding / host cell perinuclear region of cytoplasm / viral protein processing / host cell endoplasmic reticulum membrane / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / induction by virus of host autophagy / viral translational frameshifting / cysteine-type endopeptidase activity / virus-mediated perturbation of host defense response / lipid binding / SARS-CoV-2 activates/modulates innate and adaptive immune responses / proteolysis / zinc ion binding / membrane
Similarity search - Function
Non-structural protein NSP3, SUD-N (Mac2) domain, betacoronavirus / Sarbecovirus Nsp3c-N domain profile. / Non-structural protein NSP3, N-terminal, betacoronavirus / Polyprotein cleavage domain PL2pro superfamily, betacoronavirus / Non-structural protein NSP3, SUD-N (Mac2) domain superfamily, betacoronavirus / Betacoronavirus SUD-C domain / Betacoronavirus replicase NSP3, N-terminal / NSP1 globular domain superfamily, betacoronavirus / Non-structural protein 2, SARS-CoV-like / Carbamoyl-phosphate synthase subdomain signature 2. ...Non-structural protein NSP3, SUD-N (Mac2) domain, betacoronavirus / Sarbecovirus Nsp3c-N domain profile. / Non-structural protein NSP3, N-terminal, betacoronavirus / Polyprotein cleavage domain PL2pro superfamily, betacoronavirus / Non-structural protein NSP3, SUD-N (Mac2) domain superfamily, betacoronavirus / Betacoronavirus SUD-C domain / Betacoronavirus replicase NSP3, N-terminal / NSP1 globular domain superfamily, betacoronavirus / Non-structural protein 2, SARS-CoV-like / Carbamoyl-phosphate synthase subdomain signature 2. / Betacoronavirus Nsp3c-M domain profile. / NSP1, globular domain, betacoronavirus / Non-structural protein NSP3, SUD-M domain, betacoronavirus / Non-structural protein NSP3, SUD-M domain superfamily, betacoronavirus / Betacoronavirus replicase NSP1 / Betacoronavirus single-stranded poly(A) binding domain / NSP1, C-terminal domain, betacoronavirus / : / Betacoronavirus (BetaCoV) Nsp1 C-terminal domain profile. / Betacoronavirus Nsp3e group 2-specific marker (G2M) domain profile. / Betacoronavirus Nsp3c-C domain profile. / Betacoronavirus Nsp3e nucleic acid-binding (NAB) domain profile. / DPUP/SUD, C-terminal, betacoronavirus / Non-structural protein NSP3, nucleic acid-binding domain, betacoronavirus / Non-structural protein NSP3A domain-like superfamily / Non-structural protein NSP3, nucleic acid-binding domain superfamily, betacoronavirus / Non-structural protein 6, betacoronavirus / Betacoronavirus nucleic acid-binding (NAB) / Papain-like viral protease, palm and finger domains, coronavirus / Papain-like protease, N-terminal domain superfamily, coronavirus / Coronavirus replicase NSP2, N-terminal / : / Coronavirus replicase NSP2, C-terminal / NSP1, globular domain, alpha/betacoronavirus / Coronavirus (CoV) Nsp2 middle domain profile. / Coronavirus (CoV) Nsp1 globular domain profile. / Coronavirus (CoV) Nsp2 N-terminal domain profile. / Coronavirus (CoV) Nsp2 C-terminal domain profile. / Nonstructural protein 2, N-terminal domain, coronavirus / Non-structural protein 2, C-terminal domain, coronavirus / NSP3, second ubiquitin-like (Ubl) domain, coronavirus / Coronavirus Nsp3a Ubl domain profile. / Coronavirus Nsp3d Ubl domain profile. / NSP3, first ubiquitin-like (Ubl) domain, coronavirus / : / : / Coronavirus replicase NSP7 / Peptidase family C16 domain profile. / Coronavirus 3Ecto domain profile. / Coronavirus RNA-dependent RNA polymerase (RdRp) Nsp7 cofactor domain profile. / Coronavirus RNA-dependent RNA polymerase (RdRp) Nsp8 cofactor domain profile. / Coronavirus Nsp9 single-stranded RNA (ssRNA)-binding domain profile. / Coronavirus (CoV) ExoN/MTase coactivator domain profile. / Coronavirus (CoV) Nsp3 Y domain profile. / Coronavirus Nsp4 C-terminal (Nsp4C) domain profile. / Peptidase C30, coronavirus / Peptidase C16, coronavirus / Non-structural protein NSP9, coronavirus / Non-structural protein NSP7, coronavirus / Non-structural protein NSP8, coronavirus / RNA synthesis protein NSP10, coronavirus / Non-structural protein NSP4, C-terminal, coronavirus / RNA synthesis protein NSP10 superfamily, coronavirus / Non-structural protein NSP9 superfamily, coronavirus / Non-structural protein NSP7 superfamily, coronavirus / Non-structural protein NSP8 superfamily, coronavirus / Non-structural protein NSP4, C-terminal superfamily, coronavirus / Papain-like protease, thumb domain superfamily, coronavirus / Peptidase C30, domain 3, coronavirus / Non-structural protein 6, coronavirus / Coronavirus replicase NSP3, C-terminal / Non-structural protein NSP4, N-terminal, coronavirus / Coronavirus endopeptidase C30 / Coronavirus papain-like peptidase / Coronavirus replicase NSP8 / Coronavirus RNA synthesis protein NSP10 / Coronavirus replicase NSP4, C-terminal / Coronavirus replicase NSP6 / Coronavirus replicase NSP4, N-terminal / Coronavirus replicase NSP3, C-terminal / Coronavirus main protease (M-pro) domain profile. / Coronavirus replicase NSP9 / Non-structural protein 3, X-domain-like / Macro domain / Appr-1"-p processing enzyme / Macro domain / Macro domain profile. / Macro domain-like / Peptidase S1, PA clan, chymotrypsin-like fold / Peptidase S1, PA clan
Similarity search - Domain/homology
Replicase polyprotein 1a
Similarity search - Component
Biological speciesSevere acute respiratory syndrome coronavirus 2
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.08 Å
AuthorsKohaal, N. / Lewandowski, E.M. / Wang, J. / Chen, Y.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI158775 United States
CitationJournal: To Be Published
Title: Crystal Structure of SARS-CoV-2 Main Protease (Mpro) N28T Mutant
Authors: Lewandowski, E.M. / Kohaal, N. / Wang, J. / Chen, Y.
History
DepositionMay 3, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 24, 2023Provider: repository / Type: Initial release
Revision 1.1May 22, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: 3C-like proteinase nsp5


Theoretical massNumber of molelcules
Total (without water)33,8131
Polymers33,8131
Non-polymers00
Water1,11762
1
A: 3C-like proteinase nsp5

A: 3C-like proteinase nsp5


Theoretical massNumber of molelcules
Total (without water)67,6252
Polymers67,6252
Non-polymers00
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation2_555-x,y,-z1
Buried area2740 Å2
ΔGint-15 kcal/mol
Surface area25540 Å2
MethodPISA
Unit cell
Length a, b, c (Å)112.846, 53.497, 45.292
Angle α, β, γ (deg.)90.00, 101.82, 90.00
Int Tables number5
Space group name H-MC121

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Components

#1: Protein 3C-like proteinase nsp5 / 3CL-PRO / 3CLp / Main protease / Mpro / Non-structural protein 5 / nsp5 / SARS coronavirus main proteinase


Mass: 33812.547 Da / Num. of mol.: 1 / Mutation: N28T
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Severe acute respiratory syndrome coronavirus 2
Production host: Escherichia coli (E. coli)
References: UniProt: P0DTC1, SARS coronavirus main proteinase
#2: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 62 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 1.98 Å3/Da / Density % sol: 37.84 %
Crystal growTemperature: 293.15 K / Method: vapor diffusion, hanging drop
Details: 25% PEG 3350, 0.1M Potassium/Sodium Tartrate, 0.005M Magnesium Chloride

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 19-ID / Wavelength: 1 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Dec 14, 2022
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 2.08→50 Å / Num. obs: 15680 / % possible obs: 98.5 % / Redundancy: 5.8 % / Rmerge(I) obs: 0.104 / Net I/σ(I): 22.15
Reflection shellResolution: 2.08→2.13 Å / Redundancy: 4.5 % / Rmerge(I) obs: 0.543 / Mean I/σ(I) obs: 2.13 / Num. unique obs: 775 / % possible all: 96.5

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Processing

Software
NameVersionClassification
REFMAC5.8.0352refinement
HKL-2000data reduction
PDB_EXTRACTdata extraction
HKL-2000data scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 2.08→38.68 Å / Cor.coef. Fo:Fc: 0.949 / Cor.coef. Fo:Fc free: 0.94 / SU B: 15.683 / SU ML: 0.196 / Cross valid method: THROUGHOUT / ESU R: 0.313 / ESU R Free: 0.207 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflectionSelection details
Rfree0.24184 844 5.4 %RANDOM
Rwork0.21676 ---
obs0.21816 14834 97.61 %-
Solvent computationIon probe radii: 0.7 Å / Shrinkage radii: 0.7 Å / VDW probe radii: 1.3 Å / Solvent model: MASK
Displacement parametersBiso mean: 47.42 Å2
Baniso -1Baniso -2Baniso -3
1-0.69 Å2-0 Å2-2 Å2
2--2.33 Å2-0 Å2
3----2.01 Å2
Refinement stepCycle: LAST / Resolution: 2.08→38.68 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms2333 0 0 62 2395
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0060.0172385
X-RAY DIFFRACTIONr_bond_other_d0.0010.0162124
X-RAY DIFFRACTIONr_angle_refined_deg0.9721.8073242
X-RAY DIFFRACTIONr_angle_other_deg0.3561.5574951
X-RAY DIFFRACTIONr_dihedral_angle_1_deg75.176312
X-RAY DIFFRACTIONr_dihedral_angle_2_deg
X-RAY DIFFRACTIONr_dihedral_angle_3_deg12.33910383
X-RAY DIFFRACTIONr_dihedral_angle_4_deg
X-RAY DIFFRACTIONr_chiral_restr0.0510.2366
X-RAY DIFFRACTIONr_gen_planes_refined0.0030.022731
X-RAY DIFFRACTIONr_gen_planes_other0.0010.02477
X-RAY DIFFRACTIONr_nbd_refined
X-RAY DIFFRACTIONr_nbd_other
X-RAY DIFFRACTIONr_nbtor_refined
X-RAY DIFFRACTIONr_nbtor_other
X-RAY DIFFRACTIONr_xyhbond_nbd_refined
X-RAY DIFFRACTIONr_xyhbond_nbd_other
X-RAY DIFFRACTIONr_metal_ion_refined
X-RAY DIFFRACTIONr_metal_ion_other
X-RAY DIFFRACTIONr_symmetry_vdw_refined
X-RAY DIFFRACTIONr_symmetry_vdw_other
X-RAY DIFFRACTIONr_symmetry_hbond_refined
X-RAY DIFFRACTIONr_symmetry_hbond_other
X-RAY DIFFRACTIONr_symmetry_metal_ion_refined
X-RAY DIFFRACTIONr_symmetry_metal_ion_other
X-RAY DIFFRACTIONr_mcbond_it1.7852.2611207
X-RAY DIFFRACTIONr_mcbond_other1.7832.261206
X-RAY DIFFRACTIONr_mcangle_it2.9773.3841507
X-RAY DIFFRACTIONr_mcangle_other2.9773.3871508
X-RAY DIFFRACTIONr_scbond_it1.822.4211178
X-RAY DIFFRACTIONr_scbond_other1.8192.4211178
X-RAY DIFFRACTIONr_scangle_it
X-RAY DIFFRACTIONr_scangle_other2.9333.5511736
X-RAY DIFFRACTIONr_long_range_B_refined5.40633.1652591
X-RAY DIFFRACTIONr_long_range_B_other5.37533.1172586
X-RAY DIFFRACTIONr_rigid_bond_restr
X-RAY DIFFRACTIONr_sphericity_free
X-RAY DIFFRACTIONr_sphericity_bonded
LS refinement shellResolution: 2.08→2.132 Å
RfactorNum. reflection% reflection
Rfree0.344 72 -
Rwork0.305 929 -
obs--84.62 %
Refinement TLS params.Method: refined / Origin x: 12.201 Å / Origin y: 0.405 Å / Origin z: 4.685 Å
111213212223313233
T0.0992 Å20 Å20.103 Å2-0.0098 Å20.022 Å2--0.2351 Å2
L2.4427 °2-0.3026 °21.6005 °2-1.1306 °2-0.6906 °2--2.7822 °2
S0.075 Å °0.0673 Å °0.2128 Å °-0.0454 Å °-0.0234 Å °0.0397 Å °0.2056 Å °0.0375 Å °-0.0516 Å °

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