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- PDB-5e6j: Structure of SARS PLpro bound to a Lys48-linked di-ubiquitin acti... -

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Basic information

Entry
Database: PDB / ID: 5e6j
TitleStructure of SARS PLpro bound to a Lys48-linked di-ubiquitin activity based probe
Components
  • Polyubiquitin-B
  • Replicase polyprotein 1ab
  • ubiquitin
KeywordsHYDROLASE / SARS PLpro / deubiquitinating enzyme / ubiquitin / activity based probe / K48-linkage
Function / homology
Function and homology information


Assembly of the SARS-CoV-1 Replication-Transcription Complex (RTC) / Maturation of replicase proteins / Transcription of SARS-CoV-1 sgRNAs / Translation of Replicase and Assembly of the Replication Transcription Complex / Replication of the SARS-CoV-1 genome / K48-linked deubiquitinase activity / hypothalamus gonadotrophin-releasing hormone neuron development / female meiosis I / positive regulation of protein monoubiquitination / fat pad development ...Assembly of the SARS-CoV-1 Replication-Transcription Complex (RTC) / Maturation of replicase proteins / Transcription of SARS-CoV-1 sgRNAs / Translation of Replicase and Assembly of the Replication Transcription Complex / Replication of the SARS-CoV-1 genome / K48-linked deubiquitinase activity / hypothalamus gonadotrophin-releasing hormone neuron development / female meiosis I / positive regulation of protein monoubiquitination / fat pad development / mitochondrion transport along microtubule / host cell endoplasmic reticulum / K63-linked deubiquitinase activity / female gonad development / seminiferous tubule development / male meiosis I / viral transcription / SARS-CoV-1 modulates host translation machinery / positive regulation of intrinsic apoptotic signaling pathway by p53 class mediator / energy homeostasis / regulation of neuron apoptotic process / regulation of proteasomal protein catabolic process / Maturation of protein E / Maturation of protein E / ER Quality Control Compartment (ERQC) / Myoclonic epilepsy of Lafora / FLT3 signaling by CBL mutants / Prevention of phagosomal-lysosomal fusion / IRAK2 mediated activation of TAK1 complex / Alpha-protein kinase 1 signaling pathway / Glycogen synthesis / IRAK1 recruits IKK complex / IRAK1 recruits IKK complex upon TLR7/8 or 9 stimulation / Membrane binding and targetting of GAG proteins / Endosomal Sorting Complex Required For Transport (ESCRT) / Regulation of TBK1, IKKε (IKBKE)-mediated activation of IRF3, IRF7 / Negative regulation of FLT3 / PTK6 Regulates RTKs and Their Effectors AKT1 and DOK1 / Constitutive Signaling by NOTCH1 HD Domain Mutants / Regulation of TBK1, IKKε-mediated activation of IRF3, IRF7 upon TLR3 ligation / IRAK2 mediated activation of TAK1 complex upon TLR7/8 or 9 stimulation / NOTCH2 Activation and Transmission of Signal to the Nucleus / TICAM1,TRAF6-dependent induction of TAK1 complex / TICAM1-dependent activation of IRF3/IRF7 / APC/C:Cdc20 mediated degradation of Cyclin B / Regulation of FZD by ubiquitination / Downregulation of ERBB4 signaling / p75NTR recruits signalling complexes / APC-Cdc20 mediated degradation of Nek2A / InlA-mediated entry of Listeria monocytogenes into host cells / TRAF6 mediated IRF7 activation in TLR7/8 or 9 signaling / TRAF6-mediated induction of TAK1 complex within TLR4 complex / Regulation of pyruvate metabolism / Regulation of innate immune responses to cytosolic DNA / NF-kB is activated and signals survival / Downregulation of ERBB2:ERBB3 signaling / Pexophagy / NRIF signals cell death from the nucleus / Regulation of PTEN localization / VLDLR internalisation and degradation / Activated NOTCH1 Transmits Signal to the Nucleus / neuron projection morphogenesis / Synthesis of active ubiquitin: roles of E1 and E2 enzymes / Regulation of BACH1 activity / Transferases; Transferring one-carbon groups; Methyltransferases / MAP3K8 (TPL2)-dependent MAPK1/3 activation / regulation of mitochondrial membrane potential / TICAM1, RIP1-mediated IKK complex recruitment / Translesion synthesis by REV1 / Activation of IRF3, IRF7 mediated by TBK1, IKKε (IKBKE) / Translesion synthesis by POLK / InlB-mediated entry of Listeria monocytogenes into host cell / Downregulation of TGF-beta receptor signaling / Josephin domain DUBs / JNK (c-Jun kinases) phosphorylation and activation mediated by activated human TAK1 / Regulation of activated PAK-2p34 by proteasome mediated degradation / Translesion synthesis by POLI / IKK complex recruitment mediated by RIP1 / positive regulation of protein ubiquitination / Gap-filling DNA repair synthesis and ligation in GG-NER / PINK1-PRKN Mediated Mitophagy / TGF-beta receptor signaling in EMT (epithelial to mesenchymal transition) / TNFR1-induced NF-kappa-B signaling pathway / Autodegradation of Cdh1 by Cdh1:APC/C / APC/C:Cdc20 mediated degradation of Securin / TCF dependent signaling in response to WNT / N-glycan trimming in the ER and Calnexin/Calreticulin cycle / Regulation of NF-kappa B signaling / Asymmetric localization of PCP proteins / Ubiquitin-dependent degradation of Cyclin D / SCF-beta-TrCP mediated degradation of Emi1 / NIK-->noncanonical NF-kB signaling / activated TAK1 mediates p38 MAPK activation / Negative regulators of DDX58/IFIH1 signaling / TNFR2 non-canonical NF-kB pathway / AUF1 (hnRNP D0) binds and destabilizes mRNA / Regulation of signaling by CBL / NOTCH3 Activation and Transmission of Signal to the Nucleus / Vpu mediated degradation of CD4 / Assembly of the pre-replicative complex
Similarity search - Function
Papain-like viral protease, N-terminal domain / Papain-like viral protease, thumb domain / Jelly Rolls - #1680 / Non-structural protein 3, SUD-N macrodomain, SARS-CoV / Helicase, Ruva Protein; domain 3 / : / Ubiquitin domain signature. / Ubiquitin conserved site / Ubiquitin domain / Ubiquitin-like (UB roll) ...Papain-like viral protease, N-terminal domain / Papain-like viral protease, thumb domain / Jelly Rolls - #1680 / Non-structural protein 3, SUD-N macrodomain, SARS-CoV / Helicase, Ruva Protein; domain 3 / : / Ubiquitin domain signature. / Ubiquitin conserved site / Ubiquitin domain / Ubiquitin-like (UB roll) / Ubiquitin family / RNA-dependent RNA polymerase, SARS-CoV-like / Nonstructural protein 14, betacoronavirus / NSP15, NendoU domain, coronavirus / Nonstructural protein 15, middle domain, alpha/betacoronavirus / Nonstructural protein 15, N-terminal domain, alpha/beta-coronavirus / NSP14, guanine-N7-methyltransferase domain, coronavirus / Coronavirus (CoV) guanine-N7-methyltransferase (N7-MTase) domain profile. / Coronavirus (CoV) Nsp15 N-terminal oligomerization domain profile. / NSP12 RNA-dependent RNA polymerase, coronavirus / : / Coronavirus Nsp12 RNA-dependent RNA polymerase (RdRp) domain profile. / Coronavirus Nsp12 Interface domain profile. / : / : / Coronavirus Nonstructural protein 13, 1B domain / Coronavirus Non-structural protein 13, zinc-binding domain / Coronavirus Nonstructural protein 13, stalk domain / Nidovirus 2-O-methyltransferase / Nidovirus 2'-O-methyltransferase (2'-O-MTase) domain profile. / Non-structural protein NSP15, N-terminal domain superfamily, coronavirus / Non-structural protein NSP15, middle domain superfamily / Coronavirus replicase NSP15, N-terminal oligomerization / Nonstructural protein 15, middle domain, coronavirus / Nonstructural protein 13, 1B domain, coronavirus / Coronavirus replicase NSP15, middle domain / Coronavirus replicase NSP15, N-terminal oligomerisation / Nidovirus 3'-5' exoribonuclease domain / Nidovirus 3'-5' exoribonuclease (ExoN) domain profile. / Non-structural protein NSP16, coronavirus-like / Non-structural protein 14, coronavirus / RNA polymerase, N-terminal, coronavirus / Coronavirus 2'-O-methyltransferase / Coronavirus proofreading exoribonuclease / Coronavirus RNA-dependent RNA polymerase, N-terminal / Nonstructural protein 13, zinc-binding domain, coronavirus-like / Coronaviridae zinc-binding (CV ZBD) domain profile. / Arterivirus Nsp11 N-terminal/Coronavirus NSP15 middle domain / Arterivirus Nsp11 N-terminal/coronavirus NSP15 middle (AV-Nsp11N/CoV-Nsp15M) domain profile. / Nidoviral uridylate-specific endoribonuclease (NendoU) domain profile. / Nidovirus RdRp-associated nucleotidyl transferase (NiRAN) domain / Nidovirus RdRp-associated nucleotidyl transferase (NiRAN) domain profile. / Ubiquitin homologues / Endoribonuclease EndoU-like / NendoU domain, nidovirus / Coronavirus replicase NSP15, uridylate-specific endoribonuclease / : / Ubiquitin domain profile. / Ubiquitin-like domain / DNA2/NAM7 helicase-like, C-terminal / AAA domain / (+) RNA virus helicase core domain / (+)RNA virus helicase core domain profile. / 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 / 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 / Coronavirus (CoV) Nsp2 middle domain profile. / NSP1, globular domain, alpha/betacoronavirus / Coronavirus (CoV) Nsp1 globular domain profile. / Coronavirus (CoV) Nsp2 N-terminal domain profile. / Coronavirus (CoV) Nsp2 C-terminal domain profile.
Similarity search - Domain/homology
ACETATE ION / NICKEL (II) ION / Replicase polyprotein 1ab / Polyubiquitin-B / Polyubiquitin-C
Similarity search - Component
Biological speciesHuman SARS coronavirus
Homo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.85 Å
AuthorsLima, C.D. / Bekes, M.
Funding support United States, Netherlands, 5items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM065872 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)F32GM100598 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM084244 United States
Howard Hughes Medical Institute (HHMI) United States
Netherlands Organisation for Scientific Research (NWO)724013002 Netherlands
CitationJournal: Mol. Cell / Year: 2016
Title: Recognition of Lys48-Linked Di-ubiquitin and Deubiquitinating Activities of the SARS Coronavirus Papain-like Protease.
Authors: Bekes, M. / van der Heden van Noort, G.J. / Ekkebus, R. / Ovaa, H. / Huang, T.T. / Lima, C.D.
History
DepositionOct 9, 2015Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 18, 2016Provider: repository / Type: Initial release
Revision 1.1Jan 25, 2017Group: Derived calculations
Revision 1.2Sep 6, 2017Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.3Nov 1, 2017Group: Author supporting evidence / Category: pdbx_struct_assembly_auth_evidence
Revision 1.4Dec 20, 2017Group: Database references / Category: citation / citation_author
Item: _citation.journal_abbrev / _citation.journal_id_ISSN ..._citation.journal_abbrev / _citation.journal_id_ISSN / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.name
Revision 1.5Nov 20, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.6Sep 27, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_conn
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_conn.conn_type_id / _struct_conn.id / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_leaving_atom_flag / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_conn.ptnr2_label_seq_id
Revision 1.7Nov 15, 2023Group: Data collection / Derived calculations / Category: chem_comp_atom / chem_comp_bond / struct_conn
Item: _chem_comp_atom.atom_id / _chem_comp_bond.atom_id_2 / _struct_conn.pdbx_leaving_atom_flag

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Replicase polyprotein 1ab
B: ubiquitin
C: Polyubiquitin-B
D: Replicase polyprotein 1ab
E: ubiquitin
F: Polyubiquitin-B
hetero molecules


Theoretical massNumber of molelcules
Total (without water)107,7848
Polymers107,6666
Non-polymers1182
Water99155
1
A: Replicase polyprotein 1ab
B: ubiquitin
C: Polyubiquitin-B
hetero molecules


Theoretical massNumber of molelcules
Total (without water)53,9515
Polymers53,8333
Non-polymers1182
Water543
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2
D: Replicase polyprotein 1ab
E: ubiquitin
F: Polyubiquitin-B


Theoretical massNumber of molelcules
Total (without water)53,8333
Polymers53,8333
Non-polymers00
Water543
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)72.981, 68.239, 119.021
Angle α, β, γ (deg.)90.00, 103.21, 90.00
Int Tables number4
Space group name H-MP1211
DetailsTrimer confirmed by cross-linking and gel filtration

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Components

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Protein , 3 types, 6 molecules ADBECF

#1: Protein Replicase polyprotein 1ab / pp1ab / ORF1ab polyprotein


Mass: 36688.527 Da / Num. of mol.: 2 / Fragment: UNP Residues 1541-1856
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human SARS coronavirus / Gene: rep, 1a-1b / Plasmid: pET28 / Details (production host): C-terminal 6xHis-tag / Production host: Escherichia coli (E. coli) / Strain (production host): BL-21(DE3) RIL Codon Plus
References: UniProt: P0C6X7, ubiquitinyl hydrolase 1, SARS coronavirus main proteinase, Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases, RNA-directed RNA polymerase, DNA ...References: UniProt: P0C6X7, ubiquitinyl hydrolase 1, SARS coronavirus main proteinase, Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases, RNA-directed RNA polymerase, DNA helicase, RNA helicase, Transferases; Transferring one-carbon groups; Methyltransferases, Hydrolases; Acting on ester bonds; Exoribonucleases producing 5'-phosphomonoesters, Hydrolases; Acting on ester bonds
#2: Protein ubiquitin


Mass: 8624.899 Da / Num. of mol.: 2 / Mutation: K48LYQ, G76AYE / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human) / References: UniProt: P0CG48
#3: Protein Polyubiquitin-B


Mass: 8519.778 Da / Num. of mol.: 2 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human) / References: UniProt: P0CG47

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Non-polymers , 3 types, 57 molecules

#4: Chemical ChemComp-ACT / ACETATE ION


Mass: 59.044 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C2H3O2
#5: Chemical ChemComp-NI / NICKEL (II) ION


Mass: 58.693 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Ni
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 55 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION

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Sample preparation

CrystalDensity Matthews: 2.68 Å3/Da / Density % sol: 54.1 %
Crystal growTemperature: 285 K / Method: vapor diffusion, hanging drop / pH: 5.5 / Details: 0.1 M MES, 0.1 M lithium acetate, 17% PEG 6000
Temp details: Crystal moved to 277K 24-hour prior to mounting

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 24-ID-E / Wavelength: 0.9791 Å
DetectorType: ADSC QUANTUM 315 / Detector: CCD / Date: Feb 25, 2015
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9791 Å / Relative weight: 1
ReflectionResolution: 2.85→50 Å / Num. obs: 26147 / % possible obs: 98 % / Redundancy: 3.3 % / Rmerge(I) obs: 0.07 / Net I/σ(I): 13.99
Reflection shellResolution: 2.85→2.95 Å / Redundancy: 3 % / Rmerge(I) obs: 0.53 / Mean I/σ(I) obs: 1.97 / % possible all: 100

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Processing

Software
NameVersionClassification
PHENIX1.9_1692refinement
HKL-2000data reduction
HKL-2000data scaling
PHENIXphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 4MM3, 1UBQ

4mm3

1ubq


Resolution: 2.85→49.216 Å / SU ML: 0.41 / Cross valid method: FREE R-VALUE / σ(F): 0.26 / Phase error: 31.19 / Stereochemistry target values: ML
RfactorNum. reflection% reflectionSelection details
Rfree0.2643 2590 5.09 %Random selection
Rwork0.2323 ---
obs0.234 50864 97.49 %-
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 2.85→49.216 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms7306 0 5 55 7366
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0037453
X-RAY DIFFRACTIONf_angle_d0.60110091
X-RAY DIFFRACTIONf_dihedral_angle_d11.8692728
X-RAY DIFFRACTIONf_chiral_restr0.0251167
X-RAY DIFFRACTIONf_plane_restr0.0021292
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.85-2.90180.4021370.36252512X-RAY DIFFRACTION98
2.9018-2.95760.39371260.36772547X-RAY DIFFRACTION97
2.9576-3.0180.34231370.3622558X-RAY DIFFRACTION97
3.018-3.08360.34181400.34122513X-RAY DIFFRACTION98
3.0836-3.15530.42271290.30882549X-RAY DIFFRACTION96
3.1553-3.23420.33871350.30872510X-RAY DIFFRACTION96
3.2342-3.32160.34981270.28552515X-RAY DIFFRACTION97
3.3216-3.41940.31761260.2762572X-RAY DIFFRACTION98
3.4194-3.52970.34911130.26852526X-RAY DIFFRACTION97
3.5297-3.65580.30941580.26352566X-RAY DIFFRACTION98
3.6558-3.80210.22991250.23252542X-RAY DIFFRACTION97
3.8021-3.97510.31251300.21752448X-RAY DIFFRACTION94
3.9751-4.18460.23061470.212491X-RAY DIFFRACTION96
4.1846-4.44660.21211480.19362569X-RAY DIFFRACTION99
4.4466-4.78970.21711630.19532592X-RAY DIFFRACTION100
4.7897-5.27120.20321360.19012627X-RAY DIFFRACTION100
5.2712-6.03270.24921360.20952544X-RAY DIFFRACTION99
6.0327-7.59610.24181290.22612562X-RAY DIFFRACTION97
7.5961-49.22350.22771480.18262531X-RAY DIFFRACTION98

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