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- PDB-2n3u: Solution structure of the Rpn1 T1 site engaging two monoubiquitin... -

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Basic information

Entry
Database: PDB / ID: 2n3u
TitleSolution structure of the Rpn1 T1 site engaging two monoubiquitin molecules
Components
  • 26S proteasome regulatory subunit RPN1
  • Ubiquitin-60S ribosomal protein L40
KeywordsPROTEIN BINDING
Function / homology
Function and homology information


response to insecticide / proteasome regulatory particle, base subcomplex / Cross-presentation of soluble exogenous antigens (endosomes) / TNFR2 non-canonical NF-kB pathway / Ubiquitin-Mediated Degradation of Phosphorylated Cdc25A / Regulation of PTEN stability and activity / CDK-mediated phosphorylation and removal of Cdc6 / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / KEAP1-NFE2L2 pathway / Neddylation ...response to insecticide / proteasome regulatory particle, base subcomplex / Cross-presentation of soluble exogenous antigens (endosomes) / TNFR2 non-canonical NF-kB pathway / Ubiquitin-Mediated Degradation of Phosphorylated Cdc25A / Regulation of PTEN stability and activity / CDK-mediated phosphorylation and removal of Cdc6 / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / KEAP1-NFE2L2 pathway / Neddylation / Orc1 removal from chromatin / MAPK6/MAPK4 signaling / regulation of protein catabolic process / proteasome storage granule / Peptide chain elongation / Selenocysteine synthesis / Antigen processing: Ubiquitination & Proteasome degradation / Formation of a pool of free 40S subunits / Eukaryotic Translation Termination / SRP-dependent cotranslational protein targeting to membrane / Response of EIF2AK4 (GCN2) to amino acid deficiency / Viral mRNA Translation / Nonsense Mediated Decay (NMD) independent of the Exon Junction Complex (EJC) / GTP hydrolysis and joining of the 60S ribosomal subunit / L13a-mediated translational silencing of Ceruloplasmin expression / Major pathway of rRNA processing in the nucleolus and cytosol / Ub-specific processing proteases / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / enzyme regulator activity / 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 / Neutrophil degranulation / Endosomal Sorting Complex Required For Transport (ESCRT) / Membrane binding and targetting of GAG proteins / Negative regulation of FLT3 / Regulation of TBK1, IKKε (IKBKE)-mediated activation of IRF3, IRF7 / PTK6 Regulates RTKs and Their Effectors AKT1 and DOK1 / Regulation of TBK1, IKKε-mediated activation of IRF3, IRF7 upon TLR3 ligation / Constitutive Signaling by NOTCH1 HD Domain Mutants / IRAK2 mediated activation of TAK1 complex upon TLR7/8 or 9 stimulation / cytosolic ribosome / 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 / Downregulation of ERBB4 signaling / Regulation of FZD by ubiquitination / APC-Cdc20 mediated degradation of Nek2A / p75NTR recruits signalling complexes / 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 / NF-kB is activated and signals survival / Regulation of innate immune responses to cytosolic DNA / Pexophagy / Downregulation of ERBB2:ERBB3 signaling / NRIF signals cell death from the nucleus / VLDLR internalisation and degradation / Regulation of PTEN localization / proteasome complex / Regulation of BACH1 activity / Activated NOTCH1 Transmits Signal to the Nucleus / Synthesis of active ubiquitin: roles of E1 and E2 enzymes / MAP3K8 (TPL2)-dependent MAPK1/3 activation / Translesion synthesis by REV1 / TICAM1, RIP1-mediated IKK complex recruitment / InlB-mediated entry of Listeria monocytogenes into host cell / Translesion synthesis by POLK / Activation of IRF3, IRF7 mediated by TBK1, IKKε (IKBKE) / Downregulation of TGF-beta receptor signaling / JNK (c-Jun kinases) phosphorylation and activation mediated by activated human TAK1 / Josephin domain DUBs / Translesion synthesis by POLI / Gap-filling DNA repair synthesis and ligation in GG-NER / IKK complex recruitment mediated by RIP1 / Regulation of activated PAK-2p34 by proteasome mediated degradation / 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 / TCF dependent signaling in response to WNT / APC/C:Cdc20 mediated degradation of Securin / Regulation of NF-kappa B signaling / N-glycan trimming in the ER and Calnexin/Calreticulin cycle / Asymmetric localization of PCP proteins / activated TAK1 mediates p38 MAPK activation / Ubiquitin-dependent degradation of Cyclin D / SCF-beta-TrCP mediated degradation of Emi1 / NIK-->noncanonical NF-kB signaling / TNFR2 non-canonical NF-kB pathway
Similarity search - Function
26S proteasome regulatory complex, non-ATPase subcomplex, Rpn1 subunit / RPN1, N-terminal / 26S proteasome non-ATPase regulatory subunit RPN1, C-terminal / RPN1 N-terminal domain / 26S proteasome non-ATPase regulatory subunit RPN1 C-terminal / Proteasome/cyclosome repeat / Proteasome/cyclosome repeat / Ribosomal L40e family / Ribosomal_L40e / Ribosomal protein L40e ...26S proteasome regulatory complex, non-ATPase subcomplex, Rpn1 subunit / RPN1, N-terminal / 26S proteasome non-ATPase regulatory subunit RPN1, C-terminal / RPN1 N-terminal domain / 26S proteasome non-ATPase regulatory subunit RPN1 C-terminal / Proteasome/cyclosome repeat / Proteasome/cyclosome repeat / Ribosomal L40e family / Ribosomal_L40e / Ribosomal protein L40e / Ribosomal protein L40e superfamily / : / Ubiquitin domain signature. / Ubiquitin conserved site / Ubiquitin domain / Armadillo-like helical / Ubiquitin family / Ubiquitin homologues / Ubiquitin domain profile. / Ubiquitin-like domain / Armadillo-type fold / Ubiquitin-like domain superfamily
Similarity search - Domain/homology
26S proteasome regulatory subunit RPN1 / Ubiquitin-ribosomal protein eL40 fusion protein
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Homo sapiens (human)
MethodSOLUTION NMR / simulated annealing
AuthorsChen, X. / Walters, K.J.
CitationJournal: Science / Year: 2016
Title: Rpn1 provides adjacent receptor sites for substrate binding and deubiquitination by the proteasome.
Authors: Shi, Y. / Chen, X. / Elsasser, S. / Stocks, B.B. / Tian, G. / Lee, B.H. / Shi, Y. / Zhang, N. / de Poot, S.A. / Tuebing, F. / Sun, S. / Vannoy, J. / Tarasov, S.G. / Engen, J.R. / Finley, D. / Walters, K.J.
History
DepositionJun 10, 2015Deposition site: BMRB / Processing site: RCSB
Revision 1.0Feb 24, 2016Provider: repository / Type: Initial release
Revision 1.1Mar 23, 2016Group: Database references
Revision 1.2Jun 14, 2023Group: Data collection / Database references / Other
Category: database_2 / pdbx_database_status ...database_2 / pdbx_database_status / pdbx_nmr_software / pdbx_nmr_spectrometer
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_database_status.status_code_nmr_data / _pdbx_nmr_software.name / _pdbx_nmr_spectrometer.model
Revision 1.3May 15, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2 / Item: _database_2.pdbx_DOI

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: 26S proteasome regulatory subunit RPN1
B: Ubiquitin-60S ribosomal protein L40
C: Ubiquitin-60S ribosomal protein L40


Theoretical massNumber of molelcules
Total (without water)30,7903
Polymers30,7903
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area2560 Å2
ΔGint-18 kcal/mol
Surface area16540 Å2
MethodPISA
NMR ensembles
DataCriteria
Number of conformers (submitted / calculated)10 / 50
RepresentativeModel #1lowest energy

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Components

#1: Protein 26S proteasome regulatory subunit RPN1 / HMG-CoA reductase degradation protein 2 / Proteasome non-ATPase subunit 1


Mass: 13636.592 Da / Num. of mol.: 1 / Fragment: UNP residues 482-612
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Strain: ATCC 204508 / S288c / Gene: RPN1, HRD2, NAS1, RPD1, YHR027C / Plasmid: Pgex4t1 / Production host: Escherichia coli (E. coli) / References: UniProt: P38764
#2: Protein Ubiquitin-60S ribosomal protein L40 / CEP52 / Ubiquitin A-52 residue ribosomal protein fusion product 1 / Ubiquitin / 60S ribosomal protein L40


Mass: 8576.831 Da / Num. of mol.: 2 / Fragment: UNP residues 1-76
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: UBA52, UBCEP2 / Plasmid: Pet3a / Production host: Escherichia coli (E. coli) / References: UniProt: P62987

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Experimental details

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Experiment

ExperimentMethod: SOLUTION NMR
NMR experiment
Conditions-IDExperiment-IDSolution-IDType
1113D 1H-15N NOESY
1222D 1H-15N HSQC
1333D 1H-13C NOESY aliphatic
1442D 1H-13C HSQC aliphatic
1533D 13C-half filter NOESY
1643D 13C-half filter NOESY
1734D HMQC-NOESY-HMQC

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Sample preparation

Details
Solution-IDContentsSolvent system
10.6 mM [U-100% 15N] protein_1, 0.6 mM protein_2, 50 mM HEPES, 50 mM sodium chloride, 2 mM DTT, 1 mM EDTA, 0.1 % sodium azide, 95% H2O/5% D2O95% H2O/5% D2O
20.5 mM protein_1, 0.5 mM [U-100% 15N] protein_2, 50 mM HEPES, 50 mM sodium chloride, 2 mM DTT, 1 mM EDTA, 0.1 % sodium azide, 95% H2O/5% D2O95% H2O/5% D2O
30.6 mM [U-13C] protein_1, 0.6 mM protein_2, 50 mM HEPES, 50 mM sodium chloride, 2 mM DTT, 1 mM EDTA, 0.1 % sodium azide, 100% D2O100% D2O
40.6 mM protein_1, 0.6 mM [U-13C] protein_2, 50 mM HEPES, 50 mM sodium chloride, 2 mM DTT, 1 mM EDTA, 0.1 % sodium azide, 100% D2O100% D2O
Sample
Conc. (mg/ml)ComponentIsotopic labelingSolution-ID
0.6 mMentity_1-1[U-100% 15N]1
0.6 mMentity_2-21
50 mMHEPES-31
50 mMsodium chloride-41
2 mMDTT-51
1 mMEDTA-61
0.1 %sodium azide-71
0.5 mMentity_1-82
0.5 mMentity_2-9[U-100% 15N]2
50 mMHEPES-102
50 mMsodium chloride-112
2 mMDTT-122
1 mMEDTA-132
0.1 %sodium azide-142
0.6 mMentity_1-15[U-13C]3
0.6 mMentity_2-163
50 mMHEPES-173
50 mMsodium chloride-183
2 mMDTT-193
1 mMEDTA-203
0.1 %sodium azide-213
0.6 mMentity_1-224
0.6 mMentity_2-23[U-13C]4
50 mMHEPES-244
50 mMsodium chloride-254
2 mMDTT-264
1 mMEDTA-274
0.1 %sodium azide-284
Sample conditionsIonic strength: 50 / pH: 6.7 / Pressure: ambient / Temperature: 298 K

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NMR measurement

NMR spectrometerType: Bruker Avance / Manufacturer: Bruker / Model: AVANCE / Field strength: 850 MHz

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Processing

NMR software
NameDeveloperClassification
TopSpinBruker Biospincollection
NMRPipeDelaglio, Grzesiek, Vuister, Zhu, Pfeifer and Baxprocessing
XEASYBartels et al.chemical shift assignment
XEASYBartels et al.data analysis
XEASYBartels et al.peak picking
X-PLOR NIHSchwieters, Kuszewski, Tjandra and Clorerefinement
X-PLOR NIHSchwieters, Kuszewski, Tjandra and Clorestructure solution
ProcheckNMRLaskowski and MacArthurdata analysis
RefinementMethod: simulated annealing / Software ordinal: 1
NMR representativeSelection criteria: lowest energy
NMR ensembleConformers calculated total number: 50 / Conformers submitted total number: 10

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