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- PDB-8uqb: Crystal structure of RNF168 (RING)-UbcH5c fused to H2A-H2B via a ... -
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Basic information
Entry | Database: PDB / ID: 8uqb | |||||||||
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Title | Crystal structure of RNF168 (RING)-UbcH5c fused to H2A-H2B via a 20-residue linker (crystallization condition 1) | |||||||||
![]() | E3 ubiquitin-protein ligase RNF168,Ubiquitin-conjugating enzyme E2 D3,Histone H2B type 2-E,Histone H2A type 1-B/E | |||||||||
![]() | TRANSFERASE / RNF168 / UbcH5c / Histone H2A / Histone H2B / Chromatin / Ubiquitin ligase / Ubiquitin-conjugating enzyme / DNA damage response / DNA double-strand break repair / PROTEIN BINDING / PROTEIN BINDING-Transferase complex | |||||||||
Function / homology | ![]() histone H2AK15 ubiquitin ligase activity / histone ubiquitin ligase activity / (E3-independent) E2 ubiquitin-conjugating enzyme / Signaling by BMP / protein K6-linked ubiquitination / double-strand break repair via classical nonhomologous end joining / isotype switching / protein K11-linked ubiquitination / DNA repair-dependent chromatin remodeling / positive regulation of protein targeting to mitochondrion ...histone H2AK15 ubiquitin ligase activity / histone ubiquitin ligase activity / (E3-independent) E2 ubiquitin-conjugating enzyme / Signaling by BMP / protein K6-linked ubiquitination / double-strand break repair via classical nonhomologous end joining / isotype switching / protein K11-linked ubiquitination / DNA repair-dependent chromatin remodeling / positive regulation of protein targeting to mitochondrion / K63-linked polyubiquitin modification-dependent protein binding / E2 ubiquitin-conjugating enzyme / response to ionizing radiation / negative regulation of transcription elongation by RNA polymerase II / protein monoubiquitination / ubiquitin conjugating enzyme activity / protein K63-linked ubiquitination / negative regulation of BMP signaling pathway / protein localization to CENP-A containing chromatin / protein autoubiquitination / protein K48-linked ubiquitination / Replacement of protamines by nucleosomes in the male pronucleus / CENP-A containing nucleosome / interstrand cross-link repair / SUMOylation of DNA damage response and repair proteins / heterochromatin organization / epigenetic regulation of gene expression / Packaging Of Telomere Ends / ubiquitin ligase complex / Recognition and association of DNA glycosylase with site containing an affected purine / Cleavage of the damaged purine / nucleosome binding / Deposition of new CENPA-containing nucleosomes at the centromere / nucleosomal DNA binding / Recognition and association of DNA glycosylase with site containing an affected pyrimidine / Cleavage of the damaged pyrimidine / Inhibition of DNA recombination at telomere / Meiotic synapsis / positive regulation of DNA repair / RNA Polymerase I Promoter Opening / Assembly of the ORC complex at the origin of replication / TICAM1, RIP1-mediated IKK complex recruitment / DNA methylation / Condensation of Prophase Chromosomes / IKK complex recruitment mediated by RIP1 / ERCC6 (CSB) and EHMT2 (G9a) positively regulate rRNA expression / SIRT1 negatively regulates rRNA expression / Chromatin modifications during the maternal to zygotic transition (MZT) / HCMV Late Events / PRC2 methylates histones and DNA / innate immune response in mucosa / ubiquitin binding / Defective pyroptosis / Negative regulators of DDX58/IFIH1 signaling / HDACs deacetylate histones / RNA Polymerase I Promoter Escape / Peroxisomal protein import / Nonhomologous End-Joining (NHEJ) / Downregulation of SMAD2/3:SMAD4 transcriptional activity / Regulation of TNFR1 signaling / Transcriptional regulation by small RNAs / Formation of the beta-catenin:TCF transactivating complex / RUNX1 regulates genes involved in megakaryocyte differentiation and platelet function / RING-type E3 ubiquitin transferase / NoRC negatively regulates rRNA expression / Activated PKN1 stimulates transcription of AR (androgen receptor) regulated genes KLK2 and KLK3 / protein modification process / Oxygen-dependent proline hydroxylation of Hypoxia-inducible Factor Alpha / B-WICH complex positively regulates rRNA expression / G2/M DNA damage checkpoint / Inactivation of CSF3 (G-CSF) signaling / DNA Damage/Telomere Stress Induced Senescence / Metalloprotease DUBs / Meiotic recombination / RMTs methylate histone arginines / Pre-NOTCH Transcription and Translation / Activation of anterior HOX genes in hindbrain development during early embryogenesis / HCMV Early Events / double-strand break repair via nonhomologous end joining / Transcriptional regulation of granulopoiesis / protein polyubiquitination / structural constituent of chromatin / antimicrobial humoral immune response mediated by antimicrobial peptide / ubiquitin-protein transferase activity / UCH proteinases / ubiquitin protein ligase activity / nucleosome / double-strand break repair / nucleosome assembly / Antigen processing: Ubiquitination & Proteasome degradation / E3 ubiquitin ligases ubiquitinate target proteins / Recruitment and ATM-mediated phosphorylation of repair and signaling proteins at DNA double strand breaks / chromatin organization / RUNX1 regulates transcription of genes involved in differentiation of HSCs / Neddylation / site of double-strand break / HATs acetylate histones / Processing of DNA double-strand break ends / histone binding / antibacterial humoral response Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | ![]() ![]() ![]() | |||||||||
![]() | Hu, Q. / Botuyan, M.V. / Mer, G. | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Mechanisms of RNF168 nucleosome recognition and ubiquitylation. Authors: Qi Hu / Debiao Zhao / Gaofeng Cui / Janarjan Bhandari / James R Thompson / Maria Victoria Botuyan / Georges Mer / ![]() Abstract: RNF168 plays a central role in the DNA damage response (DDR) by ubiquitylating histone H2A at K13 and K15. These modifications direct BRCA1-BARD1 and 53BP1 foci formation in chromatin, essential for ...RNF168 plays a central role in the DNA damage response (DDR) by ubiquitylating histone H2A at K13 and K15. These modifications direct BRCA1-BARD1 and 53BP1 foci formation in chromatin, essential for cell-cycle-dependent DNA double-strand break (DSB) repair pathway selection. The mechanism by which RNF168 catalyzes the targeted accumulation of H2A ubiquitin conjugates to form repair foci around DSBs remains unclear. Here, using cryoelectron microscopy (cryo-EM), nuclear magnetic resonance (NMR) spectroscopy, and functional assays, we provide a molecular description of the reaction cycle and dynamics of RNF168 as it modifies the nucleosome and recognizes its ubiquitylation products. We demonstrate an interaction of a canonical ubiquitin-binding domain within full-length RNF168, which not only engages ubiquitin but also the nucleosome surface, clarifying how such site-specific ubiquitin recognition propels a signal amplification loop. Beyond offering mechanistic insights into a key DDR protein, our study aids in understanding site specificity in both generating and interpreting chromatin ubiquitylation. | |||||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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PDBx/mmCIF format | ![]() | 96.9 KB | Display | ![]() |
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PDB format | ![]() | 70 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Summary document | ![]() | 1.2 MB | Display | ![]() |
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Full document | ![]() | 1.2 MB | Display | |
Data in XML | ![]() | 16 KB | Display | |
Data in CIF | ![]() | 21.5 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 8smwC ![]() 8smxC ![]() 8smyC ![]() 8smzC ![]() 8sn0C ![]() 8sn1C ![]() 8sn2C ![]() 8sn3C ![]() 8sn4C ![]() 8sn5C ![]() 8sn6C ![]() 8sn7C ![]() 8sn8C ![]() 8sn9C ![]() 8snaC ![]() 8txvC ![]() 8txwC ![]() 8txxC ![]() 8u13C ![]() 8u14C ![]() 8upfC ![]() 8uq8C ![]() 8uq9C ![]() 8uqaC ![]() 8uqcC ![]() 8uqdC ![]() 8uqeC ![]() 4gb0S ![]() 5eggS S: Starting model for refinement 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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Unit cell |
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Components
#1: Protein | Mass: 50040.320 Da / Num. of mol.: 1 / Mutation: C85K in UbcH5c Source method: isolated from a genetically manipulated source Details: residues 1-94 of RNF168, followed by residues 2-147 of UbcH5c, followed by residues 33-123 of H2B, followed by residues 12-105 of H2A Source: (gene. exp.) ![]() Gene: RNF168, UBE2D3, UBC5C, UBCH5C, H2BC21, H2BFQ, HIST2H2BE, H2AC4, H2AFM, HIST1H2AB, H2AC8, H2AFA, HIST1H2AE Production host: ![]() ![]() References: UniProt: Q8IYW5, UniProt: P61077, UniProt: Q16778, UniProt: P04908, RING-type E3 ubiquitin transferase, E2 ubiquitin-conjugating enzyme, (E3-independent) E2 ubiquitin-conjugating enzyme | ||||||
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#2: Chemical | #3: Chemical | ChemComp-CL / #4: Water | ChemComp-HOH / | Has ligand of interest | Y | |
-Experimental details
-Experiment
Experiment | Method: ![]() |
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Sample preparation
Crystal | Density Matthews: 2.75 Å3/Da / Density % sol: 55.27 % |
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Crystal grow | Temperature: 288 K / Method: vapor diffusion, hanging drop / pH: 7 Details: Crystals were obtained by mixing 2 microliters of the protein fusion (12 mg/mL) in 10 mM HEPES, pH 7.5, 600 mM NaCl, 1 mM TCEP and 2 microliters of the reservoir solution containing 50 mM ...Details: Crystals were obtained by mixing 2 microliters of the protein fusion (12 mg/mL) in 10 mM HEPES, pH 7.5, 600 mM NaCl, 1 mM TCEP and 2 microliters of the reservoir solution containing 50 mM BIS-TRIS propane, pH 7.0, 1 M NaCl |
-Data collection
Diffraction | Mean temperature: 100 K / Serial crystal experiment: N |
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Diffraction source | Source: ![]() ![]() ![]() |
Detector | Type: ADSC QUANTUM 315r / Detector: CCD / Date: Feb 2, 2018 |
Radiation | Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray |
Radiation wavelength | Wavelength: 0.9792 Å / Relative weight: 1 |
Reflection | Resolution: 2.484→50 Å / Num. obs: 18662 / % possible obs: 99.36 % / Redundancy: 5.3 % / Biso Wilson estimate: 44.24 Å2 / CC1/2: 0.996 / CC star: 0.999 / Rmerge(I) obs: 0.113 / Rpim(I) all: 0.05322 / Rrim(I) all: 0.1255 / Net I/σ(I): 16.39 |
Reflection shell | Resolution: 2.484→2.573 Å / Redundancy: 4.6 % / Rmerge(I) obs: 0.7775 / Mean I/σ(I) obs: 2.66 / Num. unique obs: 1718 / CC1/2: 0.731 / CC star: 0.919 / Rpim(I) all: 0.3962 / Rrim(I) all: 0.8774 / % possible all: 94.24 |
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Processing
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Refinement | Method to determine structure: ![]() Starting model: 4GB0, 5EGG Resolution: 2.484→39.37 Å / SU ML: 0.34 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 26.4 / Stereochemistry target values: ML
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Solvent computation | Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement step | Cycle: LAST / Resolution: 2.484→39.37 Å
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Refine LS restraints |
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LS refinement shell |
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