+データを開く
-基本情報
登録情報 | データベース: PDB / ID: 5kgf | ||||||||||||
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タイトル | Structural model of 53BP1 bound to a ubiquitylated and methylated nucleosome, at 4.5 A resolution | ||||||||||||
要素 |
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キーワード | STRUCTURAL PROTEIN/DNA / DNA / chromatin / 53BP1 / STRUCTURAL PROTEIN-DNA complex | ||||||||||||
機能・相同性 | 機能・相同性情報 ubiquitin-modified histone reader activity / intracellular organelle / positive regulation of isotype switching / cellular response to X-ray / double-strand break repair via classical nonhomologous end joining / protein localization to site of double-strand break / hypothalamus gonadotrophin-releasing hormone neuron development / DNA repair complex / female meiosis I / positive regulation of protein monoubiquitination ...ubiquitin-modified histone reader activity / intracellular organelle / positive regulation of isotype switching / cellular response to X-ray / double-strand break repair via classical nonhomologous end joining / protein localization to site of double-strand break / hypothalamus gonadotrophin-releasing hormone neuron development / DNA repair complex / female meiosis I / positive regulation of protein monoubiquitination / mitochondrion transport along microtubule / fat pad development / telomeric DNA binding / female gonad development / seminiferous tubule development / male meiosis I / positive regulation of intrinsic apoptotic signaling pathway by p53 class mediator / SUMOylation of transcription factors / negative regulation of double-strand break repair via homologous recombination / regulation of neuron apoptotic process / Replacement of protamines by nucleosomes in the male pronucleus / regulation of proteasomal protein catabolic process / Packaging Of Telomere Ends / energy homeostasis / Recognition and association of DNA glycosylase with site containing an affected purine / Cleavage of the damaged purine / Maturation of protein E / Deposition of new CENPA-containing nucleosomes at the centromere / Maturation of protein E / ER Quality Control Compartment (ERQC) / nucleosomal DNA binding / Myoclonic epilepsy of Lafora / IRAK2 mediated activation of TAK1 complex / Alpha-protein kinase 1 signaling pathway / FLT3 signaling by CBL mutants / Recognition and association of DNA glycosylase with site containing an affected pyrimidine / Cleavage of the damaged pyrimidine / IRAK1 recruits IKK complex / IRAK1 recruits IKK complex upon TLR7/8 or 9 stimulation / methylated histone binding / Prevention of phagosomal-lysosomal fusion / Glycogen synthesis / IRAK2 mediated activation of TAK1 complex upon TLR7/8 or 9 stimulation / Inhibition of DNA recombination at telomere / Endosomal Sorting Complex Required For Transport (ESCRT) / Regulation of TBK1, IKKε (IKBKE)-mediated activation of IRF3, IRF7 / TICAM1,TRAF6-dependent induction of TAK1 complex / Membrane binding and targetting of GAG proteins / Regulation of TBK1, IKKε-mediated activation of IRF3, IRF7 upon TLR3 ligation / Negative regulation of FLT3 / Constitutive Signaling by NOTCH1 HD Domain Mutants / PTK6 Regulates RTKs and Their Effectors AKT1 and DOK1 / Regulation of FZD by ubiquitination / TICAM1-dependent activation of IRF3/IRF7 / histone reader activity / NOTCH2 Activation and Transmission of Signal to the Nucleus / p75NTR recruits signalling complexes / APC/C:Cdc20 mediated degradation of Cyclin B / VLDLR internalisation and degradation / Meiotic synapsis / Downregulation of ERBB4 signaling / TRAF6-mediated induction of TAK1 complex within TLR4 complex / TRAF6 mediated IRF7 activation in TLR7/8 or 9 signaling / APC-Cdc20 mediated degradation of Nek2A / RNA Polymerase I Promoter Opening / Regulation of innate immune responses to cytosolic DNA / NF-kB is activated and signals survival / InlA-mediated entry of Listeria monocytogenes into host cells / Regulation of pyruvate metabolism / Downregulation of ERBB2:ERBB3 signaling / Assembly of the ORC complex at the origin of replication / NRIF signals cell death from the nucleus / Pexophagy / Activated NOTCH1 Transmits Signal to the Nucleus / Regulation of PTEN localization / Regulation of BACH1 activity / Synthesis of active ubiquitin: roles of E1 and E2 enzymes / TICAM1, RIP1-mediated IKK complex recruitment / Translesion synthesis by REV1 / MAP3K8 (TPL2)-dependent MAPK1/3 activation / DNA methylation / Translesion synthesis by POLK / neuron projection morphogenesis / Condensation of Prophase Chromosomes / Activation of IRF3, IRF7 mediated by TBK1, IKKε (IKBKE) / Downregulation of TGF-beta receptor signaling / Translesion synthesis by POLI / IKK complex recruitment mediated by RIP1 / Regulation of activated PAK-2p34 by proteasome mediated degradation / JNK (c-Jun kinases) phosphorylation and activation mediated by activated human TAK1 / Gap-filling DNA repair synthesis and ligation in GG-NER / regulation of mitochondrial membrane potential / SIRT1 negatively regulates rRNA expression / Josephin domain DUBs / InlB-mediated entry of Listeria monocytogenes into host cell / HCMV Late Events / Chromatin modifications during the maternal to zygotic transition (MZT) / ERCC6 (CSB) and EHMT2 (G9a) positively regulate rRNA expression / PINK1-PRKN Mediated Mitophagy / DNA damage checkpoint signaling 類似検索 - 分子機能 | ||||||||||||
生物種 | Xenopus laevis (アフリカツメガエル) Homo sapiens (ヒト) synthetic construct (人工物) | ||||||||||||
手法 | 電子顕微鏡法 / 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 4.54 Å | ||||||||||||
データ登録者 | Wilson, M.D. / Benlekbir, S. / Sicheri, F. / Rubinstein, J.L. / Durocher, D. | ||||||||||||
資金援助 | カナダ, 3件
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引用 | ジャーナル: Nature / 年: 2016 タイトル: The structural basis of modified nucleosome recognition by 53BP1. 要旨: DNA double-strand breaks (DSBs) elicit a histone modification cascade that controls DNA repair. This pathway involves the sequential ubiquitination of histones H1 and H2A by the E3 ubiquitin ligases ...DNA double-strand breaks (DSBs) elicit a histone modification cascade that controls DNA repair. This pathway involves the sequential ubiquitination of histones H1 and H2A by the E3 ubiquitin ligases RNF8 and RNF168, respectively. RNF168 ubiquitinates H2A on lysine 13 and lysine 15 (refs 7, 8) (yielding H2AK13ub and H2AK15ub, respectively), an event that triggers the recruitment of 53BP1 (also known as TP53BP1) to chromatin flanking DSBs. 53BP1 binds specifically to H2AK15ub-containing nucleosomes through a peptide segment termed the ubiquitination-dependent recruitment motif (UDR), which requires the simultaneous engagement of histone H4 lysine 20 dimethylation (H4K20me2) by its tandem Tudor domain. How 53BP1 interacts with these two histone marks in the nucleosomal context, how it recognizes ubiquitin, and how it discriminates between H2AK13ub and H2AK15ub is unknown. Here we present the electron cryomicroscopy (cryo-EM) structure of a dimerized human 53BP1 fragment bound to a H4K20me2-containing and H2AK15ub-containing nucleosome core particle (NCP-ubme) at 4.5 Å resolution. The structure reveals that H4K20me2 and H2AK15ub recognition involves intimate contacts with multiple nucleosomal elements including the acidic patch. Ubiquitin recognition by 53BP1 is unusual and involves the sandwiching of the UDR segment between ubiquitin and the NCP surface. The selectivity for H2AK15ub is imparted by two arginine fingers in the H2A amino-terminal tail, which straddle the nucleosomal DNA and serve to position ubiquitin over the NCP-bound UDR segment. The structure of the complex between NCP-ubme and 53BP1 reveals the basis of 53BP1 recruitment to DSB sites and illuminates how combinations of histone marks and nucleosomal elements cooperate to produce highly specific chromatin responses, such as those elicited following chromosome breaks. | ||||||||||||
履歴 |
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-構造の表示
ムービー |
ムービービューア |
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構造ビューア | 分子: MolmilJmol/JSmol |
-ダウンロードとリンク
-ダウンロード
PDBx/mmCIF形式 | 5kgf.cif.gz | 358.9 KB | 表示 | PDBx/mmCIF形式 |
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PDB形式 | pdb5kgf.ent.gz | 279.5 KB | 表示 | PDB形式 |
PDBx/mmJSON形式 | 5kgf.json.gz | ツリー表示 | PDBx/mmJSON形式 | |
その他 | その他のダウンロード |
-検証レポート
文書・要旨 | 5kgf_validation.pdf.gz | 981.4 KB | 表示 | wwPDB検証レポート |
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文書・詳細版 | 5kgf_full_validation.pdf.gz | 1 MB | 表示 | |
XML形式データ | 5kgf_validation.xml.gz | 47.9 KB | 表示 | |
CIF形式データ | 5kgf_validation.cif.gz | 72.6 KB | 表示 | |
アーカイブディレクトリ | https://data.pdbj.org/pub/pdb/validation_reports/kg/5kgf ftp://data.pdbj.org/pub/pdb/validation_reports/kg/5kgf | HTTPS FTP |
-関連構造データ
-リンク
-集合体
登録構造単位 |
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-要素
-タンパク質 , 5種, 10分子 AEBFCGDHOM
#1: タンパク質 | 分子量: 15421.101 Da / 分子数: 2 / 由来タイプ: 組換発現 由来: (組換発現) Xenopus laevis (アフリカツメガエル) 発現宿主: Escherichia coli (大腸菌) / 参照: UniProt: P84233 #2: タンパク質 | 分子量: 11439.511 Da / 分子数: 2 / Mutation: K20C / 由来タイプ: 組換発現 / 詳細: cysteine alkylation at position 20 由来: (組換発現) Xenopus laevis (アフリカツメガエル) 詳細 (発現宿主): pDD2349 / 発現宿主: Escherichia coli (大腸菌) / 参照: UniProt: P62799 #3: タンパク質 | 分子量: 14163.539 Da / 分子数: 2 / Mutation: K13R, T16S, K36R / 由来タイプ: 組換発現 詳細: Isopeptide amide crosslink between K15 of H2A and G76 of ubiquitin 由来: (組換発現) Homo sapiens (ヒト) 遺伝子: HIST1H2AG, H2AFP, HIST1H2AI, H2AFC, HIST1H2AK, H2AFD, HIST1H2AL, H2AFI, HIST1H2AM, H2AFN 発現宿主: Escherichia coli (大腸菌) / 参照: UniProt: P0C0S8 #4: タンパク質 | 分子量: 13937.213 Da / 分子数: 2 / 由来タイプ: 組換発現 / 由来: (組換発現) Homo sapiens (ヒト) 遺伝子: HIST1H2BC, H2BFL, HIST1H2BE, H2BFH, HIST1H2BF, H2BFG, HIST1H2BG, H2BFA, HIST1H2BI, H2BFK 発現宿主: Escherichia coli (大腸菌) / 参照: UniProt: P62807 #8: タンパク質 | 分子量: 8576.831 Da / 分子数: 2 / 由来タイプ: 組換発現 / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: UBB / 発現宿主: Escherichia coli (大腸菌) / 参照: UniProt: P0CG47 |
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-DNA鎖 , 2種, 2分子 IJ
#5: DNA鎖 | 分子量: 44520.383 Da / 分子数: 1 / 由来タイプ: 組換発現 / 由来: (組換発現) synthetic construct (人工物) / 発現宿主: Escherichia coli (大腸菌) |
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#6: DNA鎖 | 分子量: 44991.660 Da / 分子数: 1 / 由来タイプ: 組換発現 / 由来: (組換発現) synthetic construct (人工物) / 発現宿主: Escherichia coli (大腸菌) |
-タンパク質・ペプチド , 1種, 2分子 LK
#7: タンパク質・ペプチド | 分子量: 2376.757 Da / 分子数: 2 / 由来タイプ: 組換発現 / 詳細: full protein not modeled / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: TP53BP1 / 発現宿主: Escherichia coli (大腸菌) / 参照: UniProt: H7BZY0, UniProt: Q12888*PLUS |
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-実験情報
-実験
実験 | 手法: 電子顕微鏡法 |
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EM実験 | 試料の集合状態: PARTICLE / 3次元再構成法: 単粒子再構成法 |
-試料調製
構成要素 |
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分子量 |
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由来(天然) |
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由来(組換発現) |
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緩衝液 | pH: 7.5 詳細: High concentration NCP-ubme/GST-53BP1 complex at 200 mM salt was diluted just prior to grid freezing. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
緩衝液成分 |
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試料 | 濃度: 0.6 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
試料支持 | グリッドの材料: COPPER/RHODIUM / グリッドのサイズ: 400 divisions/in. / グリッドのタイプ: electron micrsocopy sciences | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
急速凍結 | 装置: FEI VITROBOT MARK III / 凍結剤: ETHANE-PROPANE / 湿度: 100 % / 凍結前の試料温度: 277 K 詳細: Plunged into liquid ethane-propane (FEI VITROBOT MARK III) |
-電子顕微鏡撮影
実験機器 | モデル: Tecnai F20 / 画像提供: FEI Company |
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顕微鏡 | モデル: FEI TECNAI F20 |
電子銃 | 電子線源: FIELD EMISSION GUN / 加速電圧: 200 kV / 照射モード: FLOOD BEAM |
電子レンズ | モード: BRIGHT FIELD / 倍率(公称値): 25000 X / 倍率(補正後): 34483 X / Cs: 2 mm / C2レンズ絞り径: 30 µm / アライメント法: COMA FREE |
試料ホルダ | 凍結剤: NITROGEN 試料ホルダーモデル: GATAN 626 SINGLE TILT LIQUID NITROGEN CRYO TRANSFER HOLDER |
撮影 | 平均露光時間: 0.5 sec. / 電子線照射量: 36 e/Å2 / 検出モード: COUNTING フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) 撮影したグリッド数: 2 / 実像数: 319 |
画像スキャン | 動画フレーム数/画像: 30 / 利用したフレーム数/画像: 1-30 |
-解析
ソフトウェア | 名称: PHENIX / バージョン: 1.10.1_2155: / 分類: 精密化 | ||||||||||||||||||||||||||||||||||||
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EMソフトウェア |
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CTF補正 | タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||
粒子像の選択 | 選択した粒子像数: 174185 詳細: Automatically picked from roughly 3000 particles using a manually picked template | ||||||||||||||||||||||||||||||||||||
対称性 | 点対称性: C2 (2回回転対称) | ||||||||||||||||||||||||||||||||||||
3次元再構成 | 解像度: 4.54 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 45361 / アルゴリズム: FOURIER SPACE / クラス平均像の数: 9 / 対称性のタイプ: POINT | ||||||||||||||||||||||||||||||||||||
原子モデル構築 | B value: 207.5 / プロトコル: RIGID BODY FIT / 空間: REAL 詳細: The atomic models of Widom-601 DNA (PDB ID 3LZ0), octameric histones (PDB ID 1KX5), ubiquitin (PDB ID 1UBI), and H4K20me2/53BP1 tandem Tudor domain (PDB ID 2IG0) were fitted without allowing ...詳細: The atomic models of Widom-601 DNA (PDB ID 3LZ0), octameric histones (PDB ID 1KX5), ubiquitin (PDB ID 1UBI), and H4K20me2/53BP1 tandem Tudor domain (PDB ID 2IG0) were fitted without allowing flexibility into the 3D maps using UCSF Chimera. Segmentation was performed in UCSF Chimera. For the NCP-ubme structure the ubiquitin segmentation was further modified to remove obvious NCP density from the ubiquitin segment. The H2A/H2B sequence was mutated to the human H2AK13R/K36R and H2B manually in UCSF Chimera. A polyalanine model of the UDR was built within the UDR density in Coot, which compared well to predicted structures generated by Rosetta. The UDR model was mutated and fitted using UCSF Chimera, followed by iterative rounds of real-space refinement in PHENIX and model optimization in Coot. All figures were prepared in UCSF Chimera. | ||||||||||||||||||||||||||||||||||||
精密化 | 最高解像度: 4.54 Å | ||||||||||||||||||||||||||||||||||||
拘束条件 |
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