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データを開く
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基本情報
登録情報 | データベース: PDB / ID: 8qyx | ||||||
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タイトル | Human 60S ribosomal subunit | ||||||
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![]() | RIBOSOME / Large ribosomal subunit / HeLa / modifications / polyamines | ||||||
機能・相同性 | ![]() translation at presynapse / exit from mitosis / male meiosis I / eukaryotic 80S initiation complex / negative regulation of protein neddylation / optic nerve development / response to insecticide / regulation of translation involved in cellular response to UV / axial mesoderm development / negative regulation of formation of translation preinitiation complex ...translation at presynapse / exit from mitosis / male meiosis I / eukaryotic 80S initiation complex / negative regulation of protein neddylation / optic nerve development / response to insecticide / regulation of translation involved in cellular response to UV / axial mesoderm development / negative regulation of formation of translation preinitiation complex / regulation of G1 to G0 transition / ribosomal protein import into nucleus / 90S preribosome assembly / protein-DNA complex disassembly / positive regulation of intrinsic apoptotic signaling pathway in response to DNA damage by p53 class mediator / retinal ganglion cell axon guidance / GAIT complex / positive regulation of DNA damage response, signal transduction by p53 class mediator / TORC2 complex binding / alpha-beta T cell differentiation / G1 to G0 transition / middle ear morphogenesis / cytoplasmic side of rough endoplasmic reticulum membrane / negative regulation of ubiquitin protein ligase activity / homeostatic process / macrophage chemotaxis / lung morphogenesis / positive regulation of natural killer cell proliferation / Protein hydroxylation / Peptide chain elongation / Selenocysteine synthesis / Formation of a pool of free 40S subunits / Eukaryotic Translation Termination / ubiquitin ligase inhibitor activity / blastocyst development / cellular response to actinomycin D / Response of EIF2AK4 (GCN2) to amino acid deficiency / SRP-dependent cotranslational protein targeting to membrane / negative regulation of ubiquitin-dependent protein catabolic process / positive regulation of signal transduction by p53 class mediator / Viral mRNA Translation / Maturation of protein E / Maturation of protein E / Nonsense Mediated Decay (NMD) independent of the Exon Junction Complex (EJC) / protein localization to nucleus / GTP hydrolysis and joining of the 60S ribosomal subunit / ER Quality Control Compartment (ERQC) / Myoclonic epilepsy of Lafora / FLT3 signaling by CBL mutants / Prevention of phagosomal-lysosomal fusion / L13a-mediated translational silencing of Ceruloplasmin expression / IRAK2 mediated activation of TAK1 complex / Alpha-protein kinase 1 signaling pathway / Glycogen synthesis / positive regulation of protein binding / 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 / 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 / NOTCH2 Activation and Transmission of Signal to the Nucleus / Major pathway of rRNA processing in the nucleolus and cytosol / TICAM1,TRAF6-dependent induction of TAK1 complex / protein targeting / 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 / protein-RNA complex assembly / Regulation of pyruvate metabolism / TRAF6-mediated induction of TAK1 complex within TLR4 complex / TRAF6 mediated IRF7 activation in TLR7/8 or 9 signaling / Regulation of innate immune responses to cytosolic DNA / NF-kB is activated and signals survival / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / Downregulation of ERBB2:ERBB3 signaling / NRIF signals cell death from the nucleus / Pexophagy / VLDLR internalisation and degradation / maturation of LSU-rRNA / Regulation of PTEN localization / Activated NOTCH1 Transmits Signal to the Nucleus / Regulation of BACH1 activity / MAP3K8 (TPL2)-dependent MAPK1/3 activation / rough endoplasmic reticulum / Translesion synthesis by REV1 / Synthesis of active ubiquitin: roles of E1 and E2 enzymes / 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 / Josephin domain DUBs 類似検索 - 分子機能 | ||||||
生物種 | ![]() | ||||||
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: 1.78 Å | ||||||
![]() | Wiechert, F. / Schacherl, M. / Sprink, T. | ||||||
資金援助 | ![]()
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![]() | ![]() タイトル: Visualizing the modification landscape of the human 60S ribosomal subunit at close to atomic resolution. 著者: Franziska Wiechert / Anett Unbehaun / Thiemo Sprink / Helena Seibel / Jörg Bürger / Justus Loerke / Thorsten Mielke / Christoph A Diebolder / Magdalena Schacherl / Christian M T Spahn / ![]() 要旨: Chemical modifications of ribosomal RNAs (rRNAs) and proteins expand their topological repertoire, and together with the plethora of bound ligands, fine-tune ribosomal function. Detailed knowledge of ...Chemical modifications of ribosomal RNAs (rRNAs) and proteins expand their topological repertoire, and together with the plethora of bound ligands, fine-tune ribosomal function. Detailed knowledge of this natural composition provides important insights into ribosome genesis and function and clarifies some aspects of ribosomopathies. The discovery of new structural properties and functional aspects of ribosomes has gone hand in hand with cryo-electron microscopy (cryo-EM) and its technological development. In line with the ability to visualize atomic details - a prerequisite for identifying chemical modifications and ligands in cryo-EM maps - in this work we present the structure of the 60S ribosomal subunit from HeLa cells at the very high global resolution of 1.78 Å. We identified 113 rRNA modifications and four protein modifications including uL2-Hisβ-ox216, which stabilizes the local structure near the peptidyl transferase centre via an extended hydrogen-bonding network. We can differentiate metal ions Mg2+ and K+, polyamines spermine, spermidine and putrescine and identify thousands of water molecules binding to the 60S subunit. Approaching atomic resolution cryo-EM has become a powerful tool to examine fine details of macromolecular structures that will expand our knowledge about translation and other biological processes in the future and assess the variability of the chemical space due to differences between species/tissues or varying physicochemical environment. | ||||||
履歴 |
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構造の表示
構造ビューア | 分子: ![]() ![]() |
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ダウンロードとリンク
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ダウンロード
PDBx/mmCIF形式 | ![]() | 3.4 MB | 表示 | ![]() |
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PDB形式 | ![]() | 表示 | ![]() | |
PDBx/mmJSON形式 | ![]() | ツリー表示 | ![]() | |
その他 | ![]() |
-検証レポート
文書・要旨 | ![]() | 1.8 MB | 表示 | ![]() |
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文書・詳細版 | ![]() | 2 MB | 表示 | |
XML形式データ | ![]() | 238.6 KB | 表示 | |
CIF形式データ | ![]() | 459.3 KB | 表示 | |
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-関連構造データ
関連構造データ | ![]() 18765MC M: このデータのモデリングに利用したマップデータ C: 同じ文献を引用 ( |
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類似構造データ | 類似検索 - 機能・相同性 ![]() |
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リンク
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集合体
登録構造単位 | ![]()
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要素
-Large ribosomal subunit protein ... , 5種, 5分子 A1U1g1D2E2
#1: タンパク質 | 分子量: 29290.973 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() |
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#21: タンパク質 | 分子量: 16619.527 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() |
#33: タンパク質 | 分子量: 14771.411 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() |
#40: タンパク質 | 分子量: 28103.855 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() |
#41: タンパク質 | 分子量: 46224.133 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() |
+60S ribosomal protein ... , 35種, 35分子 B1C1E1F1G1H1I1J1K1L1M1N1O1P1Q1R1S1T1V1W1X1Y1Z1a1b1c1d1e1f1i1...
-タンパク質 , 1種, 1分子 D1
#4: タンパク質 | 分子量: 24570.949 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() |
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-RNA鎖 , 3種, 3分子 A2B2C2
#37: RNA鎖 | 分子量: 1640816.625 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() |
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#38: RNA鎖 | 分子量: 38691.914 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() |
#39: RNA鎖 | 分子量: 50463.840 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() |
-非ポリマー , 8種, 12074分子 














#45: 化合物 | ChemComp-MG / #46: 化合物 | ChemComp-PUT / #47: 化合物 | ChemComp-K / #48: 化合物 | ChemComp-ZN / #49: 化合物 | ChemComp-SPD / #50: 化合物 | ChemComp-SPM / #51: 化合物 | ChemComp-ATP / | #52: 水 | ChemComp-HOH / | |
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-詳細
研究の焦点であるリガンドがあるか | N |
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Has protein modification | Y |
-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 |
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試料調製
構成要素 | 名称: Human mature large ribosomal subunit / タイプ: RIBOSOME / Entity ID: #1-#44 / 由来: NATURAL |
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由来(天然) | 生物種: ![]() |
緩衝液 | pH: 7.5 |
試料 | 濃度: 0.4583 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES 詳細: Hela 60S ribosomal subunits were isolated from cytoplasmic extracts of Hela cells, involving puromycin treatment and sucrose gradient centrifugation in high salt. The gradients were manually ...詳細: Hela 60S ribosomal subunits were isolated from cytoplasmic extracts of Hela cells, involving puromycin treatment and sucrose gradient centrifugation in high salt. The gradients were manually fractionated, and separated peak fractions of 60S subunits were collected. For buffer exchange, the sample was diluted 6 x in buffer F (20 mM HEPES pH7.5, 10 mM KCl, 2 mM MgCl2, 2 mM DTT) supplemented with 8.56 % sucrose to adjust the sucrose in the storage buffer to 12%, respectively. Finally, the 60S subunit suspension was concentrated on Amicon Ultracel filter units with a 10 kDa cutoff (Merck Millipore) was snap frozen in 10 ul aliquots and stored at -80 degrees Celsius. Concentrated Hela 60S ribosomal subunits were diluted to 200 nM in buffer E (20 mM HEPES pH7.5, 2 mM MgCl2, 90 mM KCl, 0.5 mM Spermidine, 0.04 mM Spermine, 2 mM DTT) and 4 ul were spotted onto glow-discharged holey carbon grids coated with a continuous thin carbon film (Quantifoil Cu200 R2/2+) and incubated for 40 sec at 4 degrees CelsiusC, 85% RH. The excess of liquid was blotted away for 2 sec and the grids were plunge frozen using the Vitrobot Mark IV (Thermo Fischer Scientific). |
試料支持 | 詳細: Operated at 15 mA, easiGlow Discharge Cleaning system (PELCO) グリッドの材料: COPPER / グリッドのサイズ: 200 divisions/in. / グリッドのタイプ: Quantifoil R2/2 |
急速凍結 | 装置: FEI VITROBOT MARK IV / 凍結剤: ETHANE / 湿度: 85 % / 凍結前の試料温度: 277 K |
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電子顕微鏡撮影
実験機器 | ![]() モデル: Titan Krios / 画像提供: FEI Company |
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顕微鏡 | モデル: FEI TITAN KRIOS |
電子銃 | 電子線源: ![]() |
電子レンズ | モード: BRIGHT FIELD / 倍率(公称値): 105000 X / 最大 デフォーカス(公称値): 2500 nm / 最小 デフォーカス(公称値): 500 nm / Cs: 2.7 mm / C2レンズ絞り径: 50 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: FEI TITAN KRIOS AUTOGRID HOLDER 最高温度: 82 K / 最低温度: 80 K |
撮影 | 電子線照射量: 49 e/Å2 フィルム・検出器のモデル: GATAN K3 BIOQUANTUM (6k x 4k) 撮影したグリッド数: 1 / 実像数: 16983 |
画像スキャン | 横: 11520 / 縦: 8184 |
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解析
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: 1372803 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
対称性 | 点対称性: C1 (非対称) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
3次元再構成 | 解像度: 1.78 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 880876 / アルゴリズム: BACK PROJECTION / 対称性のタイプ: POINT | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
原子モデル構築 | プロトコル: OTHER / 空間: REAL / Target criteria: Cross-correlation coefficient 詳細: Interactive model building was performed in Coot using an auto sharpened map in cryoSPARC and additionally a low-pass-filtered map to 4 Angstrom for the more mobile regions of the 60S ...詳細: Interactive model building was performed in Coot using an auto sharpened map in cryoSPARC and additionally a low-pass-filtered map to 4 Angstrom for the more mobile regions of the 60S structure. Each residue of the model was checked individually for modifications and adjusted manually with a subsequent refinement in phenix.refine. Polyamines and metals were added while searching for unmodelled blobs in Coot. Water molecules were added using phenix.douse and visually validated in Coot. The model was validated using Molprobity. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
原子モデル構築 | PDB-ID: 6EKO Accession code: 6EKO / Source name: PDB / タイプ: experimental model | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
拘束条件 |
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