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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: 7bv8 | |||||||||||||||
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タイトル | Mature 50S ribosomal subunit from RrmJ knock out E.coli strain | |||||||||||||||
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![]() | RIBOSOME / ribosome assembly / ribosome biogenesis | |||||||||||||||
機能・相同性 | ![]() stringent response / transcriptional attenuation / endoribonuclease inhibitor activity / RNA-binding transcription regulator activity / positive regulation of ribosome biogenesis / negative regulation of cytoplasmic translation / translational termination / DnaA-L2 complex / translation repressor activity / negative regulation of DNA-templated DNA replication initiation ...stringent response / transcriptional attenuation / endoribonuclease inhibitor activity / RNA-binding transcription regulator activity / positive regulation of ribosome biogenesis / negative regulation of cytoplasmic translation / translational termination / DnaA-L2 complex / translation repressor activity / negative regulation of DNA-templated DNA replication initiation / ribosome assembly / mRNA regulatory element binding translation repressor activity / assembly of large subunit precursor of preribosome / cytosolic ribosome assembly / response to reactive oxygen species / regulation of cell growth / DNA-templated transcription termination / response to radiation / ribosomal large subunit assembly / mRNA 5'-UTR binding / large ribosomal subunit / ribosome binding / transferase activity / 5S rRNA binding / large ribosomal subunit rRNA binding / cytosolic large ribosomal subunit / tRNA binding / cytoplasmic translation / rRNA binding / negative regulation of translation / ribosome / structural constituent of ribosome / translation / response to antibiotic / negative regulation of DNA-templated transcription / mRNA binding / DNA binding / RNA binding / zinc ion binding / cytoplasm / cytosol 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: 3.14 Å | |||||||||||||||
![]() | Wang, W. / Li, W.Q. / Ge, X.L. / Yan, K.G. / Mandava, C.S. / Sanyal, S. / Gao, N. | |||||||||||||||
資金援助 | ![]()
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![]() | ![]() タイトル: Loss of a single methylation in 23S rRNA delays 50S assembly at multiple late stages and impairs translation initiation and elongation. 著者: Wei Wang / Wanqiu Li / Xueliang Ge / Kaige Yan / Chandra Sekhar Mandava / Suparna Sanyal / Ning Gao / ![]() ![]() ![]() 要旨: Ribosome biogenesis is a complex process, and dozens of factors are required to facilitate and regulate the subunit assembly in bacteria. The 2'-O-methylation of U2552 in 23S rRNA by ...Ribosome biogenesis is a complex process, and dozens of factors are required to facilitate and regulate the subunit assembly in bacteria. The 2'-O-methylation of U2552 in 23S rRNA by methyltransferase RrmJ is a crucial step in late-stage assembly of the 50S subunit. Its absence results in severe growth defect and marked accumulation of pre50S assembly intermediates. In the present work, we employed cryoelectron microscopy to characterize a set of late-stage pre50S particles isolated from an Δ strain. These assembly intermediates (solved at 3.2 to 3.8 Å resolution) define a collection of late-stage particles on a progressive assembly pathway. Apart from the absence of L16, L35, and L36, major structural differences between these intermediates and the mature 50S subunit are clustered near the peptidyl transferase center, such as H38, H68-71, and H89-93. In addition, the ribosomal A-loop of the mature 50S subunit from Δ strain displays large local flexibility on nucleotides next to unmethylated U2552. Fast kinetics-based biochemical assays demonstrate that the Δ 50S subunit is only 50% active and two times slower than the WT 50S subunit in rapid subunit association. While the Δ 70S ribosomes show no defect in peptide bond formation, peptide release, and ribosome recycling, they translocate with 20% slower rate than the WT ribosomes in each round of elongation. These defects amplify during synthesis of the full-length proteins and cause overall defect in protein synthesis. In conclusion, our data reveal the molecular roles of U2552 methylation in both ribosome biogenesis and protein translation. | |||||||||||||||
履歴 |
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構造の表示
ムービー |
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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 | ![]() | 2 MB | 表示 | ![]() |
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PDB形式 | ![]() | 1.5 MB | 表示 | ![]() |
PDBx/mmJSON形式 | ![]() | ツリー表示 | ![]() | |
その他 | ![]() |
-検証レポート
文書・要旨 | ![]() | 1.2 MB | 表示 | ![]() |
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文書・詳細版 | ![]() | 1.2 MB | 表示 | |
XML形式データ | ![]() | 120.1 KB | 表示 | |
CIF形式データ | ![]() | 215.9 KB | 表示 | |
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-関連構造データ
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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
-RNA鎖 , 2種, 2分子 AB
#1: RNA鎖 | 分子量: 941305.188 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
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#2: RNA鎖 | 分子量: 38790.090 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
+50S ribosomal protein ... , 30種, 30分子 CDEFGHIJKLMNOPQRSTUVWXYZabcdef
-詳細
由来についての詳細 | The E.coli. stain is BW25113, which is a derivative of E.coli. K12 strain. |
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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 |
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試料調製
構成要素 | 名称: Mature 50S ribosomal subunit from RrmJ knock out E.coli strain タイプ: RIBOSOME 詳細: Mature 50S ribosomal subunit from RrmJ knock out E.coli strain Entity ID: all / 由来: NATURAL |
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分子量 | 値: 1.5 MDa / 実験値: NO |
由来(天然) | 生物種: ![]() ![]() |
緩衝液 | pH: 7.5 詳細: Solutions were made fresh form concentrated to avoid microbial contamination. |
試料 | 濃度: 0.15 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES / 詳細: This sample was monodisperse. |
試料支持 | 詳細: The grid was coated with continuous carbon film. / グリッドの材料: COPPER / グリッドのサイズ: 300 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: 100 % / 凍結前の試料温度: 277 K / 詳細: blot for 2 seconds before plunging |
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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 / 詳細: Preliminary grid screening was performed manually. |
電子銃 | 電子線源: ![]() |
電子レンズ | モード: BRIGHT FIELD / 倍率(公称値): 75000 X / 倍率(補正後): 75000 X / 最大 デフォーカス(公称値): -2000 nm / 最小 デフォーカス(公称値): -1000 nm / Cs: 2.7 mm / 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 |
撮影 | 電子線照射量: 45 e/Å2 フィルム・検出器のモデル: FEI FALCON II (4k x 4k) 撮影したグリッド数: 1 / 実像数: 1164 |
画像スキャン | 横: 4096 / 縦: 4096 / 動画フレーム数/画像: 28 / 利用したフレーム数/画像: 3-28 |
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解析
ソフトウェア | 名称: PHENIX / バージョン: 1.16_3549: / 分類: 精密化 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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EMソフトウェア |
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画像処理 | 詳細: The images were normalized and manually selected. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
CTF補正 | 詳細: CTF amplitude correction was performed following 3D reconstruction. タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
粒子像の選択 | 選択した粒子像数: 233762 詳細: Images were binned and lowpass filtered for autopicking. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
対称性 | 点対称性: C1 (非対称) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
3次元再構成 | 解像度: 3.14 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 98194 / アルゴリズム: SIMULTANEOUS ITERATIVE (SIRT) 詳細: A soft mask was added to improve resolution at the final refinement. クラス平均像の数: 1 / 対称性のタイプ: POINT | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
原子モデル構築 |
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原子モデル構築 |
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精密化 | 交差検証法: NONE 立体化学のターゲット値: GeoStd + Monomer Library + CDL v1.2 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
原子変位パラメータ | Biso mean: 51.84 Å2 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
拘束条件 |
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