+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-29689 | |||||||||
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タイトル | Time-resolved cryo-EM study of the 70S recycling by the HflX:3rd Intermediate | |||||||||
マップデータ | i70SHflX-III (Third intermediate) | |||||||||
試料 |
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キーワード | Recycling (リサイクル) / Time-resolved Cryo-EM / 70S / HflX / RIBOSOME (リボソーム) | |||||||||
機能・相同性 | 機能・相同性情報 transcription elongation-coupled chromatin remodeling / ribosome assembly / assembly of large subunit precursor of preribosome / cytosolic ribosome assembly / regulation of DNA-templated transcription elongation / transcription antitermination / DNA-templated transcription termination / mRNA 5'-UTR binding / ribosomal large subunit assembly / small ribosomal subunit rRNA binding ...transcription elongation-coupled chromatin remodeling / ribosome assembly / assembly of large subunit precursor of preribosome / cytosolic ribosome assembly / regulation of DNA-templated transcription elongation / transcription antitermination / DNA-templated transcription termination / mRNA 5'-UTR binding / ribosomal large subunit assembly / small ribosomal subunit rRNA binding / ribosomal small subunit assembly / cytosolic small ribosomal subunit / large ribosomal subunit rRNA binding / large ribosomal subunit / small ribosomal subunit / cytoplasmic translation / 5S rRNA binding / peptidase activity / cytosolic large ribosomal subunit / transferase activity / tRNA binding / negative regulation of translation / rRNA binding / リボソーム / structural constituent of ribosome / 翻訳 (生物学) / ribonucleoprotein complex / response to antibiotic / mRNA binding / GTPase activity / GTP binding / タンパク質分解 / 生体膜 / metal ion binding / 細胞質基質 / 細胞質 類似検索 - 分子機能 | |||||||||
生物種 | Escherichia coli (大腸菌) | |||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 3.2 Å | |||||||||
データ登録者 | Bhattacharjee S / Brown PZ / Frank J | |||||||||
資金援助 | 米国, 1件
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引用 | ジャーナル: Cell / 年: 2024 タイトル: Time resolution in cryo-EM using a PDMS-based microfluidic chip assembly and its application to the study of HflX-mediated ribosome recycling. 著者: Sayan Bhattacharjee / Xiangsong Feng / Suvrajit Maji / Prikshat Dadhwal / Zhening Zhang / Zuben P Brown / Joachim Frank / 要旨: The rapid kinetics of biological processes and associated short-lived conformational changes pose a significant challenge in attempts to structurally visualize biomolecules during a reaction in real ...The rapid kinetics of biological processes and associated short-lived conformational changes pose a significant challenge in attempts to structurally visualize biomolecules during a reaction in real time. Conventionally, on-pathway intermediates have been trapped using chemical modifications or reduced temperature, giving limited insights. Here, we introduce a time-resolved cryo-EM method using a reusable PDMS-based microfluidic chip assembly with high reactant mixing efficiency. Coating of PDMS walls with SiO virtually eliminates non-specific sample adsorption and ensures maintenance of the stoichiometry of the reaction, rendering it highly reproducible. In an operating range from 10 to 1,000 ms, the device allows us to follow in vitro reactions of biological molecules at resolution levels in the range of 3 Å. By employing this method, we show the mechanism of progressive HflX-mediated splitting of the 70S E. coli ribosome in the presence of the GTP via capture of three high-resolution reaction intermediates within 140 ms. | |||||||||
履歴 |
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-構造の表示
添付画像 |
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-ダウンロードとリンク
-EMDBアーカイブ
マップデータ | emd_29689.map.gz | 133.1 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-29689-v30.xml emd-29689.xml | 63.5 KB 63.5 KB | 表示 表示 | EMDBヘッダ |
画像 | emd_29689.png | 56.9 KB | ||
Filedesc metadata | emd-29689.cif.gz | 13.2 KB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-29689 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-29689 | HTTPS FTP |
-関連構造データ
-リンク
EMDBのページ | EMDB (EBI/PDBe) / EMDataResource |
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「今月の分子」の関連する項目 |
-マップ
ファイル | ダウンロード / ファイル: emd_29689.map.gz / 形式: CCP4 / 大きさ: 149.9 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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注釈 | i70SHflX-III (Third intermediate) | ||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 1.0253 Å | ||||||||||||||||||||
密度 |
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対称性 | 空間群: 1 | ||||||||||||||||||||
詳細 | EMDB XML:
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-添付データ
-試料の構成要素
+全体 : i70SHflX-III (3rd intermediate)
+超分子 #1: i70SHflX-III (3rd intermediate)
+分子 #1: 50S ribosomal protein L32
+分子 #2: 50S ribosomal protein L33
+分子 #3: 50S ribosomal protein L34
+分子 #4: 50S ribosomal protein L35
+分子 #5: 50S ribosomal protein L36
+分子 #8: 50S ribosomal protein L2
+分子 #9: 50S ribosomal protein L3
+分子 #10: 50S ribosomal protein L4
+分子 #11: 50S ribosomal protein L5
+分子 #12: 50S ribosomal protein L6
+分子 #13: 50S ribosomal protein L13
+分子 #14: 50S ribosomal protein L14
+分子 #15: 50S ribosomal protein L15
+分子 #16: 50S ribosomal protein L16
+分子 #17: 50S ribosomal protein L17
+分子 #18: 50S ribosomal protein L18
+分子 #19: 50S ribosomal protein L19
+分子 #20: 50S ribosomal protein L20
+分子 #21: Ribosomal protein L21
+分子 #22: 50S ribosomal protein L22
+分子 #23: 50S ribosomal protein L23
+分子 #24: 50S ribosomal protein L24
+分子 #25: 50S ribosomal protein L25
+分子 #26: 50S ribosomal protein L27
+分子 #27: 50S ribosomal protein L28
+分子 #28: 50S ribosomal protein L29
+分子 #29: 50S ribosomal protein L30
+分子 #30: 30S ribosomal protein S2
+分子 #31: 30S ribosomal protein S3
+分子 #32: 30S ribosomal protein S4
+分子 #33: 30S ribosomal protein S5
+分子 #34: 30S ribosomal protein S6, non-modified isoform
+分子 #35: 30S ribosomal protein S7
+分子 #36: 30S ribosomal protein S8
+分子 #37: 30S ribosomal protein S9
+分子 #38: 30S ribosomal protein S10
+分子 #39: 30S ribosomal protein S11
+分子 #40: 30S ribosomal protein S12
+分子 #41: 30S ribosomal protein S13
+分子 #42: 30S ribosomal protein S14
+分子 #43: 30S ribosomal protein S15
+分子 #44: 30S ribosomal protein S16
+分子 #45: 30S ribosomal protein S17
+分子 #46: 30S ribosomal protein S18
+分子 #47: 30S ribosomal protein S19
+分子 #48: 30S ribosomal protein S20
+分子 #49: 30S ribosomal protein S21 (Fragment)
+分子 #50: Transcription termination/antitermination protein NusG
+分子 #52: GTPase HflX
+分子 #6: 5S
+分子 #7: 23S
+分子 #51: 16S
+分子 #53: GUANOSINE-5'-TRIPHOSPHATE
+分子 #54: water
-実験情報
-構造解析
手法 | クライオ電子顕微鏡法 |
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解析 | 単粒子再構成法 |
試料の集合状態 | cell |
-試料調製
緩衝液 | pH: 7.5 |
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凍結 | 凍結剤: ETHANE |
-電子顕微鏡法
顕微鏡 | FEI TITAN KRIOS |
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電子線 | 加速電圧: 300 kV / 電子線源: FIELD EMISSION GUN |
電子光学系 | 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELDBright-field microscopy / 最大 デフォーカス(公称値): 2.5 µm / 最小 デフォーカス(公称値): 1.0 µm |
撮影 | フィルム・検出器のモデル: GATAN K3 (6k x 4k) / 平均電子線量: 58.0 e/Å2 |
実験機器 | モデル: Titan Krios / 画像提供: FEI Company |
-画像解析
初期モデル | モデルのタイプ: NONE |
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初期 角度割当 | タイプ: ANGULAR RECONSTITUTION |
最終 角度割当 | タイプ: MAXIMUM LIKELIHOOD |
最終 再構成 | 解像度のタイプ: BY AUTHOR / 解像度: 3.2 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 使用した粒子像数: 113038 |