+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-0261 | |||||||||||||||
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タイトル | E. coli 70S d2d8 stapled ribosome | |||||||||||||||
マップデータ | map | |||||||||||||||
試料 |
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キーワード | staple / RIBOSOME | |||||||||||||||
機能・相同性 | 機能・相同性情報 negative regulation of cytoplasmic translational initiation / stringent response / ornithine decarboxylase inhibitor activity / transcription antitermination factor activity, RNA binding / misfolded RNA binding / Group I intron splicing / RNA folding / transcriptional attenuation / endoribonuclease inhibitor activity / RNA-binding transcription regulator activity ...negative regulation of cytoplasmic translational initiation / stringent response / ornithine decarboxylase inhibitor activity / transcription antitermination factor activity, RNA binding / misfolded RNA binding / Group I intron splicing / RNA folding / transcriptional attenuation / endoribonuclease inhibitor activity / RNA-binding transcription regulator activity / positive regulation of ribosome biogenesis / negative regulation of cytoplasmic translation / four-way junction DNA binding / translational termination / DnaA-L2 complex / translation repressor activity / negative regulation of translational initiation / negative regulation of DNA-templated DNA replication initiation / regulation of mRNA stability / mRNA regulatory element binding translation repressor activity / ribosome assembly / positive regulation of RNA splicing / assembly of large subunit precursor of preribosome / transcription elongation factor complex / cytosolic ribosome assembly / regulation of DNA-templated transcription elongation / DNA endonuclease activity / ribosomal large subunit assembly / transcription antitermination / response to reactive oxygen species / translational initiation / regulation of cell growth / DNA-templated transcription termination / maintenance of translational fidelity / response to radiation / mRNA 5'-UTR binding / large ribosomal subunit / ribosome biogenesis / ribosome binding / regulation of translation / ribosomal small subunit biogenesis / ribosomal small subunit assembly / small ribosomal subunit / small ribosomal subunit rRNA binding / transferase activity / 5S rRNA binding / large ribosomal subunit rRNA binding / cytosolic small ribosomal subunit / cytosolic large ribosomal subunit / cytoplasmic translation / tRNA binding / molecular adaptor activity / 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 / membrane / cytoplasm / cytosol 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: 2.96 Å | |||||||||||||||
データ登録者 | Schmied WH / Tnimov Z | |||||||||||||||
資金援助 | 英国, ベルギー, 4件
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引用 | ジャーナル: Nature / 年: 2018 タイトル: Controlling orthogonal ribosome subunit interactions enables evolution of new function. 著者: Wolfgang H Schmied / Zakir Tnimov / Chayasith Uttamapinant / Christopher D Rae / Stephen D Fried / Jason W Chin / 要旨: Orthogonal ribosomes are unnatural ribosomes that are directed towards orthogonal messenger RNAs in Escherichia coli, through an altered version of the 16S ribosomal RNA of the small subunit. ...Orthogonal ribosomes are unnatural ribosomes that are directed towards orthogonal messenger RNAs in Escherichia coli, through an altered version of the 16S ribosomal RNA of the small subunit. Directed evolution of orthogonal ribosomes has provided access to new ribosomal function, and the evolved orthogonal ribosomes have enabled the encoding of multiple non-canonical amino acids into proteins. The original orthogonal ribosomes shared the pool of 23S ribosomal RNAs, contained in the large subunit, with endogenous ribosomes. Selectively directing a new 23S rRNA to an orthogonal mRNA, by controlling the association between the orthogonal 16S rRNAs and 23S rRNAs, would enable the evolution of new function in the large subunit. Previous work covalently linked orthogonal 16S rRNA and a circularly permuted 23S rRNA to create orthogonal ribosomes with low activity; however, the linked subunits in these ribosomes do not associate specifically with each other, and mediate translation by associating with endogenous subunits. Here we discover engineered orthogonal 'stapled' ribosomes (with subunits linked through an optimized RNA staple) with activities comparable to that of the parent orthogonal ribosome; they minimize association with endogenous subunits and mediate translation of orthogonal mRNAs through the association of stapled subunits. We evolve cells with genomically encoded stapled ribosomes as the sole ribosomes, which support cellular growth at similar rates to natural ribosomes. Moreover, we visualize the engineered stapled ribosome structure by cryo-electron microscopy at 3.0 Å, revealing how the staple links the subunits and controls their association. We demonstrate the utility of controlling subunit association by evolving orthogonal stapled ribosomes which efficiently polymerize a sequence of monomers that the natural ribosome is intrinsically unable to translate. Our work provides a foundation for evolving the rRNA of the entire orthogonal ribosome for the encoded cellular synthesis of non-canonical biological polymers. | |||||||||||||||
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 | EMマップ: SurfViewMolmilJmol/JSmol |
添付画像 |
-ダウンロードとリンク
-EMDBアーカイブ
マップデータ | emd_0261.map.gz | 32.2 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-0261-v30.xml emd-0261.xml | 65.6 KB 65.6 KB | 表示 表示 | EMDBヘッダ |
FSC (解像度算出) | emd_0261_fsc.xml | 13.1 KB | 表示 | FSCデータファイル |
画像 | emd_0261.png | 82.8 KB | ||
Filedesc metadata | emd-0261.cif.gz | 13.8 KB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-0261 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-0261 | HTTPS FTP |
-検証レポート
文書・要旨 | emd_0261_validation.pdf.gz | 516 KB | 表示 | EMDB検証レポート |
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文書・詳細版 | emd_0261_full_validation.pdf.gz | 515.6 KB | 表示 | |
XML形式データ | emd_0261_validation.xml.gz | 13.5 KB | 表示 | |
CIF形式データ | emd_0261_validation.cif.gz | 18.2 KB | 表示 | |
アーカイブディレクトリ | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-0261 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-0261 | 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_0261.map.gz / 形式: CCP4 / 大きさ: 193.2 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | map | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 1.06 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
密度 |
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対称性 | 空間群: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
詳細 | EMDB XML:
CCP4マップ ヘッダ情報:
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-添付データ
-試料の構成要素
+全体 : E. coli 70S d2d8 stapled ribosome
+超分子 #1: E. coli 70S d2d8 stapled ribosome
+分子 #1: stapled 16S-23S rRNA,stapled 16S-23S rRNA,stapled 16S-23S rRNA,st...
+分子 #2: 5S ribosomal RNA
+分子 #3: 50S ribosomal protein L2
+分子 #4: 50S ribosomal protein L3
+分子 #5: 50S ribosomal protein L4
+分子 #6: 50S ribosomal protein L5
+分子 #7: 50S ribosomal protein L6
+分子 #8: 50S ribosomal protein L9
+分子 #9: 50S ribosomal protein L10
+分子 #10: 50S ribosomal protein L11
+分子 #11: 50S ribosomal protein L13
+分子 #12: 50S ribosomal protein L14
+分子 #13: 50S ribosomal protein L15
+分子 #14: 50S ribosomal protein L16
+分子 #15: 50S ribosomal protein L17
+分子 #16: 50S ribosomal protein L18
+分子 #17: 50S ribosomal protein L19
+分子 #18: 50S ribosomal protein L20
+分子 #19: 50S ribosomal protein L21
+分子 #20: 50S ribosomal protein L22
+分子 #21: 50S ribosomal protein L23
+分子 #22: 50S ribosomal protein L24
+分子 #23: 50S ribosomal protein L25
+分子 #24: 50S ribosomal protein L27
+分子 #25: 50S ribosomal protein L28
+分子 #26: 50S ribosomal protein L29
+分子 #27: 50S ribosomal protein L30
+分子 #28: 50S ribosomal protein L31
+分子 #29: 50S ribosomal protein L32
+分子 #30: 50S ribosomal protein L33
+分子 #31: 50S ribosomal protein L34
+分子 #32: 50S ribosomal protein L35
+分子 #33: 50S ribosomal protein L36
+分子 #34: 30S ribosomal protein S2
+分子 #35: 30S ribosomal protein S3
+分子 #36: 30S ribosomal protein S4
+分子 #37: 30S ribosomal protein S5
+分子 #38: 30S ribosomal protein S6
+分子 #39: 30S ribosomal protein S7
+分子 #40: 30S ribosomal protein S8
+分子 #41: 30S ribosomal protein S9
+分子 #42: 30S ribosomal protein S10
+分子 #43: 30S ribosomal protein S11
+分子 #44: 30S ribosomal protein S12
+分子 #45: 30S ribosomal protein S13
+分子 #46: 30S ribosomal protein S14
+分子 #47: 30S ribosomal protein S15
+分子 #48: 30S ribosomal protein S16
+分子 #49: 30S ribosomal protein S17
+分子 #50: 30S ribosomal protein S18
+分子 #51: 30S ribosomal protein S19
+分子 #52: 30S ribosomal protein S20
+分子 #53: 30S ribosomal protein S21
+分子 #54: MAGNESIUM ION
+分子 #55: ZINC ION
+分子 #56: 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 | particle |
-試料調製
緩衝液 | pH: 7.4 |
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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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撮影 | フィルム・検出器のモデル: FEI FALCON III (4k x 4k) 平均露光時間: 1.8 sec. / 平均電子線量: 27.0 e/Å2 |
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 FIELD |
実験機器 | モデル: Titan Krios / 画像提供: FEI Company |