ジャーナル: Nat Commun / 年: 2021 タイトル: Self-assembly and regulation of protein cages from pre-organised coiled-coil modules. 著者: Fabio Lapenta / Jana Aupič / Marco Vezzoli / Žiga Strmšek / Stefano Da Vela / Dmitri I Svergun / José María Carazo / Roberto Melero / Roman Jerala / 要旨: Coiled-coil protein origami (CCPO) is a modular strategy for the de novo design of polypeptide nanostructures. CCPO folds are defined by the sequential order of concatenated orthogonal coiled-coil ...Coiled-coil protein origami (CCPO) is a modular strategy for the de novo design of polypeptide nanostructures. CCPO folds are defined by the sequential order of concatenated orthogonal coiled-coil (CC) dimer-forming peptides, where a single-chain protein is programmed to fold into a polyhedral cage. Self-assembly of CC-based nanostructures from several chains, similarly as in DNA nanotechnology, could facilitate the design of more complex assemblies and the introduction of functionalities. Here, we show the design of a de novo triangular bipyramid fold comprising 18 CC-forming segments and define the strategy for the two-chain self-assembly of the bipyramidal cage from asymmetric and pseudo-symmetric pre-organised structural modules. In addition, by introducing a protease cleavage site and masking the interfacial CC-forming segments in the two-chain bipyramidal cage, we devise a proteolysis-mediated conformational switch. This strategy could be extended to other modular protein folds, facilitating the construction of dynamic multi-chain CC-based complexes.
Samples were applied to glow discharged carbon-coated copper grids, washed quickly with distilled water and negatively stained with 2% (w/v) uranyl acetate.
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電子顕微鏡法
顕微鏡
JEOL 1230
撮影
フィルム・検出器のモデル: TVIPS TEMCAM-F415 (4k x 4k) 平均電子線量: 20.0 e/Å2
電子線
加速電圧: 120 kV / 電子線源: TUNGSTEN HAIRPIN
電子光学系
倍率(補正後): 54926 / 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD
試料ステージ
試料ホルダーモデル: JEOL
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画像解析
初期モデル
モデルのタイプ: OTHER
最終 再構成
解像度のタイプ: BY AUTHOR / 解像度: 25.0 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 使用した粒子像数: 20000