ジャーナル: Nat Biotechnol / 年: 2017 タイトル: Design of coiled-coil protein-origami cages that self-assemble in vitro and in vivo. 著者: Ajasja Ljubetič / Fabio Lapenta / Helena Gradišar / Igor Drobnak / Jana Aupič / Žiga Strmšek / Duško Lainšček / Iva Hafner-Bratkovič / Andreja Majerle / Nuša Krivec / Mojca Benčina ...著者: Ajasja Ljubetič / Fabio Lapenta / Helena Gradišar / Igor Drobnak / Jana Aupič / Žiga Strmšek / Duško Lainšček / Iva Hafner-Bratkovič / Andreja Majerle / Nuša Krivec / Mojca Benčina / Tomaž Pisanski / Tanja Ćirković Veličković / Adam Round / José María Carazo / Roberto Melero / Roman Jerala / 要旨: Polypeptides and polynucleotides are natural programmable biopolymers that can self-assemble into complex tertiary structures. We describe a system analogous to designed DNA nanostructures in which ...Polypeptides and polynucleotides are natural programmable biopolymers that can self-assemble into complex tertiary structures. We describe a system analogous to designed DNA nanostructures in which protein coiled-coil (CC) dimers serve as building blocks for modular de novo design of polyhedral protein cages that efficiently self-assemble in vitro and in vivo. We produced and characterized >20 single-chain protein cages in three shapes-tetrahedron, four-sided pyramid, and triangular prism-with the largest containing >700 amino-acid residues and measuring 11 nm in diameter. Their stability and folding kinetics were similar to those of natural proteins. Solution small-angle X-ray scattering (SAXS), electron microscopy (EM), and biophysical analysis confirmed agreement of the expressed structures with the designs. We also demonstrated self-assembly of a tetrahedral structure in bacteria, mammalian cells, and mice without evidence of inflammation. A semi-automated computational design platform and a toolbox of CC building modules are provided to enable the design of protein cages in any polyhedral shape.
名称: TET12SN / タイプ: complex / ID: 1 / 親要素: 0 詳細: Coiled coil single chain protein origami tetrahedron composed of soluble dimeric coiled coil segments.
由来(天然)
生物種: Designed protein (人工物)
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実験情報
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構造解析
手法
ネガティブ染色法
解析
単粒子再構成法
試料の集合状態
particle
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試料調製
濃度
0.03 mg/mL
緩衝液
pH: 7.5 構成要素:
濃度
名称
式
20.0 mM
Tris
150.0 mM
Sodium Cloride
NaCl
10.0 %
Glycerol
染色
タイプ: NEGATIVE / 材質: Uranyl Acetate 詳細: 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-F416 (4k x 4k) 平均電子線量: 15.0 e/Å2
電子線
加速電圧: 100 kV / 電子線源: TUNGSTEN HAIRPIN
電子光学系
照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / Cs: 2.7 mm
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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: 28.0 Å / 解像度の算出法: FSC 0.143 CUT-OFF / ソフトウェア: (名称: Xmipp, RELION) / 使用した粒子像数: 13425