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.
ダウンロード / ファイル: emd_3781.map.gz / 形式: CCP4 / 大きさ: 3.8 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
注釈
Bioorigami coiled-coil tetrahedron TET12SN.
ボクセルのサイズ
X=Y=Z: 2.84 Å
密度
表面レベル
登録者による: 0.0559 / ムービー #1: 0.0559
最小 - 最大
-0.047377944 - 0.25644943
平均 (標準偏差)
0.0002109636 (±0.013649056)
対称性
空間群: 1
詳細
EMDB XML:
マップ形状
Axis order
X
Y
Z
Origin
0
0
0
サイズ
100
100
100
Spacing
100
100
100
セル
A=B=C: 284.0 Å α=β=γ: 90.0 °
CCP4マップ ヘッダ情報:
mode
Image stored as Reals
Å/pix. X/Y/Z
2.84
2.84
2.84
M x/y/z
100
100
100
origin x/y/z
0.000
0.000
0.000
length x/y/z
284.000
284.000
284.000
α/β/γ
90.000
90.000
90.000
start NX/NY/NZ
0
0
0
NX/NY/NZ
280
280
280
MAP C/R/S
1
2
3
start NC/NR/NS
0
0
0
NC/NR/NS
100
100
100
D min/max/mean
-0.047
0.256
0.000
-
添付データ
-
試料の構成要素
-
全体 : TET12SN
全体
名称: TET12SN
要素
複合体: TET12SN
-
超分子 #1: TET12SN
超分子
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)
-
実験情報
-
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
-
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.
-
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
-
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