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データを開く
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基本情報
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タイトル | The open state of the ASFV apo-RNA polymerase | ||||||||||||
![]() | sharpened composite map. The stalk was obtained via Multibody refinement and merged to the main map using Phenix 1.20 composite_map program using the RNAP model as guide. | ||||||||||||
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![]() | RNA polymerase / ASFV / transcription / eukaryotic virus | ||||||||||||
機能・相同性 | ![]() DNA-templated viral transcription / viral transcription / RNA polymerase III activity / tRNA transcription by RNA polymerase III / RNA polymerase I activity / DNA-directed RNA polymerase complex / RNA polymerase II activity / virion component / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity ...DNA-templated viral transcription / viral transcription / RNA polymerase III activity / tRNA transcription by RNA polymerase III / RNA polymerase I activity / DNA-directed RNA polymerase complex / RNA polymerase II activity / virion component / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / host cell cytoplasm / protein dimerization activity / DNA-templated transcription / DNA binding / zinc ion binding / metal ion binding 類似検索 - 分子機能 | ||||||||||||
生物種 | ![]() | ||||||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 2.92 Å | ||||||||||||
![]() | Pilotto S / Sykora M / Cackett G / Werner F | ||||||||||||
資金援助 | ![]()
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![]() | ![]() タイトル: Structure of the recombinant RNA polymerase from African Swine Fever Virus. 著者: Simona Pilotto / Michal Sýkora / Gwenny Cackett / Christopher Dulson / Finn Werner / ![]() 要旨: African Swine Fever Virus is a Nucleo-Cytoplasmic Large DNA Virus that causes an incurable haemorrhagic fever in pigs with a high impact on global food security. ASFV replicates in the cytoplasm of ...African Swine Fever Virus is a Nucleo-Cytoplasmic Large DNA Virus that causes an incurable haemorrhagic fever in pigs with a high impact on global food security. ASFV replicates in the cytoplasm of the infected cell and encodes its own transcription machinery that is independent of cellular factors, however, not much is known about how this system works at a molecular level. Here, we present methods to produce recombinant ASFV RNA polymerase, functional assays to screen for inhibitors, and high-resolution cryo-electron microscopy structures of the ASFV RNAP in different conformational states. The ASFV RNAP bears a striking resemblance to RNAPII with bona fide homologues of nine of its twelve subunits. Key differences include the fusion of the ASFV assembly platform subunits RPB3 and RPB11, and an unusual C-terminal domain of the stalk subunit vRPB7 that is related to the eukaryotic mRNA cap 2´-O-methyltransferase 1. Despite the high degree of structural conservation with cellular RNA polymerases, the ASFV RNAP is resistant to the inhibitors rifampicin and alpha-amanitin. The cryo-EM structures and fully recombinant RNAP system together provide an important tool for the design, development, and screening of antiviral drugs in a low biosafety containment environment. | ||||||||||||
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構造の表示
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-EMDBアーカイブ
マップデータ | ![]() | 94.4 MB | ![]() | |
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ヘッダ (付随情報) | ![]() ![]() | 27.8 KB 27.8 KB | 表示 表示 | ![]() |
FSC (解像度算出) | ![]() | 10.7 KB | 表示 | ![]() |
画像 | ![]() | 112 KB | ||
Filedesc metadata | ![]() | 9.7 KB | ||
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-検証レポート
文書・要旨 | ![]() | 564 KB | 表示 | ![]() |
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文書・詳細版 | ![]() | 563.6 KB | 表示 | |
XML形式データ | ![]() | 12.2 KB | 表示 | |
CIF形式データ | ![]() | 15.8 KB | 表示 | |
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-関連構造データ
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リンク
EMDBのページ | ![]() ![]() |
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「今月の分子」の関連する項目 |
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マップ
ファイル | ![]() | ||||||||||||||||||||
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注釈 | sharpened composite map. The stalk was obtained via Multibody refinement and merged to the main map using Phenix 1.20 composite_map program using the RNAP model as guide. | ||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 0.828 Å | ||||||||||||||||||||
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対称性 | 空間群: 1 | ||||||||||||||||||||
詳細 | EMDB XML:
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-添付データ
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試料の構成要素
+全体 : apo-form of the 8-subunit RNA polymerase from African Swine Fever...
+超分子 #1: apo-form of the 8-subunit RNA polymerase from African Swine Fever...
+分子 #1: DNA-directed RNA polymerase RPB1 homolog
+分子 #2: DNA-directed RNA polymerase RPB2 homolog
+分子 #3: DNA-directed RNA polymerase RPB3-11 homolog
+分子 #4: DNA-directed RNA polymerase RPB7 homolog
+分子 #5: DNA-directed RNA polymerase RPB5 homolog
+分子 #6: DNA-directed RNA polymerase RPB6 homolog
+分子 #7: Uncharacterized protein C122R
+分子 #8: DNA-directed RNA polymerase RPB10 homolog
+分子 #9: ZINC ION
+分子 #10: MAGNESIUM ION
-実験情報
-構造解析
手法 | クライオ電子顕微鏡法 |
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![]() | 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 | 0.05 mg/mL | ||||||||||||||||||
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緩衝液 | pH: 8 構成要素:
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凍結 | 凍結剤: ETHANE / チャンバー内湿度: 95 % / チャンバー内温度: 277 K / 装置: FEI VITROBOT MARK IV | ||||||||||||||||||
詳細 | This sample was monodisparse |
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電子顕微鏡法
顕微鏡 | FEI TITAN KRIOS |
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特殊光学系 | エネルギーフィルター - 名称: GIF Bioquantum / エネルギーフィルター - スリット幅: 20 eV |
撮影 | フィルム・検出器のモデル: GATAN K3 (6k x 4k) / 実像数: 14638 / 平均露光時間: 2.6 sec. / 平均電子線量: 48.152 e/Å2 詳細: Images were collected in movie-mode for a total of 50 frames per image. The data collection was carried out in super-resolution mode and binned 2 on-the-fly. |
電子線 | 加速電圧: 300 kV / 電子線源: ![]() |
電子光学系 | 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / Cs: 2.7 mm / 最大 デフォーカス(公称値): 2.7 µm / 最小 デフォーカス(公称値): 1.5 µm / 倍率(公称値): 105000 |
試料ステージ | 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER ホルダー冷却材: NITROGEN |
実験機器 | ![]() モデル: Titan Krios / 画像提供: FEI Company |
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画像解析
-原子モデル構築 1
初期モデル | PDB ID: Chain - Source name: Other / Chain - Initial model type: experimental model 詳細: The complete biological assembly for the PDB entry 8Q3B was used for the docking in the new map |
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精密化 | 空間: REAL / プロトコル: FLEXIBLE FIT / 温度因子: 84.5995 当てはまり具合の基準: cross-correlation coefficient |
得られたモデル | ![]() PDB-8q3k: |