ジャーナル: Nat Commun / 年: 2021 タイトル: Structural basis of RNA polymerase inhibition by viral and host factors. 著者: Simona Pilotto / Thomas Fouqueau / Natalya Lukoyanova / Carol Sheppard / Soizick Lucas-Staat / Luis Miguel Díaz-Santín / Dorota Matelska / David Prangishvili / Alan C M Cheung / Finn Werner / 要旨: RNA polymerase inhibition plays an important role in the regulation of transcription in response to environmental changes and in the virus-host relationship. Here we present the high-resolution ...RNA polymerase inhibition plays an important role in the regulation of transcription in response to environmental changes and in the virus-host relationship. Here we present the high-resolution structures of two such RNAP-inhibitor complexes that provide the structural bases underlying RNAP inhibition in archaea. The Acidianus two-tailed virus encodes the RIP factor that binds inside the DNA-binding channel of RNAP, inhibiting transcription by occlusion of binding sites for nucleic acid and the transcription initiation factor TFB. Infection with the Sulfolobus Turreted Icosahedral Virus induces the expression of the host factor TFS4, which binds in the RNAP funnel similarly to eukaryotic transcript cleavage factors. However, TFS4 allosterically induces a widening of the DNA-binding channel which disrupts trigger loop and bridge helix motifs. Importantly, the conformational changes induced by TFS4 are closely related to inactivated states of RNAP in other domains of life indicating a deep evolutionary conservation of allosteric RNAP inhibition.
Sharpened map from postprocess, relion v3.0. The map was obtained from Multibody refinement and it corresponds to the smaller body with higher flexibility
モデル: UltrAuFoil R1.2/1.3 / 材質: GOLD / メッシュ: 300 / 支持フィルム - 材質: GOLD / 支持フィルム - トポロジー: HOLEY / 前処理 - タイプ: GLOW DISCHARGE / 前処理 - 時間: 60 sec. / 前処理 - 雰囲気: AIR 詳細: the grid was coated with graphene oxide after to be glow discharged and before applying the sample
凍結
凍結剤: ETHANE / チャンバー内湿度: 94 % / チャンバー内温度: 277 K / 装置: FEI VITROBOT MARK IV
詳細
the sample was applied twice on the grid before vetrification
フィルム・検出器のモデル: GATAN K3 (6k x 4k) / 撮影したグリッド数: 1 / 実像数: 1760 / 平均露光時間: 2.0 sec. / 平均電子線量: 45.2 e/Å2 詳細: images were collected as movie-stacks of 40 frames in super resolution mode
タイプ: MAXIMUM LIKELIHOOD / ソフトウェア - 名称: RELION (ver. 3.0) / ソフトウェア - 詳細: 3D classification
最終 3次元分類
クラス数: 2 / 平均メンバー数/クラス: 170000 / ソフトウェア - 名称: RELION (ver. 3.0) / ソフトウェア - 詳細: 3D classification 詳細: All particles clustered in only one class which showed some preferred orientations later removed with a home-made script