+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-10382 | |||||||||||||||||||||||||||||||||
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タイトル | A 3.9 Angstrom structure of the EIAV CA-SP hexamer (C6) from Gag-dMA spheres assembled at pH8 | |||||||||||||||||||||||||||||||||
マップデータ | A 3.9 Angstrom structure of the EIAV CA-SP hexamer (C6) from Gag-dMA spheres assembled at pH8 | |||||||||||||||||||||||||||||||||
試料 |
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生物種 | Equine infectious anemia virus (ウマ伝染性貧血ウイルス) | |||||||||||||||||||||||||||||||||
手法 | サブトモグラム平均法 / クライオ電子顕微鏡法 / 解像度: 3.9 Å | |||||||||||||||||||||||||||||||||
データ登録者 | Dick RA / Xu C / Morado DR / Kravchuk V / Ricana CL / Lyddon TD / Broad AM / Feathers JR / Johnson MC / Vogt VM ...Dick RA / Xu C / Morado DR / Kravchuk V / Ricana CL / Lyddon TD / Broad AM / Feathers JR / Johnson MC / Vogt VM / Perilla JR / Briggs JAG / Schur FKM | |||||||||||||||||||||||||||||||||
資金援助 | ドイツ, オーストリア, 米国, 英国, 10件
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引用 | ジャーナル: PLoS Pathog / 年: 2020 タイトル: Structures of immature EIAV Gag lattices reveal a conserved role for IP6 in lentivirus assembly. 著者: Robert A Dick / Chaoyi Xu / Dustin R Morado / Vladyslav Kravchuk / Clifton L Ricana / Terri D Lyddon / Arianna M Broad / J Ryan Feathers / Marc C Johnson / Volker M Vogt / Juan R Perilla / ...著者: Robert A Dick / Chaoyi Xu / Dustin R Morado / Vladyslav Kravchuk / Clifton L Ricana / Terri D Lyddon / Arianna M Broad / J Ryan Feathers / Marc C Johnson / Volker M Vogt / Juan R Perilla / John A G Briggs / Florian K M Schur / 要旨: Retrovirus assembly is driven by the multidomain structural protein Gag. Interactions between the capsid domains (CA) of Gag result in Gag multimerization, leading to an immature virus particle that ...Retrovirus assembly is driven by the multidomain structural protein Gag. Interactions between the capsid domains (CA) of Gag result in Gag multimerization, leading to an immature virus particle that is formed by a protein lattice based on dimeric, trimeric, and hexameric protein contacts. Among retroviruses the inter- and intra-hexamer contacts differ, especially in the N-terminal sub-domain of CA (CANTD). For HIV-1 the cellular molecule inositol hexakisphosphate (IP6) interacts with and stabilizes the immature hexamer, and is required for production of infectious virus particles. We have used in vitro assembly, cryo-electron tomography and subtomogram averaging, atomistic molecular dynamics simulations and mutational analyses to study the HIV-related lentivirus equine infectious anemia virus (EIAV). In particular, we sought to understand the structural conservation of the immature lentivirus lattice and the role of IP6 in EIAV assembly. Similar to HIV-1, IP6 strongly promoted in vitro assembly of EIAV Gag proteins into virus-like particles (VLPs), which took three morphologically highly distinct forms: narrow tubes, wide tubes, and spheres. Structural characterization of these VLPs to sub-4Å resolution unexpectedly showed that all three morphologies are based on an immature lattice with preserved key structural components, highlighting the structural versatility of CA to form immature assemblies. A direct comparison between EIAV and HIV revealed that both lentiviruses maintain similar immature interfaces, which are established by both conserved and non-conserved residues. In both EIAV and HIV-1, IP6 regulates immature assembly via conserved lysine residues within the CACTD and SP. Lastly, we demonstrate that IP6 stimulates in vitro assembly of immature particles of several other retroviruses in the lentivirus genus, suggesting a conserved role for IP6 in lentiviral assembly. | |||||||||||||||||||||||||||||||||
履歴 |
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-構造の表示
ムービー |
ムービービューア |
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構造ビューア | EMマップ: SurfViewMolmilJmol/JSmol |
添付画像 |
-ダウンロードとリンク
-EMDBアーカイブ
マップデータ | emd_10382.map.gz | 25.3 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-10382-v30.xml emd-10382.xml | 19.3 KB 19.3 KB | 表示 表示 | EMDBヘッダ |
画像 | emd_10382.png | 238.8 KB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-10382 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-10382 | HTTPS FTP |
-関連構造データ
-リンク
EMDBのページ | EMDB (EBI/PDBe) / EMDataResource |
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-マップ
ファイル | ダウンロード / ファイル: emd_10382.map.gz / 形式: CCP4 / 大きさ: 27 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | A 3.9 Angstrom structure of the EIAV CA-SP hexamer (C6) from Gag-dMA spheres assembled at pH8 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 1.35 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
密度 |
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対称性 | 空間群: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
詳細 | EMDB XML:
CCP4マップ ヘッダ情報:
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-添付データ
-試料の構成要素
-全体 : Equine infectious anemia virus
全体 | 名称: Equine infectious anemia virus (ウマ伝染性貧血ウイルス) |
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要素 |
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-超分子 #1: Equine infectious anemia virus
超分子 | 名称: Equine infectious anemia virus / タイプ: virus / ID: 1 / 親要素: 0 / 含まれる分子: #1 詳細: Gag construct was expressed in E.coli and purified using the SUMO-tag system NCBI-ID: 11665 / 生物種: Equine infectious anemia virus / ウイルスタイプ: VIRUS-LIKE PARTICLE / ウイルス・単離状態: OTHER / ウイルス・エンベロープ: No / ウイルス・中空状態: Yes |
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宿主 | 生物種: Equus caballus (ウマ) |
Host system | 生物種: Escherichia coli (大腸菌) / 組換株: BL21 |
ウイルス殻 | Shell ID: 1 / 名称: Capsid / 直径: 1000.0 Å |
-実験情報
-構造解析
手法 | クライオ電子顕微鏡法 |
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解析 | サブトモグラム平均法 |
試料の集合状態 | particle |
-試料調製
緩衝液 | pH: 8 構成要素:
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グリッド | モデル: C-flat-2/2 / 材質: COPPER / メッシュ: 300 / 支持フィルム - 材質: CARBON / 支持フィルム - トポロジー: HOLEY / 前処理 - タイプ: GLOW DISCHARGE / 前処理 - 雰囲気: AIR / 詳細: 20 mA | ||||||||||||
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 90 % / チャンバー内温度: 288 K / 装置: FEI VITROBOT MARK II 詳細: 1-2 seconds blot time, offset -3mm 10 nm colloidal gold was added prior to vitrification. | ||||||||||||
詳細 | Virus-like-particles (spherical) of EIAV Gag deltaMAdeltap9 (referred to as Gag deltaMA) assembled at pH8. |
-電子顕微鏡法
顕微鏡 | FEI TITAN KRIOS |
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電子線 | 加速電圧: 300 kV / 電子線源: FIELD EMISSION GUN |
電子光学系 | C2レンズ絞り径: 50.0 µm / 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / 最大 デフォーカス(公称値): -5.0 µm / 最小 デフォーカス(公称値): -1.5 µm / 倍率(公称値): 105000 |
特殊光学系 | エネルギーフィルター - 名称: GIF Quantum LS / エネルギーフィルター - スリット幅: 20 eV |
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 |
詳細 | nanoprobe |
撮影 | フィルム・検出器のモデル: GATAN K2 QUANTUM (4k x 4k) 検出モード: SUPER-RESOLUTION / デジタル化 - サイズ - 横: 3708 pixel / デジタル化 - サイズ - 縦: 3838 pixel / 撮影したグリッド数: 1 / 平均露光時間: 1.8 sec. / 平均電子線量: 3.4 e/Å2 詳細: Data was acquired using a dose-symmetric tilt acquisition scheme, as described in Hagen et al, 2017, J. Struct. Biol, 197(2):191-8 |
実験機器 | モデル: Titan Krios / 画像提供: FEI Company |
-画像解析
抽出 | トモグラム数: 37 / 使用した粒子像数: 191612 / ソフトウェア - 名称: MATLAB ソフトウェア - 詳細: partially based on the TOM toolbox 詳細: Subtomogram extraction positions were defined in Amira using the electron microscopy toolbox by determing the radii and the center of the VLPs. Initially, positions were oversampled and ...詳細: Subtomogram extraction positions were defined in Amira using the electron microscopy toolbox by determing the radii and the center of the VLPs. Initially, positions were oversampled and subsequently cleaned during alignments using cross-correlation and distance thresholds. |
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CTF補正 | ソフトウェア: (名称: CTFFIND (ver. 4), CTFPHASEFLIP, NOVACTF) 詳細: CTF-correction was initially performed using ctfphaseflip in IMOD and NovaCTF in the final steps |
最終 角度割当 | タイプ: PROJECTION MATCHING Projection matching processing - Number reference projections: 1 Projection matching processing - Merit function: CC / ソフトウェア: (名称: AV3, TOM Toolbox) 詳細: Subtomogram alignment was performed as described in the published manuscript. |
最終 再構成 | 使用したクラス数: 1 / 想定した対称性 - 点群: C6 (6回回転対称) / 解像度のタイプ: BY AUTHOR / 解像度: 3.9 Å / 解像度の算出法: FSC 0.143 CUT-OFF / ソフトウェア: (名称: AV3, TOM Toolbox) / 使用したサブトモグラム数: 65807 |
詳細 | Tilt series were low-pass filtered according to their cumulative dose using exposure filters that were calculated using an exposure-dependent amplitude attenuation function and critical exposure constants (as published in Grant & Grigorieff, Elife, 2015). Tilt series were aligned and reconstructed in IMOD. |
-原子モデル構築 1
精密化 | 空間: REAL |
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