+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-8818 | |||||||||
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タイトル | Cryo-EM structure of the P73G mutant of cucumber necrosis virus under native conditions | |||||||||
マップデータ | Cucumber necrosis virus, P73G mutant. Mutation blocks insect vector transmission but is buried in the capsid. Map shows disruption in the putative Zn binding site that may explain loss of transmission. | |||||||||
試料 |
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機能・相同性 | Plant viruses icosahedral capsid proteins 'S' region signature. / Icosahedral viral capsid protein, S domain / Viral coat protein (S domain) / T=3 icosahedral viral capsid / Viral coat protein subunit / structural molecule activity / RNA binding / Capsid protein 機能・相同性情報 | |||||||||
生物種 | Cucumber necrosis virus (ウイルス) | |||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 4.2 Å | |||||||||
データ登録者 | Smith TJ | |||||||||
引用 | ジャーナル: J Virol / 年: 2017 タイトル: Stability of Cucumber Necrosis Virus at the Quasi-6-Fold Axis Affects Zoospore Transmission. 著者: Michael B Sherman / Kishore Kakani / D'Ann Rochon / Wen Jiang / Neil R Voss / Thomas J Smith / 要旨: (CNV) is a member of the genus and has a monopartite positive-sense RNA genome. CNV is transmitted in nature via zoospores of the fungus As with other members of the genus, the CNV capsid swells ... (CNV) is a member of the genus and has a monopartite positive-sense RNA genome. CNV is transmitted in nature via zoospores of the fungus As with other members of the genus, the CNV capsid swells when exposed to alkaline pH and EDTA. We previously demonstrated that a P73G mutation blocks the virus from zoospore transmission while not significantly affecting replication in plants (K. Kakani, R. Reade, and D. Rochon, J Mol Biol 338:507-517, 2004, https://doi.org/10.1016/j.jmb.2004.03.008). P73 lies immediately adjacent to a putative zinc binding site (M. Li et al., J Virol 87:12166-12175, 2013, https://doi.org/10.1128/JVI.01965-13) that is formed by three icosahedrally related His residues in the N termini of the C subunit at the quasi-6-fold axes. To better understand how this buried residue might affect vector transmission, we determined the cryo-electron microscopy structure of wild-type CNV in the native and swollen state and of the transmission-defective mutant, P73G, under native conditions. With the wild-type CNV, the swollen structure demonstrated the expected expansion of the capsid. However, the zinc binding region at the quasi-6-fold at the β-annulus axes remained intact. By comparison, the zinc binding region of the P73G mutant, even under native conditions, was markedly disordered, suggesting that the β-annulus had been disrupted and that this could destabilize the capsid. This was confirmed with pH and urea denaturation experiments in conjunction with electron microscopy analysis. We suggest that the P73G mutation affects the zinc binding and/or the β-annulus, making it more fragile under neutral/basic pH conditions. This, in turn, may affect zoospore transmission. (CNV), a member of the genus , is transmitted in nature via zoospores of the fungus While a number of plant viruses are transmitted via insect vectors, little is known at the molecular level as to how the viruses are recognized and transmitted. As with many spherical plant viruses, the CNV capsid swells when exposed to alkaline pH and EDTA. We previously demonstrated that a P73G mutation that lies inside the capsid immediately adjacent to a putative zinc binding site (Li et al., J Virol 87:12166-12175, 2013, https://doi.org/10.1128/JVI.01965-13) blocks the virus from zoospore transmission while not significantly affecting replication in plants (K. Kakani, R. Reade, and D. Rochon, J Mol Biol 338:507-517, 2004, https://doi.org/10.1016/j.jmb.2004.03.008). Here, we show that the P73G mutant is less stable than the wild type, and this appears to be correlated with destabilization of the β-annulus at the icosahedral 3-fold axes. Therefore, the β-annulus appears not to be essential for particle assembly but is necessary for interactions with the transmission vector. | |||||||||
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_8818.map.gz | 585.4 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-8818-v30.xml emd-8818.xml | 7.8 KB 7.8 KB | 表示 表示 | EMDBヘッダ |
画像 | emd_8818.png | 385.5 KB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-8818 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-8818 | HTTPS FTP |
-検証レポート
文書・要旨 | emd_8818_validation.pdf.gz | 78 KB | 表示 | EMDB検証レポート |
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文書・詳細版 | emd_8818_full_validation.pdf.gz | 77.1 KB | 表示 | |
XML形式データ | emd_8818_validation.xml.gz | 494 B | 表示 | |
アーカイブディレクトリ | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8818 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8818 | 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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「今月の分子」の関連する項目 |
-マップ
ファイル | ダウンロード / ファイル: emd_8818.map.gz / 形式: CCP4 / 大きさ: 634.7 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | Cucumber necrosis virus, P73G mutant. Mutation blocks insect vector transmission but is buried in the capsid. Map shows disruption in the putative Zn binding site that may explain loss of transmission. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 1.03 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 : Cucumber necrosis virus
全体 | 名称: Cucumber necrosis virus (ウイルス) |
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要素 |
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-超分子 #1: Cucumber necrosis virus
超分子 | 名称: Cucumber necrosis virus / タイプ: virus / ID: 1 / 親要素: 0 / NCBI-ID: 12143 / 生物種: Cucumber necrosis virus / ウイルスタイプ: VIRION / ウイルス・単離状態: STRAIN / ウイルス・エンベロープ: No / ウイルス・中空状態: No |
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-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 | 単粒子再構成法 |
試料の集合状態 | particle |
-試料調製
緩衝液 | pH: 5 |
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凍結 | 凍結剤: ETHANE |
-電子顕微鏡法
顕微鏡 | JEOL 2200FS |
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撮影 | フィルム・検出器のモデル: DIRECT ELECTRON DE-20 (5k x 3k) 平均電子線量: 37.0 e/Å2 |
電子線 | 加速電圧: 200 kV / 電子線源: FIELD EMISSION GUN |
電子光学系 | 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD |
-画像解析
最終 再構成 | 解像度のタイプ: BY AUTHOR / 解像度: 4.2 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 使用した粒子像数: 49525 |
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初期 角度割当 | タイプ: PROJECTION MATCHING |
最終 角度割当 | タイプ: ANGULAR RECONSTITUTION |