+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-6119 | |||||||||
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タイトル | Electron cryo-microscopy of human papillomavirus 16 and H16.V5 Fab fragments | |||||||||
マップデータ | Reconstruction of mature human papillomavirus 16 capsid complexed with Fab fragments after 1 hr incubation | |||||||||
試料 |
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キーワード | virus-Fab complex / neutralization antibody / maturation | |||||||||
生物種 | Mus musculus (ハツカネズミ) / Human papillomavirus 16 (パピローマウイルス) | |||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 14.7 Å | |||||||||
データ登録者 | Lee H / Brendle SA / Bywaters SM / Guan J / Ashley RE / Yoder JD / Makhov AM / Conway JF / Christensen ND / Hafenstein S | |||||||||
引用 | ジャーナル: J Virol / 年: 2015 タイトル: A cryo-electron microscopy study identifies the complete H16.V5 epitope and reveals global conformational changes initiated by binding of the neutralizing antibody fragment. 著者: Hyunwook Lee / Sarah A Brendle / Stephanie M Bywaters / Jian Guan / Robert E Ashley / Joshua D Yoder / Alexander M Makhov / James F Conway / Neil D Christensen / Susan Hafenstein / 要旨: Human papillomavirus 16 (HPV16) is a worldwide health threat and an etiologic agent of cervical cancer. To understand the antigenic properties of HPV16, we pursued a structural study to elucidate HPV ...Human papillomavirus 16 (HPV16) is a worldwide health threat and an etiologic agent of cervical cancer. To understand the antigenic properties of HPV16, we pursued a structural study to elucidate HPV capsids and antibody interactions. The cryo-electron microscopy (cryo-EM) structures of a mature HPV16 particle and an altered capsid particle were solved individually and as complexes with fragment of antibody (Fab) from the neutralizing antibody H16.V5. Fitted crystal structures provided a pseudoatomic model of the virus-Fab complex, which identified a precise footprint of H16.V5, including previously unrecognized residues. The altered-capsid-Fab complex map showed that binding of the Fab induced significant conformational changes that were not seen in the altered-capsid structure alone. These changes included more ordered surface loops, consolidated so-called "invading-arm" structures, and tighter intercapsomeric connections at the capsid floor. The H16.V5 Fab preferentially bound hexavalent capsomers likely with a stabilizing effect that directly correlated with the number of bound Fabs. Additional cryo-EM reconstructions of the virus-Fab complex for different incubation times and structural analysis provide a model for a hyperstabilization of the capsomer by H16.V5 Fab and showed that the Fab distinguishes subtle differences between antigenic sites. IMPORTANCE: Our analysis of the cryo-EM reconstructions of the HPV16 capsids and virus-Fab complexes has identified the entire HPV.V5 conformational epitope and demonstrated a detailed neutralization ...IMPORTANCE: Our analysis of the cryo-EM reconstructions of the HPV16 capsids and virus-Fab complexes has identified the entire HPV.V5 conformational epitope and demonstrated a detailed neutralization mechanism of this clinically important monoclonal antibody against HPV16. The Fab bound and ordered the apical loops of HPV16. This conformational change was transmitted to the lower region of the capsomer, resulting in enhanced intercapsomeric interactions evidenced by the more ordered capsid floor and "invading-arm" structures. This study advances the understanding of the neutralization mechanism used by H16.V5. | |||||||||
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_6119.map.gz | 45.1 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-6119-v30.xml emd-6119.xml | 11.3 KB 11.3 KB | 表示 表示 | EMDBヘッダ |
画像 | 400_6119.gif 80_6119.gif | 80.7 KB 6.1 KB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-6119 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-6119 | HTTPS FTP |
-検証レポート
文書・要旨 | emd_6119_validation.pdf.gz | 78.5 KB | 表示 | EMDB検証レポート |
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文書・詳細版 | emd_6119_full_validation.pdf.gz | 77.6 KB | 表示 | |
XML形式データ | emd_6119_validation.xml.gz | 495 B | 表示 | |
アーカイブディレクトリ | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-6119 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-6119 | 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_6119.map.gz / 形式: CCP4 / 大きさ: 112.1 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | Reconstruction of mature human papillomavirus 16 capsid complexed with Fab fragments after 1 hr incubation | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
投影像・断面図 | 画像のコントロール
画像は Spider により作成 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 2.86 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 : Mature HPV16 quasivirus capsid complexed with H16.V5 Fabs
全体 | 名称: Mature HPV16 quasivirus capsid complexed with H16.V5 Fabs |
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要素 |
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-超分子 #1000: Mature HPV16 quasivirus capsid complexed with H16.V5 Fabs
超分子 | 名称: Mature HPV16 quasivirus capsid complexed with H16.V5 Fabs タイプ: sample / ID: 1000 集合状態: Three hundred H16.V5 Fabs bind to one HPV16 capsid Number unique components: 2 |
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分子量 | 理論値: 42 MDa |
-超分子 #1: Human papillomavirus 16
超分子 | 名称: Human papillomavirus 16 / タイプ: virus / ID: 1 詳細: The virus sample was incubated with excessive Fab fragments for 1 hr. NCBI-ID: 337041 / 生物種: Human papillomavirus 16 / データベース: NCBI / ウイルスタイプ: VIRION / 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: No |
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宿主 | 生物種: Homo sapiens (ヒト) / 別称: VERTEBRATES |
分子量 | 理論値: 27 MDa |
ウイルス殻 | Shell ID: 1 / 名称: L1/L2 / 直径: 600 Å / T番号(三角分割数): 7 |
-分子 #1: H16.V5 Fab
分子 | 名称: H16.V5 Fab / タイプ: protein_or_peptide / ID: 1 / コピー数: 300 / 集合状態: monomer / 組換発現: No / データベース: NCBI |
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由来(天然) | 生物種: Mus musculus (ハツカネズミ) / 細胞: hybridoma |
分子量 | 理論値: 50 KDa |
-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
濃度 | 1.0 mg/mL |
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緩衝液 | pH: 7.4 詳細: 137 mM NaCl, 2.7 mM KCl, 10 mM Na2HPO4, 1.8 mM KH2PO4 |
グリッド | 詳細: glow-discharged holey carbon Quantifoil grids |
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 90 % / チャンバー内温度: 102 K / 装置: GATAN CRYOPLUNGE 3 / 手法: Blot for 0.7 seconds before plunging. |
-電子顕微鏡法
顕微鏡 | JEOL 2100 |
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温度 | 平均: 95 K |
日付 | 2014年8月13日 |
撮影 | カテゴリ: CCD フィルム・検出器のモデル: GATAN ULTRASCAN 4000 (4k x 4k) 実像数: 138 / 平均電子線量: 15 e/Å2 |
電子線 | 加速電圧: 200 kV / 電子線源: LAB6 |
電子光学系 | 照射モード: SPOT SCAN / 撮影モード: BRIGHT FIELD / Cs: 2 mm / 最大 デフォーカス(公称値): 5.67 µm / 最小 デフォーカス(公称値): 1.19 µm / 倍率(公称値): 40000 |
試料ステージ | 試料ホルダーモデル: GATAN LIQUID NITROGEN |
-画像解析
詳細 | The particles were selected using semi-automatic program e2boxer.py (EMAN2). |
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CTF補正 | 詳細: Each particle |
最終 再構成 | アルゴリズム: OTHER / 解像度のタイプ: BY AUTHOR / 解像度: 14.7 Å / 解像度の算出法: OTHER / ソフトウェア - 名称: Auto3dem 詳細: Semi-automatic particle selection was performed using e2boxer.py to obtain the particle coordinates, followed by particle boxing, linearization, normalization, and apodization of the images ...詳細: Semi-automatic particle selection was performed using e2boxer.py to obtain the particle coordinates, followed by particle boxing, linearization, normalization, and apodization of the images using Robem. Defocus and astigmatism values to perform contrast transfer function (CTF) correction were assessed using Robem for the extracted particles. The icosahedrally averaged reconstructions were initiated using a random model generated with setup_rmc and reached 14 A resolution estimated at a Fourier Shell Correlation (FSC) of 0.5. For the last step of refinement, the final maps were CTF-corrected using a B factor of 200 A2. 使用した粒子像数: 2075 |