+データを開く
-基本情報
登録情報 | データベース: PDB / ID: 6xw4 | ||||||
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タイトル | Crystal structure of murine norovirus P domain in complex with Nanobody NB-5867 | ||||||
要素 |
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キーワード | VIRAL PROTEIN (ウイルスタンパク質) / MNV / neutralizing nanobody / VHH / norovirus (ノロウイルス) | ||||||
機能・相同性 | 機能・相同性情報 Positive stranded ssRNA viruses / Nucleoplasmin-like/VP (viral coat and capsid proteins) / Positive stranded ssRNA viruses / Calicivirus coat protein C-terminal / Calicivirus coat protein C-terminal / Calicivirus coat protein / Calicivirus coat protein / Elongation Factor Tu (Ef-tu); domain 3 / Picornavirus/Calicivirus coat protein / Viral coat protein subunit ...Positive stranded ssRNA viruses / Nucleoplasmin-like/VP (viral coat and capsid proteins) / Positive stranded ssRNA viruses / Calicivirus coat protein C-terminal / Calicivirus coat protein C-terminal / Calicivirus coat protein / Calicivirus coat protein / Elongation Factor Tu (Ef-tu); domain 3 / Picornavirus/Calicivirus coat protein / Viral coat protein subunit / Βバレル / Mainly Beta 類似検索 - ドメイン・相同性 | ||||||
生物種 | Murine norovirus 1 (マウスノロウイルス 1) Vicugna pacos (アルパカ) | ||||||
手法 | X線回折 / シンクロトロン / 分子置換 / 解像度: 2.19 Å | ||||||
データ登録者 | Kilic, T. / Sabin, C. / Hansman, G. | ||||||
資金援助 | ドイツ, 1件
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引用 | ジャーナル: J Virol / 年: 2020 タイトル: Nanobody-Mediated Neutralization Reveals an Achilles Heel for Norovirus. 著者: Anna D Koromyslova / Jessica M Devant / Turgay Kilic / Charles D Sabin / Virginie Malak / Grant S Hansman / 要旨: Human norovirus frequently causes outbreaks of acute gastroenteritis. Although discovered more than five decades ago, antiviral development has, until recently, been hampered by the lack of a ...Human norovirus frequently causes outbreaks of acute gastroenteritis. Although discovered more than five decades ago, antiviral development has, until recently, been hampered by the lack of a reliable human norovirus cell culture system. Nevertheless, a lot of pathogenesis studies were accomplished using murine norovirus (MNV), which can be grown routinely in cell culture. In this study, we analyzed a sizeable library of nanobodies that were raised against the murine norovirus virion with the main purpose of developing nanobody-based inhibitors. We discovered two types of neutralizing nanobodies and analyzed the inhibition mechanisms using X-ray crystallography, cryo-electron microscopy (cryo-EM), and cell culture techniques. The first type bound on the top region of the protruding (P) domain. Interestingly, this nanobody binding region closely overlapped the MNV receptor-binding site and collectively shared numerous P domain-binding residues. In addition, we showed that these nanobodies competed with the soluble receptor, and this action blocked virion attachment to cultured cells. The second type bound at a dimeric interface on the lower side of the P dimer. We discovered that these nanobodies disrupted a structural change in the capsid associated with binding cofactors (i.e., metal cations/bile acid). Indeed, we found that capsids underwent major conformational changes following addition of Mg or Ca Ultimately, these nanobodies directly obstructed a structural modification reserved for a postreceptor attachment stage. Altogether, our new data show that nanobody-based inhibition could occur by blocking functional and structural capsid properties. This research discovered and analyzed two different types of MNV-neutralizing nanobodies. The top-binding nanobodies sterically inhibited the receptor-binding site, whereas the dimeric-binding nanobodies interfered with a structural modification associated with cofactor binding. Moreover, we found that the capsid contained a number of vulnerable regions that were essential for viral replication. In fact, the capsid appeared to be organized in a state of flux, which could be important for cofactor/receptor-binding functions. Blocking these capsid-binding events with nanobodies directly inhibited essential capsid functions. Moreover, a number of MNV-specific nanobody binding epitopes were comparable to human norovirus-specific nanobody inhibitors. Therefore, this additional structural and inhibition information could be further exploited in the development of human norovirus antivirals. | ||||||
履歴 |
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-構造の表示
構造ビューア | 分子: MolmilJmol/JSmol |
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-ダウンロードとリンク
-ダウンロード
PDBx/mmCIF形式 | 6xw4.cif.gz | 185.6 KB | 表示 | PDBx/mmCIF形式 |
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PDB形式 | pdb6xw4.ent.gz | 143.7 KB | 表示 | PDB形式 |
PDBx/mmJSON形式 | 6xw4.json.gz | ツリー表示 | PDBx/mmJSON形式 | |
その他 | その他のダウンロード |
-検証レポート
アーカイブディレクトリ | https://data.pdbj.org/pub/pdb/validation_reports/xw/6xw4 ftp://data.pdbj.org/pub/pdb/validation_reports/xw/6xw4 | HTTPS FTP |
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-関連構造データ
-リンク
-集合体
登録構造単位 |
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1 |
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単位格子 |
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-要素
#1: タンパク質 | 分子量: 33320.570 Da / 分子数: 2 / 由来タイプ: 組換発現 由来: (組換発現) Murine norovirus 1 (マウスノロウイルス 1) 発現宿主: Escherichia coli BL21 (大腸菌) / 参照: UniProt: Q80J94 #2: 抗体 | 分子量: 14195.776 Da / 分子数: 2 / 由来タイプ: 組換発現 / 由来: (組換発現) Vicugna pacos (アルパカ) / 発現宿主: Escherichia coli (大腸菌) / Variant (発現宿主): WK6 #3: 化合物 | ChemComp-EDO / #4: 水 | ChemComp-HOH / | 研究の焦点であるリガンドがあるか | N | |
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-実験情報
-実験
実験 | 手法: X線回折 / 使用した結晶の数: 1 |
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-試料調製
結晶 | マシュー密度: 3.17 Å3/Da / 溶媒含有率: 61.19 % |
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結晶化 | 温度: 291 K / 手法: 蒸気拡散法, ハンギングドロップ法 詳細: 1.0 M Lithium chloride 0.1 M Citric acid pH 5.0 20%(w/v) PEG 6000 |
-データ収集
回折 | 平均測定温度: 100 K / Serial crystal experiment: N | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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放射光源 | 由来: シンクロトロン / サイト: ESRF / ビームライン: ID30B / 波長: 0.97856 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
検出器 | タイプ: DECTRIS PILATUS3 X 6M / 検出器: PIXEL / 日付: 2018年5月13日 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
放射 | プロトコル: SINGLE WAVELENGTH / 単色(M)・ラウエ(L): M / 散乱光タイプ: x-ray | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
放射波長 | 波長: 0.97856 Å / 相対比: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
反射 | 解像度: 2.19→47.38 Å / Num. obs: 62254 / % possible obs: 99.4 % / 冗長度: 6.503 % / Biso Wilson estimate: 34.419 Å2 / CC1/2: 0.996 / Rmerge(I) obs: 0.138 / Rrim(I) all: 0.15 / Χ2: 0.959 / Net I/σ(I): 10.11 / Num. measured all: 404860 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
反射 シェル | Diffraction-ID: 1
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-解析
ソフトウェア |
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精密化 | 構造決定の手法: 分子置換 開始モデル: 3LQ6 解像度: 2.19→47.38 Å / Cor.coef. Fo:Fc: 0.958 / Cor.coef. Fo:Fc free: 0.933 / SU B: 5.071 / SU ML: 0.123 / 交差検証法: THROUGHOUT / σ(F): 0 / ESU R: 0.184 / ESU R Free: 0.168 / 立体化学のターゲット値: MAXIMUM LIKELIHOOD 詳細: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS U VALUES : REFINED INDIVIDUALLY
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溶媒の処理 | イオンプローブ半径: 0.8 Å / 減衰半径: 0.8 Å / VDWプローブ半径: 1.2 Å / 溶媒モデル: MASK | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
原子変位パラメータ | Biso max: 121.56 Å2 / Biso mean: 30.375 Å2 / Biso min: 9.08 Å2
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精密化ステップ | サイクル: final / 解像度: 2.19→47.38 Å
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拘束条件 |
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LS精密化 シェル | 解像度: 2.193→2.25 Å / Rfactor Rfree error: 0 / Total num. of bins used: 20
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