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データを開く
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基本情報
登録情報 | データベース: PDB / ID: 8pk3 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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タイトル | CryoEM reconstruction of hemagglutinin HK68 of Influenza A virus bound to an Affimer reagent | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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![]() | ANTIVIRAL PROTEIN / Complex / Inhibitor / CryoEM / Antiviral | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
機能・相同性 | ![]() viral budding from plasma membrane / clathrin-dependent endocytosis of virus by host cell / host cell surface receptor binding / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / virion attachment to host cell / host cell plasma membrane / virion membrane / membrane 類似検索 - 分子機能 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
生物種 | ![]() ![]() synthetic construct (人工物) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
手法 | 電子顕微鏡法 / 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 3.4 Å | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
![]() | Debski-Antoniak, O. / Flynn, A. / Klebl, D.P. / Tiede, C. / Muench, S. / Tomlinson, D. / Fontana, J. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
資金援助 | 1件
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![]() | ![]() タイトル: Exploiting the Affimer platform against influenza A virus. 著者: Oliver Debski-Antoniak / Alex Flynn / David P Klebl / Moisés H Rojas Rechy / Christian Tiede / Ian A Wilson / Stephen P Muench / Darren Tomlinson / Juan Fontana / ![]() ![]() 要旨: Influenza A virus (IAV) is well known for its pandemic potential. While current surveillance and vaccination strategies are highly effective, therapeutic approaches are often short-lived due to the ...Influenza A virus (IAV) is well known for its pandemic potential. While current surveillance and vaccination strategies are highly effective, therapeutic approaches are often short-lived due to the high mutation rates of IAV. Recently, monoclonal antibodies (mAbs) have emerged as a promising therapeutic approach, both against current strains and future IAV pandemics. In addition to mAbs, several antibody-like alternatives exist, which aim to improve upon mAbs. Among these, Affimers stand out for their short development time, high expression levels in , and animal-free production. In this study, we utilized the Affimer platform to isolate and produce specific and potent inhibitors of IAV. Using a monomeric version of the IAV trimeric hemagglutinin (HA) fusion protein, we isolated 12 Affimers that inhibit IAV infection . Two of these Affimers were characterized in detail and exhibited nanomolar-binding affinities to the target H3 HA protein, specifically binding to the HA1 head domain. Cryo-electron microscopy (cryo-EM), employing a novel spray approach to prepare cryo-grids, allowed us to image HA-Affimer complexes. Combined with functional assays, we determined that these Affimers inhibit IAV by blocking the interaction of HA with the host-cell receptor, sialic acid. Furthermore, these Affimers inhibited IAV strains closely related to the one used for their isolation. Overall, our results support the use of Affimers as a viable alternative to existing targeted therapies for IAV and highlight their potential as diagnostic reagents. IMPORTANCE: Influenza A virus is one of the few viruses that can cause devastating pandemics. Due to the high mutation rates of this virus, annual vaccination is required, and antivirals are short- ...IMPORTANCE: Influenza A virus is one of the few viruses that can cause devastating pandemics. Due to the high mutation rates of this virus, annual vaccination is required, and antivirals are short-lived. Monoclonal antibodies present a promising approach to tackle influenza virus infections but are associated with some limitations. To improve on this strategy, we explored the Affimer platform, which are antibody-like proteins made in bacteria. By performing phage-display against a monomeric version of influenza virus fusion protein, an established viral target, we were able to isolate Affimers that inhibit influenza virus infection . We characterized the mechanism of inhibition of the Affimers by using assays targeting different stages of the viral replication cycle. We additionally characterized HA-Affimer complex structure, using a novel approach to prepare samples for cryo-electron microscopy. Overall, these results show that Affimers are a promising tool against influenza virus infection. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
履歴 |
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構造の表示
構造ビューア | 分子: ![]() ![]() |
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ダウンロードとリンク
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ダウンロード
PDBx/mmCIF形式 | ![]() | 317.6 KB | 表示 | ![]() |
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PDB形式 | ![]() | 260.4 KB | 表示 | ![]() |
PDBx/mmJSON形式 | ![]() | ツリー表示 | ![]() | |
その他 | ![]() |
-検証レポート
文書・要旨 | ![]() | 1.6 MB | 表示 | ![]() |
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文書・詳細版 | ![]() | 1.6 MB | 表示 | |
XML形式データ | ![]() | 70.8 KB | 表示 | |
CIF形式データ | ![]() | 104 KB | 表示 | |
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-関連構造データ
関連構造データ | ![]() 17724MC M: このデータのモデリングに利用したマップデータ C: 同じ文献を引用 ( |
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類似構造データ | 類似検索 - 機能・相同性 ![]() |
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リンク
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集合体
登録構造単位 | ![]()
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要素
-Hemagglutinin ... , 2種, 6分子 ABCDEF
#1: タンパク質 | 分子量: 35349.754 Da / 分子数: 3 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 遺伝子: HA / 発現宿主: ![]() #2: タンパク質 | 分子量: 20212.350 Da / 分子数: 3 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 遺伝子: HA / 発現宿主: ![]() |
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-タンパク質 , 1種, 3分子 GHI
#3: タンパク質 | 分子量: 10491.819 Da / 分子数: 3 / 由来タイプ: 組換発現 / 由来: (組換発現) synthetic construct (人工物) / 発現宿主: ![]() ![]() |
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-糖 , 3種, 12分子 
#4: 多糖 | #5: 多糖 | #6: 糖 | ChemComp-NAG / |
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-詳細
研究の焦点であるリガンドがあるか | N |
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Has protein modification | Y |
-実験情報
-実験
実験 | 手法: 電子顕微鏡法 |
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EM実験 | 試料の集合状態: PARTICLE / 3次元再構成法: 単粒子再構成法 |
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試料調製
構成要素 | 名称: CryoEM reconstruction of hemagglutinin HK68 of Influenza A virus bound to an Affimer reagent タイプ: COMPLEX / Entity ID: #2-#3 / 由来: RECOMBINANT |
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由来(天然) | 生物種: ![]() ![]() |
由来(組換発現) | 生物種: ![]() ![]() |
緩衝液 | pH: 7.4 |
試料 | 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES |
急速凍結 | 凍結剤: ETHANE |
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電子顕微鏡撮影
実験機器 | ![]() モデル: Titan Krios / 画像提供: FEI Company |
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顕微鏡 | モデル: FEI TITAN KRIOS |
電子銃 | 電子線源: ![]() |
電子レンズ | モード: OTHER / 最大 デフォーカス(公称値): 4000 nm / 最小 デフォーカス(公称値): 2000 nm |
撮影 | 電子線照射量: 56.2 e/Å2 フィルム・検出器のモデル: FEI FALCON IV (4k x 4k) |
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解析
EMソフトウェア | 名称: PHENIX / カテゴリ: モデル精密化 | ||||||||||||||||||||||||
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CTF補正 | タイプ: PHASE FLIPPING ONLY | ||||||||||||||||||||||||
3次元再構成 | 解像度: 3.4 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 104544 / 対称性のタイプ: POINT | ||||||||||||||||||||||||
拘束条件 |
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