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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: EMDB / ID: EMD-5624 | |||||||||
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タイトル | Broadly Neutralizing Antibody PGT121 Allosterically Modulates CD4 Binding via Recognition of the HIV-1 gp120 V3 Base and Multiple Surrounding Glycans | |||||||||
![]() | Reconstruction of SOSIP.664 HIV-1 envelope trimer in complex with broadly neutralizing PGT122 Fab | |||||||||
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![]() | Single particle analysis / uranyl formate / tilt series / antibody / HIV gp120 trimer / Fab | |||||||||
生物種 | ![]() ![]() | |||||||||
手法 | 単粒子再構成法 / ネガティブ染色法 / 解像度: 14.0 Å | |||||||||
![]() | Khayat R / Lee JH / Julien JP / Wilson IA / Ward AB | |||||||||
![]() | ![]() タイトル: Broadly neutralizing antibody PGT121 allosterically modulates CD4 binding via recognition of the HIV-1 gp120 V3 base and multiple surrounding glycans. 著者: Jean-Philippe Julien / Devin Sok / Reza Khayat / Jeong Hyun Lee / Katie J Doores / Laura M Walker / Alejandra Ramos / Devan C Diwanji / Robert Pejchal / Albert Cupo / Umesh Katpally / Rafael ...著者: Jean-Philippe Julien / Devin Sok / Reza Khayat / Jeong Hyun Lee / Katie J Doores / Laura M Walker / Alejandra Ramos / Devan C Diwanji / Robert Pejchal / Albert Cupo / Umesh Katpally / Rafael S Depetris / Robyn L Stanfield / Ryan McBride / Andre J Marozsan / James C Paulson / Rogier W Sanders / John P Moore / Dennis R Burton / Pascal Poignard / Andrew B Ward / Ian A Wilson / ![]() 要旨: New broad and potent neutralizing HIV-1 antibodies have recently been described that are largely dependent on the gp120 N332 glycan for Env recognition. Members of the PGT121 family of antibodies, ...New broad and potent neutralizing HIV-1 antibodies have recently been described that are largely dependent on the gp120 N332 glycan for Env recognition. Members of the PGT121 family of antibodies, isolated from an African donor, neutralize ∼70% of circulating isolates with a median IC50 less than 0.05 µg ml(-1). Here, we show that three family members, PGT121, PGT122 and PGT123, have very similar crystal structures. A long 24-residue HCDR3 divides the antibody binding site into two functional surfaces, consisting of an open face, formed by the heavy chain CDRs, and an elongated face, formed by LCDR1, LCDR3 and the tip of the HCDR3. Alanine scanning mutagenesis of the antibody paratope reveals a crucial role in neutralization for residues on the elongated face, whereas the open face, which accommodates a complex biantennary glycan in the PGT121 structure, appears to play a more secondary role. Negative-stain EM reconstructions of an engineered recombinant Env gp140 trimer (SOSIP.664) reveal that PGT122 interacts with the gp120 outer domain at a more vertical angle with respect to the top surface of the spike than the previously characterized antibody PGT128, which is also dependent on the N332 glycan. We then used ITC and FACS to demonstrate that the PGT121 antibodies inhibit CD4 binding to gp120 despite the epitope being distal from the CD4 binding site. Together, these structural, functional and biophysical results suggest that the PGT121 antibodies may interfere with Env receptor engagement by an allosteric mechanism in which key structural elements, such as the V3 base, the N332 oligomannose glycan and surrounding glycans, including a putative V1/V2 complex biantennary glycan, are conformationally constrained. | |||||||||
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構造ビューア | EMマップ: ![]() ![]() ![]() |
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マップデータ | ![]() | 11.5 MB | ![]() | |
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ヘッダ (付随情報) | ![]() ![]() | 12.7 KB 12.7 KB | 表示 表示 | ![]() |
画像 | ![]() | 70.4 KB | ||
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
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文書・要旨 | ![]() | 78.4 KB | 表示 | ![]() |
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文書・詳細版 | ![]() | 77.5 KB | 表示 | |
XML形式データ | ![]() | 494 B | 表示 | |
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
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マップ
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注釈 | Reconstruction of SOSIP.664 HIV-1 envelope trimer in complex with broadly neutralizing PGT122 Fab | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 2.18 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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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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試料の構成要素
-全体 : Fab fragment of broadly neutralizing antibody PGT122 in complex w...
全体 | 名称: Fab fragment of broadly neutralizing antibody PGT122 in complex with HIV-1 SOSIP.664 from BG505 |
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要素 |
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-超分子 #1000: Fab fragment of broadly neutralizing antibody PGT122 in complex w...
超分子 | 名称: Fab fragment of broadly neutralizing antibody PGT122 in complex with HIV-1 SOSIP.664 from BG505 タイプ: sample / ID: 1000 / 詳細: The sample was monodisperse / 集合状態: one SOSIP.664 trimer binds 3 PGT122 Fabs / Number unique components: 2 |
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分子量 | 実験値: 350 KDa / 理論値: 350 KDa / 手法: SDS-PAGE |
-分子 #1: BG505 HIV-1 Env SOSIP.664
分子 | 名称: BG505 HIV-1 Env SOSIP.664 / タイプ: protein_or_peptide / ID: 1 / 詳細: Bound to Fab portion of PGT122 antibody / コピー数: 3 / 集合状態: Trimer / 組換発現: Yes |
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由来(天然) | 生物種: ![]() ![]() 株: BG505 / 別称: HIV-1 |
分子量 | 実験値: 220 KDa / 理論値: 220 KDa |
組換発現 | 生物種: ![]() |
-実験情報
-構造解析
手法 | ネガティブ染色法 |
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![]() | 単粒子再構成法 |
試料の集合状態 | particle |
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試料調製
濃度 | 0.1 mg/mL |
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緩衝液 | pH: 7 / 詳細: 20 mM Tris-HCl, pH 7.0, 50 mM NaCl |
染色 | タイプ: NEGATIVE 詳細: Grids were briefly adsorbed with protein, wicked, and stained with 2% Nano-W for 30 seconds. |
グリッド | 詳細: 400 Cu mesh grid with thin carob support, glow discharged in natural atmosphere. |
凍結 | 凍結剤: NONE / 装置: OTHER |
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電子顕微鏡法
顕微鏡 | FEI TECNAI F20 |
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温度 | 最低: 293 K / 最高: 294 K / 平均: 293 K |
アライメント法 | Legacy - 非点収差: Astigmatism of objective lens was corrected at 100,000x Legacy - Electron beam tilt params: -2 |
日付 | 2012年4月30日 |
撮影 | カテゴリ: CCD フィルム・検出器のモデル: GENERIC GATAN (4k x 4k) デジタル化 - サンプリング間隔: 0.109 µm / 実像数: 340 / 平均電子線量: 16 e/Å2 |
Tilt angle min | 0 |
電子線 | 加速電圧: 120 kV / 電子線源: ![]() |
電子光学系 | 倍率(補正後): 100000 / 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / 最大 デフォーカス(公称値): 0.72 µm / 最小 デフォーカス(公称値): 0.6 µm / 倍率(公称値): 100000 |
試料ステージ | 試料ホルダーモデル: SIDE ENTRY, EUCENTRIC / Tilt angle max: 55 |
実験機器 | ![]() モデル: Tecnai F20 / 画像提供: FEI Company |