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基本情報
登録情報 | データベース: EMDB / ID: EMD-8048 | |||||||||
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タイトル | Electron tomographic structure of a human plasma VLDL particle in complex with monoclonal antibody mAB012 (No.001) | |||||||||
![]() | human plasma VLDL particle in complex with monoclonal antibody mAB012 | |||||||||
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生物種 | ![]() ![]() ![]() | |||||||||
手法 | 電子線トモグラフィー法 / クライオ電子顕微鏡法 / 解像度: 80.0 Å | |||||||||
![]() | Yu Y / Kuang Y / Lei D / Zhai X / Krauss R / Ren G | |||||||||
資金援助 | ![]()
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![]() | ![]() タイトル: Polyhedral 3D structure of human plasma very low density lipoproteins by individual particle cryo-electron tomography1. 著者: Yadong Yu / Yu-Lin Kuang / Dongsheng Lei / Xiaobo Zhai / Meng Zhang / Ronald M Krauss / Gang Ren / ![]() 要旨: Human VLDLs assembled in the liver and secreted into the circulation supply energy to peripheral tissues. VLDL lipolysis yields atherogenic LDLs and VLDL remnants that strongly correlate with CVD. ...Human VLDLs assembled in the liver and secreted into the circulation supply energy to peripheral tissues. VLDL lipolysis yields atherogenic LDLs and VLDL remnants that strongly correlate with CVD. Although the composition of VLDL particles has been well-characterized, their 3D structure is elusive because of their variations in size, heterogeneity in composition, structural flexibility, and mobility in solution. Here, we employed cryo-electron microscopy and individual-particle electron tomography to study the 3D structure of individual VLDL particles (without averaging) at both below and above their lipid phase transition temperatures. The 3D reconstructions of VLDL and VLDL bound to antibodies revealed an unexpected polyhedral shape, in contrast to the generally accepted model of a spherical emulsion-like particle. The smaller curvature of surface lipids compared with HDL may also reduce surface hydrophobicity, resulting in lower binding affinity to the hydrophobic distal end of the N-terminal β-barrel domain of cholesteryl ester transfer protein (CETP) compared with HDL. The directional binding of CETP to HDL and VLDL may explain the function of CETP in transferring TGs and cholesteryl esters between these particles. This first visualization of the 3D structure of VLDL could improve our understanding of the role of VLDL in atherogenesis. | |||||||||
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画像 | ![]() | 48.1 KB | ||
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文書・要旨 | ![]() | 78.6 KB | 表示 | ![]() |
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文書・詳細版 | ![]() | 77.8 KB | 表示 | |
XML形式データ | ![]() | 494 B | 表示 | |
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マップ
ファイル | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | human plasma VLDL particle in complex with monoclonal antibody mAB012 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 4.8 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
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試料の構成要素
-全体 : VLDL in complex with antibody mAB012
全体 | 名称: VLDL in complex with antibody mAB012 |
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要素 |
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-超分子 #1: VLDL in complex with antibody mAB012
超分子 | 名称: VLDL in complex with antibody mAB012 / タイプ: complex / ID: 1 / 親要素: 0 詳細: VLDL was isolated from human plasma by density gradient centrifugation. mAB012 was purchased from EMD Millipore. The monoclonal IgG antibody was designed to recognize the first 10 amino acid ...詳細: VLDL was isolated from human plasma by density gradient centrifugation. mAB012 was purchased from EMD Millipore. The monoclonal IgG antibody was designed to recognize the first 10 amino acid residues of apoB-100. IgG fraction was purified from mouse ascites by DEAE column chromatography. Equimolar VLDL and mAB012 were mixed and kept at 4 degC overnight to prepare the complexes. |
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由来(天然) | 生物種: ![]() |
分子量 | 実験値: 5.0 MDa |
-超分子 #2: human plasma VLDL
超分子 | 名称: human plasma VLDL / タイプ: complex / ID: 2 / 親要素: 1 詳細: VLDL was isolated from human plasma by density gradient centrifugation. |
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由来(天然) | 生物種: ![]() |
分子量 | 実験値: 5 MDa |
-超分子 #3: Monoclonal IgG antibody mAB012
超分子 | 名称: Monoclonal IgG antibody mAB012 / タイプ: complex / ID: 3 / 親要素: 1 詳細: mAB012 was purchased from EMD Millipore. The monoclonal IgG antibody was designed to recognize the first 10 amino acid residues of apoB-100. IgG fraction was purified from mouse ascites by ...詳細: mAB012 was purchased from EMD Millipore. The monoclonal IgG antibody was designed to recognize the first 10 amino acid residues of apoB-100. IgG fraction was purified from mouse ascites by DEAE column chromatography. |
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由来(天然) | 生物種: ![]() ![]() |
組換発現 | 生物種: ![]() ![]() |
分子量 | 理論値: 160 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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![]() | 電子線トモグラフィー法 |
試料の集合状態 | particle |
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試料調製
濃度 | 0.4 mg/mL | |||||||||||||||
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緩衝液 | pH: 7.3 構成要素:
詳細: DPBS | |||||||||||||||
グリッド | モデル: EMS Lacey carbon / 材質: COPPER / メッシュ: 200 / 支持フィルム - 材質: CARBON / 支持フィルム - トポロジー: LACEY / 支持フィルム - Film thickness: 50.0 nm / 前処理 - タイプ: GLOW DISCHARGE / 前処理 - 雰囲気: AIR / 前処理 - 気圧: 0.039 kPa | |||||||||||||||
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 90 % / チャンバー内温度: 277 K / 装置: LEICA EM GP 詳細: 3 microliter of specimen blotted for 3 seconds before plunging. | |||||||||||||||
詳細 | This sample was monodisperse. | |||||||||||||||
切片作成 | その他: NO SECTIONING |
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電子顕微鏡法
顕微鏡 | ZEISS LIBRA120PLUS |
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温度 | 最低: 90.0 K / 最高: 93.0 K |
特殊光学系 | エネルギーフィルター - 名称: Zeiss in-column Omega エネルギーフィルター - エネルギー下限: 0 eV エネルギーフィルター - エネルギー上限: 20 eV |
撮影 | フィルム・検出器のモデル: GATAN ULTRASCAN 4000 (4k x 4k) デジタル化 - サイズ - 横: 2048 pixel / デジタル化 - サイズ - 縦: 2048 pixel / デジタル化 - サンプリング間隔: 30.0 µm / 撮影したグリッド数: 1 / 実像数: 85 / 平均露光時間: 0.5 sec. / 平均電子線量: 1.0 e/Å2 詳細: Images were collected by using the Gatan Tomography module of Digital Micrograph. XY tracking and focusing were done manually. |
電子線 | 加速電圧: 120 kV / 電子線源: LAB6 |
電子光学系 | C2レンズ絞り径: 75.0 µm / 最大 デフォーカス(補正後): 2.5 µm / 最小 デフォーカス(補正後): 1.5 µm / 倍率(補正後): 50000 / 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / Cs: 2.2 mm / 最大 デフォーカス(公称値): 2.5 µm / 最小 デフォーカス(公称値): 1.5 µm / 倍率(公称値): 50000 |
試料ステージ | 試料ホルダーモデル: GATAN 626 SINGLE TILT LIQUID NITROGEN CRYO TRANSFER HOLDER ホルダー冷却材: NITROGEN |
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画像解析
詳細 | Defects in images such as bad pixels and X-rays were removed prior to alignment and 3D reconstruction. |
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最終 再構成 | アルゴリズム: BACK PROJECTION / 解像度のタイプ: BY AUTHOR / 解像度: 80.0 Å / 解像度の算出法: FSC 0.5 CUT-OFF / ソフトウェア - 名称: IPET 詳細: The 3D reconstruction was conducted by using Individual Particle Electron Tomography (IPET) 使用した粒子像数: 85 |
CTF補正 | ソフトウェア - 名称: TOMOCTF (ver. V. October 2012) |