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データを開く
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基本情報
登録情報 | ![]() | |||||||||
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タイトル | Local refinement of MCE ring and needle in Msmeg Mce1-LucB complex (Map1c) | |||||||||
![]() | Constituent Map1c used to in composite Map1. Map was generated by locally refining Class1 particles that were re-centered and extracted with a smaller boxsize (360 px). | |||||||||
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![]() | Membrane protein complex / ABC transporter / Virulence factor / Lipid transport / MEMBRANE PROTEIN | |||||||||
生物種 | ![]() | |||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 2.97 Å | |||||||||
![]() | Chen J / Bhabha G / Ekiert DC | |||||||||
資金援助 | ![]()
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![]() | ![]() タイトル: Structure of an endogenous mycobacterial MCE lipid transporter. 著者: James Chen / Alice Fruhauf / Catherine Fan / Jackeline Ponce / Beatrix Ueberheide / Gira Bhabha / Damian C Ekiert / ![]() 要旨: To replicate inside macrophages and cause tuberculosis, Mycobacterium tuberculosis must scavenge a variety of nutrients from the host. The mammalian cell entry (MCE) proteins are important virulence ...To replicate inside macrophages and cause tuberculosis, Mycobacterium tuberculosis must scavenge a variety of nutrients from the host. The mammalian cell entry (MCE) proteins are important virulence factors in M. tuberculosis, where they are encoded by large gene clusters and have been implicated in the transport of fatty acids and cholesterol across the impermeable mycobacterial cell envelope. Very little is known about how cargos are transported across this barrier, and it remains unclear how the approximately ten proteins encoded by a mycobacterial mce gene cluster assemble to transport cargo across the cell envelope. Here we report the cryo-electron microscopy (cryo-EM) structure of the endogenous Mce1 lipid-import machine of Mycobacterium smegmatis-a non-pathogenic relative of M. tuberculosis. The structure reveals how the proteins of the Mce1 system assemble to form an elongated ABC transporter complex that is long enough to span the cell envelope. The Mce1 complex is dominated by a curved, needle-like domain that appears to be unrelated to previously described protein structures, and creates a protected hydrophobic pathway for lipid transport across the periplasm. Our structural data revealed the presence of a subunit of the Mce1 complex, which we identified using a combination of cryo-EM and AlphaFold2, and name LucB. Our data lead to a structural model for Mce1-mediated lipid import across the mycobacterial cell envelope. | |||||||||
履歴 |
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構造の表示
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ダウンロードとリンク
-EMDBアーカイブ
マップデータ | ![]() | 168.1 MB | ![]() | |
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ヘッダ (付随情報) | ![]() ![]() | 30.3 KB 30.3 KB | 表示 表示 | ![]() |
FSC (解像度算出) | ![]() | 11.9 KB | 表示 | ![]() |
画像 | ![]() | 69.5 KB | ||
その他 | ![]() ![]() | 165.1 MB 165.1 MB | ||
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-関連構造データ
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リンク
EMDBのページ | ![]() ![]() |
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マップ
ファイル | ![]() | ||||||||||||||||||||
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注釈 | Constituent Map1c used to in composite Map1. Map was generated by locally refining Class1 particles that were re-centered and extracted with a smaller boxsize (360 px). | ||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 0.8255 Å | ||||||||||||||||||||
密度 |
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対称性 | 空間群: 1 | ||||||||||||||||||||
詳細 | EMDB XML:
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-添付データ
-ハーフマップ: Half map A for Map1c
ファイル | emd_29235_half_map_1.map | ||||||||||||
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注釈 | Half map A for Map1c | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: Half map B for Map1c
ファイル | emd_29235_half_map_2.map | ||||||||||||
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注釈 | Half map B for Map1c | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
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試料の構成要素
+全体 : Mce1 lipid transporter composed of Mce1 MCE proteins (Mce1ABCDEF)...
+超分子 #1: Mce1 lipid transporter composed of Mce1 MCE proteins (Mce1ABCDEF)...
+分子 #1: Virulence factor Mce family protein Mce1A
+分子 #2: Virulence factor Mce family protein Mce1B
+分子 #3: Virulence factor Mce family protein Mce1C
+分子 #4: Virulence factor Mce family protein Mce1D
+分子 #5: Virulence factor Mce family protein Mce1E
+分子 #6: Virulence factor Mce family protein Mce1F
+分子 #7: ATP-binding protein MceG
+分子 #8: ATP-binding protein MceG
+分子 #9: ABC transporter permease protein YrbE1A
+分子 #10: ABC transporter permease protein YrbE1B
+分子 #11: Mce binding protein LucB/MSMEG_3032
-実験情報
-構造解析
手法 | クライオ電子顕微鏡法 |
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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 | 1.7 mg/mL | ||||||||||||||||||
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緩衝液 | pH: 7.5 構成要素:
詳細: 50 mM Tris-HCl pH 7.5, 5 mM MgSO4, 150 mM NaCl, 1 mM DDM, 1 mM DTT | ||||||||||||||||||
グリッド | モデル: Quantifoil R2/2 / 材質: COPPER / メッシュ: 300 / 支持フィルム - 材質: CARBON / 支持フィルム - トポロジー: CONTINUOUS / 支持フィルム - Film thickness: 2 | ||||||||||||||||||
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 100 % / チャンバー内温度: 295.15 K / 装置: FEI VITROBOT MARK IV | ||||||||||||||||||
詳細 | This sample contains a mixture of MCE proteins endogenously purified from Mycobacterium smegmatis. |
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電子顕微鏡法
顕微鏡 | FEI TITAN KRIOS |
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特殊光学系 | エネルギーフィルター - スリット幅: 20 eV |
撮影 | フィルム・検出器のモデル: GATAN K3 BIOQUANTUM (6k x 4k) デジタル化 - サイズ - 横: 11520 pixel / デジタル化 - サイズ - 縦: 8184 pixel / 撮影したグリッド数: 2 / 実像数: 43925 / 平均露光時間: 2.0 sec. / 平均電子線量: 60.0 e/Å2 詳細: Images were collected in super resolution mode (nominal pixel size of 0.8255 A/pix after binning by 2). |
電子線 | 加速電圧: 300 kV / 電子線源: ![]() |
電子光学系 | 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / Cs: 2.7 mm / 最大 デフォーカス(公称値): 2.4 µm / 最小 デフォーカス(公称値): 0.8 µm / 倍率(公称値): 105000 |
試料ステージ | 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER ホルダー冷却材: NITROGEN |
実験機器 | ![]() モデル: Titan Krios / 画像提供: FEI Company |
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画像解析
-原子モデル構築 1
詳細 | Model was initial fitted into the map using Chimera follow-by rigid body refinement in PHENIX. Models were further refined using PHENIX real-space refinement and then manually inspected in Coot. |
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精密化 | 空間: REAL / プロトコル: FLEXIBLE FIT 当てはまり具合の基準: Cross-correlation coefficient |