+データを開く
-基本情報
登録情報 | データベース: PDB / ID: 7nd2 | ||||||
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タイトル | Cryo-EM structure of the human FERRY complex | ||||||
要素 |
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キーワード | RNA BINDING PROTEIN / Human FERRY complex / Five-subunit Early endosome RNA and Ribosome intermediarY complex / Intracellular RNA transport / Early Endosome-associated transport of RNA | ||||||
機能・相同性 | 機能・相同性情報 quinone metabolic process / glyoxalase III activity / NADPH:quinone reductase activity / methylglyoxal catabolic process to D-lactate via S-lactoyl-glutathione / 酸化還元酵素 / NADP binding / early endosome / extracellular exosome / identical protein binding / membrane ...quinone metabolic process / glyoxalase III activity / NADPH:quinone reductase activity / methylglyoxal catabolic process to D-lactate via S-lactoyl-glutathione / 酸化還元酵素 / NADP binding / early endosome / extracellular exosome / identical protein binding / membrane / cytoplasm / cytosol 類似検索 - 分子機能 | ||||||
生物種 | Homo sapiens (ヒト) | ||||||
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: 4 Å | ||||||
データ登録者 | Quentin, D. / Klink, B.U. / Raunser, S. | ||||||
資金援助 | 1件
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引用 | ジャーナル: Mol Cell / 年: 2023 タイトル: Structural basis of mRNA binding by the human FERRY Rab5 effector complex. 著者: Dennis Quentin / Jan S Schuhmacher / Björn U Klink / Jeni Lauer / Tanvir R Shaikh / Pim J Huis In 't Veld / Luisa M Welp / Henning Urlaub / Marino Zerial / Stefan Raunser / 要旨: The pentameric FERRY Rab5 effector complex is a molecular link between mRNA and early endosomes in mRNA intracellular distribution. Here, we determine the cryo-EM structure of human FERRY. It reveals ...The pentameric FERRY Rab5 effector complex is a molecular link between mRNA and early endosomes in mRNA intracellular distribution. Here, we determine the cryo-EM structure of human FERRY. It reveals a unique clamp-like architecture that bears no resemblance to any known structure of Rab effectors. A combination of functional and mutational studies reveals that while the Fy-2 C-terminal coiled-coil acts as binding region for Fy-1/3 and Rab5, both coiled-coils and Fy-5 concur to bind mRNA. Mutations causing truncations of Fy-2 in patients with neurological disorders impair Rab5 binding or FERRY complex assembly. Thus, Fy-2 serves as a binding hub connecting all five complex subunits and mediating the binding to mRNA and early endosomes via Rab5. Our study provides mechanistic insights into long-distance mRNA transport and demonstrates that the particular architecture of FERRY is closely linked to a previously undescribed mode of RNA binding, involving coiled-coil domains. | ||||||
履歴 |
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-構造の表示
ムービー |
ムービービューア |
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構造ビューア | 分子: MolmilJmol/JSmol |
-ダウンロードとリンク
-ダウンロード
PDBx/mmCIF形式 | 7nd2.cif.gz | 395.5 KB | 表示 | PDBx/mmCIF形式 |
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PDB形式 | pdb7nd2.ent.gz | 311.2 KB | 表示 | PDB形式 |
PDBx/mmJSON形式 | 7nd2.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
文書・要旨 | 7nd2_validation.pdf.gz | 789.3 KB | 表示 | wwPDB検証レポート |
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文書・詳細版 | 7nd2_full_validation.pdf.gz | 825.9 KB | 表示 | |
XML形式データ | 7nd2_validation.xml.gz | 62.6 KB | 表示 | |
CIF形式データ | 7nd2_validation.cif.gz | 95.8 KB | 表示 | |
アーカイブディレクトリ | https://data.pdbj.org/pub/pdb/validation_reports/nd/7nd2 ftp://data.pdbj.org/pub/pdb/validation_reports/nd/7nd2 | HTTPS FTP |
-関連構造データ
-リンク
-集合体
登録構造単位 |
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-要素
#1: タンパク質 | 分子量: 88782.836 Da / 分子数: 2 / 由来タイプ: 組換発現 / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: PPP1R21, CCDC128, KLRAQ1 発現宿主: Spodoptera frugiperda (ツマジロクサヨトウ) 株 (発現宿主): Sf-9 / 参照: UniProt: Q6ZMI0 #2: タンパク質 | 分子量: 39661.414 Da / 分子数: 2 / 由来タイプ: 組換発現 / 詳細: N-terminal His-6 tag / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: CRYZL1, 4P11 / 発現宿主: Escherichia coli BL21(DE3) (大腸菌) / 参照: UniProt: O95825, 酸化還元酵素 #3: タンパク質 | 分子量: 24237.488 Da / 分子数: 4 / 由来タイプ: 組換発現 / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: GATD1, PDDC1 / 発現宿主: Escherichia coli BL21(DE3) (大腸菌) / 参照: UniProt: Q8NB37 |
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-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 |
-試料調製
構成要素 |
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由来(天然) |
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由来(組換発現) |
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緩衝液 | pH: 7.5 | ||||||||||||||||||||||||||||||
緩衝液成分 |
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試料 | 濃度: 0.7 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES | ||||||||||||||||||||||||||||||
試料支持 | グリッドの材料: GOLD / グリッドのサイズ: 300 divisions/in. / グリッドのタイプ: UltrAuFoil R1.2/1.3 | ||||||||||||||||||||||||||||||
急速凍結 | 装置: FEI VITROBOT MARK III / 凍結剤: ETHANE / 湿度: 100 % / 凍結前の試料温度: 286 K / 詳細: 3s blotting time |
-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: TFS KRIOS |
電子銃 | 電子線源: FIELD EMISSION GUN / 加速電圧: 300 kV / 照射モード: FLOOD BEAM |
電子レンズ | モード: BRIGHT FIELD / Calibrated defocus min: 1600 nm / 最大 デフォーカス(補正後): 2800 nm |
試料ホルダ | 凍結剤: NITROGEN 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER |
撮影 | 平均露光時間: 15 sec. / 電子線照射量: 75.8 e/Å2 フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) 実像数: 1879 |
電子光学装置 | エネルギーフィルター名称: GIF Bioquantum / エネルギーフィルタースリット幅: 20 eV |
画像スキャン | 動画フレーム数/画像: 40 |
-解析
ソフトウェア | 名称: PHENIX / バージョン: 1.18.2_3874: / 分類: 精密化 | |||||||||||||||||||||||||||||||||
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EMソフトウェア |
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CTF補正 | タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||
粒子像の選択 | 選択した粒子像数: 1800000 | |||||||||||||||||||||||||||||||||
対称性 | 点対称性: C2 (2回回転対称) | |||||||||||||||||||||||||||||||||
3次元再構成 | 解像度: 4 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 18300 / 対称性のタイプ: POINT | |||||||||||||||||||||||||||||||||
原子モデル構築 | 詳細: To build the model for the (CRYZL1)2(PPP1r21)2(GATD1)4 core of the FERRY complex, the obtained crystal structures of CRYZL1 and GATD1 were initially fitted into the corresponding density ...詳細: To build the model for the (CRYZL1)2(PPP1r21)2(GATD1)4 core of the FERRY complex, the obtained crystal structures of CRYZL1 and GATD1 were initially fitted into the corresponding density using the rigid body fitting tool in Chimera. trRosetta, a de novo protein structure prediction algorithm that is based on direct energy minimization with restrained Rosetta, was used to obtain initial models for PPP1r21. The predicted model for the 6-helix bundle domain, containing residues 246 to 498, that matched our experimental density best was subsequently fitted similar as CRYZL1 and GATD1 using rigid body fit. Manual model building for the regions N- and C-terminal 6-helix bundle, which comprise residues 218 to 245 and 499 to 552, respectively, was further guided by secondary structure predictions of individual trRosetta runs for these regions, that include the vertical helix as well as the beginning of the two terminal coiled-coils of PPP1r21. With the resulting combined model, containing residues 2 to 349, 218 to 552 and 8 to 217 of CRYZL1, PPP1r21 and GATD1, respectively, a restrained refinement in PHENIX was performed. In the next step, the model was further refined using a combination of manual building in COOT and real-space refinement in PHENIX. | |||||||||||||||||||||||||||||||||
拘束条件 |
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