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基本情報
登録情報 | データベース: PDB / ID: 7tpt | ||||||
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タイトル | Single-particle Cryo-EM structure of Arp2/3 complex at branched-actin junction. | ||||||
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![]() | STRUCTURAL PROTEIN / Arp2/3 / actin / cytoskeletal protein / actin regulator | ||||||
機能・相同性 | ![]() EPHB-mediated forward signaling / Regulation of actin dynamics for phagocytic cup formation / RHO GTPases Activate WASPs and WAVEs / Arp2/3 protein complex / Arp2/3 complex-mediated actin nucleation / regulation of actin filament polymerization / Clathrin-mediated endocytosis / Neutrophil degranulation / cytoskeletal motor activator activity / tropomyosin binding ...EPHB-mediated forward signaling / Regulation of actin dynamics for phagocytic cup formation / RHO GTPases Activate WASPs and WAVEs / Arp2/3 protein complex / Arp2/3 complex-mediated actin nucleation / regulation of actin filament polymerization / Clathrin-mediated endocytosis / Neutrophil degranulation / cytoskeletal motor activator activity / tropomyosin binding / mesenchyme migration / troponin I binding / myosin heavy chain binding / filamentous actin / actin filament bundle / skeletal muscle thin filament assembly / striated muscle thin filament / actin filament bundle assembly / skeletal muscle myofibril / actin monomer binding / positive regulation of double-strand break repair via homologous recombination / cilium assembly / skeletal muscle fiber development / positive regulation of lamellipodium assembly / stress fiber / titin binding / actin filament polymerization / filopodium / cell projection / actin filament / 加水分解酵素; 酸無水物に作用; 酸無水物に作用・細胞または細胞小器官の運動に関与 / structural constituent of cytoskeleton / actin filament binding / calcium-dependent protein binding / cell migration / lamellipodium / site of double-strand break / actin binding / cell cortex / cell body / hydrolase activity / neuron projection / protein domain specific binding / synapse / calcium ion binding / positive regulation of gene expression / magnesium ion binding / positive regulation of transcription by RNA polymerase II / ATP binding / identical protein binding / nucleus / cytoplasm / cytosol 類似検索 - 分子機能 | ||||||
生物種 | ![]() ![]() ![]() ![]() ![]() | ||||||
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: 3.9 Å | ||||||
![]() | Ding, B. / Narvaez-Ortiz, H.Y. / Nolen, B.J. / Chowdhury, S. | ||||||
資金援助 | ![]()
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![]() | ![]() タイトル: Structure of Arp2/3 complex at a branched actin filament junction resolved by single-particle cryo-electron microscopy. 著者: Bojian Ding / Heidy Y Narvaez-Ortiz / Yuvraj Singh / Glen M Hocky / Saikat Chowdhury / Brad J Nolen / ![]() ![]() 要旨: Arp2/3 complex nucleates branched actin filaments that provide pushing forces to drive cellular processes such as lamellipodial protrusion and endocytosis. Arp2/3 complex is intrinsically inactive, ...Arp2/3 complex nucleates branched actin filaments that provide pushing forces to drive cellular processes such as lamellipodial protrusion and endocytosis. Arp2/3 complex is intrinsically inactive, and multiple classes of nucleation promoting factors (NPFs) stimulate its nucleation activity. When activated by WASP family NPFs, the complex must bind to the side of a preexisting (mother) filament of actin to complete the nucleation process, ensuring that WASP-mediated activation creates branched rather than linear actin filaments. How actin filaments contribute to activation is currently not understood, largely due to the lack of high-resolution structures of activated Arp2/3 complex bound to the side of a filament. Here, we present the 3.9-Å cryo-electron microscopy structure of the Arp2/3 complex at a branch junction. The structure reveals contacts between Arp2/3 complex and the side of the mother actin filament that likely stimulate subunit flattening, a conformational change that allows the actin-related protein subunits in the complex (Arp2 and Arp3) to mimic filamentous actin subunits. In contrast, limited contact between the bottom half of the complex and the mother filament suggests that clamp twisting, a second major conformational change observed in the active state, is not stimulated by actin filaments, potentially explaining why actin filaments are required but insufficient to trigger nucleation during WASP-mediated activation. Along with biochemical and live-cell imaging data and molecular dynamics simulations, the structure reveals features critical for the interaction of Arp2/3 complex with actin filaments and regulated assembly of branched actin filament networks in cells. | ||||||
履歴 |
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構造の表示
構造ビューア | 分子: ![]() ![]() |
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PDBx/mmCIF形式 | ![]() | 1.1 MB | 表示 | ![]() |
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PDB形式 | ![]() | 946.2 KB | 表示 | ![]() |
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その他 | ![]() |
-検証レポート
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
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-関連構造データ
関連構造データ | ![]() 26063MC 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
-Actin-related protein ... , 7種, 7分子 ABCDEFG
#1: タンパク質 | 分子量: 47428.031 Da / 分子数: 1 / 由来タイプ: 天然 詳細: Missing sequences correspond to unmodeled regions due to poor density map 由来: (天然) ![]() ![]() |
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#2: タンパク質 | 分子量: 44818.711 Da / 分子数: 1 / 由来タイプ: 天然 詳細: Missing sequences correspond to unmodelled regions due to poor density map 由来: (天然) ![]() ![]() |
#3: タンパク質 | 分子量: 41030.766 Da / 分子数: 1 / 由来タイプ: 天然 詳細: Missing sequences correspond to unmodeled regions due to poor density map 由来: (天然) ![]() ![]() |
#4: タンパク質 | 分子量: 34402.043 Da / 分子数: 1 / 由来タイプ: 天然 詳細: Missing sequences correspond to unmodeled regions due to poor density map 由来: (天然) ![]() ![]() |
#5: タンパク質 | 分子量: 20572.666 Da / 分子数: 1 / 由来タイプ: 天然 詳細: Missing sequences correspond to unmodeled regions due to poor density map 由来: (天然) ![]() ![]() |
#6: タンパク質 | 分子量: 19697.047 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
#7: タンパク質 | 分子量: 16251.308 Da / 分子数: 1 / 由来タイプ: 天然 詳細: Missing sequences correspond to unmodeled regions due to poor density map 由来: (天然) ![]() ![]() |
-タンパク質 / タンパク質・ペプチド , 2種, 29分子 HIJKLMNOPQRSTUabcdefghijklmno
#8: タンパク質 | 分子量: 42109.973 Da / 分子数: 14 / 由来タイプ: 天然 詳細: Missing sequences correspond to unmodeled regions due to poor density map. Actin subunits corresponding to chains J,K,L,M,T and U have been trimmed to c-beta due to lack of density for ...詳細: Missing sequences correspond to unmodeled regions due to poor density map. Actin subunits corresponding to chains J,K,L,M,T and U have been trimmed to c-beta due to lack of density for modeling sidechains. These subunits were rigid body fitted into the map. 由来: (天然) ![]() ![]() #9: タンパク質・ペプチド | タイプ: Peptide-like / クラス: 毒素 / 分子量: 808.899 Da / 分子数: 15 / 由来タイプ: 天然 詳細: Phalloidin from Amanita phalloides is a rigid bicyclic heptapeptide. This is a small molecule and does not have conventional planar peptide bonds present in proteins. 由来: (天然) ![]() 参照: BIRD: PRD_002366 |
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-非ポリマー , 2種, 32分子 ![](data/chem/img/MG.gif)
![](data/chem/img/ADP.gif)
![](data/chem/img/ADP.gif)
#10: 化合物 | ChemComp-MG / #11: 化合物 | ChemComp-ADP / |
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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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-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: Complex consisting of Arp2/3 complex-mediated branched actin junction タイプ: COMPLEX / Entity ID: #1-#9 / 由来: NATURAL |
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分子量 | 値: 0.834 MDa / 実験値: NO |
由来(天然) | 生物種: ![]() ![]() |
緩衝液 | pH: 7.5 |
試料 | 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES |
試料支持 | 詳細: 30 mA / 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: HOMEMADE PLUNGER / 凍結剤: ETHANE / 湿度: 95 % / 凍結前の試料温度: 277.15 K |
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電子顕微鏡撮影
実験機器 | ![]() モデル: Talos Arctica / 画像提供: FEI Company |
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顕微鏡 | モデル: FEI TALOS ARCTICA 詳細: Data were collected by shifting of stage to targeted exposure position. Stage was tilted to different angles: including 40 degree, 36 degree, 30 degree, 25 degree, 15 degree, 0 degree, -20 ...詳細: Data were collected by shifting of stage to targeted exposure position. Stage was tilted to different angles: including 40 degree, 36 degree, 30 degree, 25 degree, 15 degree, 0 degree, -20 degree, and -33 degree during data collection. |
電子銃 | 電子線源: ![]() |
電子レンズ | モード: BRIGHT FIELD / 倍率(公称値): 92000 X / 最大 デフォーカス(公称値): 1200 nm / 最小 デフォーカス(公称値): 800 nm / Cs: 2.7 mm / C2レンズ絞り径: 70 µm / アライメント法: BASIC |
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: 70 sec. / 電子線照射量: 41.97 e/Å2 / 検出モード: COUNTING フィルム・検出器のモデル: FEI FALCON III (4k x 4k) 撮影したグリッド数: 3 / 実像数: 7436 詳細: Each micrograph was acquired as dose-fractionated movies consisting of 82 frames per movie. |
画像スキャン | サンプリングサイズ: 14 µm / 横: 4096 / 縦: 4096 |
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解析
EMソフトウェア |
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CTF補正 | 詳細: Particles were CTF-corrected during projection matching and back projection タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||||||||
粒子像の選択 | 選択した粒子像数: 2290562 | ||||||||||||||||||||||||||||||||||||||||||||||||||
対称性 | 点対称性: C1 (非対称) | ||||||||||||||||||||||||||||||||||||||||||||||||||
3次元再構成 | 解像度: 3.9 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 127093 / アルゴリズム: BACK PROJECTION / 対称性のタイプ: POINT | ||||||||||||||||||||||||||||||||||||||||||||||||||
原子モデル構築 | プロトコル: FLEXIBLE FIT / 空間: REAL |