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基本情報
登録情報 | データベース: EMDB / ID: EMD-2533 | |||||||||
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タイトル | Electron cryo-microscopy of microtubule-bound human kinesin-5 motor domain in the ADP.AlFx state. | |||||||||
![]() | Reconstruction of microtubule-bound human kinesin-5 motor domain in presence of ADP.AlFx (ATP hydrolysis transition state) | |||||||||
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![]() | cryo-electron microscopy / kinesins / mechanochemistry / microtubules / mitosis | |||||||||
機能・相同性 | ![]() spindle elongation / regulation of mitotic centrosome separation / Kinesins / plus-end-directed microtubule motor activity / mitotic centrosome separation / positive regulation of axon guidance / COPI-dependent Golgi-to-ER retrograde traffic / microtubule motor activity / kinesin complex / spindle organization ...spindle elongation / regulation of mitotic centrosome separation / Kinesins / plus-end-directed microtubule motor activity / mitotic centrosome separation / positive regulation of axon guidance / COPI-dependent Golgi-to-ER retrograde traffic / microtubule motor activity / kinesin complex / spindle organization / microtubule-based movement / microtubule-based process / mitotic spindle assembly / MHC class II antigen presentation / mitotic spindle organization / 加水分解酵素; 酸無水物に作用; GTPに作用・細胞または細胞小器官の運動に関与 / structural constituent of cytoskeleton / mitotic spindle / spindle pole / microtubule cytoskeleton organization / spindle / microtubule cytoskeleton / mitotic cell cycle / nervous system development / microtubule binding / microtubule / hydrolase activity / protein heterodimerization activity / cell division / GTPase activity / GTP binding / protein kinase binding / protein-containing complex / ATP binding / membrane / nucleus / metal ion binding / cytosol / cytoplasm 類似検索 - 分子機能 | |||||||||
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: 9.2 Å | |||||||||
![]() | Goulet A / Major J / Jun Y / Gross SP / Rosenfeld SS / Moores CA | |||||||||
![]() | ![]() タイトル: Comprehensive structural model of the mechanochemical cycle of a mitotic motor highlights molecular adaptations in the kinesin family. 著者: Adeline Goulet / Jennifer Major / Yonggun Jun / Steven P Gross / Steven S Rosenfeld / Carolyn A Moores / ![]() 要旨: Kinesins are responsible for a wide variety of microtubule-based, ATP-dependent functions. Their motor domain drives these activities, but the molecular adaptations that specify these diverse and ...Kinesins are responsible for a wide variety of microtubule-based, ATP-dependent functions. Their motor domain drives these activities, but the molecular adaptations that specify these diverse and essential cellular activities are poorly understood. It has been assumed that the first identified kinesin--the transport motor kinesin-1--is the mechanistic paradigm for the entire superfamily, but accumulating evidence suggests otherwise. To address the deficits in our understanding of the molecular basis of functional divergence within the kinesin superfamily, we studied kinesin-5s, which are essential mitotic motors whose inhibition blocks cell division. Using cryo-electron microscopy and determination of structure at subnanometer resolution, we have visualized conformations of microtubule-bound human kinesin-5 motor domain at successive steps in its ATPase cycle. After ATP hydrolysis, nucleotide-dependent conformational changes in the active site are allosterically propagated into rotations of the motor domain and uncurling of the drug-binding loop L5. In addition, the mechanical neck-linker element that is crucial for motor stepping undergoes discrete, ordered displacements. We also observed large reorientations of the motor N terminus that indicate its importance for kinesin-5 function through control of neck-linker conformation. A kinesin-5 mutant lacking this N terminus is enzymatically active, and ATP-dependent neck-linker movement and motility are defective, although not ablated. All these aspects of kinesin-5 mechanochemistry are distinct from kinesin-1. Our findings directly demonstrate the regulatory role of the kinesin-5 N terminus in collaboration with the motor's structured neck-linker and highlight the multiple adaptations within kinesin motor domains that tune their mechanochemistries according to distinct functional requirements. | |||||||||
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構造ビューア | EMマップ: ![]() ![]() ![]() |
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マップデータ | ![]() | 133.1 KB | ![]() | |
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ヘッダ (付随情報) | ![]() ![]() | 12.2 KB 12.2 KB | 表示 表示 | ![]() |
画像 | ![]() | 272.5 KB | ||
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-関連構造データ
関連構造データ | ![]() 4ck6MC ![]() 4ck7MC ![]() 2534C ![]() 2535C ![]() 2536C ![]() 2537C ![]() 2538C ![]() 2539C ![]() 2540C ![]() 2541C ![]() 2542C ![]() 4ck5C M: このマップから作成された原子モデル C: 同じ文献を引用 ( |
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「今月の分子」の関連する項目 |
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ファイル | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | Reconstruction of microtubule-bound human kinesin-5 motor domain in presence of ADP.AlFx (ATP hydrolysis transition state) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 2.8 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 : 13-protofilament microtubule-bound human kinesin-5 motor domain w...
全体 | 名称: 13-protofilament microtubule-bound human kinesin-5 motor domain with ADP.AlFx |
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要素 |
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-超分子 #1000: 13-protofilament microtubule-bound human kinesin-5 motor domain w...
超分子 | 名称: 13-protofilament microtubule-bound human kinesin-5 motor domain with ADP.AlFx タイプ: sample / ID: 1000 集合状態: 13-protofilament microtubule with one kineisn-5 motor domain bound every tubulin heterodimers Number unique components: 3 |
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-分子 #1: TUBULIN ALPHA-1D CHAIN
分子 | 名称: TUBULIN ALPHA-1D CHAIN / タイプ: protein_or_peptide / ID: 1 / Name.synonym: ALPHA TUBULIN / 組換発現: No |
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由来(天然) | 生物種: ![]() ![]() |
配列 | UniProtKB: Tubulin alpha-1D chain |
-分子 #2: TUBULIN BETA-2B CHAIN
分子 | 名称: TUBULIN BETA-2B CHAIN / タイプ: protein_or_peptide / ID: 2 / Name.synonym: BETA TUBULIN / 組換発現: No |
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由来(天然) | 生物種: ![]() ![]() |
配列 | UniProtKB: Tubulin beta-2B chain |
-分子 #3: Kinesin-5 motor domain
分子 | 名称: Kinesin-5 motor domain / タイプ: protein_or_peptide / ID: 3 / 詳細: cys-lite mutant containing the substitution T126C / 組換発現: Yes |
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由来(天然) | 生物種: ![]() |
組換発現 | 生物種: ![]() ![]() |
-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
緩衝液 | pH: 6.8 / 詳細: 80 mM PIPES, 5 mM MgCl2, 1 mM EGTA, 2 mM ADPAlFx |
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グリッド | 詳細: 400 mesh holey carbon grids |
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 100 % / 装置: FEI VITROBOT MARK I 手法: chamber at 24 degrees C, 100% humidity, blot 3.5 sec |
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電子顕微鏡法
顕微鏡 | FEI TECNAI F20 |
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温度 | 平均: 90 K |
アライメント法 | Legacy - 非点収差: Objective lens astigmatism was corrected at 150,000 times magnification |
日付 | 2011年3月10日 |
撮影 | カテゴリ: FILM / フィルム・検出器のモデル: KODAK SO-163 FILM / デジタル化 - スキャナー: ZEISS SCAI / デジタル化 - サンプリング間隔: 7 µm / 実像数: 125 / 平均電子線量: 18 e/Å2 / ビット/ピクセル: 8 |
電子線 | 加速電圧: 200 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: 2.0 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): 0.7 µm / 倍率(公称値): 50000 |
試料ステージ | 試料ホルダーモデル: GATAN LIQUID NITROGEN |
実験機器 | ![]() モデル: Tecnai F20 / 画像提供: FEI Company |
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画像解析
詳細 | The particles were selected along individual microtubules. |
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CTF補正 | 詳細: FREALIGN |
最終 再構成 | 想定した対称性 - 点群: C1 (非対称) / 解像度のタイプ: BY AUTHOR / 解像度: 9.2 Å / 解像度の算出法: OTHER / ソフトウェア - 名称: SPIDER, FREALIGN 詳細: Approximately 139,000 asymmetric units were averaged in the final reconstruction. 使用した粒子像数: 10692 |
-原子モデル構築 1
初期モデル | PDB ID: Chain - Chain ID: C |
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ソフトウェア | 名称: Chimera, FlexEM |
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 |
得られたモデル | ![]() PDB-4ck6: ![]() PDB-4ck7: |
-原子モデル構築 2
初期モデル | PDB ID: Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B |
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ソフトウェア | 名称: ![]() |
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 |
得られたモデル | ![]() PDB-4ck6: ![]() PDB-4ck7: |