+データを開く
-基本情報
登録情報 | データベース: PDB / ID: 7k57 | ||||||
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タイトル | Structure of apo VCP dodecamer generated from bacterially recombinant VCP/p97 | ||||||
要素 | Transitional endoplasmic reticulum ATPase | ||||||
キーワード | CYTOSOLIC PROTEIN / HYDROLASE / AAA ATPase / ATP hydrolysis / segregase | ||||||
機能・相同性 | 機能・相同性情報 positive regulation of Lys63-specific deubiquitinase activity / flavin adenine dinucleotide catabolic process / positive regulation of oxidative phosphorylation / VCP-NSFL1C complex / endoplasmic reticulum stress-induced pre-emptive quality control / endosome to lysosome transport via multivesicular body sorting pathway / cellular response to arsenite ion / Derlin-1 retrotranslocation complex / BAT3 complex binding / protein-DNA covalent cross-linking repair ...positive regulation of Lys63-specific deubiquitinase activity / flavin adenine dinucleotide catabolic process / positive regulation of oxidative phosphorylation / VCP-NSFL1C complex / endoplasmic reticulum stress-induced pre-emptive quality control / endosome to lysosome transport via multivesicular body sorting pathway / cellular response to arsenite ion / Derlin-1 retrotranslocation complex / BAT3 complex binding / protein-DNA covalent cross-linking repair / positive regulation of protein K63-linked deubiquitination / deubiquitinase activator activity / mitotic spindle disassembly / aggresome assembly / VCP-NPL4-UFD1 AAA ATPase complex / regulation of protein localization to chromatin / vesicle-fusing ATPase / ubiquitin-modified protein reader activity / NADH metabolic process / cellular response to misfolded protein / stress granule disassembly / positive regulation of mitochondrial membrane potential / negative regulation of protein localization to chromatin / K48-linked polyubiquitin modification-dependent protein binding / retrograde protein transport, ER to cytosol / regulation of aerobic respiration / positive regulation of ATP biosynthetic process / regulation of synapse organization / ATPase complex / ubiquitin-specific protease binding / ubiquitin-like protein ligase binding / MHC class I protein binding / autophagosome maturation / RHOH GTPase cycle / polyubiquitin modification-dependent protein binding / HSF1 activation / proteasomal protein catabolic process / endoplasmic reticulum to Golgi vesicle-mediated transport / Protein methylation / translesion synthesis / interstrand cross-link repair / ERAD pathway / negative regulation of smoothened signaling pathway / ATP metabolic process / endoplasmic reticulum unfolded protein response / Attachment and Entry / proteasome complex / viral genome replication / lipid droplet / Josephin domain DUBs / N-glycan trimming in the ER and Calnexin/Calreticulin cycle / macroautophagy / Hh mutants are degraded by ERAD / Hedgehog ligand biogenesis / Defective CFTR causes cystic fibrosis / Translesion Synthesis by POLH / positive regulation of protein-containing complex assembly / establishment of protein localization / ABC-family proteins mediated transport / ADP binding / autophagy / Aggrephagy / cytoplasmic stress granule / positive regulation of non-canonical NF-kappaB signal transduction / positive regulation of protein catabolic process / activation of cysteine-type endopeptidase activity involved in apoptotic process / azurophil granule lumen / KEAP1-NFE2L2 pathway / double-strand break repair / Ovarian tumor domain proteases / positive regulation of canonical Wnt signaling pathway / positive regulation of proteasomal ubiquitin-dependent protein catabolic process / E3 ubiquitin ligases ubiquitinate target proteins / Neddylation / site of double-strand break / cellular response to heat / ubiquitin-dependent protein catabolic process / regulation of apoptotic process / protein phosphatase binding / proteasome-mediated ubiquitin-dependent protein catabolic process / secretory granule lumen / ficolin-1-rich granule lumen / Attachment and Entry / protein ubiquitination / protein domain specific binding / intracellular membrane-bounded organelle / DNA repair / lipid binding / glutamatergic synapse / ubiquitin protein ligase binding / DNA damage response / Neutrophil degranulation / endoplasmic reticulum membrane / perinuclear region of cytoplasm / endoplasmic reticulum / ATP hydrolysis activity / protein-containing complex / RNA binding / extracellular exosome / extracellular region 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: 3.7 Å | ||||||
データ登録者 | Yu, G. / Bai, Y. / Li, K. / Jiang, W. / Zhang, Z.Y. | ||||||
資金援助 | 米国, 1件
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引用 | ジャーナル: iScience / 年: 2021 タイトル: Cryo-electron microscopy structures of VCP/p97 reveal a new mechanism of oligomerization regulation. 著者: Guimei Yu / Yunpeng Bai / Kunpeng Li / Ovini Amarasinghe / Wen Jiang / Zhong-Yin Zhang / 要旨: VCP/p97 is an evolutionarily conserved AAA+ ATPase important for cellular homeostasis. Previous studies suggest that VCP predominantly exists as a homohexamer. Here, we performed structural and ...VCP/p97 is an evolutionarily conserved AAA+ ATPase important for cellular homeostasis. Previous studies suggest that VCP predominantly exists as a homohexamer. Here, we performed structural and biochemical characterization of VCP dodecamer, an understudied state of VCP. The structure revealed an apo nucleotide status that has rarely been captured, a tail-to-tail assembly of two hexamers, and the up-elevated N-terminal domains akin to that seen in the ATP-bound hexamer. Further analyses elucidated a nucleotide status-dependent dodecamerization mechanism, where nucleotide dissociation from the D2 AAA domains induces and promotes VCP dodecamerization. In contrast, nucleotide-free D1 AAA domains are associated with the up-rotation of N-terminal domains, which may prime D1 for ATP binding. These results therefore reveal new nucleotide status-dictated intra- and interhexamer conformational changes and suggest that modulation of D2 domain nucleotide occupancy may serve as a mechanism in controlling VCP oligomeric states. | ||||||
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 | 7k57.cif.gz | 1.4 MB | 表示 | PDBx/mmCIF形式 |
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PDB形式 | pdb7k57.ent.gz | 1.2 MB | 表示 | PDB形式 |
PDBx/mmJSON形式 | 7k57.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
アーカイブディレクトリ | https://data.pdbj.org/pub/pdb/validation_reports/k5/7k57 ftp://data.pdbj.org/pub/pdb/validation_reports/k5/7k57 | HTTPS FTP |
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-関連構造データ
-リンク
-集合体
登録構造単位 |
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1 |
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-要素
#1: タンパク質 | 分子量: 89464.828 Da / 分子数: 12 / 由来タイプ: 組換発現 / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: VCP / プラスミド: PET28a / 詳細 (発現宿主): PET28a-VCP/p97 / 発現宿主: Escherichia coli BL21(DE3) (大腸菌) / 参照: UniProt: P55072, vesicle-fusing ATPase |
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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 | 名称: VCP/p97 / タイプ: COMPLEX 詳細: VCP/p97 was overexpressed and purified from E.coli BL21(DE3). Purified protein was treated with apyrase to remove prebound ADP and generate apo VCP/p97. Entity ID: all / 由来: RECOMBINANT | ||||||||||||||||||||
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分子量 | 値: 1.08 MDa / 実験値: YES | ||||||||||||||||||||
由来(天然) | 生物種: Homo sapiens (ヒト) | ||||||||||||||||||||
由来(組換発現) | 生物種: Escherichia coli (大腸菌) / 株: BL21(DE3) / プラスミド: PET28a | ||||||||||||||||||||
緩衝液 | pH: 7.4 / 詳細: 20mM Hepes pH7.4, 150mM NaCl, 1mM TCEP | ||||||||||||||||||||
緩衝液成分 |
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試料 | 濃度: 1 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES 詳細: VCP/p97 was overexpressed and purified with E.coli BL21(DE3). Purified proteins were treated with apyrase to remove prebound ADP and generate apo VCP/p97. | ||||||||||||||||||||
試料支持 | 詳細: 15mA for 40s with Emitech. / グリッドの材料: COPPER / グリッドのサイズ: 400 divisions/in. グリッドのタイプ: PELCO Ultrathin Carbon with Lacey Carbon | ||||||||||||||||||||
急速凍結 | 装置: GATAN CRYOPLUNGE 3 / 凍結剤: ETHANE / 湿度: 70 % / 凍結前の試料温度: 293 K 詳細: 3.5ul sample was applied to a lacy carbon grid coated with graphene oxide. 7 seconds blot with filter paper was performed using Gatan Cp3. |
-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: FEI TITAN 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 / Cs: 2.7 mm / C2レンズ絞り径: 100 µm / アライメント法: COMA FREE |
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: 8 sec. / 電子線照射量: 40 e/Å2 / 検出モード: SUPER-RESOLUTION フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) 撮影したグリッド数: 1 / 実像数: 807 詳細: A frame rate of 5 frames per second, a dose rate of 7.6 eps and a total exposure of 8 seconds were used. |
電子光学装置 | エネルギーフィルター名称: GIF Quantum LS / エネルギーフィルタースリット幅: 20 eV |
-解析
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: 71600 | |||||||||||||||||||||||||||
対称性 | 点対称性: D6 (2回x6回 2面回転対称) | |||||||||||||||||||||||||||
3次元再構成 | 解像度: 3.7 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 38312 / クラス平均像の数: 1 / 対称性のタイプ: POINT |