+データを開く
-基本情報
登録情報 | データベース: PDB / ID: 7otm | ||||||
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タイトル | Cryo-EM structure of DNA-PKcs in complex with NU7441 | ||||||
要素 | DNA-dependent protein kinase catalytic subunit,DNA-PKcs | ||||||
キーワード | DNA BINDING PROTEIN / complex / inhibitor / DNA repair | ||||||
機能・相同性 | 機能・相同性情報 positive regulation of platelet formation / T cell receptor V(D)J recombination / pro-B cell differentiation / small-subunit processome assembly / positive regulation of lymphocyte differentiation / DNA-dependent protein kinase activity / histone H2AXS139 kinase activity / DNA-dependent protein kinase complex / immature B cell differentiation / DNA-dependent protein kinase-DNA ligase 4 complex ...positive regulation of platelet formation / T cell receptor V(D)J recombination / pro-B cell differentiation / small-subunit processome assembly / positive regulation of lymphocyte differentiation / DNA-dependent protein kinase activity / histone H2AXS139 kinase activity / DNA-dependent protein kinase complex / immature B cell differentiation / DNA-dependent protein kinase-DNA ligase 4 complex / immunoglobulin V(D)J recombination / nonhomologous end joining complex / regulation of smooth muscle cell proliferation / Cytosolic sensors of pathogen-associated DNA / regulation of epithelial cell proliferation / IRF3-mediated induction of type I IFN / telomere capping / regulation of hematopoietic stem cell differentiation / U3 snoRNA binding / T cell lineage commitment / negative regulation of cGAS/STING signaling pathway / double-strand break repair via alternative nonhomologous end joining / maturation of 5.8S rRNA / B cell lineage commitment / positive regulation of double-strand break repair via nonhomologous end joining / mitotic G1 DNA damage checkpoint signaling / ectopic germ cell programmed cell death / somitogenesis / activation of innate immune response / telomere maintenance / positive regulation of erythrocyte differentiation / negative regulation of protein phosphorylation / Nonhomologous End-Joining (NHEJ) / small-subunit processome / positive regulation of translation / regulation of circadian rhythm / protein-DNA complex / response to gamma radiation / peptidyl-threonine phosphorylation / protein destabilization / protein modification process / brain development / cellular response to insulin stimulus / double-strand break repair via nonhomologous end joining / rhythmic process / intrinsic apoptotic signaling pathway in response to DNA damage / double-strand break repair / E3 ubiquitin ligases ubiquitinate target proteins / double-stranded DNA binding / heart development / peptidyl-serine phosphorylation / chromosome, telomeric region / T cell differentiation in thymus / transcription regulator complex / RNA polymerase II-specific DNA-binding transcription factor binding / non-specific serine/threonine protein kinase / protein kinase activity / protein domain specific binding / positive regulation of apoptotic process / protein phosphorylation / protein serine kinase activity / innate immune response / protein serine/threonine kinase activity / DNA damage response / chromatin / negative regulation of apoptotic process / nucleolus / enzyme binding / positive regulation of transcription by RNA polymerase II / protein-containing complex / RNA binding / nucleoplasm / ATP binding / membrane / nucleus / 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: 3.33 Å | ||||||
データ登録者 | Liang, S. / Thomas, S.E. / Blundell, T.L. | ||||||
資金援助 | 英国, 1件
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引用 | ジャーナル: Nature / 年: 2022 タイトル: Structural insights into inhibitor regulation of the DNA repair protein DNA-PKcs. 著者: Shikang Liang / Sherine E Thomas / Amanda K Chaplin / Steven W Hardwick / Dimitri Y Chirgadze / Tom L Blundell / 要旨: The DNA-dependent protein kinase catalytic subunit (DNA-PKcs) has a central role in non-homologous end joining, one of the two main pathways that detect and repair DNA double-strand breaks (DSBs) in ...The DNA-dependent protein kinase catalytic subunit (DNA-PKcs) has a central role in non-homologous end joining, one of the two main pathways that detect and repair DNA double-strand breaks (DSBs) in humans. DNA-PKcs is of great importance in repairing pathological DSBs, making DNA-PKcs inhibitors attractive therapeutic agents for cancer in combination with DSB-inducing radiotherapy and chemotherapy. Many of the selective inhibitors of DNA-PKcs that have been developed exhibit potential as treatment for various cancers. Here we report cryo-electron microscopy (cryo-EM) structures of human DNA-PKcs natively purified from HeLa cell nuclear extracts, in complex with adenosine-5'-(γ-thio)-triphosphate (ATPγS) and four inhibitors (wortmannin, NU7441, AZD7648 and M3814), including drug candidates undergoing clinical trials. The structures reveal molecular details of ATP binding at the active site before catalysis and provide insights into the modes of action and specificities of the competitive inhibitors. Of note, binding of the ligands causes movement of the PIKK regulatory domain (PRD), revealing a connection between the p-loop and PRD conformations. Electrophoretic mobility shift assay and cryo-EM studies on the DNA-dependent protein kinase holoenzyme further show that ligand binding does not have a negative allosteric or inhibitory effect on assembly of the holoenzyme complex and that inhibitors function through direct competition with ATP. Overall, the structures described in this study should greatly assist future efforts in rational drug design targeting DNA-PKcs, demonstrating the potential of cryo-EM in structure-guided drug development for large and challenging targets. | ||||||
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 | 7otm.cif.gz | 650.5 KB | 表示 | PDBx/mmCIF形式 |
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PDB形式 | pdb7otm.ent.gz | 522.9 KB | 表示 | PDB形式 |
PDBx/mmJSON形式 | 7otm.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/ot/7otm ftp://data.pdbj.org/pub/pdb/validation_reports/ot/7otm | 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: 471375.406 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) 参照: UniProt: P78527, non-specific serine/threonine protein kinase |
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#2: 化合物 | ChemComp-2R4 / |
研究の焦点であるリガンドがあるか | Y |
-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: DNA-PKcs in complex with NU7441 / タイプ: COMPLEX / Entity ID: #1 / 由来: NATURAL |
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由来(天然) | 生物種: Homo sapiens (ヒト) |
緩衝液 | pH: 7.6 |
試料 | 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES |
急速凍結 | 凍結剤: ETHANE |
-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 |
撮影 | 電子線照射量: 47.3 e/Å2 / フィルム・検出器のモデル: GATAN K3 (6k x 4k) |
-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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CTF補正 | タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3次元再構成 | 解像度: 3.33 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 115505 / 対称性のタイプ: POINT | ||||||||||||||||||||||||
拘束条件 |
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