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基本情報
登録情報 | ![]() | |||||||||
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タイトル | Structure of p110 gamma bound to the Ras inhibitory nanobody NB7 | |||||||||
![]() | human p110 gamma bound to Nanobody NB8 | |||||||||
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![]() | PI3K / p110 / PIK3CG / phosphoinositide 3-kinase / PIP3 / TRANSFERASE-IMMUNE SYSTEM complex | |||||||||
機能・相同性 | ![]() negative regulation of cardiac muscle contraction / negative regulation of triglyceride catabolic process / secretory granule localization / natural killer cell chemotaxis / neutrophil extravasation / phosphatidylinositol-4-phosphate 3-kinase / positive regulation of acute inflammatory response / respiratory burst involved in defense response / T cell chemotaxis / negative regulation of fibroblast apoptotic process ...negative regulation of cardiac muscle contraction / negative regulation of triglyceride catabolic process / secretory granule localization / natural killer cell chemotaxis / neutrophil extravasation / phosphatidylinositol-4-phosphate 3-kinase / positive regulation of acute inflammatory response / respiratory burst involved in defense response / T cell chemotaxis / negative regulation of fibroblast apoptotic process / phosphatidylinositol 3-kinase complex, class IB / regulation of calcium ion transmembrane transport / 1-phosphatidylinositol-4-phosphate 3-kinase activity / Co-stimulation by ICOS / sphingosine-1-phosphate receptor signaling pathway / phosphatidylinositol 3-kinase complex, class IA / phosphatidylinositol-3-phosphate biosynthetic process / 1-phosphatidylinositol-4,5-bisphosphate 3-kinase activity / phosphatidylinositol-4,5-bisphosphate 3-kinase / dendritic cell chemotaxis / phosphatidylinositol 3-kinase / 1-phosphatidylinositol-3-kinase activity / mast cell degranulation / Erythropoietin activates Phosphoinositide-3-kinase (PI3K) / hepatocyte apoptotic process / phosphatidylinositol-mediated signaling / regulation of cell adhesion mediated by integrin / phosphatidylinositol phosphate biosynthetic process / Synthesis of PIPs at the plasma membrane / positive regulation of MAP kinase activity / positive regulation of Rac protein signal transduction / CD28 dependent PI3K/Akt signaling / regulation of angiogenesis / T cell proliferation / GPVI-mediated activation cascade / ephrin receptor binding / neutrophil chemotaxis / positive regulation of endothelial cell migration / T cell activation / cellular response to cAMP / positive regulation of cytokine production / phosphatidylinositol 3-kinase/protein kinase B signal transduction / platelet aggregation / endocytosis / Constitutive Signaling by Aberrant PI3K in Cancer / G beta:gamma signalling through PI3Kgamma / cell migration / PIP3 activates AKT signaling / positive regulation of cytosolic calcium ion concentration / PI5P, PP2A and IER3 Regulate PI3K/AKT Signaling / angiogenesis / phospholipase C-activating G protein-coupled receptor signaling pathway / adaptive immune response / positive regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / non-specific serine/threonine protein kinase / protein kinase activity / immune response / G protein-coupled receptor signaling pathway / inflammatory response / innate immune response / protein serine kinase activity / protein serine/threonine kinase activity / ATP binding / identical protein binding / membrane / plasma membrane / 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: 2.96 Å | |||||||||
![]() | Burke JE / Nam SE / Rathinaswamy MK / Yip CK | |||||||||
資金援助 | ![]()
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![]() | ![]() タイトル: Allosteric activation or inhibition of PI3Kγ mediated through conformational changes in the p110γ helical domain. 著者: Noah J Harris / Meredith L Jenkins / Sung-Eun Nam / Manoj K Rathinaswamy / Matthew A H Parson / Harish Ranga-Prasad / Udit Dalwadi / Brandon E Moeller / Eleanor Sheeky / Scott D Hansen / ...著者: Noah J Harris / Meredith L Jenkins / Sung-Eun Nam / Manoj K Rathinaswamy / Matthew A H Parson / Harish Ranga-Prasad / Udit Dalwadi / Brandon E Moeller / Eleanor Sheeky / Scott D Hansen / Calvin K Yip / John E Burke / ![]() ![]() 要旨: PI3Kγ is a critical immune signaling enzyme activated downstream of diverse cell surface molecules, including Ras, PKCβ activated by the IgE receptor, and Gβγ subunits released from activated ...PI3Kγ is a critical immune signaling enzyme activated downstream of diverse cell surface molecules, including Ras, PKCβ activated by the IgE receptor, and Gβγ subunits released from activated GPCRs. PI3Kγ can form two distinct complexes, with the p110γ catalytic subunit binding to either a p101 or p84 regulatory subunit, with these complexes being differentially activated by upstream stimuli. Here, using a combination of cryo electron microscopy, HDX-MS, and biochemical assays, we have identified novel roles of the helical domain of p110γ in regulating lipid kinase activity of distinct PI3Kγ complexes. We defined the molecular basis for how an allosteric inhibitory nanobody potently inhibits kinase activity through rigidifying the helical domain and regulatory motif of the kinase domain. The nanobody did not block either p110γ membrane recruitment or Ras/Gβγ binding, but instead decreased ATP turnover. We also identified that p110γ can be activated by dual PKCβ helical domain phosphorylation leading to partial unfolding of an N-terminal region of the helical domain. PKCβ phosphorylation is selective for p110γ-p84 compared to p110γ-p101, driven by differential dynamics of the helical domain of these different complexes. Nanobody binding prevented PKCβ-mediated phosphorylation. Overall, this work shows an unexpected allosteric regulatory role of the helical domain of p110γ that is distinct between p110γ-p84 and p110γ-p101 and reveals how this can be modulated by either phosphorylation or allosteric inhibitory binding partners. This opens possibilities of future allosteric inhibitor development for therapeutic intervention. | |||||||||
履歴 |
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構造の表示
添付画像 |
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ダウンロードとリンク
-EMDBアーカイブ
マップデータ | ![]() | 188.7 MB | ![]() | |
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ヘッダ (付随情報) | ![]() ![]() | 18.3 KB 18.3 KB | 表示 表示 | ![]() |
画像 | ![]() | 70.5 KB | ||
Filedesc metadata | ![]() | 7.4 KB | ||
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-検証レポート
文書・要旨 | ![]() | 470.1 KB | 表示 | ![]() |
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文書・詳細版 | ![]() | 469.7 KB | 表示 | |
XML形式データ | ![]() | 7.2 KB | 表示 | |
CIF形式データ | ![]() | 8.3 KB | 表示 | |
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 | ![]() 8dp0MC 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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類似構造データ | 類似検索 - 機能・相同性 ![]() |
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リンク
EMDBのページ | ![]() ![]() |
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「今月の分子」の関連する項目 |
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マップ
ファイル | ![]() | ||||||||||||||||||||||||||||||||||||
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注釈 | human p110 gamma bound to Nanobody NB8 | ||||||||||||||||||||||||||||||||||||
投影像・断面図 | 画像のコントロール
画像は Spider により作成 | ||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 0.83 Å | ||||||||||||||||||||||||||||||||||||
密度 |
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対称性 | 空間群: 1 | ||||||||||||||||||||||||||||||||||||
詳細 | EMDB XML:
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-添付データ
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試料の構成要素
-全体 : Complex of p110 gamma with NB7
全体 | 名称: Complex of p110 gamma with NB7 |
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要素 |
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-超分子 #1: Complex of p110 gamma with NB7
超分子 | 名称: Complex of p110 gamma with NB7 / タイプ: complex / ID: 1 / 親要素: 0 / 含まれる分子: all |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 126.454 KDa |
-分子 #1: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit ...
分子 | 名称: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit gamma isoform タイプ: protein_or_peptide / ID: 1 / コピー数: 1 / 光学異性体: LEVO / EC番号: phosphatidylinositol 3-kinase |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 126.627406 KDa |
組換発現 | 生物種: ![]() ![]() |
配列 | 文字列: MELENYKQPV VLREDNCRRR RRMKPRSAAA SLSSMELIPI EFVLPTSQRK CKSPETALLH VAGHGNVEQM KAQVWLRALE TSVAADFYH RLGPHHFLLL YQKKGQWYEI YDKYQVVQTL DCLRYWKATH RSPGQIHLVQ RHPPSEESQA FQRQLTALIG Y DVTDVSNV ...文字列: MELENYKQPV VLREDNCRRR RRMKPRSAAA SLSSMELIPI EFVLPTSQRK CKSPETALLH VAGHGNVEQM KAQVWLRALE TSVAADFYH RLGPHHFLLL YQKKGQWYEI YDKYQVVQTL DCLRYWKATH RSPGQIHLVQ RHPPSEESQA FQRQLTALIG Y DVTDVSNV HDDELEFTRR GLVTPRMAEV ASRDPKLYAM HPWVTSKPLP EYLWKKIANN CIFIVIHRST TSQTIKVSPD DT PGAILQS FFTKMAKKKS LMDIPESQSE QDFVLRVCGR DEYLVGETPI KNFQWVRHCL KNGEEIHVVL DTPPDPALDE VRK EEWPLV DDCTGVTGYH EQLTIHGKDH ESVFTVSLWD CDRKFRVKIR GIDIPVLPRN TDLTVFVEAN IQHGQQVLCQ RRTS PKPFT EEVLWNVWLE FSIKIKDLPK GALLNLQIYC GKAPALSSKA SAESPSSESK GKVQLLYYVN LLLIDHRFLL RRGEY VLHM WQISGKGEDQ GSFNADKLTS ATNPDKENSM SISILLDNYC HPIALPKHQP TPDPEGDRVR AEMPNQLRKQ LEAIIA TDP LNPLTAEDKE LLWHFRYESL KHPKAYPKLF SSVKWGQQEI VAKTYQLLAR REVWDQSALD VGLTMQLLDC NFSDENV RA IAVQKLESLE DDDVLHYLLQ LVQAVKFEPY HDSALARFLL KRGLRNKRIG HFLFWFLRSE IAQSRHYQQR FAVILEAY L RGCGTAMLHD FTQQVQVIEM LQKVTLDIKS LSAEKYDVSS QVISQLKQKL ENLQNSQLPE SFRVPYDPGL KAGALAIEK CKVMASKKKP LWLEFKCADP TALSNETIGI IFKHGDDLRQ DMLILQILRI MESIWETESL DLCLLPYGCI STGDKIGMIE IVKDATTIA KIQQSTVGNT GAFKDEVLNH WLKEKSPTEE KFQAAVERFV YSCAGYCVAT FVLGIGDRHN DNIMITETGN L FHIDFGHI LGNYKSFLGI NKERVPFVLT PDFLFVMGTS GKKTSPHFQK FQDICVKAYL ALRHHTNLLI ILFSMMLMTG MP QLTSKED IEYIRDALTV GKNEEDAKKY FLDQIEVCRD KGWTVQFNWF LHLVLGIKQG EKHSA UniProtKB: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit gamma isoform |
-分子 #2: Nanobody NB7
分子 | 名称: Nanobody NB7 / タイプ: protein_or_peptide / ID: 2 / コピー数: 1 / 光学異性体: LEVO |
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由来(天然) | 生物種: ![]() ![]() |
分子量 | 理論値: 14.574995 KDa |
組換発現 | 生物種: ![]() ![]() |
配列 | 文字列: QVQLVESGGG LVQPGGSLRL SCAASGSIFS INAMGWYRQA PGKQRELVAH ITSGGSTNYA DSVKGRFTIS RDNAKNTVYL QMNSLKPED TAVYYCNEAG DPFLGSTWNG PPAFGSWGQG TQVTVSSHHH HHHEPEA |
-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
濃度 | 0.45 mg/mL | |||||||||||||||
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緩衝液 | pH: 8.5 構成要素:
詳細: Freshly prepared gel filtration buffer, filtered through 0.22 um filter and degassed | |||||||||||||||
グリッド | モデル: C-flat-2/2 / 材質: COPPER / メッシュ: 300 / 前処理 - タイプ: GLOW DISCHARGE / 前処理 - 時間: 25 sec. / 前処理 - 雰囲気: AIR / 前処理 - 気圧: 0.039 kPa | |||||||||||||||
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 100 % / チャンバー内温度: 277.15 K / 装置: FEI VITROBOT MARK IV 詳細: 3 uL of specimen was loaded onto grid and immediately blotted for 1.5 seconds with -5 force before plunging into liquid ethane.. | |||||||||||||||
詳細 | Specimen was p110 gamma purified with stabilizing nanobody, NB7, to homogeneity by gel filtration. |
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電子顕微鏡法
顕微鏡 | FEI TITAN KRIOS |
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撮影 | フィルム・検出器のモデル: GATAN K3 BIOQUANTUM (6k x 4k) 撮影したグリッド数: 1 / 実像数: 7344 / 平均電子線量: 50.0 e/Å2 |
電子線 | 加速電圧: 300 kV / 電子線源: ![]() |
電子光学系 | 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / 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): 0.5 µm |
試料ステージ | 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER ホルダー冷却材: NITROGEN |
実験機器 | ![]() モデル: Titan Krios / 画像提供: FEI Company |