+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-12665 | |||||||||
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タイトル | Cryo-EM structure of pre-dephosphorylation complex of phosphorylated eIF2alpha with trapped holophosphatase (PP1A_D64A/PPP1R15A/G-actin/DNase I) | |||||||||
マップデータ | Unsharpened GS-FSC map after corrected FSC-mask auto-tightening from non-uniform refinement in CryoSPARC. The contour level was set when opend in UCSF Chimera. | |||||||||
試料 |
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機能・相同性 | 機能・相同性情報 fatty acid derivative binding / positive regulation of translational initiation in response to stress / : / regulation of translational initiation by eIF2 alpha dephosphorylation / regulation of neutrophil mediated cytotoxicity / regulation of acute inflammatory response / zymogen granule / : / protein phosphatase type 1 complex / : ...fatty acid derivative binding / positive regulation of translational initiation in response to stress / : / regulation of translational initiation by eIF2 alpha dephosphorylation / regulation of neutrophil mediated cytotoxicity / regulation of acute inflammatory response / zymogen granule / : / protein phosphatase type 1 complex / : / regulation of translation in response to endoplasmic reticulum stress / translation initiation ternary complex / glial limiting end-foot / deoxyribonuclease I / HRI-mediated signaling / response to kainic acid / Cellular response to mitochondrial stress / response to manganese-induced endoplasmic reticulum stress / PTW/PP1 phosphatase complex / positive regulation of type B pancreatic cell apoptotic process / negative regulation of translational initiation in response to stress / Response of EIF2AK1 (HRI) to heme deficiency / Recycling of eIF2:GDP / PERK-mediated unfolded protein response / peptidyl-serine dephosphorylation / PERK regulates gene expression / regulation of translational initiation in response to stress / eukaryotic translation initiation factor 2 complex / : / protein localization to endoplasmic reticulum / deoxyribonuclease I activity / neutrophil activation involved in immune response / protein phosphatase 1 binding / protein phosphatase regulator activity / eukaryotic 48S preinitiation complex / DNA catabolic process / Formation of the ternary complex, and subsequently, the 43S complex / cytoskeletal motor activator activity / Ribosomal scanning and start codon recognition / glycogen metabolic process / protein serine/threonine phosphatase activity / Translation initiation complex formation / myosin phosphatase activity / tropomyosin binding / protein-serine/threonine phosphatase / myosin heavy chain binding / negative regulation of PERK-mediated unfolded protein response / mesenchyme migration / detection of maltose stimulus / entrainment of circadian clock by photoperiod / troponin I binding / maltose transport complex / protein phosphatase activator activity / filamentous actin / actin filament bundle / phosphatase activity / carbohydrate transport / : / skeletal muscle thin filament assembly / actin filament bundle assembly / phosphoprotein phosphatase activity / striated muscle thin filament / maltose binding / transition metal ion binding / Response of EIF2AK4 (GCN2) to amino acid deficiency / maltose transport / intrinsic apoptotic signaling pathway in response to endoplasmic reticulum stress / skeletal muscle myofibril / maltodextrin transmembrane transport / actin monomer binding / carbohydrate transmembrane transporter activity / mitophagy / GTP hydrolysis and joining of the 60S ribosomal subunit / L13a-mediated translational silencing of Ceruloplasmin expression / ATP-binding cassette (ABC) transporter complex, substrate-binding subunit-containing / skeletal muscle fiber development / stress fiber / stress granule assembly / titin binding / response to endoplasmic reticulum stress / protein dephosphorylation / translation initiation factor activity / cellular response to amino acid starvation / actin filament polymerization / ATP-binding cassette (ABC) transporter complex / cell chemotaxis / Downregulation of TGF-beta receptor signaling / filopodium / actin filament / translational initiation / circadian regulation of gene expression / 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 / regulation of circadian rhythm / ABC-family proteins mediated transport / PKR-mediated signaling / cytoplasmic stress granule / calcium-dependent protein binding / cellular response to UV / positive regulation of canonical Wnt signaling pathway / lamellipodium 類似検索 - 分子機能 | |||||||||
生物種 | Homo sapiens (ヒト) / Oryctolagus cuniculus (ウサギ) / Rabbit (ウサギ) / Bovine (ウシ) / Escherichia coli (strain K12) (大腸菌) | |||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 3.96 Å | |||||||||
データ登録者 | Yan Y / Hardwick S / Ron D | |||||||||
資金援助 | 英国, 1件
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引用 | ジャーナル: Nat Struct Mol Biol / 年: 2021 タイトル: Higher-order phosphatase-substrate contacts terminate the integrated stress response. 著者: Yahui Yan / Heather P Harding / David Ron / 要旨: Many regulatory PPP1R subunits join few catalytic PP1c subunits to mediate phosphoserine and phosphothreonine dephosphorylation in metazoans. Regulatory subunits engage the surface of PP1c, locally ...Many regulatory PPP1R subunits join few catalytic PP1c subunits to mediate phosphoserine and phosphothreonine dephosphorylation in metazoans. Regulatory subunits engage the surface of PP1c, locally affecting flexible access of the phosphopeptide to the active site. However, catalytic efficiency of holophosphatases towards their phosphoprotein substrates remains unexplained. Here we present a cryo-EM structure of the tripartite PP1c-PPP1R15A-G-actin holophosphatase that terminates signaling in the mammalian integrated stress response (ISR) in the pre-dephosphorylation complex with its substrate, translation initiation factor 2α (eIF2α). G-actin, whose essential role in eIF2α dephosphorylation is supported crystallographically, biochemically and genetically, aligns the catalytic and regulatory subunits, creating a composite surface that engages the N-terminal domain of eIF2α to position the distant phosphoserine-51 at the active site. Substrate residues that mediate affinity for the holophosphatase also make critical contacts with eIF2α kinases. Thus, a convergent process of higher-order substrate recognition specifies functionally antagonistic phosphorylation and dephosphorylation in the ISR. | |||||||||
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 | EMマップ: SurfViewMolmilJmol/JSmol |
添付画像 |
-ダウンロードとリンク
-EMDBアーカイブ
マップデータ | emd_12665.map.gz | 73.8 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-12665-v30.xml emd-12665.xml | 27.7 KB 27.7 KB | 表示 表示 | EMDBヘッダ |
FSC (解像度算出) | emd_12665_fsc.xml | 15.7 KB | 表示 | FSCデータファイル |
画像 | emd_12665.png | 205.4 KB | ||
マスクデータ | emd_12665_msk_1.map | 149.9 MB | マスクマップ | |
その他 | emd_12665_additional_1.map.gz emd_12665_half_map_1.map.gz emd_12665_half_map_2.map.gz | 13.4 MB 139 MB 139 MB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-12665 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-12665 | HTTPS FTP |
-検証レポート
文書・要旨 | emd_12665_validation.pdf.gz | 516.9 KB | 表示 | EMDB検証レポート |
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文書・詳細版 | emd_12665_full_validation.pdf.gz | 516.5 KB | 表示 | |
XML形式データ | emd_12665_validation.xml.gz | 19.7 KB | 表示 | |
CIF形式データ | emd_12665_validation.cif.gz | 25.6 KB | 表示 | |
アーカイブディレクトリ | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-12665 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-12665 | HTTPS FTP |
-関連構造データ
-リンク
EMDBのページ | EMDB (EBI/PDBe) / EMDataResource |
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「今月の分子」の関連する項目 |
-マップ
ファイル | ダウンロード / ファイル: emd_12665.map.gz / 形式: CCP4 / 大きさ: 149.9 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | Unsharpened GS-FSC map after corrected FSC-mask auto-tightening from non-uniform refinement in CryoSPARC. The contour level was set when opend in UCSF Chimera. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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: 0.652 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
-マスク #1
ファイル | emd_12665_msk_1.map | ||||||||||||
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投影像・断面図 |
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密度ヒストグラム |
-追加マップ: ResolveCyroEM map; 1.81rmsd (0.443e/A^3) in COOT
ファイル | emd_12665_additional_1.map | ||||||||||||
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注釈 | ResolveCyroEM map; 1.81rmsd (0.443e/A^3) in COOT | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: half (A) map of the main map
ファイル | emd_12665_half_map_1.map | ||||||||||||
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注釈 | half (A) map of the main map | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: half (B) map of the main map
ファイル | emd_12665_half_map_2.map | ||||||||||||
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注釈 | half (B) map of the main map | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-試料の構成要素
-全体 : pre-dephosphorylation complex of phosphorylated eIF2alpha_2-187 w...
全体 | 名称: pre-dephosphorylation complex of phosphorylated eIF2alpha_2-187 with trapped holophosphatase (PP1A_D64A/PPP1R15A_553-624/G-actin) |
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要素 |
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-超分子 #1: pre-dephosphorylation complex of phosphorylated eIF2alpha_2-187 w...
超分子 | 名称: pre-dephosphorylation complex of phosphorylated eIF2alpha_2-187 with trapped holophosphatase (PP1A_D64A/PPP1R15A_553-624/G-actin) タイプ: complex / ID: 1 / 親要素: 0 / 含まれる分子: #1-#5 詳細: One copy of each component was present in the complex: phosphorylated eIF2alpha_2-187, PP1A_D64A, PPP1R15A_553-624, G-actin and DNase I. The full complex was purified by size exclusion chromatography. |
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分子量 | 実験値: 181 KDa |
-分子 #1: Eukaryotic translation initiation factor 2 subunit 1
分子 | 名称: Eukaryotic translation initiation factor 2 subunit 1 タイプ: protein_or_peptide / ID: 1 / コピー数: 1 / 光学異性体: LEVO |
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由来(天然) | 生物種: Homo sapiens (ヒト) |
分子量 | 理論値: 21.817863 KDa |
組換発現 | 生物種: Escherichia coli (大腸菌) |
配列 | 文字列: PGLSCRFYQH KFPEVEDVVM VNVRSIAEMG AYVSLLEYNN IEGMILLSEL (SEP)RRRIRSINK LIRIGRNECV VVIRVD KEK GYIDLSKRRV SPEEAIKCED KFTKSKTVYS ILRHVAEVLE YTKDEQLESL FQRTAWVFDD KYKRPGYGAY DAFKHAV SD ...文字列: PGLSCRFYQH KFPEVEDVVM VNVRSIAEMG AYVSLLEYNN IEGMILLSEL (SEP)RRRIRSINK LIRIGRNECV VVIRVD KEK GYIDLSKRRV SPEEAIKCED KFTKSKTVYS ILRHVAEVLE YTKDEQLESL FQRTAWVFDD KYKRPGYGAY DAFKHAV SD PSILDSLDLN EDEREVLINN INRRLTPQ |
-分子 #2: Serine/threonine-protein phosphatase PP1-alpha catalytic subunit
分子 | 名称: Serine/threonine-protein phosphatase PP1-alpha catalytic subunit タイプ: protein_or_peptide / ID: 2 / コピー数: 1 / 光学異性体: LEVO / EC番号: protein-serine/threonine phosphatase |
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由来(天然) | 生物種: Oryctolagus cuniculus (ウサギ) |
分子量 | 理論値: 33.647621 KDa |
組換発現 | 生物種: Escherichia coli (大腸菌) |
配列 | 文字列: LNLDSIIGRL LEVQGSRPGK NVQLTENEIR GLCLKSREIF LSQPILLELE APLKICGAIH GQYYDLLRLF EYGGFPPESN YLFLGDYVD RGKQSLETIC LLLAYKIKYP ENFFLLRGNH ECASINRIYG FYDECKRRYN IKLWKTFTDC FNCLPIAAIV D EKIFCCHG ...文字列: LNLDSIIGRL LEVQGSRPGK NVQLTENEIR GLCLKSREIF LSQPILLELE APLKICGAIH GQYYDLLRLF EYGGFPPESN YLFLGDYVD RGKQSLETIC LLLAYKIKYP ENFFLLRGNH ECASINRIYG FYDECKRRYN IKLWKTFTDC FNCLPIAAIV D EKIFCCHG GLSPDLQSME QIRRIMRPTD VPDQGLLCDL LWSDPDKDVQ GWGENDRGVS FTFGAEVVAK FLHKHDLDLI CR AHQVVED GYEFFAKRQL VTLFSAPNYC GEFDNAGAMM SVDETLMCSF QILKPAD |
-分子 #3: Actin, alpha skeletal muscle, intermediate form
分子 | 名称: Actin, alpha skeletal muscle, intermediate form / タイプ: protein_or_peptide / ID: 3 / コピー数: 1 / 光学異性体: LEVO |
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由来(天然) | 生物種: Rabbit (ウサギ) |
分子量 | 理論値: 41.862613 KDa |
配列 | 文字列: DEDETTALVC DNGSGLVKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IEHGIITNWD DMEKIWHHT FYNELRVAPE EHPTLLTEAP LNPKANREKM TQIMFETFNV PAMYVAIQAV LSLYASGRTT GIVLDSGDGV T HNVPIYEG ...文字列: DEDETTALVC DNGSGLVKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IEHGIITNWD DMEKIWHHT FYNELRVAPE EHPTLLTEAP LNPKANREKM TQIMFETFNV PAMYVAIQAV LSLYASGRTT GIVLDSGDGV T HNVPIYEG YALPHAIMRL DLAGRDLTDY LMKILTERGY SFVTTAEREI VRDIKEKLCY VALDFENEMA TAASSSSLEK SY ELPDGQV ITIGNERFRC PETLFQPSFI GMESAGIHET TYNSIMKCDI DIRKDLYANN VMSGGTTMYP GIADRMQKEI TAL APSTMK IKIIAPPERK YSVWIGGSIL ASLSTFQQMW ITKQEYDEAG PSIVHRKCF |
-分子 #4: Deoxyribonuclease-1
分子 | 名称: Deoxyribonuclease-1 / タイプ: protein_or_peptide / ID: 4 / コピー数: 1 / 光学異性体: LEVO / EC番号: deoxyribonuclease I |
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由来(天然) | 生物種: Bovine (ウシ) |
分子量 | 理論値: 29.092574 KDa |
配列 | 文字列: LKIAAFNIRT FGETKMSNAT LASYIVRIVR RYDIVLIQEV RDSHLVAVGK LLDYLNQDDP NTYHYVVSEP LGRNSYKERY LFLFRPNKV SVLDTYQYDD GCESCGNDSF SREPAVVKFS SHSTKVKEFA IVALHSAPSD AVAEINSLYD VYLDVQQKWH L NDVMLMGD ...文字列: LKIAAFNIRT FGETKMSNAT LASYIVRIVR RYDIVLIQEV RDSHLVAVGK LLDYLNQDDP NTYHYVVSEP LGRNSYKERY LFLFRPNKV SVLDTYQYDD GCESCGNDSF SREPAVVKFS SHSTKVKEFA IVALHSAPSD AVAEINSLYD VYLDVQQKWH L NDVMLMGD FNADCSYVTS SQWSSIRLRT SSTFQWLIPD SADTTATSTN CAYDRIVVAG SLLQSSVVPG SAAPFDFQAA YG LSNEMAL AISDHYPVEV TLT |
-分子 #5: Protein phosphatase 1 regulatory subunit 15A,Maltose/maltodextrin...
分子 | 名称: Protein phosphatase 1 regulatory subunit 15A,Maltose/maltodextrin-binding periplasmic protein タイプ: protein_or_peptide / ID: 5 / コピー数: 1 / 光学異性体: LEVO |
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由来(天然) | 生物種: Escherichia coli (strain K12) (大腸菌) / 株: K12 |
分子量 | 理論値: 49.169797 KDa |
組換発現 | 生物種: Escherichia coli (大腸菌) |
配列 | 文字列: ARKVRFSEKV TVHFLAVWAG PAQAARQGPW EQLARDRSRF ARRITQAQEE LSPCLTPAAR ARAWARLRNP PLQAKIEEGK LVIWINGDK GYNGLAEVGK KFEKDTGIKV TVEHPDKLEE KFPQVAATGD GPDIIFWAHD RFGGYAQSGL LAEITPDKAF Q DKLYPFTW ...文字列: ARKVRFSEKV TVHFLAVWAG PAQAARQGPW EQLARDRSRF ARRITQAQEE LSPCLTPAAR ARAWARLRNP PLQAKIEEGK LVIWINGDK GYNGLAEVGK KFEKDTGIKV TVEHPDKLEE KFPQVAATGD GPDIIFWAHD RFGGYAQSGL LAEITPDKAF Q DKLYPFTW DAVRYNGKLI AYPIAVEALS LIYNKDLLPN PPKTWEEIPA LDKELKAKGK SALMFNLQEP YFTWPLIAAD GG YAFKYEN GKYDIKDVGV DNAGAKAGLT FLVDLIKNKH MNADTDYSIA EAAFNKGETA MTINGPWAWS NIDTSKVNYG VTV LPTFKG QPSKPFVGVL SAGINAASPN KELAKEFLEN YLLTDEGLEA VNKDKPLGAV ALKSYEEELA KDPRIAATME NAQK GEIMP NIPQMSAFWY AVRTAVINAA SGRQTVDEAL KDAQTRITK |
-分子 #7: MANGANESE (II) ION
分子 | 名称: MANGANESE (II) ION / タイプ: ligand / ID: 7 / コピー数: 1 / 式: MN |
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分子量 | 理論値: 54.938 Da |
-分子 #8: ADENOSINE-5'-TRIPHOSPHATE
分子 | 名称: ADENOSINE-5'-TRIPHOSPHATE / タイプ: ligand / ID: 8 / コピー数: 1 / 式: ATP |
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分子量 | 理論値: 507.181 Da |
Chemical component information | ChemComp-ATP: |
-実験情報
-構造解析
手法 | クライオ電子顕微鏡法 |
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解析 | 単粒子再構成法 |
試料の集合状態 | particle |
-試料調製
濃度 | 5 mg/mL | ||||||||||||||||
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緩衝液 | pH: 7.4 構成要素:
詳細: 0.22mM Triton X-100 was added into the solution before plunging. | ||||||||||||||||
グリッド | モデル: UltrAuFoil R0.6/1 / 材質: GOLD / メッシュ: 300 / 前処理 - タイプ: GLOW DISCHARGE / 前処理 - 雰囲気: OTHER / 詳細: current 25mA at Pelco EasiGLOW | ||||||||||||||||
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 100 % / チャンバー内温度: 277 K / 装置: FEI VITROBOT MARK IV |
-電子顕微鏡法
顕微鏡 | FEI TITAN KRIOS |
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特殊光学系 | 位相板: VOLTA PHASE PLATE / エネルギーフィルター - 名称: GIF Bioquantum / エネルギーフィルター - スリット幅: 20 eV |
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: 4025 / 平均電子線量: 46.84 e/Å2 |
電子線 | 加速電圧: 300 kV / 電子線源: FIELD EMISSION GUN |
電子光学系 | C2レンズ絞り径: 50.0 µm / 照射モード: SPOT SCAN / 撮影モード: BRIGHT FIELD / Cs: 2.7 mm 最大 デフォーカス(公称値): -2.8000000000000003 µm 最小 デフォーカス(公称値): -1.0 µm / 倍率(公称値): 130000 |
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 |