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基本情報
登録情報 | ![]() | |||||||||
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タイトル | Cryo-EM structure of LKB1-STRADalpha-MO25alpha heterocomplex | |||||||||
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![]() | serine/threonine kinase / pseudokinase / complex / TRANSFERASE | |||||||||
機能・相同性 | ![]() positive regulation of vesicle transport along microtubule / intracellular protein-containing complex / Golgi localization / LRR domain binding / negative regulation of potassium ion transmembrane transporter activity / AMPK inhibits chREBP transcriptional activation activity / negative regulation of potassium ion transmembrane transport / dendrite extension / negative regulation of epithelial cell proliferation involved in prostate gland development / regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction ...positive regulation of vesicle transport along microtubule / intracellular protein-containing complex / Golgi localization / LRR domain binding / negative regulation of potassium ion transmembrane transporter activity / AMPK inhibits chREBP transcriptional activation activity / negative regulation of potassium ion transmembrane transport / dendrite extension / negative regulation of epithelial cell proliferation involved in prostate gland development / regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / serine/threonine protein kinase complex / cellular hypotonic response / response to lipid / tissue homeostasis / epithelial cell proliferation involved in prostate gland development / response to thyroid hormone / Energy dependent regulation of mTOR by LKB1-AMPK / positive thymic T cell selection / vasculature development / regulation of Wnt signaling pathway / anoikis / response to glucagon / negative regulation of cold-induced thermogenesis / FOXO-mediated transcription of cell death genes / positive regulation of axonogenesis / establishment of cell polarity / cellular response to UV-B / regulation of dendrite morphogenesis / response to ionizing radiation / G1 to G0 transition / activation of protein kinase activity / protein kinase activator activity / positive regulation of transforming growth factor beta receptor signaling pathway / intrinsic apoptotic signaling pathway by p53 class mediator / protein localization to nucleus / positive regulation of autophagy / positive regulation of gluconeogenesis / negative regulation of TORC1 signaling / protein export from nucleus / protein dephosphorylation / protein serine/threonine kinase activator activity / axonogenesis / positive regulation of peptidyl-threonine phosphorylation / regulation of signal transduction by p53 class mediator / response to activity / regulation of cell growth / kinase binding / peptidyl-threonine phosphorylation / negative regulation of canonical Wnt signaling pathway / positive regulation of protein serine/threonine kinase activity / negative regulation of cell growth / p53 binding / Z disc / autophagy / positive regulation of protein localization to nucleus / glucose homeostasis / peptidyl-serine phosphorylation / T cell receptor signaling pathway / spermatogenesis / Regulation of TP53 Activity through Phosphorylation / secretory granule lumen / protein autophosphorylation / ficolin-1-rich granule lumen / regulation of cell cycle / non-specific serine/threonine protein kinase / intracellular signal transduction / protein phosphorylation / negative regulation of cell population proliferation / protein serine kinase activity / protein serine/threonine kinase activity / DNA damage response / Neutrophil degranulation / protein-containing complex binding / magnesium ion binding / signal transduction / mitochondrion / extracellular exosome / extracellular region / nucleoplasm / ATP binding / membrane / nucleus / cytoplasm / cytosol 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: 2.86 Å | |||||||||
![]() | Chan LM / Courteau BJ / Verba KA | |||||||||
資金援助 | 1件
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![]() | ![]() タイトル: High-resolution single-particle imaging at 100-200 keV with the Gatan Alpine direct electron detector. 著者: Lieza M Chan / Brandon J Courteau / Allison Maker / Mengyu Wu / Benjamin Basanta / Hev Mehmood / David Bulkley / David Joyce / Brian C Lee / Stephen Mick / Cory Czarnik / Sahil Gulati / ...著者: Lieza M Chan / Brandon J Courteau / Allison Maker / Mengyu Wu / Benjamin Basanta / Hev Mehmood / David Bulkley / David Joyce / Brian C Lee / Stephen Mick / Cory Czarnik / Sahil Gulati / Gabriel C Lander / Kliment A Verba / ![]() 要旨: Developments in direct electron detector technology have played a pivotal role in enabling high-resolution structural studies by cryo-EM at 200 and 300 keV. Yet, theory and recent experiments ...Developments in direct electron detector technology have played a pivotal role in enabling high-resolution structural studies by cryo-EM at 200 and 300 keV. Yet, theory and recent experiments indicate advantages to imaging at 100 keV, energies for which the current detectors have not been optimized. In this study, we evaluated the Gatan Alpine detector, designed for operation at 100 and 200 keV. Compared to the Gatan K3, Alpine demonstrated a significant DQE improvement at these energies, specifically a ∼ 4-fold improvement at Nyquist at 100 keV. In single-particle cryo-EM experiments, Alpine datasets yielded better than 2 Å resolution reconstructions of apoferritin at 120 and 200 keV on a ThermoFisher Scientific (TFS) Glacios microscope fitted with a non-standard SP-Twin lens. We also achieved a ∼ 3.2 Å resolution reconstruction of a 115 kDa asymmetric protein complex, proving Alpine's effectiveness with complex biological samples. In-depth analysis revealed that Alpine reconstructions are comparable to K3 reconstructions at 200 keV, and remarkably, reconstruction from Alpine at 120 keV on a TFS Glacios surpassed all but the 300 keV data from a TFS Titan Krios with GIF/K3. Additionally, we show Alpine's capability for high-resolution data acquisition and screening on lower-end systems by obtaining ∼ 3 Å resolution reconstructions of apoferritin and aldolase at 100 keV and detailed 2D averages of a 55 kDa sample using a side-entry cryo holder. Overall, we show that Gatan Alpine performs well with the standard 200 keV imaging systems and may potentially capture the benefits of lower accelerating voltages, bringing smaller sized particles within the scope of cryo-EM. | |||||||||
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FSC (解像度算出) | ![]() | 8.8 KB | 表示 | ![]() |
画像 | ![]() | 69.5 KB | ||
Filedesc metadata | ![]() | 7.1 KB | ||
その他 | ![]() ![]() ![]() | 31.9 MB 59.5 MB 59.5 MB | ||
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-検証レポート
文書・要旨 | ![]() | 889.1 KB | 表示 | ![]() |
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文書・詳細版 | ![]() | 888.7 KB | 表示 | |
XML形式データ | ![]() | 16.4 KB | 表示 | |
CIF形式データ | ![]() | 21.3 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 | ![]() 8vsuMC C: 同じ文献を引用 ( M: このマップから作成された原子モデル |
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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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ボクセルのサイズ | X=Y=Z: 0.835 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
-追加マップ: #1
ファイル | emd_43506_additional_1.map | ||||||||||||
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投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: #2
ファイル | emd_43506_half_map_1.map | ||||||||||||
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投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: #1
ファイル | emd_43506_half_map_2.map | ||||||||||||
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投影像・断面図 |
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密度ヒストグラム |
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試料の構成要素
-全体 : Heterotrimeric complex of serine/threonine kinase LKB1 with psued...
全体 | 名称: Heterotrimeric complex of serine/threonine kinase LKB1 with psuedokinase STRADa and scaffolding MO25a |
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要素 |
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-超分子 #1: Heterotrimeric complex of serine/threonine kinase LKB1 with psued...
超分子 | 名称: Heterotrimeric complex of serine/threonine kinase LKB1 with psuedokinase STRADa and scaffolding MO25a タイプ: complex / ID: 1 / 親要素: 0 / 含まれる分子: #1-#3 |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 140.5 KDa |
-分子 #1: Calcium-binding protein 39
分子 | 名称: Calcium-binding protein 39 / 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: ![]() |
分子量 | 理論値: 41.997109 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: MPFPFGKSHK SPADIVKNLK ESMAVLEKQD ISDKKAEKAT EEVSKNLVAM KEILYGTNEK EPQTEAVAQL AQELYNSGLL STLVADLQL IDFEGKKDVA QIFNNILRRQ IGTRTPTVEY ICTQQNILFM LLKGYESPEI ALNCGIMLRE CIRHEPLAKI I LWSEQFYD ...文字列: MPFPFGKSHK SPADIVKNLK ESMAVLEKQD ISDKKAEKAT EEVSKNLVAM KEILYGTNEK EPQTEAVAQL AQELYNSGLL STLVADLQL IDFEGKKDVA QIFNNILRRQ IGTRTPTVEY ICTQQNILFM LLKGYESPEI ALNCGIMLRE CIRHEPLAKI I LWSEQFYD FFRYVEMSTF DIASDAFATF KDLLTRHKLL SAEFLEQHYD RFFSEYEKLL HSENYVTKRQ SLKLLGELLL DR HNFTIMT KYISKPENLK LMMNLLRDKS RNIQFEAFHV FKVFVANPNK TQPILDILLK NQAKLIEFLS KFQNDRTEDE QFN DEKTYL VKQIRDLKRP AQQEAGSGAT NFSLLKQAGD VEENPG UniProtKB: Calcium-binding protein 39 |
-分子 #2: Serine/threonine-protein kinase STK11
分子 | 名称: Serine/threonine-protein kinase STK11 / タイプ: protein_or_peptide / ID: 2 / コピー数: 1 / 光学異性体: LEVO / EC番号: non-specific serine/threonine protein kinase |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 50.649785 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: PDYKDDDDKE NLYFQGMEVV DPQQLGMFTE GELMSVGMDT FIHRIDSTEV IYQPRRKRAK LIGKYLMGDL LGEGSYGKVK EVLDSETLC RRAVKILKKK KLRRIPNGEA NVKKEIQLLR RLRHKNVIQL VDVLYNEEKQ KMYMVMEYCV CGMQEMLDSV P EKRFPVCQ ...文字列: PDYKDDDDKE NLYFQGMEVV DPQQLGMFTE GELMSVGMDT FIHRIDSTEV IYQPRRKRAK LIGKYLMGDL LGEGSYGKVK EVLDSETLC RRAVKILKKK KLRRIPNGEA NVKKEIQLLR RLRHKNVIQL VDVLYNEEKQ KMYMVMEYCV CGMQEMLDSV P EKRFPVCQ AHGYFCQLID GLEYLHSQGI VHKDIKPGNL LLTTGGTLKI SDLGVAEALH PFAADDTCRT SQGSPAFQPP EI ANGLDTF SGFKVDIWSA GVTLYNITTG LYPFEGDNIY KLFENIGKGS YAIPGDCGPP LSDLLKGMLE YEPAKRFSIR QIR QHSWFR KKHPPAEAPV PIPPSPDTKD RWRSMTVVPY LEDLHGADED EDLFDIEDDI IYTQDFTVPG QVPEEEASHN GQRR GLPKA VCMNGTEAAQ LSTKSRAEGR APNPARKACS ASSKIRRLSA CKQQ UniProtKB: Serine/threonine-protein kinase STK11 |
-分子 #3: Isoform 3 of STE20-related kinase adapter protein alpha
分子 | 名称: Isoform 3 of STE20-related kinase adapter protein alpha 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: ![]() |
分子量 | 理論値: 47.861875 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: PHHHHHHENL YFQGMSFLTN DASSESIASF SKQEVMSSFL PEGGCYELLT VIGKGFEDLM TVNLARYKPT GEYVTVRRIN LEACSNEMV TFLQGELHVS KLFNHPNIVP YRATFIADNE LWVVTSFMAY GSAKDLICTH FMDGMNELAI AYILQGVLKA L DYIHHMGY ...文字列: PHHHHHHENL YFQGMSFLTN DASSESIASF SKQEVMSSFL PEGGCYELLT VIGKGFEDLM TVNLARYKPT GEYVTVRRIN LEACSNEMV TFLQGELHVS KLFNHPNIVP YRATFIADNE LWVVTSFMAY GSAKDLICTH FMDGMNELAI AYILQGVLKA L DYIHHMGY VHRSVKASHI LISVDGKVYL SGLRSNLSMI SHGQRQRVVH DFPKYSVKVL PWLSPEVLQQ NLQGYDAKSD IY SVGITAC ELANGHVPFK DMPATQMLLE KLNGTVPCLL DTSTIPAEEL TMSPSRSVAN SGLSDSLTTS TPRPSNGDSP SHP YHRTFS PHFHHFVEQC LQRNPDARPS ASTLLNHSFF KQIKRRASEA LPELLRPVTP ITNFEGSQSQ DHSGIFGLVT NLEE LEVDD WEFGSGATNF SLLKQAGDVE ENPG UniProtKB: STE20-related kinase adapter protein alpha |
-分子 #4: ADENOSINE-5'-DIPHOSPHATE
分子 | 名称: ADENOSINE-5'-DIPHOSPHATE / タイプ: ligand / ID: 4 / コピー数: 1 / 式: ADP |
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分子量 | 理論値: 427.201 Da |
Chemical component information | ![]() ChemComp-ADP: |
-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: 7.6 構成要素:
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グリッド | モデル: Quantifoil R1.2/1.3 / 材質: GOLD / メッシュ: 400 / 支持フィルム - 材質: CARBON / 支持フィルム - トポロジー: HOLEY / 前処理 - タイプ: GLOW DISCHARGE / 前処理 - 時間: 30 sec. / 前処理 - 雰囲気: AIR / 前処理 - 気圧: 0.033 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 K / 装置: FEI VITROBOT MARK III |
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電子顕微鏡法
顕微鏡 | TFS KRIOS |
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特殊光学系 | 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 (6k x 4k) / 撮影したグリッド数: 1 / 実像数: 1293 / 平均電子線量: 45.8 e/Å2 |
電子線 | 加速電圧: 300 kV / 電子線源: ![]() |
電子光学系 | C2レンズ絞り径: 70.0 µm / 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / Cs: 2.7 mm / 最大 デフォーカス(公称値): 2.0 µm / 最小 デフォーカス(公称値): 1.0 µm / 倍率(公称値): 105000 |
試料ステージ | 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER |
実験機器 | ![]() モデル: Titan Krios / 画像提供: FEI Company |
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画像解析
-原子モデル構築 1
初期モデル | PDB ID: Chain - Source name: PDB / Chain - Initial model type: experimental model |
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精密化 | プロトコル: RIGID BODY FIT |
得られたモデル | ![]() PDB-8vsu: |
-原子モデル構築 2
精密化 | プロトコル: FLEXIBLE FIT |
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得られたモデル | ![]() PDB-8vsu: |