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基本情報
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タイトル | CryoEM structure of beta-2-adrenergic receptor in complex with GTP-bound Gs heterotrimer (Class K) | |||||||||
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![]() | GPCR / Adrenergic / Receptor / G protein / SIGNALING PROTEIN | |||||||||
機能・相同性 | ![]() beta2-adrenergic receptor activity / positive regulation of mini excitatory postsynaptic potential / positive regulation of cAMP-dependent protein kinase activity / positive regulation of AMPA receptor activity / norepinephrine binding / positive regulation of autophagosome maturation / Adrenoceptors / heat generation / norepinephrine-epinephrine-mediated vasodilation involved in regulation of systemic arterial blood pressure / activation of transmembrane receptor protein tyrosine kinase activity ...beta2-adrenergic receptor activity / positive regulation of mini excitatory postsynaptic potential / positive regulation of cAMP-dependent protein kinase activity / positive regulation of AMPA receptor activity / norepinephrine binding / positive regulation of autophagosome maturation / Adrenoceptors / heat generation / norepinephrine-epinephrine-mediated vasodilation involved in regulation of systemic arterial blood pressure / activation of transmembrane receptor protein tyrosine kinase activity / negative regulation of smooth muscle contraction / positive regulation of lipophagy / response to psychosocial stress / negative regulation of multicellular organism growth / negative regulation of G protein-coupled receptor signaling pathway / endosome to lysosome transport / adrenergic receptor signaling pathway / diet induced thermogenesis / neuronal dense core vesicle / positive regulation of protein kinase A signaling / PKA activation in glucagon signalling / hair follicle placode formation / adenylate cyclase binding / smooth muscle contraction / developmental growth / D1 dopamine receptor binding / intracellular transport / potassium channel regulator activity / Hedgehog 'off' state / bone resorption / positive regulation of bone mineralization / adenylate cyclase-activating adrenergic receptor signaling pathway / activation of adenylate cyclase activity / brown fat cell differentiation / regulation of sodium ion transport / adenylate cyclase activator activity / response to cold / receptor-mediated endocytosis / trans-Golgi network membrane / clathrin-coated endocytic vesicle membrane / G-protein beta/gamma-subunit complex binding / bone development / Olfactory Signaling Pathway / positive regulation of protein serine/threonine kinase activity / Activation of the phototransduction cascade / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / G beta:gamma signalling through PLC beta / Presynaptic function of Kainate receptors / Thromboxane signalling through TP receptor / adenylate cyclase-activating G protein-coupled receptor signaling pathway / G protein-coupled acetylcholine receptor signaling pathway / G-protein activation / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / Prostacyclin signalling through prostacyclin receptor / Glucagon signaling in metabolic regulation / G beta:gamma signalling through CDC42 / cognition / platelet aggregation / ADP signalling through P2Y purinoceptor 12 / G beta:gamma signalling through BTK / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / Sensory perception of sweet, bitter, and umami (glutamate) taste / photoreceptor disc membrane / Adrenaline,noradrenaline inhibits insulin secretion / Glucagon-type ligand receptors / Vasopressin regulates renal water homeostasis via Aquaporins / G alpha (z) signalling events / cellular response to catecholamine stimulus / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / ADORA2B mediated anti-inflammatory cytokines production / sensory perception of taste / ADP signalling through P2Y purinoceptor 1 / adenylate cyclase-activating dopamine receptor signaling pathway / cellular response to amyloid-beta / G beta:gamma signalling through PI3Kgamma / cellular response to prostaglandin E stimulus / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / GPER1 signaling / Cargo recognition for clathrin-mediated endocytosis / G-protein beta-subunit binding / Inactivation, recovery and regulation of the phototransduction cascade / heterotrimeric G-protein complex / G alpha (12/13) signalling events / extracellular vesicle / sensory perception of smell / Clathrin-mediated endocytosis / signaling receptor complex adaptor activity / Thrombin signalling through proteinase activated receptors (PARs) / GTPase binding / retina development in camera-type eye / Ca2+ pathway / positive regulation of cold-induced thermogenesis / phospholipase C-activating G protein-coupled receptor signaling pathway / amyloid-beta binding / G alpha (i) signalling events / fibroblast proliferation / G alpha (s) signalling events / Ras protein signal transduction / G alpha (q) signalling events 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: 3.3 Å | |||||||||
![]() | Papasergi-Scott MM / Skiniotis G | |||||||||
資金援助 | ![]()
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![]() | ![]() タイトル: Time-resolved cryo-EM of G-protein activation by a GPCR. 著者: Makaía M Papasergi-Scott / Guillermo Pérez-Hernández / Hossein Batebi / Yang Gao / Gözde Eskici / Alpay B Seven / Ouliana Panova / Daniel Hilger / Marina Casiraghi / Feng He / Luis Maul / ...著者: Makaía M Papasergi-Scott / Guillermo Pérez-Hernández / Hossein Batebi / Yang Gao / Gözde Eskici / Alpay B Seven / Ouliana Panova / Daniel Hilger / Marina Casiraghi / Feng He / Luis Maul / Peter Gmeiner / Brian K Kobilka / Peter W Hildebrand / Georgios Skiniotis / ![]() ![]() ![]() 要旨: G-protein-coupled receptors (GPCRs) activate heterotrimeric G proteins by stimulating guanine nucleotide exchange in the Gα subunit. To visualize this mechanism, we developed a time-resolved cryo-EM ...G-protein-coupled receptors (GPCRs) activate heterotrimeric G proteins by stimulating guanine nucleotide exchange in the Gα subunit. To visualize this mechanism, we developed a time-resolved cryo-EM approach that examines the progression of ensembles of pre-steady-state intermediates of a GPCR-G-protein complex. By monitoring the transitions of the stimulatory G protein in complex with the β-adrenergic receptor at short sequential time points after GTP addition, we identified the conformational trajectory underlying G-protein activation and functional dissociation from the receptor. Twenty structures generated from sequential overlapping particle subsets along this trajectory, compared to control structures, provide a high-resolution description of the order of main events driving G-protein activation in response to GTP binding. Structural changes propagate from the nucleotide-binding pocket and extend through the GTPase domain, enacting alterations to Gα switch regions and the α5 helix that weaken the G-protein-receptor interface. Molecular dynamics simulations with late structures in the cryo-EM trajectory support that enhanced ordering of GTP on closure of the α-helical domain against the nucleotide-bound Ras-homology domain correlates with α5 helix destabilization and eventual dissociation of the G protein from the GPCR. These findings also highlight the potential of time-resolved cryo-EM as a tool for mechanistic dissection of GPCR signalling events. | |||||||||
履歴 |
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構造の表示
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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 | ![]() | 127.2 MB | ![]() | |
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ヘッダ (付随情報) | ![]() ![]() | 25.4 KB 25.4 KB | 表示 表示 | ![]() |
FSC (解像度算出) | ![]() | 10.8 KB | 表示 | ![]() |
画像 | ![]() | 104.8 KB | ||
マスクデータ | ![]() | 134.6 MB | ![]() | |
Filedesc metadata | ![]() | 7.7 KB | ||
その他 | ![]() ![]() ![]() | 67.3 MB 125 MB 125 MB | ||
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 | ![]() 8unvMC ![]() 8gdzC ![]() 8ge1C ![]() 8ge2C ![]() 8ge3C ![]() 8ge4C ![]() 8ge5C ![]() 8ge6C ![]() 8ge7C ![]() 8ge8C ![]() 8ge9C ![]() 8geaC ![]() 8gebC ![]() 8gecC ![]() 8gedC ![]() 8geeC ![]() 8gefC ![]() 8gegC ![]() 8gehC ![]() 8geiC ![]() 8gejC ![]() 8gfvC ![]() 8gfwC ![]() 8gfxC ![]() 8gfyC ![]() 8gfzC ![]() 8gg0C ![]() 8gg1C ![]() 8gg2C ![]() 8gg3C ![]() 8gg4C ![]() 8gg5C ![]() 8gg6C ![]() 8gg7C ![]() 8gg8C ![]() 8gg9C ![]() 8ggaC ![]() 8ggbC ![]() 8ggcC ![]() 8ggeC ![]() 8ggfC ![]() 8ggiC ![]() 8ggjC ![]() 8ggkC ![]() 8gglC ![]() 8ggmC ![]() 8ggnC ![]() 8ggoC ![]() 8ggpC ![]() 8ggqC ![]() 8ggrC ![]() 8ggsC ![]() 8ggtC ![]() 8gguC ![]() 8ggvC ![]() 8ggwC ![]() 8ggxC ![]() 8ggyC ![]() 8ggzC ![]() 8gh0C ![]() 8gh1C ![]() 8unlC ![]() 8unmC ![]() 8unnC ![]() 8unoC ![]() 8unpC ![]() 8unqC ![]() 8unrC ![]() 8unsC ![]() 8untC ![]() 8unuC ![]() 8unwC ![]() 8unxC ![]() 8unyC ![]() 8unzC ![]() 8uo0C ![]() 8uo1C ![]() 8uo2C ![]() 8uo3C ![]() 8uo4C 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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ボクセルのサイズ | X=Y=Z: 0.8677 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
-マスク #1
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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: #1
ファイル | emd_42418_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_42418_half_map_1.map | ||||||||||||
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密度ヒストグラム |
-ハーフマップ: #1
ファイル | emd_42418_half_map_2.map | ||||||||||||
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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 : Complex of beta-2 adrenergic receptor and Gs heterotrimer with GTP
全体 | 名称: Complex of beta-2 adrenergic receptor and Gs heterotrimer with GTP |
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要素 |
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-超分子 #1: Complex of beta-2 adrenergic receptor and Gs heterotrimer with GTP
超分子 | 名称: Complex of beta-2 adrenergic receptor and Gs heterotrimer with GTP タイプ: complex / ID: 1 / 親要素: 0 / 含まれる分子: #1-#4 詳細: Combined datasets of complex plunge frozen at 5 sec, 10 sec, or 17 sec after GTP addition to the sample. |
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由来(天然) | 生物種: ![]() |
-分子 #1: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
分子 | 名称: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short 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: 44.32616 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: MGCLGNSKTE DQRNEEKAQR EANKKIEKQL QKDKQVYRAT HRLLLLGAGE SGKSTIVKQM RILHVNGFNG DSEKATKVQD IKNNLKEAI ETIVAAMSNL VPPVELANPE NQFRVDYILS VMNVPDFDFP PEFYEHAKAL WEDEGVRACY ERSNEYQLID C AQYFLDKI ...文字列: MGCLGNSKTE DQRNEEKAQR EANKKIEKQL QKDKQVYRAT HRLLLLGAGE SGKSTIVKQM RILHVNGFNG DSEKATKVQD IKNNLKEAI ETIVAAMSNL VPPVELANPE NQFRVDYILS VMNVPDFDFP PEFYEHAKAL WEDEGVRACY ERSNEYQLID C AQYFLDKI DVIKQADYVP SDQDLLRCRV LTSGIFETKF QVDKVNFHMF DVGGQRDERR KWIQCFNDVT AIIFVVASSS YN MVIREDN QTNRLQEALN LFKSIWNNRW LRTISVILFL NKQDLLAEKV LAGKSKIEDY FPEFARYTTP EDATPEPGED PRV TRAKYF IRDEFLRIST ASGDGRHYCY PHFTCAVDTE NIRRVFNDCR DIIQRMHLRQ YELL UniProtKB: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short |
-分子 #2: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
分子 | 名称: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 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: 37.573988 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: GSSGSELDQL RQEAEQLKNQ IRDARKACAD ATLSQITNNI DPVGRIQMRT RRTLRGHLAK IYAMHWGTDS RLLVSASQDG KLIIWDSYT TNKVHAIPLR SSWVMTCAYA PSGNYVACGG LDNICSIYNL KTREGNVRVS RELAGHTGYL SCCRFLDDNQ I VTSSGDTT ...文字列: GSSGSELDQL RQEAEQLKNQ IRDARKACAD ATLSQITNNI DPVGRIQMRT RRTLRGHLAK IYAMHWGTDS RLLVSASQDG KLIIWDSYT TNKVHAIPLR SSWVMTCAYA PSGNYVACGG LDNICSIYNL KTREGNVRVS RELAGHTGYL SCCRFLDDNQ I VTSSGDTT CALWDIETGQ QTTTFTGHTG DVMSLSLAPD TRLFVSGACD ASAKLWDVRE GMCRQTFTGH ESDINAICFF PN GNAFATG SDDATCRLFD LRADQELMTY SHDNIICGIT SVSFSKSGRL LLAGYDDFNC NVWDALKADR AGVLAGHDNR VSC LGVTDD GMAVATGSWD SFLKIWN UniProtKB: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 |
-分子 #3: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
分子 | 名称: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 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: 7.861143 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: MASNNTASIA QARKLVEQLK MEANIDRIKV SKAAADLMAY CEAHAKEDPL LTPVPASENP FREKKFFCAI L UniProtKB: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 |
-分子 #4: Beta-2 adrenergic receptor
分子 | 名称: Beta-2 adrenergic receptor / タイプ: protein_or_peptide / ID: 4 / 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: 51.767242 KDa |
組換発現 | 生物種: ![]() ![]() |
配列 | 文字列: MKTIIALSYI FCLVFADYKD DDDAMGQPGN GSAFLLAPNR SHAPDHDVEN LYFQGTQQRD EVWVVGMGIV MSLIVLAIVF GNVLVITAI AKFERLQTVT NYFITSLACA DLVMGLAVVP FGAAHILTKT WTFGNFWCEF WTSIDVLCVT ASIETLCVIA V DRYFAITS ...文字列: MKTIIALSYI FCLVFADYKD DDDAMGQPGN GSAFLLAPNR SHAPDHDVEN LYFQGTQQRD EVWVVGMGIV MSLIVLAIVF GNVLVITAI AKFERLQTVT NYFITSLACA DLVMGLAVVP FGAAHILTKT WTFGNFWCEF WTSIDVLCVT ASIETLCVIA V DRYFAITS PFKYQSLLTK NKARVIILMV WIVSGLTSFL PIQMHWYRAT HQEAINCYAE ETCCDFFTNQ AYAIASSIVS FY VPLVIMV FVYSRVFQEA KRQLQKIDKS EGRFHVQNLS QVEQDGRTGH GLRRSSKFCL KEHKALKTLG IIMGTFTLCW LPF FIVNIV HVIQDNLIRK EVYILLNWIG YVNSGFNPLI YCRSPDFRIA FQELLCLRRS SLKAYGNGYS SNGNTGEQSG LEVL FQGPY HVEQEKENKL LAEDLPGTED FVGHQGTVPS DNIDSQGRNA STNDSLLETS QVAPA UniProtKB: Beta-2 adrenergic receptor |
-分子 #5: GUANOSINE-5'-TRIPHOSPHATE
分子 | 名称: GUANOSINE-5'-TRIPHOSPHATE / タイプ: ligand / ID: 5 / コピー数: 1 / 式: GTP |
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分子量 | 理論値: 523.18 Da |
Chemical component information | ![]() ChemComp-GTP: |
-分子 #6: (5R,6R)-6-(methylamino)-5,6,7,8-tetrahydronaphthalene-1,2,5-triol
分子 | 名称: (5R,6R)-6-(methylamino)-5,6,7,8-tetrahydronaphthalene-1,2,5-triol タイプ: ligand / ID: 6 / コピー数: 1 / 式: G1I |
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分子量 | 理論値: 209.242 Da |
Chemical component information | ![]() ChemComp-G1I: |
-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: 7.5 詳細: GTP was added just prior to freezing at 5 sec, 10 sec, or 17 sec before plunging. |
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グリッド | 材質: GOLD / 支持フィルム - 材質: GOLD / 支持フィルム - トポロジー: HOLEY |
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: GTP was added just prior to freezing at 5 sec, 10 sec, or 17 sec before plunging.. |
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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 (6k x 4k) / 検出モード: COUNTING / 平均電子線量: 50.0 e/Å2 詳細: The cryoEM map is the result of combining multiple datasets: cryo-EM imaging of the beta-2AR-Gs + GTP (5 sec) complex was performed on a Titan Krios electron microscope equipped with a K3 ...詳細: The cryoEM map is the result of combining multiple datasets: cryo-EM imaging of the beta-2AR-Gs + GTP (5 sec) complex was performed on a Titan Krios electron microscope equipped with a K3 Summit direct electron detector (Gatan). The microscope was operated at 300 kV accelerating voltage, with a nominal magnification of 105,000x in counting mode resulting in a magnified pixel size of 0.8677 Angstrom. A total exposure of 60.48 electrons/ Angstrom^2 over 63 frames with defocus ranging from -1.0 - -2.0 micrometers was used. Cryo-EM imaging of beta-2AR-Gs + GTP (10 sec) complex was performed on four separate grids over three collection sessions. The microscope was operated at 300 kV accelerating voltage, with a magnification at the camera of 58,679x in counting mode resulting in a magnified pixel size of 0.8521 Angstrom. For the first and second grids, movies were obtained at an exposure rate of 21.13 electrons/Angstrum^2/sec with defocus ranging from -0.4 - -2.0 micrometers. The total exposure time was 2.717 sec over 77 frames per movie stack. For an additional collection of the first grid, movies were obtained at an exposure rate of 20.95 electrons/ Angstrum^2/sec with defocus ranging from -0.4 -2.0 micrometers. The total exposure time was 2.717 sec over 77 frames per movie stack. For the third and fourth grids, movies were obtained at an exposure rate of 30.71 electrons/ Angstrum^2/sec with defocus ranging from -0.5 - -1.6 micrometers. The total exposure time was 2.008 sec over 79 frames per movie stack. Cryo-EM imaging of beta-2AR-Gs + GTP (17 sec) was performed on a Titan Krios equipped with a post-column energy filter, with a magnification of 105,000x in counting mode resulting in a magnified pixel size of 0.8677 Angstrom. Movies were obtained at an exposure rate of 32.46 electrons/Angstrum^2/sec with defocus ranging from -0.4 - -0.9 micrometers. The total exposure time was 1.999 sec over 79 frames per movie stack. |
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): 2.0 µm / 最小 デフォーカス(公称値): 0.4 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 |