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Yorodumi- EMDB-16249: Structure of Adenylyl cyclase 8 bound to stimulatory G protein, F... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-16249 | |||||||||
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Title | Structure of Adenylyl cyclase 8 bound to stimulatory G protein, Forskolin, ATPalphaS, and Ca2+/Calmodulin in lipid nanodisc | |||||||||
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Sample |
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Keywords | Adenylyl Cyclase / cAMP signaling / G proteins / Calmodulin / MEMBRANE PROTEIN | |||||||||
Function / homology | Function and homology information cellular response to morphine / Adenylate cyclase activating pathway / Adenylate cyclase inhibitory pathway / positive regulation of long-term synaptic depression / glucose mediated signaling pathway / calcium- and calmodulin-responsive adenylate cyclase activity / regulation of cellular response to stress / PKA activation / neuroinflammatory response / sensory perception of chemical stimulus ...cellular response to morphine / Adenylate cyclase activating pathway / Adenylate cyclase inhibitory pathway / positive regulation of long-term synaptic depression / glucose mediated signaling pathway / calcium- and calmodulin-responsive adenylate cyclase activity / regulation of cellular response to stress / PKA activation / neuroinflammatory response / sensory perception of chemical stimulus / adenylate cyclase / positive regulation of synaptic plasticity / Hedgehog 'off' state / cAMP biosynthetic process / adenylate cyclase activity / G alpha (z) signalling events / neuronal cell body membrane / positive regulation of ryanodine-sensitive calcium-release channel activity / cellular response to glucagon stimulus / presynaptic active zone / positive regulation of insulin secretion involved in cellular response to glucose stimulus / excitatory synapse / mu-type opioid receptor binding / corticotropin-releasing hormone receptor 1 binding / negative regulation of ryanodine-sensitive calcium-release channel activity / D1 dopamine receptor binding / long-term memory / regulation of release of sequestered calcium ion into cytosol by sarcoplasmic reticulum / beta-2 adrenergic receptor binding / adenylate cyclase-activating adrenergic receptor signaling pathway / regulation of cytosolic calcium ion concentration / adenylate cyclase activator activity / hippocampal mossy fiber to CA3 synapse / positive regulation of long-term synaptic potentiation / locomotory behavior / insulin-like growth factor receptor binding / ionotropic glutamate receptor binding / Schaffer collateral - CA1 synapse / G-protein beta/gamma-subunit complex binding / adenylate cyclase-activating G protein-coupled receptor signaling pathway / spindle pole / adenylate cyclase-activating dopamine receptor signaling pathway / heterotrimeric G-protein complex / presynaptic membrane / basolateral plasma membrane / postsynaptic density / intracellular signal transduction / apical plasma membrane / protein domain specific binding / axon / GTPase activity / glutamatergic synapse / calcium ion binding / dendrite / GTP binding / protein-containing complex / ATP binding / metal ion binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Bos taurus (cattle) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.97 Å | |||||||||
Authors | Khanppnavar B / Korkhov VM | |||||||||
Funding support | Switzerland, 2 items
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Citation | Journal: EMBO Rep / Year: 2024 Title: Regulatory sites of CaM-sensitive adenylyl cyclase AC8 revealed by cryo-EM and structural proteomics. Authors: Basavraj Khanppnavar / Dina Schuster / Pia Lavriha / Federico Uliana / Merve Özel / Ved Mehta / Alexander Leitner / Paola Picotti / Volodymyr M Korkhov / Abstract: Membrane adenylyl cyclase AC8 is regulated by G proteins and calmodulin (CaM), mediating the crosstalk between the cAMP pathway and Ca signalling. Despite the importance of AC8 in physiology, the ...Membrane adenylyl cyclase AC8 is regulated by G proteins and calmodulin (CaM), mediating the crosstalk between the cAMP pathway and Ca signalling. Despite the importance of AC8 in physiology, the structural basis of its regulation by G proteins and CaM is not well defined. Here, we report the 3.5 Å resolution cryo-EM structure of the bovine AC8 bound to the stimulatory Gαs protein in the presence of Ca/CaM. The structure reveals the architecture of the ordered AC8 domains bound to Gαs and the small molecule activator forskolin. The extracellular surface of AC8 features a negatively charged pocket, a potential site for unknown interactors. Despite the well-resolved forskolin density, the captured state of AC8 does not favour tight nucleotide binding. The structural proteomics approaches, limited proteolysis and crosslinking mass spectrometry (LiP-MS and XL-MS), allowed us to identify the contact sites between AC8 and its regulators, CaM, Gαs, and Gβγ, as well as to infer the conformational changes induced by these interactions. Our results provide a framework for understanding the role of flexible regions in the mechanism of AC regulation. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_16249.map.gz | 24.9 MB | EMDB map data format | |
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Header (meta data) | emd-16249-v30.xml emd-16249.xml | 18.4 KB 18.4 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_16249_fsc.xml | 15.7 KB | Display | FSC data file |
Images | emd_16249.png | 69.6 KB | ||
Masks | emd_16249_msk_1.map | 325 MB | Mask map | |
Filedesc metadata | emd-16249.cif.gz | 6.2 KB | ||
Others | emd_16249_half_map_1.map.gz emd_16249_half_map_2.map.gz | 260.8 MB 260.7 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-16249 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-16249 | HTTPS FTP |
-Validation report
Summary document | emd_16249_validation.pdf.gz | 836.8 KB | Display | EMDB validaton report |
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Full document | emd_16249_full_validation.pdf.gz | 836.4 KB | Display | |
Data in XML | emd_16249_validation.xml.gz | 23.1 KB | Display | |
Data in CIF | emd_16249_validation.cif.gz | 30.6 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-16249 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-16249 | HTTPS FTP |
-Related structure data
Related structure data | 8buzC 8bv5C C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_16249.map.gz / Format: CCP4 / Size: 325 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.66 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_16249_msk_1.map | ||||||||||||
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Density Histograms |
-Half map: half1
File | emd_16249_half_map_1.map | ||||||||||||
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Annotation | half1 | ||||||||||||
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Density Histograms |
-Half map: half2
File | emd_16249_half_map_2.map | ||||||||||||
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Annotation | half2 | ||||||||||||
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Density Histograms |
-Sample components
-Entire : Structure of Adenylyl cyclase 8 bound to stimulatory G protein, F...
Entire | Name: Structure of Adenylyl cyclase 8 bound to stimulatory G protein, Forskolin, ATPalphaS, and Ca2+/Calmodulin in lipid nanodisc |
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Components |
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-Supramolecule #1: Structure of Adenylyl cyclase 8 bound to stimulatory G protein, F...
Supramolecule | Name: Structure of Adenylyl cyclase 8 bound to stimulatory G protein, Forskolin, ATPalphaS, and Ca2+/Calmodulin in lipid nanodisc type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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-Supramolecule #2: Adenylyl cyclase 8 and G protein alpha S
Supramolecule | Name: Adenylyl cyclase 8 and G protein alpha S / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#2 |
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Source (natural) | Organism: Bos taurus (cattle) |
-Supramolecule #3: Calmodulin
Supramolecule | Name: Calmodulin / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #3 |
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Source (natural) | Organism: Bos taurus (cattle) |
-Macromolecule #1: Adenylyl cyclase 8
Macromolecule | Name: Adenylyl cyclase 8 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO / EC number: adenylate cyclase |
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Source (natural) | Organism: Bos taurus (cattle) |
Recombinant expression | Organism: Homo sapiens (human) |
Sequence | String: MELSDVRCLS GSEELYTIHP TPPAGDSGSG SRPQRLLWQT AVRHITEQRF IHGHRGGGGG GGNGSSSKA SDPGGGGPNH HHASQLSGDS ALPLYALGPG ERAHGTGGPK VFPERSGSGS A SGSGGGGD LGFLHLDCAP SNSDFFLNGG YSYRGVIFPT LRNSFKSRDL ...String: MELSDVRCLS GSEELYTIHP TPPAGDSGSG SRPQRLLWQT AVRHITEQRF IHGHRGGGGG GGNGSSSKA SDPGGGGPNH HHASQLSGDS ALPLYALGPG ERAHGTGGPK VFPERSGSGS A SGSGGGGD LGFLHLDCAP SNSDFFLNGG YSYRGVIFPT LRNSFKSRDL ERLYQRYFLG QR RKSEVVM NVLDVLTKLT LLVLHLSLAS APMDPLKGIL LGFFTGIEVV ICALVVVRKD TTS HTYLQY SGVVTWVAMT TQILAAGLGY GLLGDGIGYV LFTLFATYSM LPLPLTWAIL AGLG TSLMQ VVLQAVIPRL AVISINQVVA QAVLFMCMNT AGIFISYLSD RAQRQAFLET RRCVE ARLR LETENQRQER LVLSVLPRFV VLEMINDMTN VEDEHLQHQF HRIYIHRYEN VSILFA DVK GFTNLSTTLS AQELVRMLNE LFARFDRLAH EHHCLRIKIL GDCYYCVSGL PEPRQDH AH CCVEMGLSMI KTIRYVRSRT KHDVDMRIGI HSGSVLCGVL GLRKWQFDVW SWDVDIAN K LESGGIPGRI HISKATLDCL NGDYNVEEGH GKERNEFLRK HNIETYLIKQ PEESLLSLP EDIVKESVSS SDRRNSGATF TEGSWSPELP FDNIVGKQNT LAALTRNSIN LLPNHLAQAL HVQSGPEEI NKRIEHTIDL RSGDKLRREH IKPFSLMFKD SSLEHKYSQM RDEVFKSNLV C AFIVLLFI TAIQSLLPSS RVMPMAIQFS ILIMLHSALV LITTAEDYKC LPLVLRKTCC WI NETYLAR NVIIFASILI NFLGAILNIL WCDFDKSIPL KNLTFNSSAV FTDICSYPEY FVF TGVLAM VTCAVFLRLN SVLKLAVLLI MIAIYALLTE TIYAGLFLRY DNLNHSGEDF LGTK EASLL LMAMFLLAVF YHGQQLEYTA RLDFLWRVQA KEEINEMKEL REHNENMLRN ILPSH VARH FLEKDRDNEE LYSQSYDAVG VMFASIPGFA DFYSQTEMNN QGVECLRLLN EIIADF DEL LGEDRFQDIE KIKTIGSTYM AVSGLSPEKQ QCEDKWGHLC ALADFSLALT ESIQEIN KH SFNNFELRIG ISHGSVVAGV IGAKKPQYDI WGKTVNLASR MDSTGVSGRI QVPEETYL I LKDQGFAFDY RGEIYVKGIS EQEGKIKTYF LLGRVQPNPF ILPPRRLPGQ YSLAAVVLG LVQSLNRQRQ KQLLNENNNT GIIKGHYNRR TLLTPSGPEP GAPAEGADKP ELP UniProtKB: Adenylate cyclase type 8 |
-Macromolecule #2: G protein alpha S
Macromolecule | Name: G protein alpha S / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: Bos taurus (cattle) |
Recombinant expression | Organism: Trichoplusia ni (cabbage looper) |
Sequence | String: MGCLGNSKTE DQRNEEKAQR EANKKIEKQL QKDKQVYRAT HRLLLLGAGE SGKSTIVKQM RILHVNGFNG GEGGEEDPNA AKSNSDGEKA TKVQDIKNNL KEAIETIVAA MSNLVPPVEL ANPENQFRVD YILSVMNVPD FDFPPEFYEH AKALWEDEGV RACYERSNEY ...String: MGCLGNSKTE DQRNEEKAQR EANKKIEKQL QKDKQVYRAT HRLLLLGAGE SGKSTIVKQM RILHVNGFNG GEGGEEDPNA AKSNSDGEKA TKVQDIKNNL KEAIETIVAA MSNLVPPVEL ANPENQFRVD YILSVMNVPD FDFPPEFYEH AKALWEDEGV RACYERSNEY QLIDCAQYFL DKIDVIKQDD YVPSDQDLLR CRVLTSGIFE TKFQVDKVNF HMFDVGGQRD ERRKWIQCFN DVTAIIFVVA SSSYNMVIRE DNQTNRLQEA LNLFKSIWNN RWLRTISVIL FLNKQDLLAE KVLAGKSKIE DYFPEFARYT TPEDATPEPG EDPRVTRAKY FIRDEFLRIS TASGDGRHYC YPHFTCAVDT ENIRRVFNDC RDIIQRMHLR QYELLGGHHH HHHHH UniProtKB: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short |
-Macromolecule #3: Calmodulin
Macromolecule | Name: Calmodulin / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO |
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Source (natural) | Organism: Bos taurus (cattle) |
Sequence | String: MADQLTEEQI AEFKEAFSLF DKDGDGTITT KELGTVMRSL GQNPTEAELQ DMINEVDADG NGTIDFPEF LTMMARKMKD TDSEEEIREA FRVFDKDGNG YISAAELRHV MTNLGEKLTD E EVDEMIRE ADIDGDGQVN YEEFVQMMTA K UniProtKB: Calmodulin |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 5 mg/mL |
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Buffer | pH: 8 |
Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 55.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.6 µm |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |