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- EMDB-4723: Soluble domain of membrane adenylyl cyclase bound to an activated... -

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Basic information

Entry
Database: EMDB / ID: EMD-4723
TitleSoluble domain of membrane adenylyl cyclase bound to an activated stimulatory G protein, in the presence of MANT-GTP (SOL-M)
Map data
SampleSoluble domain of adenylyl cyclase AC9 bound to GalphaS, in the presence of MANT-GTP (SOL-M)
  • Adenylate cyclase 9Adenylyl cyclase
  • Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
  • (nucleic-acidNucleic acid) x 2
SourceBos taurus (cattle)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsQi C / Sorrentino S / Medalia O / Korkhov VM
CitationJournal: Science / Year: 2019
Title: The structure of a membrane adenylyl cyclase bound to an activated stimulatory G protein.
Authors: Chao Qi / Simona Sorrentino / Ohad Medalia / Volodymyr M Korkhov /
Abstract: Membrane-integral adenylyl cyclases (ACs) are key enzymes in mammalian heterotrimeric GTP-binding protein (G protein)-dependent signal transduction, which is important in many cellular processes. ...Membrane-integral adenylyl cyclases (ACs) are key enzymes in mammalian heterotrimeric GTP-binding protein (G protein)-dependent signal transduction, which is important in many cellular processes. Signals received by the G protein-coupled receptors are conveyed to ACs through G proteins to modulate the levels of cellular cyclic adenosine monophosphate (cAMP). Here, we describe the cryo-electron microscopy structure of the bovine membrane AC9 bound to an activated G protein αs subunit at 3.4-angstrom resolution. The structure reveals the organization of the membrane domain and helical domain that spans between the membrane and catalytic domains of AC9. The carboxyl-terminal extension of the catalytic domain occludes both the catalytic and the allosteric sites of AC9, inducing a conformation distinct from the substrate- and activator-bound state, suggesting a regulatory role in cAMP production.
DateDeposition: Mar 22, 2019 / Header (metadata) release: May 8, 2019 / Map release: May 8, 2019 / Update: May 8, 2019

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.042
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.042
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_4723.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.09 Å/pix.
x 300 pix.
= 326.7 Å
1.09 Å/pix.
x 300 pix.
= 326.7 Å
1.09 Å/pix.
x 300 pix.
= 326.7 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.089 Å
Density
Contour LevelBy AUTHOR: 0.042 / Movie #1: 0.042
Minimum - Maximum-0.08217961 - 0.15531243
Average (Standard dev.)0.00012704714 (±0.0026195599)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 326.69998 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0891.0891.089
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z326.700326.700326.700
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-0.0820.1550.000

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Supplemental data

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Sample components

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Entire Soluble domain of adenylyl cyclase AC9 bound to GalphaS, in the p...

EntireName: Soluble domain of adenylyl cyclase AC9 bound to GalphaS, in the presence of MANT-GTP (SOL-M)
Number of components: 5

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Component #1: protein, Soluble domain of adenylyl cyclase AC9 bound to GalphaS,...

ProteinName: Soluble domain of adenylyl cyclase AC9 bound to GalphaS, in the presence of MANT-GTP (SOL-M)
Recombinant expression: No

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Component #2: protein, Adenylate cyclase 9

ProteinName: Adenylate cyclase 9Adenylyl cyclase / Recombinant expression: No
SourceSpecies: Bos taurus (cattle)
Source (engineered)Expression System: Homo sapiens (human) / Vector: pEZT-BM / Cell of expression system: HEK293F

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Component #3: protein, Guanine nucleotide-binding protein G(s) subunit alpha is...

ProteinName: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
Recombinant expression: No
SourceSpecies: Bos taurus (cattle)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper) / Vector: pFastbac / Cell of expression system: High FIve

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Component #4: nucleic-acid, Adenylyl cyclase AC9

nucleic acidName: Adenylyl cyclase AC9 / Class: OTHER / Structure: OTHER / Synthetic: No
Sequence:
MASPPHQQLL QHHSTEVSCD SSGDSNSVRV RINPKQPSSN SHPKHCKYSI SSSCSSSGDS GGVPRRMGAG GRLRRRKKLP QLFERASSRW WDPKFDSVNL EEACMERCFP QTQRRFRYAL FYIGFACLLW SIYFGVHMKS KLIVMVAPAL CFLVVCVGFF LFTFTKLYAR HYVWTSLVLT LLVFALTLAA QFQVLTPLSG RVDNFNHTRA ARPTDTCLSQ VGSFSMCIEV LFLLYTVMHL PLYLSLILGV AYSVLFETFG YHFQDEACFA SPGAEALHWE LLSRALLHLC IHAIGIHLFI MSQVRSRSTF LKVGQSIMHG KDLEVEKALK ERMIHSVMPR IIADDLMKQG DEESENSVKR HATSSPKNRK KKSSIQKAPI AFRPFKMQQI EEVSILFADI VGFTKMSANK SAHALVGLLN DLFGRFDRLC EETKCEKIST LGDCYYCVAG CPEPRADHAY CCIEMGLGMI RAIEQFCQEK KEMVNMRVGV HTGTVLCGIL GMRRFKFDVW SNDVNLANLM EQLGVAGKVH ISEATAKYLD DRYEMEDGKV TERLGQSVVA DQLKGLKTYL IAGQRAKESH CSCSEALLSG FEVLDGSRVS SGPRGQGTAS PGSVSDLAQT VKTFDNLKTC PSCGITFTPK PEAGAEGGAV QNGCQEEPKN SAKASGGPSS KTQNGLLSPP PEEKLTNSQT SLCEILQEKG RWAGVSLDQS ALLPLRFKNI REKTDAHFVD VIKEDSLMKD YFFKPPINQF SLNFLDPELE RAYRTSYQEE VVKSSPVRTF ASATFSSLLD VLLSTTVFLI LSITCFLRYG AASTPPPPAA LAVFGAALLL EILSLVVSVR MVFFLEDVMT CTKRLLEWIA GWLPRHFIGA ILVSLPALAV YSHVTSEFET NIHSTMFTGS AVLTAVVQYC NFCQLSSWMR SSLATVVGAG PLLLLLYVSL CPDSSTVISH LDAVQNFSST RKLCNASLPH DGRSPASLIG QEVILVFFLL LLLVWFLNRE FEVSYRLHYH GDVEADLHRT KIQSMRDQAD WLLRNIIPYH VAEQLKVSQT YSKNHDSGGV IFASIVNFSE FYEENYEGGK ECYRVLNELI GDFDELLSKP DYSSIEKIKT IGATYMAASG LNATQCRDGS HPQEHLQILF EFAKEMMRVV DDFNNNMLWF NFKLRVGFNH GPLTAGVIGT TKLLYDIWGD TVNIASRMDT TGVECRIQVS EESYRVLSKM GYEFDYRGTV NVKGKGQMKT YLYPKCTDSG LVPQHQLSIS PDIRVQVDGS IGRSPTDEIA SLVPSVQNPD QVPPGSENNA QTRDAHPSAK RPWKEPVRAE ERCRFGKAIE KSDCEEVGME EANELTKLNV SERAAAALEV LFQGPGGVSK GEELFTGVVP ILVELDGDVN GHKFSVSGEG EGDATYGKLT LKFICTTGKL PVPWPTLVTT FGYGLQCFAR YPDHMKQHDF FKSAMPEGYV QERTIFFKDD GNYKTRAEVK FEGDTLVNRI ELKGIDFKED GNILGHKLEY NYNSHNVYIM ADKQKNGIKV NFKIRHNIED GSVQLADHYQ QNTPIGDGPV LLPDNHYLSY QSALSKDPNE KRDHMVLLEF VTAAGITLGM DELYKAASAW SHPQFEKGGG SGGGSGGSAW SHPQFEK

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Component #5: nucleic-acid, Guanine nucleotide-binding protein G(s) subunit alp...

nucleic acidName: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
Class: OTHER / Structure: OTHER / Synthetic: No
Sequence:
MGCLGNSKTE DQRNEEKAQR EANKKIEKQL QKDKQVYRAT HRLLLLGAGE SGKSTIVKQM RILHVNGFNG GEGGEEDPNA KSNSDGEKAT KVQDIKNNLK EAIETIVAAM SNLVPPVELA NPENQFRVDY ILSVMNVPDF DFPPEFYEHA KALWEDEGVR ACYERSNEYQ LIDCAQYFLD KIDVIKQDDY VPSDQDLLRC RVLTSGIFET KFQVDKVNFH MFDVGGQRDE RRKWIQCFND VTAIIFVVAS SSYNMVIRED NQTNRLQEAL NLFKSIWNNR WLRTISVILF LNKQDLLAEK VLAGKSKIED YFPEFARYTT PEDATPEPGE DPRVTRAKYF IRDEFLRIST ASGDGRHYCY PHFTCAVDTE NIRRVFNDCR DIIQRMHLRQ YELLGGHHHH HHHH
SourceSpecies: Bos taurus (cattle)

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionpH: 8
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 278 K / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 60 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD / Defocus: 1000.0 - 3000.0 nm
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 3233

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 53394
3D reconstructionSoftware: RELION / Resolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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Atomic model buiding

Modeling #1Target criteria: Correlation coefficient / Refinement space: REAL / Overall bvalue: 125.16

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