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- EMDB-20812: Cryo-EM structure of mammalian Ric-8A:Galpha(i):nanobody complex -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-20812 | ||||||||||||
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Title | Cryo-EM structure of mammalian Ric-8A:Galpha(i):nanobody complex | ||||||||||||
![]() | mammalian Ric-8A:Galpha(i):nanobody complex | ||||||||||||
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Function / homology | ![]() cell-cell adhesion involved in gastrulation / cell migration involved in gastrulation / Extra-nuclear estrogen signaling / Adenylate cyclase inhibitory pathway / basement membrane organization / vasculature development / Adrenaline,noradrenaline inhibits insulin secretion / ADP signalling through P2Y purinoceptor 12 / GTPase activating protein binding / negative regulation of synaptic transmission ...cell-cell adhesion involved in gastrulation / cell migration involved in gastrulation / Extra-nuclear estrogen signaling / Adenylate cyclase inhibitory pathway / basement membrane organization / vasculature development / Adrenaline,noradrenaline inhibits insulin secretion / ADP signalling through P2Y purinoceptor 12 / GTPase activating protein binding / negative regulation of synaptic transmission / G alpha (i) signalling events / G-protein alpha-subunit binding / positive regulation of protein localization to cell cortex / regulation of cAMP-mediated signaling / D2 dopamine receptor binding / G protein-coupled serotonin receptor binding / regulation of mitotic spindle organization / cellular response to forskolin / adenylate cyclase-inhibiting G protein-coupled receptor signaling pathway / guanyl-nucleotide exchange factor activity / G protein-coupled receptor binding / G-protein beta/gamma-subunit complex binding / visual learning / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / GDP binding / heterotrimeric G-protein complex / cell cortex / midbody / in utero embryonic development / G protein-coupled receptor signaling pathway / cell cycle / cell division / GTPase activity / centrosome / GTP binding / magnesium ion binding / protein-containing complex / nucleus / plasma membrane / cytoplasm Similarity search - Function | ||||||||||||
Biological species | ![]() ![]() ![]() ![]() | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.87 Å | ||||||||||||
![]() | Mou TC / Zhang K / Johnston JD / Chiu W / Sprang SR | ||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Structure of the G protein chaperone and guanine nucleotide exchange factor Ric-8A bound to Gαi1. Authors: Levi J McClelland / Kaiming Zhang / Tung-Chung Mou / Jake Johnston / Cindee Yates-Hansen / Shanshan Li / Celestine J Thomas / Tzanko I Doukov / Sarah Triest / Alexandre Wohlkonig / Gregory G ...Authors: Levi J McClelland / Kaiming Zhang / Tung-Chung Mou / Jake Johnston / Cindee Yates-Hansen / Shanshan Li / Celestine J Thomas / Tzanko I Doukov / Sarah Triest / Alexandre Wohlkonig / Gregory G Tall / Jan Steyaert / Wah Chiu / Stephen R Sprang / ![]() ![]() Abstract: Ric-8A is a cytosolic Guanine Nucleotide exchange Factor (GEF) that activates heterotrimeric G protein alpha subunits (Gα) and serves as an essential Gα chaperone. Mechanisms by which Ric-8A ...Ric-8A is a cytosolic Guanine Nucleotide exchange Factor (GEF) that activates heterotrimeric G protein alpha subunits (Gα) and serves as an essential Gα chaperone. Mechanisms by which Ric-8A catalyzes these activities, which are stimulated by Casein Kinase II phosphorylation, are unknown. We report the structure of the nanobody-stabilized complex of nucleotide-free Gα bound to phosphorylated Ric-8A at near atomic resolution by cryo-electron microscopy and X-ray crystallography. The mechanism of Ric-8A GEF activity differs considerably from that employed by G protein-coupled receptors at the plasma membrane. Ric-8A engages a specific conformation of Gα at multiple interfaces to form a complex that is stabilized by phosphorylation within a Ric-8A segment that connects two Gα binding sites. The C-terminus of Gα is ejected from its beta sheet core, thereby dismantling the GDP binding site. Ric-8A binds to the exposed Gα beta sheet and switch II to stabilize the nucleotide-free state of Gα. | ||||||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 39.6 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 21.1 KB 21.1 KB | Display Display | ![]() |
Images | ![]() | 260 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 430.2 KB | Display | ![]() |
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Full document | ![]() | 429.7 KB | Display | |
Data in XML | ![]() | 6.1 KB | Display | |
Data in CIF | ![]() | 6.9 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6uktMC ![]() 6tylC M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | mammalian Ric-8A:Galpha(i):nanobody complex | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.06 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : RIC-8A:Galpha(1):NB8109:NB8117:NB8119:NB9156 complex
Entire | Name: RIC-8A:Galpha(1):NB8109:NB8117:NB8119:NB9156 complex |
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Components |
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-Supramolecule #1: RIC-8A:Galpha(1):NB8109:NB8117:NB8119:NB9156 complex
Supramolecule | Name: RIC-8A:Galpha(1):NB8109:NB8117:NB8119:NB9156 complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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-Supramolecule #2: RIC-8A:Galpha(1)
Supramolecule | Name: RIC-8A:Galpha(1) / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#2 |
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Source (natural) | Organism: ![]() ![]() |
Recombinant expression | Organism: ![]() ![]() |
-Supramolecule #3: NB8109:NB8117:NB8119:NB9156
Supramolecule | Name: NB8109:NB8117:NB8119:NB9156 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #3-#6 |
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Source (natural) | Organism: ![]() ![]() |
Recombinant expression | Organism: ![]() ![]() |
-Macromolecule #1: Resistance to inhibitors of cholinesterase 8 homolog A (C. elegans)
Macromolecule | Name: Resistance to inhibitors of cholinesterase 8 homolog A (C. elegans) type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 55.836926 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: GMEPRAVADA LETGEEDAVT EALRSFNREH SQSFTFDDAQ QEDRKRLAKL LVSVLEQGLS PKHRVTWLQT IRILSRDRSC LDSFASRQS LHALACYADI AISEEPIPQP PDMDVLLESL KCLCNLVLSS PTAQMLAAEA RLVVRLAERV GLYRKRSYPH E VQFFDLRL ...String: GMEPRAVADA LETGEEDAVT EALRSFNREH SQSFTFDDAQ QEDRKRLAKL LVSVLEQGLS PKHRVTWLQT IRILSRDRSC LDSFASRQS LHALACYADI AISEEPIPQP PDMDVLLESL KCLCNLVLSS PTAQMLAAEA RLVVRLAERV GLYRKRSYPH E VQFFDLRL LFLLTALRTD VRQQLFQELH GVRLLTDALE LTLGVAPKEN PLVILPAQET ERAMEILKVL FNITFDSVKR EV DEEDAAL YRYLGTLLRH CVMADAAGDR TEEFHGHTVN LLGNLPLKCL DVLLALELHE GSLEFMGVNM DVINALLAFL EKR LHQTHR LKECVAPVLS VLTECARMHR PARKFLKAQV LPPLRDVRTR PEVGDLLRNK LVRLMTHLDT DVKRVAAEFL FVLC SESVP RFIKYTGYGN AAGLLAARGL MAGGRPEGQY (SEP)EDED(TPO)DTEE YREAKASINP VTGRVEEKPP NPMEGMT EE QKEHEAMKLV NMFDKLSR |
-Macromolecule #2: Guanine nucleotide-binding protein G(i) subunit alpha-1
Macromolecule | Name: Guanine nucleotide-binding protein G(i) subunit alpha-1 type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 37.015219 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: REVKLLLLGA GESGKSTIVK QMKIIHEAGY SEEECKQYKA VVYSNTIQSI IAIIRAMGRL KIDFGDAARA DDARQLFVLA GAAEEGFMT AELAGVIKRL WKDSGVQACF NRSREYQLND SAAYYLNDLD RIAQPNYIPT QQDVLRTRVK TTGIVETHFT F KDLHFKMF ...String: REVKLLLLGA GESGKSTIVK QMKIIHEAGY SEEECKQYKA VVYSNTIQSI IAIIRAMGRL KIDFGDAARA DDARQLFVLA GAAEEGFMT AELAGVIKRL WKDSGVQACF NRSREYQLND SAAYYLNDLD RIAQPNYIPT QQDVLRTRVK TTGIVETHFT F KDLHFKMF DVGGQRSERK KWIHCFEGVT AIIFCVALSD YDLVLAEDEE MNRMHESMKL FDSICNNKWF TDTSIILFLN KK DLFEEKI KKSPLTICYP EYAGSNTYEE AAAYIQCQFE DLNKRKDTKE IYTHFTCATD TKNVQFVFDA VTDVIIKNNL KDC GLF |
-Macromolecule #3: NB8109
Macromolecule | Name: NB8109 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 13.609938 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: QVQLQESGGG LVQPGGSLRL SCAASGIIFR SNGMAWYRQA PGKEREWVAS ITSFGDAIYR DSVKGRFTIS RDNARNAVSL QTNSLKTED TAVYYCNTYP VNSAWGQGTQ VTVSSHHHHH HEPEA |
-Macromolecule #4: NB8117
Macromolecule | Name: NB8117 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 14.8944 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: QVQLQESGGG LEQAGDSLRL SCAASGLIVS NYAMGWFRQA PGKEREFVAY INWNGGVTYY TNSVKGRFTI SRDNAKNTVY LQMNSLKPE DTAVYYCART SRASVTTRVA DFGYWGQGTQ VTVSSHHHHH HEPEA |
-Macromolecule #5: NB8119
Macromolecule | Name: NB8119 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 14.247603 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: QVQLQESGGG LVQAGGSLRL SCAASGGIVH ISSMGWFRQA PGKQRELVAT SPSNGDIRYA DSVKGRFTLS RDNAKNTVSL QMNSLEPED TAVYYCHSFL RHTASASYNN YYGQGTQVTV SSHHHHHHEP EA |
-Macromolecule #6: NB9156
Macromolecule | Name: NB9156 / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 14.993406 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: QVQLQESGGG LVQAGGSLRL SCAASVRTSD TDGMAWFRQA PGKEREFVGG IRWNSATWYA DFVKGRFTIS RDNAKNTLYL QMNSLKPED TALYYCARRA YGFDTDSRES AYSNWGQGTQ VTVSSHHHHH HEPEA |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.4 mg/mL | ||||||||||||
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Buffer | pH: 8 Component:
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Grid | Model: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: LACEY / Pretreatment - Type: GLOW DISCHARGE | ||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV | ||||||||||||
Details | monodisperse |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 11.5 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |