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Yorodumi- EMDB-20308: Single Particle Reconstruction of Phosphatidylinositol (3,4,5) tr... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-20308 | ||||||||||||
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Title | Single Particle Reconstruction of Phosphatidylinositol (3,4,5) trisphosphate-dependent Rac exchanger 1 bound to G protein beta gamma subunits | ||||||||||||
Map data | |||||||||||||
Sample |
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Keywords | RhoGEF / G protein / Complex / Phosphatase fold / SIGNALING PROTEIN | ||||||||||||
Function / homology | Function and homology information regulation of signaling / regulation of dendrite development / Olfactory Signaling Pathway / Sensory perception of sweet, bitter, and umami (glutamate) taste / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / neutrophil activation / regulation of actin filament polymerization / Activation of the phototransduction cascade / regulation of small GTPase mediated signal transduction / Activation of G protein gated Potassium channels ...regulation of signaling / regulation of dendrite development / Olfactory Signaling Pathway / Sensory perception of sweet, bitter, and umami (glutamate) taste / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / neutrophil activation / regulation of actin filament polymerization / Activation of the phototransduction cascade / regulation of small GTPase mediated signal transduction / Activation of G protein gated Potassium channels / G-protein activation / G beta:gamma signalling through PI3Kgamma / Prostacyclin signalling through prostacyclin receptor / G beta:gamma signalling through PLC beta / ADP signalling through P2Y purinoceptor 1 / Thromboxane signalling through TP receptor / Presynaptic function of Kainate receptors / G beta:gamma signalling through CDC42 / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / Glucagon-type ligand receptors / Adrenaline,noradrenaline inhibits insulin secretion / G alpha (12/13) signalling events / G beta:gamma signalling through BTK / ADP signalling through P2Y purinoceptor 12 / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / Thrombin signalling through proteinase activated receptors (PARs) / Ca2+ pathway / G alpha (z) signalling events / Extra-nuclear estrogen signaling / G alpha (s) signalling events / RHOB GTPase cycle / G alpha (q) signalling events / NRAGE signals death through JNK / G alpha (i) signalling events / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / Vasopressin regulates renal water homeostasis via Aquaporins / superoxide metabolic process / RHOC GTPase cycle / RHOJ GTPase cycle / RHOQ GTPase cycle / CDC42 GTPase cycle / T cell differentiation / RHOG GTPase cycle / RHOA GTPase cycle / RAC2 GTPase cycle / RAC3 GTPase cycle / positive regulation of substrate adhesion-dependent cell spreading / RAC1 GTPase cycle / neutrophil chemotaxis / actin filament polymerization / GTPase activator activity / guanyl-nucleotide exchange factor activity / dendritic shaft / phospholipid binding / photoreceptor disc membrane / cellular response to catecholamine stimulus / adenylate cyclase-activating dopamine receptor signaling pathway / cellular response to prostaglandin E stimulus / G-protein beta-subunit binding / heterotrimeric G-protein complex / G alpha (12/13) signalling events / signaling receptor complex adaptor activity / growth cone / intracellular signal transduction / positive regulation of cell migration / G protein-coupled receptor signaling pathway / GTPase activity / perinuclear region of cytoplasm / enzyme binding / membrane / plasma membrane / cytosol / cytoplasm Similarity search - Function | ||||||||||||
Biological species | Homo sapiens (human) / Bos taurus (cattle) | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.2 Å | ||||||||||||
Authors | Cash JN / Cianfrocco MA | ||||||||||||
Funding support | United States, 3 items
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Citation | Journal: Sci Adv / Year: 2019 Title: Cryo-electron microscopy structure and analysis of the P-Rex1-Gβγ signaling scaffold. Authors: Jennifer N Cash / Sarah Urata / Sheng Li / Sandeep K Ravala / Larisa V Avramova / Michael D Shost / J Silvio Gutkind / John J G Tesmer / Michael A Cianfrocco / Abstract: PIP-dependent Rac exchanger 1 (P-Rex1) is activated downstream of G protein-coupled receptors to promote neutrophil migration and metastasis. The structure of more than half of the enzyme and its ...PIP-dependent Rac exchanger 1 (P-Rex1) is activated downstream of G protein-coupled receptors to promote neutrophil migration and metastasis. The structure of more than half of the enzyme and its regulatory G protein binding site are unknown. Our 3.2 Å cryo-EM structure of the P-Rex1-Gβγ complex reveals that the carboxyl-terminal half of P-Rex1 adopts a complex fold most similar to those of phosphoinositide phosphatases. Although catalytically inert, the domain coalesces with a DEP domain and two PDZ domains to form an extensive docking site for Gβγ. Hydrogen-deuterium exchange mass spectrometry suggests that Gβγ binding induces allosteric changes in P-Rex1, but functional assays indicate that membrane localization is also required for full activation. Thus, a multidomain assembly is key to the regulation of P-Rex1 by Gβγ and the formation of a membrane-localized scaffold optimized for recruitment of other signaling proteins such as PKA and PTEN. | ||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_20308.map.gz | 52.5 MB | EMDB map data format | |
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Header (meta data) | emd-20308-v30.xml emd-20308.xml | 27.5 KB 27.5 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_20308_fsc.xml | 12.5 KB | Display | FSC data file |
Images | emd_20308.png | 215.7 KB | ||
Masks | emd_20308_msk_1.map | 103 MB | Mask map | |
Filedesc metadata | emd-20308.cif.gz | 7.5 KB | ||
Others | emd_20308_additional_1.map.gz emd_20308_additional_2.map.gz emd_20308_half_map_1.map.gz emd_20308_half_map_2.map.gz | 97.2 MB 51.9 MB 95.5 MB 95.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-20308 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-20308 | HTTPS FTP |
-Validation report
Summary document | emd_20308_validation.pdf.gz | 1 MB | Display | EMDB validaton report |
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Full document | emd_20308_full_validation.pdf.gz | 1 MB | Display | |
Data in XML | emd_20308_validation.xml.gz | 15.6 KB | Display | |
Data in CIF | emd_20308_validation.cif.gz | 21 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-20308 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-20308 | HTTPS FTP |
-Related structure data
Related structure data | 6pcvMC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | |
EM raw data | EMPIAR-10285 (Title: Cryo-electron microscopy structure of the P-Rex1–G-beta-gamma signaling scaffold Data size: 3.0 TB Data #1: Movie files (.tif) for P-Rex1-Gbg [micrographs - multiframe] Data #2: Micrograph files (.mrc) & CTF log files for P-Rex1-Gbg [micrographs - single frame] Data #3: Extracted particles from Warp for P-Rex1-Gbg [picked particles - single frame - processed]) |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_20308.map.gz / Format: CCP4 / Size: 103 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: 1 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_20308_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: Sharpened map
File | emd_20308_additional_1.map | ||||||||||||
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Annotation | Sharpened map | ||||||||||||
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-Additional map: Unsharpened map
File | emd_20308_additional_2.map | ||||||||||||
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Annotation | Unsharpened map | ||||||||||||
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Density Histograms |
-Half map: #2
File | emd_20308_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_20308_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : P-Rex1 bound to G protein beta gamma subunits
Entire | Name: P-Rex1 bound to G protein beta gamma subunits |
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Components |
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-Supramolecule #1: P-Rex1 bound to G protein beta gamma subunits
Supramolecule | Name: P-Rex1 bound to G protein beta gamma subunits / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 9 KDa |
-Supramolecule #2: Phosphatidylinositol (3,4,5) trisphosphate-dependent Rac exchanger 1
Supramolecule | Name: Phosphatidylinositol (3,4,5) trisphosphate-dependent Rac exchanger 1 type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1 |
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-Supramolecule #3: G protein beta-1 subunit
Supramolecule | Name: G protein beta-1 subunit / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2 |
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-Supramolecule #4: G protein gamma-2 subunit
Supramolecule | Name: G protein gamma-2 subunit / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #3 |
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-Macromolecule #1: Phosphatidylinositol (3,4,5) trisphosphate-dependent Rac exchanger 1
Macromolecule | Name: Phosphatidylinositol (3,4,5) trisphosphate-dependent Rac exchanger 1 type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 184.840891 KDa |
Recombinant expression | Organism: Homo sapiens (human) |
Sequence | String: GEFAAARESE RQLRLRLCVL NEILGTERDY VGTLRFLQSA FLHRIRQNVA DSVEKGLTEE NVKVLFSNIE DILEVHKDFL AALEYCLHP EPQSQHELGN VFLKFKDKFC VYEEYCSNHE KALRLLVELN KIPTVRAFLL SCMLLGGRKT TDIPLEGYLL S PIQRICKY ...String: GEFAAARESE RQLRLRLCVL NEILGTERDY VGTLRFLQSA FLHRIRQNVA DSVEKGLTEE NVKVLFSNIE DILEVHKDFL AALEYCLHP EPQSQHELGN VFLKFKDKFC VYEEYCSNHE KALRLLVELN KIPTVRAFLL SCMLLGGRKT TDIPLEGYLL S PIQRICKY PLLLKELAKR TPGKHPDHPA VQSALQAMKT VCSNINETKR QMEKLEALEQ LQSHIEGWEG SNLTDICTQL LL QGTLLKI SAGNIQERAF FLFDNLLVYC KRKSRVTGSK KSTKRTKSIN GSLYIFRGRI NTEVMEVENV EDGTADYHSN GYT VTNGWK IHNTAKNKWF VCMAKTAEEK QKWLDAIIRE REQRESLKLG MERDAYVMIA EKGEKLYHMM MNKKVNLIKD RRRK LSTVP KCFLGNEFVA WLLEIGEISK TEEGVNLGQA LLENGIIHHV SDKHQFKNEQ VMYRFRYDDG TYKARSELED IMSKG VRLY CRLHSLYTPV IKDRDYHLKT YKSVLPGSKL VDWLLAQGDC QTREEAVALG VGLCNNGFMH HVLEKSEFRD ESQYFR FHA DEEMEGTSSK NKQLRNDFKL VENILAKRLL ILPQEEDYGF DIEEKNKAVV VKSVQRGSLA EVAGLQVGRK IYSINED LV FLRPFSEVES ILNQSFCSRR PLRLLVATKA KEIIKIPDQP DTLCFQIRGA APPYVYAVGR GSEAMAAGLC AGQCILKV N GSNVMNDGAP EVLEHFQAFR SRREEALGLY QWIYHTHEDA QEARASQEAS TEDPSGEQAQ EEDQADSAFP LLSLGPRLS LCEDSPMVTL TVDNVHLEHG VVYEYVSTAG VRCHVLEKIV EPRGCFGLTA KILEAFAAND SVFVENCRRL MALSSAIVTM PHFEFRNIC DTKLESIGQR IACYQEFAAQ LKSRVSPPFK QAPLEPHPLC GLDFCPTNCH INLMEVSYPK TTPSVGRSFS I RFGRKPSL IGLDPEQGHL NPMSYTQHCI TTMAAPSWKC LPAAEGDPQG QGLHDGSFGP ASGTLGQEDR GLSFLLKQED RE IQDAYLQ LFTKLDVALK EMKQYVTQIN RLLSTITEPT SGGSCDASLA EEASSLPLVS EESEMDRSDH GGIKKVCFKV AEE DQEDSG HDTMSYRDSY SECNSNRDSV LSYTSVRSNS SYLGSDEMGS GDELPCDMRI PSDKQDKLHG CLEHLFNQVD SINA LLKGP VMSRAFEETK HFPMNHSLQE FKQKEECTIR GRSLIQISIQ EDPWNLPNSI KTLVDNIQRY VEDGKNQLLL ALLKC TDTE LQLRRDAIFC QALVAAVCTF SEQLLAALGY RYNNNGEYEE SSRDASRKWL EQVAATGVLL HCQSLLSPAT VKEERT MLE DIWVTLSELD NVTFSFKQLD ENYVANTNVF YHIEGSRQAL KVIFYLDSYH FSKLPSRLEG GASLRLHTAL FTKVLEN VE GLPSPGSQAA EDLQQDINAQ SLEKVQQYYR KLRAFYLERS NLPTDASTTA VKIDQLIRPI NALDELCRLM KSFVHPKP G AAGSVGAGLI PISSELCYRL GACQMVMCGT GMQRSTLSVS LEQAAILARS HGLLPKCIMQ ATDIMRKQGP RVEILAKNL RVKDQMPQGA PRLYRLCQPP VDGDLHHHHH HHHHH UniProtKB: PREX1 |
-Macromolecule #2: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
Macromolecule | Name: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Bos taurus (cattle) |
Molecular weight | Theoretical: 37.41693 KDa |
Recombinant expression | Organism: Trichoplusia ni (cabbage looper) |
Sequence | String: MSELDQLRQE AEQLKNQIRD ARKACADATL SQITNNIDPV GRIQMRTRRT LRGHLAKIYA MHWGTDSRLL VSASQDGKLI IWDSYTTNK VHAIPLRSSW VMTCAYAPSG NYVACGGLDN ICSIYNLKTR EGNVRVSREL AGHTGYLSCC RFLDDNQIVT S SGDTTCAL ...String: MSELDQLRQE AEQLKNQIRD ARKACADATL SQITNNIDPV GRIQMRTRRT LRGHLAKIYA MHWGTDSRLL VSASQDGKLI IWDSYTTNK VHAIPLRSSW VMTCAYAPSG NYVACGGLDN ICSIYNLKTR EGNVRVSREL AGHTGYLSCC RFLDDNQIVT S SGDTTCAL WDIETGQQTT TFTGHTGDVM SLSLAPDTRL FVSGACDASA KLWDVREGMC RQTFTGHESD INAICFFPNG NA FATGSDD ATCRLFDLRA DQELMTYSHD NIICGITSVS FSKSGRLLLA GYDDFNCNVW DALKADRAGV LAGHDNRVSC LGV TDDGMA VATGSWDSFL KIWN UniProtKB: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 |
-Macromolecule #3: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
Macromolecule | Name: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Bos taurus (cattle) |
Molecular weight | Theoretical: 9.226547 KDa |
Recombinant expression | Organism: Trichoplusia ni (cabbage looper) |
Sequence | String: HHHHHHHHHH MASNNTASIA QARKLVEQLK MEANIDRIKV SKAAADLMAY CEAHAKEDPL LTPVPASENP FREKKFFSAI L UniProtKB: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.7 mg/mL |
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Buffer | pH: 8 |
Grid | Model: Quantifoil, UltrAuFoil, R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 75 sec. |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV |
-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 / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Number grids imaged: 2 / Number real images: 6746 / Average electron dose: 47.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Illumination mode: OTHER / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal magnification: 29000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Initial model |
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Refinement | Space: REAL / Protocol: AB INITIO MODEL / Overall B value: 83 | ||||||||
Output model | PDB-6pcv: |