+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-16660 | |||||||||
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Title | cryo-EM Structure of Craf:14-3-3:Mek1 | |||||||||
Map data | ||||||||||
Sample |
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Keywords | Kinase / STRUCTURAL PROTEIN | |||||||||
Function / homology | Function and homology information death-inducing signaling complex assembly / epithelial cell proliferation involved in lung morphogenesis / positive regulation of endodermal cell differentiation / intermediate filament cytoskeleton organization / placenta blood vessel development / regulation of Rho protein signal transduction / regulation of axon regeneration / mitogen-activated protein kinase kinase / labyrinthine layer development / MAP-kinase scaffold activity ...death-inducing signaling complex assembly / epithelial cell proliferation involved in lung morphogenesis / positive regulation of endodermal cell differentiation / intermediate filament cytoskeleton organization / placenta blood vessel development / regulation of Rho protein signal transduction / regulation of axon regeneration / mitogen-activated protein kinase kinase / labyrinthine layer development / MAP-kinase scaffold activity / type B pancreatic cell proliferation / cerebellar cortex formation / SHOC2 M1731 mutant abolishes MRAS complex function / Gain-of-function MRAS complexes activate RAF signaling / Signaling by MAP2K mutants / Rap1 signalling / regulation of cell motility / insulin secretion involved in cellular response to glucose stimulus / regulation of Golgi inheritance / spindle pole body / trachea formation / Negative feedback regulation of MAPK pathway / regulation of early endosome to late endosome transport / positive regulation of axonogenesis / regulation of stress-activated MAPK cascade / IFNG signaling activates MAPKs / GP1b-IX-V activation signalling / Frs2-mediated activation / activation of adenylate cyclase activity / ERBB2-ERBB3 signaling pathway / regulation of cell differentiation / protein kinase activator activity / endodermal cell differentiation / MAPK3 (ERK1) activation / pseudopodium / face development / somatic stem cell population maintenance / MAP kinase kinase activity / Bergmann glial cell differentiation / neurotrophin TRK receptor signaling pathway / thyroid gland development / Uptake and function of anthrax toxins / extrinsic apoptotic signaling pathway via death domain receptors / MAP kinase kinase kinase activity / negative regulation of protein-containing complex assembly / negative regulation of extrinsic apoptotic signaling pathway via death domain receptors / type II interferon-mediated signaling pathway / Schwann cell development / keratinocyte differentiation / response to muscle stretch / myelination / protein serine/threonine/tyrosine kinase activity / ERK1 and ERK2 cascade / CD209 (DC-SIGN) signaling / protein serine/threonine kinase activator activity / MAP3K8 (TPL2)-dependent MAPK1/3 activation / insulin-like growth factor receptor signaling pathway / negative regulation of cysteine-type endopeptidase activity involved in apoptotic process / thymus development / Signal transduction by L1 / cell motility / RAF activation / Signaling by high-kinase activity BRAF mutants / wound healing / MAP2K and MAPK activation / positive regulation of protein serine/threonine kinase activity / neuron differentiation / Stimuli-sensing channels / Negative regulation of MAPK pathway / Signaling by RAF1 mutants / Signaling by moderate kinase activity BRAF mutants / Paradoxical activation of RAF signaling by kinase inactive BRAF / Signaling downstream of RAS mutants / chemotaxis / cellular senescence / MAPK cascade / Signaling by BRAF and RAF1 fusions / late endosome / insulin receptor signaling pathway / positive regulation of peptidyl-serine phosphorylation / heart development / scaffold protein binding / protein tyrosine kinase activity / regulation of apoptotic process / mitochondrial outer membrane / positive regulation of MAPK cascade / positive regulation of ERK1 and ERK2 cascade / early endosome / non-specific serine/threonine protein kinase / protein kinase activity / protein phosphorylation / negative regulation of cell population proliferation / protein serine kinase activity / focal adhesion / protein serine/threonine kinase activity / centrosome / positive regulation of gene expression / negative regulation of apoptotic process / apoptotic process / positive regulation of DNA-templated transcription Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) / Spodoptera frugiperda (fall armyworm) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.25 Å | |||||||||
Authors | Dedden D / Ulrich G | |||||||||
Funding support | Germany, 1 items
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Citation | Journal: J Mol Biol / Year: 2024 Title: Cryo-EM Structures of CRAF/14-3-3 and CRAF/14-3-3/MEK1 Complexes. Authors: Dirk Dedden / Julius Nitsche / Elisabeth V Schneider / Maren Thomsen / Daniel Schwarz / Birgitta Leuthner / Ulrich Grädler / Abstract: RAF protein kinases are essential effectors in the MAPK pathway and are important cancer drug targets. Structural understanding of RAF activation is so far based on cryo-electron microscopy (cryo-EM) ...RAF protein kinases are essential effectors in the MAPK pathway and are important cancer drug targets. Structural understanding of RAF activation is so far based on cryo-electron microscopy (cryo-EM) and X-ray structures of BRAF in different conformational states as inactive or active complexes with KRAS, 14-3-3 and MEK1. In this study, we have solved the first cryo-EM structures of CRAF/14-3-3 at 3.4 Å resolution and CRAF/14-3-3/MEK1 at 4.2 Å resolution using CRAF kinase domain expressed as constitutively active Y340D/Y341D mutant in insect cells. The overall architecture of our CRAF/14-3-3 and CRAF/14-3-3/MEK1 cryo-EM structures is highly similar to corresponding BRAF structures in complex with 14-3-3 or 14-3-3/MEK1 and represent the activated dimeric RAF conformation. Our CRAF cryo-EM structures provide additional insights into structural understanding of the activated CRAF/14-3-3/MEK1 complex. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_16660.map.gz | 55.2 MB | EMDB map data format | |
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Header (meta data) | emd-16660-v30.xml emd-16660.xml | 18.5 KB 18.5 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_16660_fsc.xml | 9.2 KB | Display | FSC data file |
Images | emd_16660.png | 65.2 KB | ||
Filedesc metadata | emd-16660.cif.gz | 6.9 KB | ||
Others | emd_16660_half_map_1.map.gz emd_16660_half_map_2.map.gz | 48.7 MB 48.7 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-16660 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-16660 | HTTPS FTP |
-Validation report
Summary document | emd_16660_validation.pdf.gz | 823.3 KB | Display | EMDB validaton report |
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Full document | emd_16660_full_validation.pdf.gz | 822.9 KB | Display | |
Data in XML | emd_16660_validation.xml.gz | 16 KB | Display | |
Data in CIF | emd_16660_validation.cif.gz | 21.1 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-16660 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-16660 | HTTPS FTP |
-Related structure data
Related structure data | 8chfMC 8cpdC M: atomic model generated by this map 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_16660.map.gz / Format: CCP4 / Size: 64 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: 0.91366 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #2
File | emd_16660_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_16660_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Complex of dimer phosphorylated C-raf kinase domain with 14-3-3 d...
Entire | Name: Complex of dimer phosphorylated C-raf kinase domain with 14-3-3 dimer and 2 subunits of Mek1 containing ligand GDC-0623 |
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Components |
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-Supramolecule #1: Complex of dimer phosphorylated C-raf kinase domain with 14-3-3 d...
Supramolecule | Name: Complex of dimer phosphorylated C-raf kinase domain with 14-3-3 dimer and 2 subunits of Mek1 containing ligand GDC-0623 type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 |
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Source (natural) | Organism: Homo sapiens (human) |
-Macromolecule #1: RAF proto-oncogene serine/threonine-protein kinase
Macromolecule | Name: RAF proto-oncogene serine/threonine-protein kinase / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: non-specific serine/threonine protein kinase |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 73.184477 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: MEHIQGAWKT ISNGFGFKDA VFDGSSCISP TIVQQFGYQR RASDDGKLTD PSKTSNTIRV FLPNKQRTVV NVRNGMSLHD CLMKALKVR GLQPECCAVF RLLHEHKGKK ARLDWNTDAA SLIGEELQVD FLDHVPLTTH NFARKTFLKL AFCDICQKFL L NGFRCQTC ...String: MEHIQGAWKT ISNGFGFKDA VFDGSSCISP TIVQQFGYQR RASDDGKLTD PSKTSNTIRV FLPNKQRTVV NVRNGMSLHD CLMKALKVR GLQPECCAVF RLLHEHKGKK ARLDWNTDAA SLIGEELQVD FLDHVPLTTH NFARKTFLKL AFCDICQKFL L NGFRCQTC GYKFHEHCST KVPTMCVDWS NIRQLLLFPN STIGDSGVPA LPSLTMRRMR ESVSRMPVSS QHRYSTPHAF TF NTSSPSS EGSLSQRQRS TSTPNVHMVS TTLPVDSRMI EDAIRSHSES ASPSALSSSP NNLSPTGWSQ PKTPVPAQRE RAP VSGTQE KNKIRPRGQR DSSYDWEIEA SEVMLSTRIG SGSFGTVYKG KWHGDVAVKI LKVVDPTPEQ FQAFRNEVAV LRKT RHVNI LLFMGYMTKD NLAIVTQWCE GSSLYKHLHV QETKFQMFQL IDIARQTAQG MDYLHAKNII HRDMKSNNIF LHEGL TVKI GDFGLATVKS RWSGSQQVEQ PTGSVLWMAP EVIRMQDNNP FSFQSDVYSY GIVLYELMTG ELPYSHINNR DQIIFM VGR GYASPDLSKL YKNCPKAMKR LVADCVKKVK EERPLFPQIL SSIELLQHSL PKINRSA(SEP)EP SLHRAAHTED INA CTLTTS PRLPVF UniProtKB: RAF proto-oncogene serine/threonine-protein kinase |
-Macromolecule #2: 14-3-3 protein zeta isoform X1
Macromolecule | Name: 14-3-3 protein zeta isoform X1 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: Spodoptera frugiperda (fall armyworm) |
Molecular weight | Theoretical: 28.108514 KDa |
Sequence | String: MSVDKEELVQ RAKLAEQAER YDDMAAAMKE VTETGVELSN EERNLLSVAY KNVVGARRSS WRVISSIEQK TEGSERKQQM AKEYRVKVE KELREICYDV LGLLDKHLIP KASNPESKVF YLKMKGDYYR YLAEVATGET RNSVVEDSQK AYQDAFEISK A KMQPTHPI ...String: MSVDKEELVQ RAKLAEQAER YDDMAAAMKE VTETGVELSN EERNLLSVAY KNVVGARRSS WRVISSIEQK TEGSERKQQM AKEYRVKVE KELREICYDV LGLLDKHLIP KASNPESKVF YLKMKGDYYR YLAEVATGET RNSVVEDSQK AYQDAFEISK A KMQPTHPI RLGLALNFSV FYYEILNSPD KACQLAKQAF DDAIAELDTL NEDSYKDSTL IMQLLRDNLT LWTSDTQGDG DE PAEGGDN UniProtKB: 14-3-3 protein zeta isoform X1 |
-Macromolecule #3: Dual specificity mitogen-activated protein kinase kinase 1
Macromolecule | Name: Dual specificity mitogen-activated protein kinase kinase 1 type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO / EC number: mitogen-activated protein kinase kinase |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 43.461938 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: MPKKKPTPIQ LNPAPDGSAV NGTSSAETNL EALQKKLEEL ELDEQQRKRL EAFLTQKQKV GELKDDDFEK ISELGAGNGG VVFKVSHKP SGLVMARKLI HLEIKPAIRN QIIRELQVLH ECNSPYIVGF YGAFYSDGEI SICMEHMDGG SLDQVLKKAG R IPEQILGK ...String: MPKKKPTPIQ LNPAPDGSAV NGTSSAETNL EALQKKLEEL ELDEQQRKRL EAFLTQKQKV GELKDDDFEK ISELGAGNGG VVFKVSHKP SGLVMARKLI HLEIKPAIRN QIIRELQVLH ECNSPYIVGF YGAFYSDGEI SICMEHMDGG SLDQVLKKAG R IPEQILGK VSIAVIKGLT YLREKHKIMH RDVKPSNILV NSRGEIKLCD FGVSGQLIDA MANAFVGTRS YMSPERLQGT HY SVQSDIW SMGLSLVEMA VGRYPIPPPD AKELELMFGC QVEGDAAETP PRPRTPGRPL SSYGMDSRPP MAIFELLDYI VNE PPPKLP SGVFSLEFQD FVNKCLIKNP AERADLKQLM VHAFIKRSDA EEVDFAGWLC STIGLNQPST PTHAAGV UniProtKB: Dual specificity mitogen-activated protein kinase kinase 1 |
-Macromolecule #4: 2-{4-[(1E)-1-(hydroxyimino)-2,3-dihydro-1H-inden-5-yl]-3-(pyridin...
Macromolecule | Name: 2-{4-[(1E)-1-(hydroxyimino)-2,3-dihydro-1H-inden-5-yl]-3-(pyridin-4-yl)-1H-pyrazol-1-yl}ethanol type: ligand / ID: 4 / Number of copies: 2 / Formula: 29L |
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Molecular weight | Theoretical: 334.372 Da |
Chemical component information | ChemComp-29L: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.4 mg/mL |
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Buffer | pH: 7.5 |
Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE |
Vitrification | Cryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | TFS GLACIOS |
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Image recording | Film or detector model: FEI FALCON IV (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 2732 / Average exposure time: 7.0 sec. / Average electron dose: 60.0 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.3000000000000003 µm / Nominal defocus min: 0.7000000000000001 µm / Nominal magnification: 150000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
+Image processing
-Atomic model buiding 1
Refinement | Protocol: RIGID BODY FIT |
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Output model | PDB-8chf: |