+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-40253 | ||||||||||||
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Title | Truncated Braf/Mek/14-3-3 complex | ||||||||||||
Map data | (del1-155)Braf/Mek/14-3-3 complex | ||||||||||||
Sample |
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Keywords | kinase complex / TRANSFERASE | ||||||||||||
Biological species | Homo sapiens (human) | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.1 Å | ||||||||||||
Authors | Eck MJ / Jeon H / Park E / Rawson S | ||||||||||||
Funding support | United States, 3 items
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Citation | Journal: Nat Commun / Year: 2023 Title: Cryo-EM structure of a RAS/RAF recruitment complex. Authors: Eunyoung Park / Shaun Rawson / Anna Schmoker / Byeong-Won Kim / Sehee Oh / Kangkang Song / Hyesung Jeon / Michael J Eck / Abstract: RAF-family kinases are activated by recruitment to the plasma membrane by GTP-bound RAS, whereupon they initiate signaling through the MAP kinase cascade. Prior structural studies of KRAS with RAF ...RAF-family kinases are activated by recruitment to the plasma membrane by GTP-bound RAS, whereupon they initiate signaling through the MAP kinase cascade. Prior structural studies of KRAS with RAF have focused on the isolated RAS-binding and cysteine-rich domains of RAF (RBD and CRD, respectively), which interact directly with RAS. Here we describe cryo-EM structures of a KRAS bound to intact BRAF in an autoinhibited state with MEK1 and a 14-3-3 dimer. Analysis of this KRAS/BRAF/MEK1/14-3-3 complex reveals KRAS bound to the RAS-binding domain of BRAF, captured in two orientations. Core autoinhibitory interactions in the complex are unperturbed by binding of KRAS and in vitro activation studies confirm that KRAS binding is insufficient to activate BRAF, absent membrane recruitment. These structures illustrate the separability of binding and activation of BRAF by RAS and suggest stabilization of this pre-activation intermediate as an alternative therapeutic strategy to blocking binding of KRAS. | ||||||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_40253.map.gz | 28.8 MB | EMDB map data format | |
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Header (meta data) | emd-40253-v30.xml emd-40253.xml | 18.1 KB 18.1 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_40253_fsc.xml | 6.6 KB | Display | FSC data file |
Images | emd_40253.png | 50 KB | ||
Masks | emd_40253_msk_1.map | 30.5 MB | Mask map | |
Others | emd_40253_additional_1.map.gz emd_40253_half_map_1.map.gz emd_40253_half_map_2.map.gz | 21.1 MB 28.3 MB 28.3 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-40253 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-40253 | HTTPS FTP |
-Validation report
Summary document | emd_40253_validation.pdf.gz | 802.5 KB | Display | EMDB validaton report |
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Full document | emd_40253_full_validation.pdf.gz | 802.1 KB | Display | |
Data in XML | emd_40253_validation.xml.gz | 14.2 KB | Display | |
Data in CIF | emd_40253_validation.cif.gz | 18 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-40253 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-40253 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_40253.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Annotation | (del1-155)Braf/Mek/14-3-3 complex | ||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.06 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_40253_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: (del1-155)Braf/Mek/14-3-3 complex
File | emd_40253_additional_1.map | ||||||||||||
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Annotation | (del1-155)Braf/Mek/14-3-3 complex | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: (del1-155)Braf/Mek/14-3-3 complex
File | emd_40253_half_map_1.map | ||||||||||||
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Annotation | (del1-155)Braf/Mek/14-3-3 complex | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: (del1-155)Braf/Mek/14-3-3 complex
File | emd_40253_half_map_2.map | ||||||||||||
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Annotation | (del1-155)Braf/Mek/14-3-3 complex | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Truncated Braf/Mek/14-3-3 complex
Entire | Name: Truncated Braf/Mek/14-3-3 complex |
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Components |
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-Supramolecule #1: Truncated Braf/Mek/14-3-3 complex
Supramolecule | Name: Truncated Braf/Mek/14-3-3 complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 / Details: (del1-155)Braf/Mek/14-3-3 complex |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 185 KDa |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.1 mg/mL |
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Buffer | pH: 7.4 / Component - Concentration: 150.0 mM / Component - Formula: NaCl / Component - Name: sodium chloride Details: 50 mM Tris pH 7.4, 150 mM NaCl, 2 mM MgCl2, 0.5 mM TCEP, 50 uM ATP-gammaS |
Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV |
Details | monodisperse |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Phase plate: VOLTA PHASE PLATE |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 1 / Number real images: 3252 / Average electron dose: 50.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.2 µm / Nominal defocus min: 0.2 µm / Nominal magnification: 130000 |
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
Refinement | Space: REAL / Protocol: OTHER |
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