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- PDB-6q0k: Structure of a MAPK pathway complex -

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Basic information

Entry
Database: PDB / ID: 6q0k
TitleStructure of a MAPK pathway complex
Components
  • 14-3-3 protein zeta/delta
  • Serine/threonine-protein kinase B-raf
KeywordsSIGNALING PROTEIN/Transferase / TRANSFERASE / SIGNALING PROTEIN-Transferase complex
Function / homology
Function and homology information


Translocation of SLC2A4 (GLUT4) to the plasma membrane / Activation of BAD and translocation to mitochondria / ARMS-mediated activation / Frs2-mediated activation / Spry regulation of FGF signaling / TP53 Regulates Metabolic Genes / Interleukin-3, Interleukin-5 and GM-CSF signaling / RAF activation / KSRP (KHSRP) binds and destabilizes mRNA / MAP2K and MAPK activation ...Translocation of SLC2A4 (GLUT4) to the plasma membrane / Activation of BAD and translocation to mitochondria / ARMS-mediated activation / Frs2-mediated activation / Spry regulation of FGF signaling / TP53 Regulates Metabolic Genes / Interleukin-3, Interleukin-5 and GM-CSF signaling / RAF activation / KSRP (KHSRP) binds and destabilizes mRNA / MAP2K and MAPK activation / Negative feedback regulation of MAPK pathway / GP1b-IX-V activation signalling / Negative regulation of MAPK pathway / Signaling by moderate kinase activity BRAF mutants / Signaling by high-kinase activity BRAF mutants / Signaling by RAS mutants / Signaling by BRAF and RAF fusions / Rap1 signalling / Paradoxical activation of RAF signaling by kinase inactive BRAF / Deactivation of the beta-catenin transactivating complex / Chk1/Chk2(Cds1) mediated inactivation of Cyclin B:Cdk1 complex / NOTCH4 Activation and Transmission of Signal to the Nucleus / Negative regulation of NOTCH4 signaling / Signalling to p38 via RIT and RIN / Regulation of localization of FOXO transcription factors / RHO GTPases activate PKNs / regulation of synapse maturation / synaptic target recognition / Golgi reassembly / trehalose metabolism in response to stress / establishment of Golgi localization / positive regulation of glucose transmembrane transport / establishment of protein localization to membrane / protein targeting / positive regulation of protein insertion into mitochondrial membrane involved in apoptotic signaling pathway / cellular response to calcium ion / hippocampal mossy fiber to CA3 synapse / animal organ morphogenesis / melanosome / regulation of mRNA stability / membrane organization / platelet activation / ion channel binding / positive regulation of peptidyl-serine phosphorylation / vesicle / blood microparticle / positive regulation of ERK1 and ERK2 cascade / non-specific serine/threonine protein kinase / MAPK cascade / cadherin binding / protein kinase activity / intracellular membrane-bounded organelle / glutamatergic synapse / cytokine-mediated signaling pathway / focal adhesion / transcription factor binding / protein domain specific binding / ubiquitin protein ligase binding / protein serine/threonine kinase activity / positive regulation of gene expression / protein phosphorylation / calcium ion binding / protein kinase binding / negative regulation of apoptotic process / signal transduction / mitochondrion / RNA binding / extracellular space / extracellular exosome / nucleoplasm / ATP binding / identical protein binding / plasma membrane / nucleus / cytosol / cytoplasm
Protein kinase domain / Protein kinase, ATP binding site / Protein kinase domain profile. / Ras-binding domain (RBD) profile. / 14-3-3 protein / Zinc finger phorbol-ester/DAG-type profile. / Serine-threonine/tyrosine-protein kinase, catalytic domain / Protein kinase C-like, phorbol ester/diacylglycerol-binding domain / Raf-like Ras-binding / Serine/threonine-protein kinase, active site ...Protein kinase domain / Protein kinase, ATP binding site / Protein kinase domain profile. / Ras-binding domain (RBD) profile. / 14-3-3 protein / Zinc finger phorbol-ester/DAG-type profile. / Serine-threonine/tyrosine-protein kinase, catalytic domain / Protein kinase C-like, phorbol ester/diacylglycerol-binding domain / Raf-like Ras-binding / Serine/threonine-protein kinase, active site / Protein kinase-like domain superfamily / Diacylglycerol/phorbol-ester binding / 14-3-3 protein, conserved site / 14-3-3 domain / Ubiquitin-like domain superfamily / 14-3-3 domain superfamily / Phorbol esters/diacylglycerol binding domain (C1 domain) / 14-3-3 protein / Raf-like Ras-binding domain / Protein tyrosine kinase / Protein kinases ATP-binding region signature. / Serine/Threonine protein kinases active-site signature. / Zinc finger phorbol-ester/DAG-type signature. / 14-3-3 proteins signature 1. / 14-3-3 proteins signature 2.
Serine/threonine-protein kinase B-raf / 14-3-3 protein zeta/delta
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 6.8 Å
AuthorsPark, E. / Rawson, S. / Jeon, H. / Eck, M.J.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Human Genome Research InstituteP50CA165962 United States
National Institutes of Health/National Cancer InstituteR50CA221830 United States
CitationJournal: Nature / Year: 2019
Title: Architecture of autoinhibited and active BRAF-MEK1-14-3-3 complexes.
Authors: Eunyoung Park / Shaun Rawson / Kunhua Li / Byeong-Won Kim / Scott B Ficarro / Gonzalo Gonzalez-Del Pino / Humayun Sharif / Jarrod A Marto / Hyesung Jeon / Michael J Eck /
Abstract: RAF family kinases are RAS-activated switches that initiate signaling through the MAP kinase cascade to control cellular proliferation, differentiation and survival. RAF activity is tightly ...RAF family kinases are RAS-activated switches that initiate signaling through the MAP kinase cascade to control cellular proliferation, differentiation and survival. RAF activity is tightly regulated, and inappropriate activation is a frequent cause of cancer. At present, the structural basis of RAF regulation is poorly understood. Here we describe autoinhibited and active state structures of full-length BRAF in complexes with MEK1 and a 14-3-3 dimer, determined using cryo-electron microscopy (cryo-EM). A 4.1 Å resolution cryo-EM reconstruction reveals an inactive BRAF-MEK1 complex restrained in a cradle formed by the 14-3-3 dimer, which binds the phosphorylated S365 and S729 sites that flank the BRAF kinase domain. The BRAF cysteine-rich domain (CRD) occupies a central position that stabilizes this assembly, but the adjacent RAS-binding domain (RBD) is poorly ordered and peripheral. The 14-3-3 cradle maintains autoinhibition by sequestering the membrane-binding CRD and blocking dimerization of the BRAF kinase domain. In the active state, these inhibitory interactions are released and a single 14-3-3 dimer rearranges to bridge the C-terminal pS729 binding sites of two BRAFs, driving formation of an active, back-to-back BRAF dimer. Our structural snapshots provide a foundation for understanding normal RAF regulation and its mutational disruption in cancer and developmental syndromes.
Validation Report
SummaryFull reportAbout validation report
History
DepositionAug 1, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 9, 2019Provider: repository / Type: Initial release

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Structure visualization

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Assembly

Deposited unit
A: Serine/threonine-protein kinase B-raf
B: Serine/threonine-protein kinase B-raf
X: 14-3-3 protein zeta/delta
Y: 14-3-3 protein zeta/delta


Theoretical massNumber of molelcules
Total (without water)234,2004
Polymers234,2004
Non-polymers00
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein/peptide Serine/threonine-protein kinase B-raf / Proto-oncogene B-Raf / p94 / v-Raf murine sarcoma viral oncogene homolog B1


Mass: 89322.812 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: BRAF, BRAF1, RAFB1 / Production host: Homo sapiens (human)
References: UniProt: P15056, non-specific serine/threonine protein kinase
#2: Protein/peptide 14-3-3 protein zeta/delta / Protein kinase C inhibitor protein 1 / KCIP-1


Mass: 27777.092 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P63104
Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: ERK pathway complexMAPK/ERK pathway / Type: COMPLEX / Entity ID: 1, 2 / Source: MULTIPLE SOURCES
Molecular weightValue: 233 kDa/nm / Experimental value: YES
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: unspecified
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: OTHER
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: NONE
3D reconstructionResolution: 6.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 66215 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model building

3D fitting-ID: 1

IDPDB-IDPdb chain-ID
14MNEA
24MNEB
33NKXX
43NKXY

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