|Entry||Database: EMDB / ID: 2712|
|Title||Structure of the RET receptor tyrosine kinase extracellular domain|
|Map data||Reconstruction of a reconstituted mammalian RETecd-GDNF-GFRa1 ternary (mTC) complex|
|Sample||Reconstituted mammalian RETecd-GDNF-GFRa1 ternary complex|
|Keywords||vertebrate development / human diseases / RET-GFL-GFRa complex|
|Function / homology||Cadherin domain / Glial cell line-derived neurotrophic factor receptor, alpha 1 / Protein kinases ATP-binding region signature. / Protein tyrosine kinase / Tyrosine protein kinases specific active-site signature. / Protein kinase domain / Serine-threonine/tyrosine-protein kinase, catalytic domain / Transforming growth factor-beta, C-terminal / Cadherin-like / Protein kinase domain profile. ...Cadherin domain / Glial cell line-derived neurotrophic factor receptor, alpha 1 / Protein kinases ATP-binding region signature. / Protein tyrosine kinase / Tyrosine protein kinases specific active-site signature. / Protein kinase domain / Serine-threonine/tyrosine-protein kinase, catalytic domain / Transforming growth factor-beta, C-terminal / Cadherin-like / Protein kinase domain profile. / GDNF/GAS1 domain / TGF-beta family profile. / Glial cell line-derived neurotrophic factor receptor / NCAM1 interactions / RAF/MAP kinase cascade / RET signaling / Cadherins domain profile. / Tyrosine-protein kinase, active site / Protein kinase, ATP binding site / Protein kinase-like domain superfamily / Transforming growth factor beta like domain / GDNF receptor alpha / Cystine-knot cytokine / Tyrosine-protein kinase, catalytic domain / Glial cell line-derived neurotrophic factor receptor, alpha 1/2 / Glial cell line-derived neurotrophic factor / Tyrosine-protein kinase, Ret receptor / GDNF/GAS1 / Cadherin-like superfamily / Glial cell line-derived neurotrophic factor-like / postsynaptic membrane organization / chemoattractant activity involved in axon guidance / mesenchymal to epithelial transition involved in metanephros morphogenesis / dorsal spinal cord development / positive regulation of mesenchymal to epithelial transition involved in metanephros morphogenesis / postganglionic parasympathetic fiber development / positive regulation of ureteric bud formation / regulation of dopamine uptake involved in synaptic transmission / ureteric bud formation / regulation of semaphorin-plexin signaling pathway / embryonic epithelial tube formation / lymphocyte migration into lymphoid organs / glial cell-derived neurotrophic factor receptor signaling pathway / regulation of stem cell differentiation / positive regulation of metanephric glomerulus development / Peyer's patch morphogenesis / posterior midgut development / enteric nervous system development / ureter maturation / membrane protein proteolysis / peristalsis / positive regulation of monooxygenase activity / positive regulation of dopamine secretion / regulation of morphogenesis of a branching structure / positive regulation of peptidyl-serine phosphorylation of STAT protein / neurotrophin receptor activity / neuron cell-cell adhesion / innervation / plasma membrane protein complex / sympathetic nervous system development / positive regulation of neuron maturation / positive regulation of branching involved in ureteric bud morphogenesis / neuron maturation / metanephros development / positive regulation of extrinsic apoptotic signaling pathway in absence of ligand / mRNA stabilization / commissural neuron axon guidance / positive regulation of cell adhesion mediated by integrin / neurogenesis / positive regulation of cell size / neural crest cell migration / organ induction / response to pain / embryonic organ development / ureteric bud development / regulation of axonogenesis / branching involved in ureteric bud morphogenesis / homophilic cell adhesion via plasma membrane adhesion molecules / regulation of cell adhesion / retina development in camera-type eye / negative regulation of extrinsic apoptotic signaling pathway in absence of ligand / adult locomotory behavior / anchored component of membrane / cellular response to retinoic acid / positive regulation of neuron projection development / positive regulation of cell differentiation / neuron differentiation / receptor protein-tyrosine kinase / transmembrane receptor protein tyrosine kinase activity / neuron projection development / activation of cysteine-type endopeptidase activity involved in apoptotic process / nervous system development / growth factor activity / male gonad development / axon guidance / Ras guanyl-nucleotide exchange factor activity / kidney development / positive regulation of peptidyl-tyrosine phosphorylation / integrin binding / transmembrane receptor protein tyrosine kinase signaling pathway|
Function and homology information
|Source||Homo sapiens (human) / Rattus norvegicus (Norway rat)|
|Method||single particle reconstruction / negative staining / 24 Å resolution|
|Authors||Goodman K / Kjaer S / Beuron F / Knowles P / Nawrotek A / Burns E / Purkiss A / George R / Santoro M / Morris EP / McDonald NQ|
|Citation||Journal: Cell Rep / Year: 2014|
Title: RET recognition of GDNF-GFRα1 ligand by a composite binding site promotes membrane-proximal self-association.
Authors: Kerry M Goodman / Svend Kjær / Fabienne Beuron / Phillip P Knowles / Agata Nawrotek / Emily M Burns / Andrew G Purkiss / Roger George / Massimo Santoro / Edward P Morris / Neil Q McDonald
Abstract: The RET receptor tyrosine kinase is essential to vertebrate development and implicated in multiple human diseases. RET binds a cell surface bipartite ligand comprising a GDNF family ligand and a ...The RET receptor tyrosine kinase is essential to vertebrate development and implicated in multiple human diseases. RET binds a cell surface bipartite ligand comprising a GDNF family ligand and a GFRα coreceptor, resulting in RET transmembrane signaling. We present a hybrid structural model, derived from electron microscopy (EM) and low-angle X-ray scattering (SAXS) data, of the RET extracellular domain (RET(ECD)), GDNF, and GFRα1 ternary complex, defining the basis for ligand recognition. RET(ECD) envelopes the dimeric ligand complex through a composite binding site comprising four discrete contact sites. The GFRα1-mediated contacts are crucial, particularly close to the invariant RET calcium-binding site, whereas few direct contacts are made by GDNF, explaining how distinct ligand/coreceptor pairs are accommodated. The RET(ECD) cysteine-rich domain (CRD) contacts both ligand components and makes homotypic membrane-proximal interactions occluding three different antibody epitopes. Coupling of these CRD-mediated interactions suggests models for ligand-induced RET activation and ligand-independent oncogenic deregulation.
|Validation Report||PDB-ID: 4ux8|
SummaryFull reportAbout validation report
|Date||Deposition: Jul 17, 2014 / Header (metadata) release: Jul 23, 2014 / Map release: Oct 1, 2014 / Last update: Oct 8, 2014|
|Structure viewer||EM map: |
Downloads & links
|File||emd_2712.map.gz (map file in CCP4 format, 3457 KB)|
|Projections & slices|
Images are generated by Spider.
|Voxel size||X=Y=Z: 4.34 Å|
CCP4 map header:
-Entire Reconstituted mammalian RETecd-GDNF-GFRa1 ternary complex
|Entire||Name: Reconstituted mammalian RETecd-GDNF-GFRa1 ternary complex|
Details: Monodisperse. Measured mass difference with theoretical MW corresponds to glycosylation.
Number of components: 3 / Oligomeric State: Hexamer
|Mass||Theoretical: 260 kDa / Experimental: 340 kDa / Measured by: SEC-MALS|
-Component #1: protein, RET receptor tyrosine kinase
|Protein||Name: RET receptor tyrosine kinase / a.k.a: RET / Number of Copies: 2 / Recombinant expression: Yes|
|Mass||Theoretical: 80 kDa|
|Source||Species: Homo sapiens (human)|
|Source (engineered)||Expression System: Chinese Hamster Ovary / Strain: Lec8|
-Component #2: protein, GDNF receptor alpha
|Protein||Name: GDNF receptor alphaGFRα / Recombinant expression: Yes / Number of Copies: 2|
|Mass||Theoretical: 40 kDa|
|Source||Species: Rattus norvegicus (Norway rat)|
|Source (engineered)||Expression System: Chinese Hamster Ovary|
-Component #3: protein, glial-cell-line-derived neurotrophic factor
|Protein||Name: glial-cell-line-derived neurotrophic factor / a.k.a: GDNF / Details: from Amgen (USA) / Number of Copies: 2 / Recombinant expression: Yes|
|Mass||Theoretical: 10 kDa|
|Source||Species: Homo sapiens (human)|
|Specimen||Specimen state: particle / Method: negative staining|
|Sample solution||Specimen conc.: 0.03 mg/ml / Buffer solution: 20 mM Tris HCl, 300 mM NaCl, 1 mM Ca++ / pH: 8|
|Support film||quantifoil (R1.2/1.3) coated with a thin carbon layer, glow discharge in air.|
|Staining||Samples were applied to glow-discharged grids and stained with 2% uranyl acetate|
|Vitrification||Instrument: NONE / Cryogen name: NONE|
-Electron microscopy imaging
Model: Tecnai F20 / Image courtesy: FEI Company
|Imaging||Microscope: FEI TECNAI F20 / Date: Nov 8, 2012|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Electron dose: 100 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Magnification: 80000 X (nominal) / Imaging mode: BRIGHT FIELD / Defocus: 900 - nm|
|Specimen Holder||Model: SIDE ENTRY, EUCENTRIC|
|Camera||Detector: TVIPS TEMCAM-F415 (4k x 4k)|
|Image acquisition||Number of digital images: 1200 / Bit depth: 16|
|Processing||Method: single particle reconstruction / Number of class averages: 975 / Applied symmetry: C2 (2 fold cyclic) / Number of projections: 8519|
Details: The particles were selected manually. The starting model was calculated from reference-free classes using angular reconstitution methods and further refined by projection matching.
|3D reconstruction||Algorithm: angular reconstitution / Software: IMAGIC, SPIDER, in-house, software / Resolution: 24 Å / Resolution method: FSC 0.5, semi-independent|
-Atomic model buiding
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