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Open data
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Basic information
Entry | Database: PDB / ID: 7sj4 | |||||||||
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Title | Human Trio residues 1284-1959 in complex with Rac1 | |||||||||
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![]() | SIGNALING PROTEIN / guanine nucleotide exchange factor / GTPase / dbl homology / pleckstrin homology | |||||||||
Function / homology | ![]() cell surface receptor protein tyrosine phosphatase signaling pathway / embryonic olfactory bulb interneuron precursor migration / anatomical structure arrangement / regulation of ERK5 cascade / angiotensin-activated signaling pathway involved in heart process / positive regulation of ovarian follicle development / cerebral cortex GABAergic interneuron development / regulation of respiratory burst / auditory receptor cell morphogenesis / cerebral cortex radially oriented cell migration ...cell surface receptor protein tyrosine phosphatase signaling pathway / embryonic olfactory bulb interneuron precursor migration / anatomical structure arrangement / regulation of ERK5 cascade / angiotensin-activated signaling pathway involved in heart process / positive regulation of ovarian follicle development / cerebral cortex GABAergic interneuron development / regulation of respiratory burst / auditory receptor cell morphogenesis / cerebral cortex radially oriented cell migration / erythrocyte enucleation / regulation of neutrophil migration / negative regulation of interleukin-23 production / localization within membrane / Activated NTRK2 signals through CDK5 / interneuron migration / regulation of hydrogen peroxide metabolic process / kinocilium / regulation of cell adhesion involved in heart morphogenesis / negative regulation of receptor-mediated endocytosis / engulfment of apoptotic cell / ruffle assembly / NTRK2 activates RAC1 / NADPH oxidase complex / cochlea morphogenesis / Inactivation of CDC42 and RAC1 / regulation of neuron maturation / respiratory burst / WNT5:FZD7-mediated leishmania damping / cortical cytoskeleton organization / SEMA3A-Plexin repulsion signaling by inhibiting Integrin adhesion / positive regulation of skeletal muscle acetylcholine-gated channel clustering / hepatocyte growth factor receptor signaling pathway / GTP-dependent protein binding / midbrain dopaminergic neuron differentiation / ruffle organization / epithelial cell morphogenesis / cell projection assembly / positive regulation of bicellular tight junction assembly / thioesterase binding / regulation of lamellipodium assembly / regulation of stress fiber assembly / regulation of neuron migration / negative regulation of fibroblast migration / RHO GTPases activate CIT / motor neuron axon guidance / Nef and signal transduction / sphingosine-1-phosphate receptor signaling pathway / regulation of nitric oxide biosynthetic process / PCP/CE pathway / Activation of RAC1 / RHO GTPases activate KTN1 / positive regulation of neutrophil chemotaxis / MET activates RAP1 and RAC1 / DCC mediated attractive signaling / cell-cell junction organization / Azathioprine ADME / regulation of small GTPase mediated signal transduction / hyperosmotic response / Sema4D mediated inhibition of cell attachment and migration / positive regulation of cell-substrate adhesion / Ephrin signaling / CD28 dependent Vav1 pathway / positive regulation of ruffle assembly / superoxide anion generation / Wnt signaling pathway, planar cell polarity pathway / lamellipodium assembly / extrinsic component of membrane / regulation of receptor signaling pathway via JAK-STAT / Activation of RAC1 downstream of NMDARs / small GTPase-mediated signal transduction / NRAGE signals death through JNK / regulation of cell size / dendrite morphogenesis / negative regulation of fat cell differentiation / positive regulation of Rho protein signal transduction / Rho GDP-dissociation inhibitor binding / establishment or maintenance of cell polarity / RHOJ GTPase cycle / synaptic transmission, GABAergic / Rac protein signal transduction / positive regulation of actin filament polymerization / positive regulation of dendritic spine development / RHO GTPases activate PAKs / pericentriolar material / semaphorin-plexin signaling pathway / presynaptic active zone / CDC42 GTPase cycle / ficolin-1-rich granule membrane / RHOG GTPase cycle / Sema3A PAK dependent Axon repulsion / RHO GTPases Activate NADPH Oxidases / EPH-ephrin mediated repulsion of cells / regulation of postsynapse assembly / regulation of synaptic vesicle endocytosis / RHOA GTPase cycle / anatomical structure morphogenesis / regulation of neuronal synaptic plasticity / positive regulation of focal adhesion assembly / RHO GTPases Activate WASPs and WAVEs Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.86 Å | |||||||||
![]() | Chen, C.-L. / Ravala, S.K. / Bandekar, S.J. / Cash, J. / Tesmer, J.J.G. | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural/functional studies of Trio provide insights into its configuration and show that conserved linker elements enhance its activity for Rac1. Authors: Sumit J Bandekar / Chun-Liang Chen / Sandeep K Ravala / Jennifer N Cash / Larisa V Avramova / Mariya V Zhalnina / J Silvio Gutkind / Sheng Li / John J G Tesmer / ![]() Abstract: Trio is a large and highly conserved metazoan signaling scaffold that contains two Dbl family guanine nucleotide exchange factor (GEF) modules, TrioN and TrioC, selective for Rac and RhoA GTPases, ...Trio is a large and highly conserved metazoan signaling scaffold that contains two Dbl family guanine nucleotide exchange factor (GEF) modules, TrioN and TrioC, selective for Rac and RhoA GTPases, respectively. The GEF activities of TrioN and TrioC are implicated in several cancers, especially uveal melanoma. However, little is known about how these modules operate in the context of larger fragments of Trio. Here we show via negative stain electron microscopy that the N-terminal region of Trio is extended and could thus serve as a rigid spacer between the N-terminal putative lipid-binding domain and TrioN, whereas the C-terminal half of Trio seems globular. We found that regions C-terminal to TrioN enhance its Rac1 GEF activity and thus could play a regulatory role. We went on to characterize a minimal, well-behaved Trio fragment with enhanced activity, Trio, in complex with Rac1 using cryo-electron microscopy and hydrogen-deuterium exchange mass spectrometry and found that the region conferring enhanced activity is disordered. Deletion of two different strongly conserved motifs in this region eliminated this enhancement, suggesting that they form transient intramolecular interactions that promote GEF activity. Because Dbl family RhoGEF modules have been challenging to directly target with small molecules, characterization of accessory Trio domains such as these may provide alternate routes for the development of therapeutics that inhibit Trio activity in human cancer. | |||||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 119.8 KB | Display | ![]() |
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PDB format | ![]() | 83.9 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 834.2 KB | Display | ![]() |
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Full document | ![]() | 835.8 KB | Display | |
Data in XML | ![]() | 22.8 KB | Display | |
Data in CIF | ![]() | 32.4 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 25153MC M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 75592.969 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() References: UniProt: O75962, non-specific serine/threonine protein kinase |
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#2: Protein | Mass: 21811.453 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() |
#3: Water | ChemComp-HOH / |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: Human Trio residues 1284-1959 in complex with Rac1 / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT | ||||||||||||||||||||
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Molecular weight | Experimental value: NO | ||||||||||||||||||||
Source (natural) | Organism: ![]() | ||||||||||||||||||||
Source (recombinant) | Organism: ![]() ![]() | ||||||||||||||||||||
Buffer solution | pH: 8 | ||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 0.25 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3 | ||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K / Details: Blot force 10 |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 81000 X / Nominal defocus max: 3000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Residual tilt: 0.01 mradians |
Image recording | Average exposure time: 3.12 sec. / Electron dose: 55 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of real images: 2817 Details: 3514 movies were initially collected. After motion correction and CTF estimation, 697 micrographs were rejected due to poor CTF fitting. |
Image scans | Width: 11520 / Height: 8184 / Movie frames/image: 40 |
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Processing
Software | Name: PHENIX / Version: 1.20_4459: / Classification: refinement | ||||||||||||||||||||||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.86 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 922202 / Algorithm: BACK PROJECTION / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT / Space: REAL / Target criteria: correlation coefficient Details: The initial model was generated from the pdb structure (2NZ8) using the SWISS-MODEL server and rigid-body fitted using Chimera. Several runs of structure refinement were done using the coot ...Details: The initial model was generated from the pdb structure (2NZ8) using the SWISS-MODEL server and rigid-body fitted using Chimera. Several runs of structure refinement were done using the coot and phenix real-space refinement. | ||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | 3D fitting-ID: 1 / Accession code: 2NZ8 / Initial refinement model-ID: 1 / PDB-ID: 2NZ8 / Source name: PDB / Type: experimental model
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Refine LS restraints |
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