+Open data
-Basic information
Entry | Database: PDB / ID: 7sj4 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | Human Trio residues 1284-1959 in complex with Rac1 | |||||||||
Components |
| |||||||||
Keywords | SIGNALING PROTEIN / guanine nucleotide exchange factor / GTPase / dbl homology / pleckstrin homology | |||||||||
Function / homology | Function and homology information cell surface receptor protein tyrosine phosphatase signaling pathway / regulation of respiratory burst / negative regulation of interleukin-23 production / regulation of neutrophil migration / localization within membrane / Activated NTRK2 signals through CDK5 / negative regulation of receptor-mediated endocytosis / regulation of hydrogen peroxide metabolic process / ruffle assembly / NTRK2 activates RAC1 ...cell surface receptor protein tyrosine phosphatase signaling pathway / regulation of respiratory burst / negative regulation of interleukin-23 production / regulation of neutrophil migration / localization within membrane / Activated NTRK2 signals through CDK5 / negative regulation of receptor-mediated endocytosis / regulation of hydrogen peroxide metabolic process / ruffle assembly / NTRK2 activates RAC1 / engulfment of apoptotic cell / Inactivation of CDC42 and RAC1 / NADPH oxidase complex / respiratory burst / WNT5:FZD7-mediated leishmania damping / SEMA3A-Plexin repulsion signaling by inhibiting Integrin adhesion / cortical cytoskeleton organization / hepatocyte growth factor receptor signaling pathway / ruffle organization / thioesterase binding / regulation of stress fiber assembly / negative regulation of fibroblast migration / cell projection assembly / RHO GTPases activate CIT / sphingosine-1-phosphate receptor signaling pathway / Nef and signal transduction / PCP/CE pathway / positive regulation of neutrophil chemotaxis / motor neuron axon guidance / regulation of nitric oxide biosynthetic process / RHO GTPases activate KTN1 / Activation of RAC1 / regulation of lamellipodium assembly / Azathioprine ADME / MET activates RAP1 and RAC1 / regulation of small GTPase mediated signal transduction / DCC mediated attractive signaling / positive regulation of cell-substrate adhesion / Wnt signaling pathway, planar cell polarity pathway / Sema4D mediated inhibition of cell attachment and migration / CD28 dependent Vav1 pathway / Ephrin signaling / lamellipodium assembly / establishment or maintenance of cell polarity / regulation of cell size / DSCAM interactions / Activation of RAC1 downstream of NMDARs / small GTPase-mediated signal transduction / postsynaptic modulation of chemical synaptic transmission / Rho GDP-dissociation inhibitor binding / extrinsic component of membrane / positive regulation of Rho protein signal transduction / NRAGE signals death through JNK / negative regulation of fat cell differentiation / Rac protein signal transduction / RHOJ GTPase cycle / presynaptic active zone / RHO GTPases activate PAKs / positive regulation of focal adhesion assembly / CDC42 GTPase cycle / semaphorin-plexin signaling pathway / Sema3A PAK dependent Axon repulsion / ficolin-1-rich granule membrane / RHOG GTPase cycle / RHOA GTPase cycle / EPH-ephrin mediated repulsion of cells / RHO GTPases Activate NADPH Oxidases / localization / RAC2 GTPase cycle / RAC3 GTPase cycle / RHO GTPases Activate WASPs and WAVEs / RHO GTPases activate IQGAPs / anatomical structure morphogenesis / PTK6 Regulates RHO GTPases, RAS GTPase and MAP kinases / positive regulation of lamellipodium assembly / positive regulation of substrate adhesion-dependent cell spreading / RHO GTPases activate PKNs / positive regulation of microtubule polymerization / positive regulation of stress fiber assembly / regulation of cell migration / GPVI-mediated activation cascade / EPHB-mediated forward signaling / RAC1 GTPase cycle / actin filament polymerization / positive regulation of endothelial cell migration / substrate adhesion-dependent cell spreading / cell-matrix adhesion / neuron projection morphogenesis / cell chemotaxis / small monomeric GTPase / guanyl-nucleotide exchange factor activity / secretory granule membrane / VEGFR2 mediated vascular permeability / G protein activity / Signal transduction by L1 / cell projection / Translocation of SLC2A4 (GLUT4) to the plasma membrane / RHO GTPases Activate Formins / cell motility / regulation of actin cytoskeleton organization Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.86 Å | |||||||||
Authors | Chen, C.-L. / Ravala, S.K. / Bandekar, S.J. / Cash, J. / Tesmer, J.J.G. | |||||||||
Funding support | United States, 2items
| |||||||||
Citation | Journal: J Biol Chem / Year: 2022 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 |
|
-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
---|
-Downloads & links
-Download
PDBx/mmCIF format | 7sj4.cif.gz | 119.8 KB | Display | PDBx/mmCIF format |
---|---|---|---|---|
PDB format | pdb7sj4.ent.gz | 83.9 KB | Display | PDB format |
PDBx/mmJSON format | 7sj4.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7sj4_validation.pdf.gz | 834.2 KB | Display | wwPDB validaton report |
---|---|---|---|---|
Full document | 7sj4_full_validation.pdf.gz | 835.8 KB | Display | |
Data in XML | 7sj4_validation.xml.gz | 22.8 KB | Display | |
Data in CIF | 7sj4_validation.cif.gz | 32.4 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/sj/7sj4 ftp://data.pdbj.org/pub/pdb/validation_reports/sj/7sj4 | HTTPS FTP |
-Related structure data
Related structure data | 25153MC M: map data used to model this data C: citing same article (ref.) |
---|---|
Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
|
---|---|
1 |
|
-Components
#1: Protein | Mass: 75592.969 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: TRIO / Production host: Escherichia coli (E. coli) / Strain (production host): Rosetta (DE3) pLysS References: UniProt: O75962, non-specific serine/threonine protein kinase |
---|---|
#2: Protein | Mass: 21811.453 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: RAC1, TC25, MIG5 / Production host: Escherichia coli (E. coli) / Strain (production host): Rosetta (DE3) pLysS / References: UniProt: P63000 |
#3: Water | ChemComp-HOH / |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Human Trio residues 1284-1959 in complex with Rac1 / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT | ||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Molecular weight | Experimental value: NO | ||||||||||||||||||||
Source (natural) | Organism: Homo sapiens (human) | ||||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli (E. coli) / Strain: Rosetta (DE3) pLysS | ||||||||||||||||||||
Buffer solution | pH: 8 | ||||||||||||||||||||
Buffer component |
| ||||||||||||||||||||
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 |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
---|---|
Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
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 |
-Processing
Software | Name: PHENIX / Version: 1.20_4459: / Classification: refinement | ||||||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
EM software |
| ||||||||||||||||||||||||||||||||||||||||||||||||
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
| ||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
|