+Open data
-Basic information
Entry | Database: PDB / ID: 8ugs | ||||||
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Title | Cryo-EM structure of bovine phosphodiesterase 6 bound to cGMP | ||||||
Components |
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Keywords | SIGNALING PROTEIN/INHIBITOR / phosphodiesterase / GPCR effector enzyme / SIGNALING PROTEIN / inhibitor / SIGNALING PROTEIN-INHIBITOR complex | ||||||
Function / homology | Function and homology information 3',5'-cyclic-GMP phosphodiesterase / Inactivation, recovery and regulation of the phototransduction cascade / positive regulation of G protein-coupled receptor signaling pathway / Activation of the phototransduction cascade / Ca2+ pathway / positive regulation of epidermal growth factor receptor signaling pathway / photoreceptor outer segment membrane / entrainment of circadian clock by photoperiod / cGMP binding / 3',5'-cyclic-GMP phosphodiesterase activity ...3',5'-cyclic-GMP phosphodiesterase / Inactivation, recovery and regulation of the phototransduction cascade / positive regulation of G protein-coupled receptor signaling pathway / Activation of the phototransduction cascade / Ca2+ pathway / positive regulation of epidermal growth factor receptor signaling pathway / photoreceptor outer segment membrane / entrainment of circadian clock by photoperiod / cGMP binding / 3',5'-cyclic-GMP phosphodiesterase activity / 3',5'-cyclic-AMP phosphodiesterase activity / cAMP-mediated signaling / visual perception / photoreceptor disc membrane / retina development in camera-type eye / molecular adaptor activity / zinc ion binding / metal ion binding Similarity search - Function | ||||||
Biological species | Bos taurus (cattle) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å | ||||||
Authors | Aplin, C. / Cerione, R.A. | ||||||
Funding support | United States, 1items
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Citation | Journal: J Biol Chem / Year: 2024 Title: Probing the mechanism by which the retinal G protein transducin activates its biological effector PDE6. Authors: Cody Aplin / Richard A Cerione / Abstract: Phototransduction in retinal rods occurs when the G protein-coupled photoreceptor rhodopsin triggers the activation of phosphodiesterase 6 (PDE6) by GTP-bound alpha subunits of the G protein ...Phototransduction in retinal rods occurs when the G protein-coupled photoreceptor rhodopsin triggers the activation of phosphodiesterase 6 (PDE6) by GTP-bound alpha subunits of the G protein transducin (Gα). Recently, we presented a cryo-EM structure for a complex between two GTP-bound recombinant Gα subunits and native PDE6, that included a bivalent antibody bound to the C-terminal ends of Gα and the inhibitor vardenafil occupying the active sites on the PDEα and PDEβ subunits. We proposed Gα-activated PDE6 by inducing a striking reorientation of the PDEγ subunits away from the catalytic sites. However, questions remained including whether in the absence of the antibody Gα binds to PDE6 in a similar manner as observed when the antibody is present, does Gα activate PDE6 by enabling the substrate cGMP to access the catalytic sites, and how does the lipid membrane enhance PDE6 activation? Here, we demonstrate that 2:1 Gα-PDE6 complexes form with either recombinant or retinal Gα in the absence of the Gα antibody. We show that Gα binding is not necessary for cGMP nor competitive inhibitors to access the active sites; instead, occupancy of the substrate binding sites enables Gα to bind and reposition the PDE6γ subunits to promote catalytic activity. Moreover, we demonstrate by reconstituting Gα-stimulated PDE6 activity in lipid bilayer nanodiscs that the membrane-induced enhancement results from an increase in the apparent binding affinity of Gα for PDE6. These findings provide new insights into how the retinal G protein stimulates rapid catalytic turnover by PDE6 required for dim light vision. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8ugs.cif.gz | 436.4 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8ugs.ent.gz | 283.2 KB | Display | PDB format |
PDBx/mmJSON format | 8ugs.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8ugs_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 8ugs_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 8ugs_validation.xml.gz | 58.9 KB | Display | |
Data in CIF | 8ugs_validation.cif.gz | 86.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ug/8ugs ftp://data.pdbj.org/pub/pdb/validation_reports/ug/8ugs | HTTPS FTP |
-Related structure data
Related structure data | 42234MC 8ufiC 8ugbC 8ulgC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Rod cGMP-specific 3',5'-cyclic phosphodiesterase subunit ... , 2 types, 2 molecules AB
#1: Protein | Mass: 99461.789 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Bos taurus (cattle) References: UniProt: P11541, 3',5'-cyclic-GMP phosphodiesterase |
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#2: Protein | Mass: 98449.648 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Bos taurus (cattle) References: UniProt: P23439, 3',5'-cyclic-GMP phosphodiesterase |
-Protein , 1 types, 2 molecules CD
#3: Protein | Mass: 9684.229 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Bos taurus (cattle) References: UniProt: P04972, 3',5'-cyclic-GMP phosphodiesterase |
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-Non-polymers , 3 types, 8 molecules
#4: Chemical | ChemComp-PCG / #5: Chemical | #6: Chemical | |
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-Details
Has ligand of interest | Y |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Bovine rod phosphodiesterase 6 bound to cGMP / Type: COMPLEX / Entity ID: #1-#3 / Source: NATURAL |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Bos taurus (cattle) |
Buffer solution | pH: 8 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |
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Microscopy | Model: FEI TALOS ARCTICA |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 63000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 1400 nm |
Image recording | Electron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
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3D reconstruction | Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 788557 / Symmetry type: POINT | ||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||
Displacement parameters | Biso mean: 92.07 Å2 | ||||||||||||||||||||||||
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