Journal: Structure / Year: 2020 Title: Lipid Interactions of a Ciliary Membrane TRP Channel: Simulation and Structural Studies of Polycystin-2. Authors: Qinrui Wang / Robin A Corey / George Hedger / Prafulla Aryal / Mariana Grieben / Chady Nasrallah / Agnese Baronina / Ashley C W Pike / Jiye Shi / Elisabeth P Carpenter / Mark S P Sansom / Abstract: Polycystin-2 (PC2) is a transient receptor potential (TRP) channel present in ciliary membranes of the kidney. PC2 shares a transmembrane fold with other TRP channels, in addition to an extracellular ...Polycystin-2 (PC2) is a transient receptor potential (TRP) channel present in ciliary membranes of the kidney. PC2 shares a transmembrane fold with other TRP channels, in addition to an extracellular domain found in TRPP and TRPML channels. Using molecular dynamics (MD) simulations and cryoelectron microscopy we identify and characterize PIP and cholesterol interactions with PC2. PC2 is revealed to have a PIP binding site close to the equivalent vanilloid/lipid binding site in the TRPV1 channel. A 3.0-Å structure reveals a binding site for cholesterol on PC2. Cholesterol interactions with the channel at this site are characterized by MD simulations. The two classes of lipid binding sites are compared with sites observed in other TRPs and in Kv channels. These findings suggest PC2, in common with other ion channels, may be modulated by both PIPs and cholesterol, and position PC2 within an emerging model of the roles of lipids in the regulation and organization of ciliary membranes.
History
Deposition
Oct 28, 2019
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Header (metadata) release
Nov 20, 2019
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Map release
Nov 20, 2019
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Update
Dec 2, 2020
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Current status
Dec 2, 2020
Processing site: PDBe / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
Name: CALCIUM ION / type: ligand / ID: 6 / Number of copies: 1 / Formula: CA
Molecular weight
Theoretical: 40.078 Da
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Experimental details
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Structure determination
Method
cryo EM
Processing
single particle reconstruction
Aggregation state
particle
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Sample preparation
Concentration
4.5 mg/mL
Buffer
pH: 7.5 Component:
Concentration
Formula
Name
20.0 mM
C8H18N2O4S
HEPES
200.0 mM
NaCl
Sodium chloride
20.0 mM
CaCl2
Calcium chloride
0.035 % (w/v)
C23H44O11
N-Undecyl-beta-D-maltopyranoside
Grid
Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR
Vitrification
Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 278 K / Instrument: FEI VITROBOT MARK IV
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Electron microscopy
Microscope
FEI TITAN KRIOS
Image recording
Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-20 / Number grids imaged: 1 / Number real images: 1597 / Average exposure time: 8.0 sec. / Average electron dose: 52.2 e/Å2
Electron beam
Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Images were corrected for local motion using patch-based methods with MotionCor2
Particle selection
Number selected: 98113
CTF correction
Software - Name: CTFFIND (ver. v.4.1.10)
Startup model
Type of model: OTHER / Details: Ab-initio model generated in RELION
Final reconstruction
Number classes used: 1 / Applied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.96 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3) Details: RELION autorefinement. Unmasked resolution is 3.12. After masking to remove the detergent micelle resolution is 2.96A Number images used: 51694
Initial angle assignment
Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3)
Final angle assignment
Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3)
Final 3D classification
Number classes: 10 / Avg.num./class: 7400 / Software - Name: RELION (ver. 3) Details: Final classification carried out after 3D autorefinement with particles split into 10 classes without image alignment
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