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- EMDB-3523: cryoEM Structure of Polycystin-2 in complex with cations and lipids -

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Basic information

Entry
Database: EMDB / ID: EMD-3523
TitlecryoEM Structure of Polycystin-2 in complex with cations and lipids
Map data
Sample
  • Complex: Polycystin-2Polycystin 2
    • Protein or peptide: Polycystin-2Polycystin 2
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: 4-AMINO-5-CYCLOHEXYL-3-HYDROXY-PENTANOIC ACID
  • Ligand: 1,2-DIPALMITOYL-SN-GLYCERO-3-PHOSPHATE
  • Ligand: PALMITIC ACID
  • Ligand: CALCIUM IONCalcium
Function / homology
Function and homology information


detection of nodal flow / metanephric smooth muscle tissue development / metanephric cortex development / metanephric cortical collecting duct development / metanephric distal tubule development / polycystin complex / mesonephric tubule development / mesonephric duct development / : / metanephric part of ureteric bud development ...detection of nodal flow / metanephric smooth muscle tissue development / metanephric cortex development / metanephric cortical collecting duct development / metanephric distal tubule development / polycystin complex / mesonephric tubule development / mesonephric duct development / : / metanephric part of ureteric bud development / determination of liver left/right asymmetry / renal tubule morphogenesis / metanephric ascending thin limb development / HLH domain binding / basal cortex / metanephric mesenchyme development / metanephric S-shaped body morphogenesis / renal artery morphogenesis / positive regulation of inositol 1,4,5-trisphosphate-sensitive calcium-release channel activity / migrasome / cilium organization / VxPx cargo-targeting to cilium / detection of mechanical stimulus / regulation of calcium ion import / cation channel complex / calcium-induced calcium release activity / muscle alpha-actinin binding / placenta blood vessel development / voltage-gated monoatomic ion channel activity / cellular response to hydrostatic pressure / outward rectifier potassium channel activity / voltage-gated monoatomic cation channel activity / non-motile cilium / cellular response to fluid shear stress / cellular response to osmotic stress / voltage-gated sodium channel activity / inorganic cation transmembrane transport / actinin binding / transcription regulator inhibitor activity / motile cilium / determination of left/right symmetry / neural tube development / aorta development / protein heterotetramerization / ciliary membrane / branching involved in ureteric bud morphogenesis / negative regulation of G1/S transition of mitotic cell cycle / spinal cord development / heart looping / cytoplasmic side of endoplasmic reticulum membrane / voltage-gated potassium channel activity / cell surface receptor signaling pathway via JAK-STAT / potassium channel activity / centrosome duplication / sodium ion transmembrane transport / negative regulation of ryanodine-sensitive calcium-release channel activity / voltage-gated calcium channel activity / embryonic placenta development / monoatomic cation channel activity / cellular response to cAMP / release of sequestered calcium ion into cytosol / potassium ion transmembrane transport / cellular response to calcium ion / cytoskeletal protein binding / basal plasma membrane / ciliary basal body / liver development / establishment of localization in cell / lumenal side of endoplasmic reticulum membrane / calcium ion transmembrane transport / protein tetramerization / phosphoprotein binding / cytoplasmic vesicle membrane / cilium / intracellular calcium ion homeostasis / mitotic spindle / Wnt signaling pathway / cellular response to reactive oxygen species / calcium ion transport / positive regulation of nitric oxide biosynthetic process / cell-cell junction / lamellipodium / regulation of cell population proliferation / heart development / ATPase binding / positive regulation of cytosolic calcium ion concentration / protein homotetramerization / basolateral plasma membrane / transmembrane transporter binding / regulation of cell cycle / negative regulation of cell population proliferation / signaling receptor binding / calcium ion binding / endoplasmic reticulum membrane / positive regulation of gene expression / Golgi apparatus / endoplasmic reticulum / protein homodimerization activity / positive regulation of transcription by RNA polymerase II / extracellular exosome
Similarity search - Function
Ferredoxin I 4Fe-4S cluster domain / Polycystin domain / Polycystin domain / Polycystic kidney disease type 2 protein / Polycystin cation channel, PKD1/PKD2 / Polycystin cation channel / Voltage-dependent channel domain superfamily / EF-hand calcium-binding domain profile. / EF-hand domain / EF-hand domain pair
Similarity search - Domain/homology
Biological specieshomo sapiens (human) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.3 Å
AuthorsWilkes M / Madej MG / Ziegler C
CitationJournal: Nat Struct Mol Biol / Year: 2017
Title: Molecular insights into lipid-assisted Ca regulation of the TRP channel Polycystin-2.
Authors: Martin Wilkes / M Gregor Madej / Lydia Kreuter / Daniel Rhinow / Veronika Heinz / Silvia De Sanctis / Sabine Ruppel / Rebecca M Richter / Friederike Joos / Marina Grieben / Ashley C W Pike / ...Authors: Martin Wilkes / M Gregor Madej / Lydia Kreuter / Daniel Rhinow / Veronika Heinz / Silvia De Sanctis / Sabine Ruppel / Rebecca M Richter / Friederike Joos / Marina Grieben / Ashley C W Pike / Juha T Huiskonen / Elisabeth P Carpenter / Werner Kühlbrandt / Ralph Witzgall / Christine Ziegler /
Abstract: Polycystin-2 (PC2), a calcium-activated cation TRP channel, is involved in diverse Ca signaling pathways. Malfunctioning Ca regulation in PC2 causes autosomal-dominant polycystic kidney disease. Here ...Polycystin-2 (PC2), a calcium-activated cation TRP channel, is involved in diverse Ca signaling pathways. Malfunctioning Ca regulation in PC2 causes autosomal-dominant polycystic kidney disease. Here we report two cryo-EM structures of distinct channel states of full-length human PC2 in complex with lipids and cations. The structures reveal conformational differences in the selectivity filter and in the large exoplasmic domain (TOP domain), which displays differing N-glycosylation. The more open structure has one cation bound below the selectivity filter (single-ion mode, PC2), whereas multiple cations are bound along the translocation pathway in the second structure (multi-ion mode, PC2). Ca binding at the entrance of the selectivity filter suggests Ca blockage in PC2, and we observed density for the Ca-sensing C-terminal EF hand in the unblocked PC2 state. The states show altered interactions of lipids with the pore loop and TOP domain, thus reflecting the functional diversity of PC2 at different locations, owing to different membrane compositions.
History
DepositionDec 4, 2016-
Header (metadata) releaseDec 28, 2016-
Map releaseJan 18, 2017-
UpdateJul 29, 2020-
Current statusJul 29, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0345
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.0345
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-5mke
  • Surface level: 0.0345
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3523.png

Downloads & links

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Map

FileDownload / File: emd_3523.map.gz / Format: CCP4 / Size: 18.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.14 Å
Density
Contour LevelBy AUTHOR: 0.0345 / Movie #1: 0.0345
Minimum - Maximum-0.06904477 - 0.09819269
Average (Standard dev.)0.001177011 (±0.0061609545)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions168168168
Spacing168168168
CellA=B=C: 191.52 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.141.141.14
M x/y/z168168168
origin x/y/z0.0000.0000.000
length x/y/z191.520191.520191.520
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS168168168
D min/max/mean-0.0690.0980.001

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Supplemental data

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Sample components

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Entire : Polycystin-2

EntireName: Polycystin-2Polycystin 2
Components
  • Complex: Polycystin-2Polycystin 2
    • Protein or peptide: Polycystin-2Polycystin 2
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: 4-AMINO-5-CYCLOHEXYL-3-HYDROXY-PENTANOIC ACID
  • Ligand: 1,2-DIPALMITOYL-SN-GLYCERO-3-PHOSPHATE
  • Ligand: PALMITIC ACID
  • Ligand: CALCIUM IONCalcium

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Supramolecule #1: Polycystin-2

SupramoleculeName: Polycystin-2 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: homo sapiens (human)
Recombinant expressionOrganism: homo sapiens (human)

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Macromolecule #1: Polycystin-2

MacromoleculeName: Polycystin-2 / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 109.820086 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MVNSSRVQPQ QPGDAKRPPA PRAPDPGRLM AGCAAVGASL AAPGGLCEQR GLEIEMQRIR QAAARDPPAG AAASPSPPLS SCSRQAWSR DNPGFEAEEE EEEVEGEEGG MVVEMDVEWR PGSRRSAASS AVSSVGARSR GLGGYHGAGH PSGRRRRRED Q GPPCPSPV ...String:
MVNSSRVQPQ QPGDAKRPPA PRAPDPGRLM AGCAAVGASL AAPGGLCEQR GLEIEMQRIR QAAARDPPAG AAASPSPPLS SCSRQAWSR DNPGFEAEEE EEEVEGEEGG MVVEMDVEWR PGSRRSAASS AVSSVGARSR GLGGYHGAGH PSGRRRRRED Q GPPCPSPV GGGDPLHRHL PLEGQPPRVA WAERLVRGLR GLWGTRLMEE SSTNREKYLK SVLRELVTYL LFLIVLCILT YG MMSSNVY YYTRMMSQLF LDTPVSKTEK TNFKTLSSME DFWKFTEGSL LDGLYWKMQP SNQTEADNRS FIFYENLLLG VPR IRQLRV RNGSCSIPQD LRDEIKECYD VYSVSSEDRA PFGPRNGTAW IYTSEKDLNG SSHWGIIATY SGAGYYLDLS RTRE ETAAQ VASLKKNVWL DRGTRATFID FSVYNANINL FCVVRLLVEF PATGGVIPSW QFQPLKLIRY VTTFDFFLAA CEIIF CFFI FYYVVEEILE IRIHKLHYFR SFWNCLDVVI VVLSVVAIGI NIYRTSNVEV LLQFLEDQNT FPNFEHLAYW QIQFNN IAA VTVFFVWIKL FKFINFNRTM SQLSTTMSRC AKDLFGFAIM FFIIFLAYAQ LAYLVFGTQV DDFSTFQECI FTQFRII LG DINFAEIEEA NRVLGPIYFT TFVFFMFFIL LNMFLAIIND TYSEVKSDLA QQKAEMELSD LIRKGYHKAL VKLKLKKN T VDDISESLRQ GGGKLNFDEL RQDLKGKGHT DAEIEAIFTK YDQDGDQELT EHEHQQMRDD LEKEREDLDL DHSSLPRPM SSRSFPRSLD DSEEDDDEDS GHSSRRRGSI SSGVSYEEFQ VLVRRVDRME HSIGSIVSKI DAVIVKLEIM ERAKLKRREV LGRLLDGVA EDERLGRDSE IHREQMERLV REELERWESD DAASQISHGL GTPVGLNGQP RPRSSRPSSS QSTEGMEGAG G NGSSNVHV

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Macromolecule #3: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 3 / Number of copies: 10 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose / N-Acetylglucosamine

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Macromolecule #4: 4-AMINO-5-CYCLOHEXYL-3-HYDROXY-PENTANOIC ACID

MacromoleculeName: 4-AMINO-5-CYCLOHEXYL-3-HYDROXY-PENTANOIC ACID / type: ligand / ID: 4 / Number of copies: 8 / Formula: CHS
Molecular weightTheoretical: 215.289 Da
Chemical component information

ChemComp-CHS:
4-AMINO-5-CYCLOHEXYL-3-HYDROXY-PENTANOIC ACID

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Macromolecule #5: 1,2-DIPALMITOYL-SN-GLYCERO-3-PHOSPHATE

MacromoleculeName: 1,2-DIPALMITOYL-SN-GLYCERO-3-PHOSPHATE / type: ligand / ID: 5 / Number of copies: 4 / Formula: PX6
Molecular weightTheoretical: 647.883 Da
Chemical component information

ChemComp-PX6:
1,2-DIPALMITOYL-SN-GLYCERO-3-PHOSPHATE

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Macromolecule #6: PALMITIC ACID

MacromoleculeName: PALMITIC ACID / type: ligand / ID: 6 / Number of copies: 12 / Formula: PLM
Molecular weightTheoretical: 256.424 Da
Chemical component information

ChemComp-PLM:
PALMITIC ACID / Palmitic acid

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Macromolecule #7: CALCIUM ION

MacromoleculeName: CALCIUM ION / type: ligand / ID: 7 / Number of copies: 2 / Formula: CA
Molecular weightTheoretical: 40.078 Da

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeJEOL 3200FSC
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 4.1 mm
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 1.8 e/Å2

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

3j5q

Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: PROJECTION MATCHING
Final reconstructionApplied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 42268

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Atomic model buiding 1

DetailsWe used for comparative structure modeling TRPA1 (pdb entry code 3J9P) as template for S1 and S3-S5, TRPV1 (pdb entry code 3J5Q) for S5-S6, and the TRPV2 (pdb entry code 5AN8) fitted best for S2-S3 to obtain an initial model. The soluble domain was build based on pdbID: 5K47. But we had no search model for molecular replacement. Although we had a good idea what the architecture would be like, we build the model de novo with COOT.
Output model

PDB-5mke:
cryoEM Structure of Polycystin-2 in complex with cations and lipids

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