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- EMDB-3524: cryoEM Structure of Polycystin-2 in complex with calcium and lipids -

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Basic information

Entry
Database: EMDB / ID: EMD-3524
TitlecryoEM Structure of Polycystin-2 in complex with calcium and lipids
Map data
Sample
  • Complex: Polycystin-2
    • Protein or peptide: Polycystin-2
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: 1,2-DIPALMITOYL-SN-GLYCERO-3-PHOSPHATE
  • Ligand: PALMITIC ACID
  • Ligand: 4-AMINO-5-CYCLOHEXYL-3-HYDROXY-PENTANOIC ACID
  • Ligand: CALCIUM ION
KeywordsCa2+ signaling / cryoEM / membrane protein structure / Polycystin-2 / TRP channel / transport protein
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 / renal tubule morphogenesis ...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 / renal tubule morphogenesis / determination of liver left/right asymmetry / HLH domain binding / metanephric ascending thin limb development / metanephric mesenchyme development / metanephric S-shaped body morphogenesis / basal cortex / renal artery morphogenesis / positive regulation of inositol 1,4,5-trisphosphate-sensitive calcium-release channel activity / calcium-induced calcium release activity / migrasome / cilium organization / VxPx cargo-targeting to cilium / detection of mechanical stimulus / muscle alpha-actinin binding / regulation of calcium ion import / voltage-gated monoatomic ion channel activity / placenta blood vessel development / cellular response to hydrostatic pressure / cation channel complex / cellular response to fluid shear stress / outward rectifier potassium channel activity / actinin binding / cellular response to osmotic stress / non-motile cilium / determination of left/right symmetry / inorganic cation transmembrane transport / voltage-gated monoatomic cation channel activity / aorta development / neural tube development / motile cilium / voltage-gated sodium channel activity / ciliary membrane / branching involved in ureteric bud morphogenesis / protein heterotetramerization / negative regulation of G1/S transition of mitotic cell cycle / spinal cord development / cytoplasmic side of endoplasmic reticulum membrane / heart looping / centrosome duplication / voltage-gated potassium channel activity / cell surface receptor signaling pathway via JAK-STAT / potassium channel activity / embryonic placenta development / voltage-gated calcium channel activity / transcription regulator inhibitor activity / monoatomic cation channel activity / cytoskeletal protein binding / cellular response to cAMP / release of sequestered calcium ion into cytosol / potassium ion transmembrane transport / sodium ion transmembrane transport / cytoplasmic vesicle membrane / cellular response to calcium ion / liver development / basal plasma membrane / lumenal side of endoplasmic reticulum membrane / cellular response to reactive oxygen species / establishment of localization in cell / phosphoprotein binding / protein tetramerization / calcium ion transmembrane transport / Wnt signaling pathway / intracellular calcium ion homeostasis / calcium ion transport / mitotic spindle / 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 / basolateral plasma membrane / protein homotetramerization / transmembrane transporter binding / cell surface receptor signaling pathway / regulation of cell cycle / ciliary basal body / cilium / signaling receptor binding / negative regulation of cell population proliferation / calcium ion binding / positive regulation of gene expression / endoplasmic reticulum membrane / endoplasmic reticulum / Golgi apparatus / protein homodimerization activity / positive regulation of transcription by RNA polymerase II / extracellular exosome / identical protein binding
Similarity search - Function
Ferredoxin I 4Fe-4S cluster domain / : / Polycystic kidney disease type 2 protein / Polycystin domain / Polycystin domain / 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)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsWilkes M / Madej MG
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-
UpdateNov 13, 2024-
Current statusNov 13, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0354
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.0354
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-5mkf
  • Surface level: 0.0354
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3524.png

Downloads & links

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Map

FileDownload / File: emd_3524.map.gz / Format: CCP4 / Size: 18.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.14 Å/pix.
x 168 pix.
= 191.52 Å		1.14 Å/pix.
x 168 pix.
= 191.52 Å		1.14 Å/pix.
x 168 pix.
= 191.52 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.14 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0354 / Movie #1: 0.0354
Minimum - Maximum-0.07413489 - 0.1353984
Average (Standard dev.)0.0011936896 (±0.0071791406)
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-0.0740.1350.001

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Supplemental data

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Sample components

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Entire : Polycystin-2

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

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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)

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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

UniProtKB: Polycystin-2

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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: 12 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Macromolecule #4: 1,2-DIPALMITOYL-SN-GLYCERO-3-PHOSPHATE

MacromoleculeName: 1,2-DIPALMITOYL-SN-GLYCERO-3-PHOSPHATE / type: ligand / ID: 4 / 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 #5: PALMITIC ACID

MacromoleculeName: PALMITIC ACID / type: ligand / ID: 5 / Number of copies: 12 / Formula: PLM
Molecular weightTheoretical: 256.424 Da
Chemical component information

ChemComp-PLM:
PALMITIC ACID

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Macromolecule #6: 4-AMINO-5-CYCLOHEXYL-3-HYDROXY-PENTANOIC ACID

MacromoleculeName: 4-AMINO-5-CYCLOHEXYL-3-HYDROXY-PENTANOIC ACID / type: ligand / ID: 6 / 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 #7: CALCIUM ION

MacromoleculeName: CALCIUM ION / type: ligand / ID: 7 / Number of copies: 5 / 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
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 1.8 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 4.1 mm

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Image processing

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

3j5q

Final reconstructionApplied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 35318
Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: PROJECTION MATCHING
FSC plot (resolution estimation)

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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.
RefinementSpace: REAL
Output model

PDB-5mkf:
cryoEM Structure of Polycystin-2 in complex with calcium and lipids

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