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- EMDB-22150: CryoEM structure of GIRK2PIP2* - G protein-gated inwardly rectify... -

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Basic information

Entry
Database: EMDB / ID: EMD-22150
TitleCryoEM structure of GIRK2PIP2* - G protein-gated inwardly rectifying potassium channel GIRK2 with PIP2
Map dataCryoEM structure of GIRK2PIP2* - G protein-gated inwardly rectifying potassium channel GIRK2 with PIP2
Sample
  • Complex: G protein-gated inwardly rectifying potassium channel (GIRK2)
    • Protein or peptide: G protein-activated inward rectifier potassium channel 2
  • Ligand: [(2R)-2-octanoyloxy-3-[oxidanyl-[(1R,2R,3S,4R,5R,6S)-2,3,6-tris(oxidanyl)-4,5-diphosphonooxy-cyclohexyl]oxy-phosphoryl]oxy-propyl] octanoate
  • Ligand: POTASSIUM ION
  • Ligand: SODIUM ION
KeywordsGIRK / inwardly rectifying potassium channel / cholesterol / lipids / PIP2 / TRANSPORT PROTEIN
Function / homology
Function and homology information


G-protein activated inward rectifier potassium channel activity / inward rectifier potassium channel complex
Similarity search - Function
Potassium channel, inwardly rectifying, Kir3.2 / Potassium channel, inwardly rectifying, transmembrane domain / Inward rectifier potassium channel transmembrane domain / Potassium channel, inwardly rectifying, Kir, cytoplasmic / Potassium channel, inwardly rectifying, Kir / Inward rectifier potassium channel, C-terminal / Inward rectifier potassium channel C-terminal domain / Immunoglobulin E-set
Similarity search - Domain/homology
G protein-activated inward rectifier potassium channel 2
Similarity search - Component
Biological speciesMus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsMathiharan YK / Glaaser IW / Skiniotis G / Slesinger PA
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute on Alcohol Abuse and Alcoholism (NIH/NIAAA)AA018734 United States
CitationJournal: Cell Rep / Year: 2021
Title: Structural insights into GIRK2 channel modulation by cholesterol and PIP.
Authors: Yamuna Kalyani Mathiharan / Ian W Glaaser / Yulin Zhao / Michael J Robertson / Georgios Skiniotis / Paul A Slesinger /
Abstract: G-protein-gated inwardly rectifying potassium (GIRK) channels are important for determining neuronal excitability. In addition to G proteins, GIRK channels are potentiated by membrane cholesterol, ...G-protein-gated inwardly rectifying potassium (GIRK) channels are important for determining neuronal excitability. In addition to G proteins, GIRK channels are potentiated by membrane cholesterol, which is elevated in the brains of people with neurodegenerative diseases such as Alzheimer's dementia and Parkinson's disease. The structural mechanism of cholesterol modulation of GIRK channels is not well understood. In this study, we present cryo- electron microscopy (cryoEM) structures of GIRK2 in the presence and absence of the cholesterol analog cholesteryl hemisuccinate (CHS) and phosphatidylinositol 4,5-bisphosphate (PIP). The structures reveal that CHS binds near PIP in lipid-facing hydrophobic pockets of the transmembrane domain. Our structural analysis suggests that CHS stabilizes PIP interaction with the channel and promotes engagement of the cytoplasmic domain onto the transmembrane region. Mutagenesis of one of the CHS binding pockets eliminates cholesterol-dependent potentiation of GIRK2. Elucidating the structural mechanisms underlying cholesterol modulation of GIRK2 channels could facilitate the development of therapeutics for treating neurological diseases. VIDEO ABSTRACT.
History
DepositionJun 13, 2020-
Header (metadata) releaseSep 1, 2021-
Map releaseSep 1, 2021-
UpdateNov 6, 2024-
Current statusNov 6, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0144
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.0144
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6xeu
  • Surface level: 0.0144
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_22150.png

Downloads & links

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Map

FileDownload / File: emd_22150.map.gz / Format: CCP4 / Size: 70.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryoEM structure of GIRK2PIP2* - G protein-gated inwardly rectifying potassium channel GIRK2 with PIP2
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.85 Å/pix.
x 264 pix.
= 224.4 Å		0.85 Å/pix.
x 264 pix.
= 224.4 Å		0.85 Å/pix.
x 264 pix.
= 224.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.85 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0144 / Movie #1: 0.0144
Minimum - Maximum-0.01722162 - 0.048360944
Average (Standard dev.)0.00012183783 (±0.0020044043)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions264264264
Spacing264264264
CellA=B=C: 224.40001 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.850.850.85
M x/y/z264264264
origin x/y/z0.0000.0000.000
length x/y/z224.400224.400224.400
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS264264264
D min/max/mean-0.0170.0480.000

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Supplemental data

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Sample components

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Entire : G protein-gated inwardly rectifying potassium channel (GIRK2)

EntireName: G protein-gated inwardly rectifying potassium channel (GIRK2)
Components
  • Complex: G protein-gated inwardly rectifying potassium channel (GIRK2)
    • Protein or peptide: G protein-activated inward rectifier potassium channel 2
  • Ligand: [(2R)-2-octanoyloxy-3-[oxidanyl-[(1R,2R,3S,4R,5R,6S)-2,3,6-tris(oxidanyl)-4,5-diphosphonooxy-cyclohexyl]oxy-phosphoryl]oxy-propyl] octanoate
  • Ligand: POTASSIUM ION
  • Ligand: SODIUM ION

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Supramolecule #1: G protein-gated inwardly rectifying potassium channel (GIRK2)

SupramoleculeName: G protein-gated inwardly rectifying potassium channel (GIRK2)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: The cryoEM structure obtained in the presence of modulator PIP2.
Source (natural)Organism: Mus musculus (house mouse)

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Macromolecule #1: G protein-activated inward rectifier potassium channel 2

MacromoleculeName: G protein-activated inward rectifier potassium channel 2
type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 39.061957 KDa
Recombinant expressionOrganism: Komagataella pastoris (fungus)
SequenceString: MAKRKIQRYV RKDGKCNVHH GNVRETYRYL TDIFTTLVDL KWRFNLLIFV MVYTVTWLFF GMIWWLIAYI RGDMDHIEDP SWTPCVTNL NGFVSAFLFS IETETTIGYG YRVITDKCPE GIILLLIQSV LGSIVNAFMV GCMFVKISQP KKRAETLVFS T HAVISMRD ...String:
MAKRKIQRYV RKDGKCNVHH GNVRETYRYL TDIFTTLVDL KWRFNLLIFV MVYTVTWLFF GMIWWLIAYI RGDMDHIEDP SWTPCVTNL NGFVSAFLFS IETETTIGYG YRVITDKCPE GIILLLIQSV LGSIVNAFMV GCMFVKISQP KKRAETLVFS T HAVISMRD GKLCLMFRVG DLRNSHIVEA SIRAKLIKSK QTSEGEFIPL NQTDINVGYY TGDDRLFLVS PLIISHEINQ QS PFWEISK AQLPKEELEI VVILEGMVEA TGMTCQARSS YITSEILWGY RFTPVLTLED GFYEVDYNSF HETYETSTPS LSA KELAEL ANRAESNSLE VLFQ

UniProtKB: G protein-activated inward rectifier potassium channel 2

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Macromolecule #2: [(2R)-2-octanoyloxy-3-[oxidanyl-[(1R,2R,3S,4R,5R,6S)-2,3,6-tris(o...

MacromoleculeName: [(2R)-2-octanoyloxy-3-[oxidanyl-[(1R,2R,3S,4R,5R,6S)-2,3,6-tris(oxidanyl)-4,5-diphosphonooxy-cyclohexyl]oxy-phosphoryl]oxy-propyl] octanoate
type: ligand / ID: 2 / Number of copies: 4 / Formula: PIO
Molecular weightTheoretical: 746.566 Da
Chemical component information

ChemComp-PIO:
[(2R)-2-octanoyloxy-3-[oxidanyl-[(1R,2R,3S,4R,5R,6S)-2,3,6-tris(oxidanyl)-4,5-diphosphonooxy-cyclohexyl]oxy-phosphoryl]oxy-propyl] octanoate

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Macromolecule #3: POTASSIUM ION

MacromoleculeName: POTASSIUM ION / type: ligand / ID: 3 / Number of copies: 8 / Formula: K
Molecular weightTheoretical: 39.098 Da

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Macromolecule #4: SODIUM ION

MacromoleculeName: SODIUM ION / type: ligand / ID: 4 / Number of copies: 4
Molecular weightTheoretical: 22.99 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

Concentration7 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
20.0 mMC4H11NO3Tris
150.0 mMKClPotassium Chloride
20.0 mMC4H10O2S2DTT
3.0 mMC9H15O6PTCEP
1.0 mMC10H16N2O8EDTA
0.025 %C24H46O11DDM
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV
DetailsGIRK2PIP2* with the modulator PIP2

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number real images: 6480 / Average exposure time: 3.0 sec. / Average electron dose: 83.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.8 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 2058148
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

3sya

Final reconstructionNumber classes used: 2 / Applied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 154071
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

3sya


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementProtocol: FLEXIBLE FIT
Output model

PDB-6xeu:
CryoEM structure of GIRK2PIP2* - G protein-gated inwardly rectifying potassium channel GIRK2 with PIP2

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