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- EMDB-22152: CryoEM structure of GIRK2-PIP2*** - G protein-gated inwardly rect... -

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Basic information

Entry
Database: EMDB / ID: EMD-22152
TitleCryoEM structure of GIRK2-PIP2*** - G protein-gated inwardly rectifying potassium channel GIRK2 with PIP2
Map dataCryoEM structure of GIRK2-PIP2*** - 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-gated inwardly rectifying potassium channel GIRK2
Function / homology
Function and homology information


G-protein activated inward rectifier potassium channel activity / monoatomic ion 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: 7.7 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 14, 2020-
Header (metadata) releaseSep 1, 2021-
Map releaseSep 1, 2021-
UpdateSep 1, 2021-
Current statusSep 1, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0079
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.0079
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_22152.png

Downloads & links

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Map

FileDownload / File: emd_22152.map.gz / Format: CCP4 / Size: 70.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryoEM structure of GIRK2-PIP2*** - G protein-gated inwardly rectifying potassium channel GIRK2 with PIP2
Voxel sizeX=Y=Z: 0.85 Å
Density
Contour LevelBy AUTHOR: 0.0079 / Movie #1: 0.0079
Minimum - Maximum-0.004878668 - 0.016713854
Average (Standard dev.)0.00014259935 (±0.0014532987)
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-0.0050.0170.000

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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-gated inwardly rectifying potassium channel GIRK2

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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: all
Details: The cryoEM structure obtained in the presence of modulator PIP2.
Source (natural)Organism: Mus musculus (house mouse)
Recombinant expressionOrganism: Komagataella pastoris (fungus)

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Macromolecule #1: G protein-gated inwardly rectifying potassium channel GIRK2

MacromoleculeName: G protein-gated inwardly rectifying potassium channel GIRK2
type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Recombinant expressionOrganism: Komagataella pastoris (fungus)
SequenceString: MAKRKIQRYV RKDGKCNVH H GNVRETYR YL TDIFTTL VDL KWRFNL LIFV MVYTV TWLFF GMIW WLIAYI RGD MDHIEDP SW TPCVTNLN G FVSAFLFSI ETETTIGYGY RVITDKCPE G IILLLIQS VL GSIVNAF MVG CMFVKI SQPK KRAET ...String:
MAKRKIQRYV RKDGKCNVH H GNVRETYR YL TDIFTTL VDL KWRFNL LIFV MVYTV TWLFF GMIW WLIAYI RGD MDHIEDP SW TPCVTNLN G FVSAFLFSI ETETTIGYGY RVITDKCPE G IILLLIQS VL GSIVNAF MVG CMFVKI SQPK KRAET LVFST HAVI SMRDGK LCL MFRVGDL RN SHIVEASI R AKLIKSKQT SEGEFIPLNQ TDINVGYYT G DDRLFLVS PL IISHEIN QQS PFWEIS KAQL PKEEL EIVVI LEGM VEATGM TCQ ARSSYIT SE ILWGYRFT P VLTLEDGFY EVDYNSFHET YETSTPSLS A KELAELAN RA ESNSLEV LFQ

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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 mMC10H16N2O8EDTAEthylenediaminetetraacetic acid
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
DetailsGIRK2 with modulator PIP2

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / 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
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
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 2058148
CTF correctionSoftware - Name: CTFFIND
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

3sya

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 7.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 40463
FSC plot (resolution estimation)

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