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- EMDB-22200: Cryo-EM structure of the G protein-gated inward rectifier K+ chan... -

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Basic information

Entry
Database: EMDB / ID: EMD-22200
TitleCryo-EM structure of the G protein-gated inward rectifier K+ channel GIRK2 (Kir3.2) in complex with PIP2
Map dataCryo-EM structure of the G protein-gated inward rectifier K+ channel GIRK2
Sample
  • Complex: Homo-tetrameric assembly of G protein-activated inward rectifier potassium channel 2
    • 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
KeywordsG protein-coupled inwardly rectifying potassium channels / PIP2 / MEMBRANE PROTEIN
Function / homology
Function and homology information


G-protein activated inward rectifier potassium channel activity / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / voltage-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / inward rectifier potassium channel activity / regulation of monoatomic ion transmembrane transport / parallel fiber to Purkinje cell synapse / monoatomic ion channel complex / neuronal cell body membrane / potassium ion import across plasma membrane ...G-protein activated inward rectifier potassium channel activity / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / voltage-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / inward rectifier potassium channel activity / regulation of monoatomic ion transmembrane transport / parallel fiber to Purkinje cell synapse / monoatomic ion channel complex / neuronal cell body membrane / potassium ion import across plasma membrane / potassium channel activity / G-protein alpha-subunit binding / negative regulation of insulin secretion / presynapse / presynaptic membrane / postsynapse / axon / dendrite / cell surface / plasma membrane
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 / G protein-activated inward rectifier potassium channel 2
Similarity search - Component
Biological speciesMus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsNiu Y / Tao X
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM43949 United States
CitationJournal: Elife / Year: 2020
Title: Cryo-EM analysis of PIP regulation in mammalian GIRK channels.
Authors: Yiming Niu / Xiao Tao / Kouki K Touhara / Roderick MacKinnon /
Abstract: G-protein-gated inward rectifier potassium (GIRK) channels are regulated by G proteins and PIP. Here, using cryo-EM single particle analysis we describe the equilibrium ensemble of structures of ...G-protein-gated inward rectifier potassium (GIRK) channels are regulated by G proteins and PIP. Here, using cryo-EM single particle analysis we describe the equilibrium ensemble of structures of neuronal GIRK2 as a function of the C8-PIP concentration. We find that PIP shifts the equilibrium between two distinguishable structures of neuronal GIRK (GIRK2), extended and docked, towards the docked form. In the docked form the cytoplasmic domain, to which G binds, becomes accessible to the cytoplasmic membrane surface where G resides. Furthermore, PIP binding reshapes the G binding surface on the cytoplasmic domain, preparing it to receive G. We find that cardiac GIRK (GIRK1/4) can also exist in both extended and docked conformations. These findings lead us to conclude that PIP influences GIRK channels in a structurally similar manner to Kir2.2 channels. In Kir2.2 channels, the PIP-induced conformational changes open the pore. In GIRK channels, they prepare the channel for activation by G.
History
DepositionJun 21, 2020-
Header (metadata) releaseOct 7, 2020-
Map releaseOct 7, 2020-
UpdateOct 23, 2024-
Current statusOct 23, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.011
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.011
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6xit
  • Surface level: 0.011
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_22200.png

Downloads & links

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Map

FileDownload / File: emd_22200.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM structure of the G protein-gated inward rectifier K+ channel GIRK2
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1 Å/pix.
x 400 pix.
= 400. Å		1 Å/pix.
x 400 pix.
= 400. Å		1 Å/pix.
x 400 pix.
= 400. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0096 / Movie #1: 0.011
Minimum - Maximum-0.015782911 - 0.03328726
Average (Standard dev.)0.00000040459517 (±0.00105614)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 400.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z111
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z400.000400.000400.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ510510510
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-0.0160.0330.000

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Supplemental data

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Sample components

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Entire : Homo-tetrameric assembly of G protein-activated inward rectifier ...

EntireName: Homo-tetrameric assembly of G protein-activated inward rectifier potassium channel 2
Components
  • Complex: Homo-tetrameric assembly of G protein-activated inward rectifier potassium channel 2
    • 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

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Supramolecule #1: Homo-tetrameric assembly of G protein-activated inward rectifier ...

SupramoleculeName: Homo-tetrameric assembly of G protein-activated inward rectifier potassium channel 2
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
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: 3 / Formula: K
Molecular weightTheoretical: 39.098 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

Concentration6 mg/mL
BufferpH: 7.5
Details: 20 mM Tris-HCl pH 7.5, 150 mM KCl, 10 mM DTT, 1 mM DETA, 0.2% DM and 0.01% CHS
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 1-50 / Number grids imaged: 1 / Number real images: 11349 / Average exposure time: 10.0 sec. / Average electron dose: 8.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
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

Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 155128
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software: (Name: RELION (ver. 3.0), cryoSPARC (ver. 2.9.0))
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final 3D classificationNumber classes: 5 / Software - Name: RELION (ver. 3.0)

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Atomic model buiding 1

Initial modelPDB ID:

3sya


Chain - Chain ID: A / Chain - Residue range: 53-380 / Chain - Source name: PDB / Chain - Initial model type: experimental model
Output model

PDB-6xit:
Cryo-EM structure of the G protein-gated inward rectifier K+ channel GIRK2 (Kir3.2) in complex with PIP2

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