+Open data
-Basic information
Entry | Database: PDB / ID: 8ghf | |||||||||
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Title | cryo-EM structure of hSlo1 in plasma membrane vesicles | |||||||||
Components | Calcium-activated potassium channel subunit alpha-1 | |||||||||
Keywords | TRANSPORT PROTEIN / Slo1 / BK channel / Ca2+- and voltage-activated K+ channel / ion channel / plasma membrane vesicles | |||||||||
Function / homology | Function and homology information Acetylcholine inhibits contraction of outer hair cells / micturition / Ca2+ activated K+ channels / large conductance calcium-activated potassium channel activity / response to carbon monoxide / calcium-activated potassium channel activity / negative regulation of cell volume / smooth muscle contraction involved in micturition / intracellular potassium ion homeostasis / Sensory processing of sound by inner hair cells of the cochlea ...Acetylcholine inhibits contraction of outer hair cells / micturition / Ca2+ activated K+ channels / large conductance calcium-activated potassium channel activity / response to carbon monoxide / calcium-activated potassium channel activity / negative regulation of cell volume / smooth muscle contraction involved in micturition / intracellular potassium ion homeostasis / Sensory processing of sound by inner hair cells of the cochlea / response to osmotic stress / cGMP effects / voltage-gated potassium channel activity / voltage-gated potassium channel complex / potassium ion transmembrane transport / potassium ion transport / regulation of membrane potential / caveola / response to calcium ion / vasodilation / actin binding / postsynaptic membrane / response to hypoxia / apical plasma membrane / positive regulation of apoptotic process / identical protein binding / membrane / metal ion binding / plasma membrane Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.7 Å | |||||||||
Authors | Tao, X. / Zhao, C. / MacKinnon, R. | |||||||||
Funding support | United States, 2items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2023 Title: Membrane protein isolation and structure determination in cell-derived membrane vesicles. Authors: Xiao Tao / Chen Zhao / Roderick MacKinnon / Abstract: Integral membrane protein structure determination traditionally requires extraction from cell membranes using detergents or polymers. Here, we describe the isolation and structure determination of ...Integral membrane protein structure determination traditionally requires extraction from cell membranes using detergents or polymers. Here, we describe the isolation and structure determination of proteins in membrane vesicles derived directly from cells. Structures of the ion channel Slo1 from total cell membranes and from cell plasma membranes were determined at 3.8 Å and 2.7 Å resolution, respectively. The plasma membrane environment stabilizes Slo1, revealing an alteration of global helical packing, polar lipid, and cholesterol interactions that stabilize previously unresolved regions of the channel and an additional ion binding site in the Ca regulatory domain. The two methods presented enable structural analysis of both internal and plasma membrane proteins without disrupting weakly interacting proteins, lipids, and cofactors that are essential to biological function. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8ghf.cif.gz | 814.4 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8ghf.ent.gz | 659 KB | Display | PDB format |
PDBx/mmJSON format | 8ghf.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/gh/8ghf ftp://data.pdbj.org/pub/pdb/validation_reports/gh/8ghf | HTTPS FTP |
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-Related structure data
Related structure data | 40044MC 8gh9C 8ghgC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Protein , 1 types, 4 molecules ABCD
#1: Protein | Mass: 122440.102 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: KCNMA1, KCNMA, SLO / Production host: Homo sapiens (human) / References: UniProt: Q12791 |
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-Non-polymers , 5 types, 128 molecules
#2: Chemical | ChemComp-MG / #3: Chemical | ChemComp-CA / #4: Chemical | ChemComp-POV / ( #5: Chemical | ChemComp-CLR / #6: Chemical | ChemComp-NA / |
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-Details
Has ligand of interest | Y |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: ALFA-hSlo1-eGFP ion channel / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Homo sapiens (human) |
Buffer solution | pH: 7.4 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: GOLD / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295.15 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 60 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k) |
EM imaging optics | Energyfilter slit width: 6 eV |
-Processing
Software | Name: PHENIX / Version: 1.20.1_4487: / Classification: refinement | ||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
Symmetry | Point symmetry: C4 (4 fold cyclic) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 32063 / Algorithm: BACK PROJECTION / Symmetry type: POINT | ||||||||||||||||||||||||
Refine LS restraints |
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