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- PDB-8wa5: Cryo-EM structure of the gastric proton pump Y799W/E936Q mutant i... -
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Basic information
Entry | Database: PDB / ID: 8wa5 | ||||||
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Title | Cryo-EM structure of the gastric proton pump Y799W/E936Q mutant in K+-occluded (K+)E2-AlF state | ||||||
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![]() | MEMBRANE PROTEIN / P-type ATPase / gastric proton pump / primary transporter / transporter | ||||||
Function / homology | ![]() regulation of proton transport / pH reduction / potassium:proton exchanging ATPase complex / P-type potassium:proton transporter activity / Ion transport by P-type ATPases / P-type sodium:potassium-exchanging transporter activity / sodium:potassium-exchanging ATPase complex / sodium ion export across plasma membrane / intracellular potassium ion homeostasis / intracellular sodium ion homeostasis ...regulation of proton transport / pH reduction / potassium:proton exchanging ATPase complex / P-type potassium:proton transporter activity / Ion transport by P-type ATPases / P-type sodium:potassium-exchanging transporter activity / sodium:potassium-exchanging ATPase complex / sodium ion export across plasma membrane / intracellular potassium ion homeostasis / intracellular sodium ion homeostasis / potassium ion import across plasma membrane / ATPase activator activity / potassium ion transmembrane transport / proton transmembrane transport / cell adhesion / response to xenobiotic stimulus / apical plasma membrane / magnesium ion binding / ATP hydrolysis activity / ATP binding Similarity search - Function | ||||||
Biological species | ![]() ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.51 Å | ||||||
![]() | Abe, K. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Specific protonation of acidic residues confers K selectivity to the gastric proton pump. Authors: Hridya Valia Madapally / Kazuhiro Abe / Vikas Dubey / Himanshu Khandelia / ![]() ![]() Abstract: The gastric proton pump (H,K-ATPase) transports a proton into the stomach lumen for every K ion exchanged in the opposite direction. In the lumen-facing state of the pump (E2), the pump selectively ...The gastric proton pump (H,K-ATPase) transports a proton into the stomach lumen for every K ion exchanged in the opposite direction. In the lumen-facing state of the pump (E2), the pump selectively binds K despite the presence of a 10-fold higher concentration of Na. The molecular basis for the ion selectivity of the pump is unknown. Using molecular dynamics simulations, free energy calculations, and Na and K-dependent ATPase activity assays, we demonstrate that the K selectivity of the pump depends upon the simultaneous protonation of the acidic residues E343 and E795 in the ion-binding site. We also show that when E936 is protonated, the pump becomes Na sensitive. The protonation-mimetic mutant E936Q exhibits weak Na-activated ATPase activity. A 2.5-Å resolution cryo-EM structure of the E936Q mutant in the K-occluded E2-Pi form shows, however, no significant structural difference compared with wildtype except less-than-ideal coordination of K in the mutant. The selectivity toward a specific ion correlates with a more rigid and less fluctuating ion-binding site. Despite being exposed to a pH of 1, the fundamental principle driving the K ion selectivity of H,K-ATPase is similar to that of Na,K-ATPase: the ionization states of the acidic residues in the ion-binding sites determine ion selectivity. Unlike the Na,K-ATPase, however, protonation of an ion-binding glutamate residue (E936) confers Na sensitivity. | ||||||
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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PDBx/mmCIF format | ![]() | 360.4 KB | Display | ![]() |
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PDB format | ![]() | 238.5 KB | Display | ![]() |
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-Validation report
Summary document | ![]() | 1.3 MB | Display | ![]() |
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Full document | ![]() | 1.3 MB | Display | |
Data in XML | ![]() | 53.8 KB | Display | |
Data in CIF | ![]() | 79.3 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 37391MC M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Components
-Protein , 2 types, 2 molecules AB
#1: Protein | Mass: 114282.594 Da / Num. of mol.: 1 / Mutation: Y799W/E936Q Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() |
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#2: Protein | Mass: 33113.844 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() |
-Sugars , 1 types, 5 molecules ![](data/chem/img/NAG.gif)
#8: Sugar | ChemComp-NAG / |
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-Non-polymers , 6 types, 13 molecules ![](data/chem/img/K.gif)
![](data/chem/img/ALF.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/CLR.gif)
![](data/chem/img/PCW.gif)
![](data/chem/img/HOH.gif)
![](data/chem/img/ALF.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/CLR.gif)
![](data/chem/img/PCW.gif)
![](data/chem/img/HOH.gif)
#3: Chemical | #4: Chemical | ChemComp-ALF / | #5: Chemical | ChemComp-MG / | #6: Chemical | ChemComp-CLR / | #7: Chemical | ChemComp-PCW / | #9: Water | ChemComp-HOH / | |
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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 |
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Sample preparation
Component | Name: alpha-beta complex of the gastric proton pump / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT |
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Molecular weight | Value: 0.135 MDa / Experimental value: YES |
Source (natural) | Organism: ![]() ![]() |
Source (recombinant) | Organism: ![]() |
Buffer solution | pH: 6.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
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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: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 1600 nm / Nominal defocus min: 800 nm |
Image recording | Electron dose: 64 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
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Processing
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
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3D reconstruction | Resolution: 2.51 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 302608 / Symmetry type: POINT | ||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||
Displacement parameters | Biso mean: 54.08 Å2 | ||||||||||||||||||||||||
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