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- PDB-7et1: Cryo-EM structure of the gastric proton pump K791S/E820D/Y340N/E9... -
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Basic information
Entry | Database: PDB / ID: 7et1 | ||||||
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Title | Cryo-EM structure of the gastric proton pump K791S/E820D/Y340N/E936V/Y799W 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 | ![]() potassium:proton exchanging ATPase complex / P-type potassium:proton transporter activity / Ion transport by P-type ATPases / 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 ...potassium:proton exchanging ATPase complex / P-type potassium:proton transporter activity / Ion transport by P-type ATPases / 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 / 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: 2.6 Å | ||||||
![]() | Abe, K. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Gastric proton pump with two occluded K engineered with sodium pump-mimetic mutations. Authors: Kazuhiro Abe / Kenta Yamamoto / Katsumasa Irie / Tomohiro Nishizawa / Atsunori Oshima / ![]() Abstract: The gastric H,K-ATPase mediates electroneutral exchange of 1H/1K per ATP hydrolysed across the membrane. Previous structural analysis of the K-occluded E2-P transition state of H,K-ATPase showed a ...The gastric H,K-ATPase mediates electroneutral exchange of 1H/1K per ATP hydrolysed across the membrane. Previous structural analysis of the K-occluded E2-P transition state of H,K-ATPase showed a single bound K at cation-binding site II, in marked contrast to the two K ions occluded at sites I and II of the closely-related Na,K-ATPase which mediates electrogenic 3Na/2K translocation across the membrane. The molecular basis of the different K stoichiometry between these K-counter-transporting pumps is elusive. We show a series of crystal structures and a cryo-EM structure of H,K-ATPase mutants with changes in the vicinity of site I, based on the structure of the sodium pump. Our step-wise and tailored construction of the mutants finally gave a two-K bound H,K-ATPase, achieved by five mutations, including amino acids directly coordinating K (Lys791Ser, Glu820Asp), indirectly contributing to cation-binding site formation (Tyr340Asn, Glu936Val), and allosterically stabilizing K-occluded conformation (Tyr799Trp). This quintuple mutant in the K-occluded E2-P state unambiguously shows two separate densities at the cation-binding site in its 2.6 Å resolution cryo-EM structure. These results offer new insights into how two closely-related cation pumps specify the number of K accommodated at their cation-binding site. | ||||||
History |
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Structure visualization
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Structure viewer | Molecule: ![]() ![]() |
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PDBx/mmCIF format | ![]() | 322.4 KB | Display | ![]() |
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PDB format | ![]() | 245.7 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Summary document | ![]() | 1.3 MB | Display | ![]() |
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Full document | ![]() | 1.4 MB | Display | |
Data in XML | ![]() | 49.8 KB | Display | |
Data in CIF | ![]() | 73.7 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 31294MC ![]() 7eflC ![]() 7efmC ![]() 7efnC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Assembly
Deposited unit | ![]()
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Components
-Protein , 2 types, 2 molecules AB
#1: Protein | Mass: 109099.547 Da / Num. of mol.: 1 / Mutation: K791S/E820D/Y340N/E936V/Y799W Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() |
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#2: Protein | Mass: 29762.107 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, 10 molecules ![](data/chem/img/MG.gif)
![](data/chem/img/PCW.gif)
![](data/chem/img/K.gif)
![](data/chem/img/ALF.gif)
![](data/chem/img/CLR.gif)
![](data/chem/img/HOH.gif)
![](data/chem/img/PCW.gif)
![](data/chem/img/K.gif)
![](data/chem/img/ALF.gif)
![](data/chem/img/CLR.gif)
![](data/chem/img/HOH.gif)
#3: Chemical | ChemComp-MG / | ||||||||
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#4: Chemical | #5: Chemical | #6: Chemical | ChemComp-ALF / | #7: Chemical | ChemComp-CLR / | #9: Water | ChemComp-HOH / | |
-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 |
Specimen support | Grid material: COPPER/RHODIUM / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
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 |
Image recording | Electron dose: 64 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
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Processing
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 2.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 72308 / Symmetry type: POINT | ||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||
Displacement parameters | Biso mean: 63.43 Å2 | ||||||||||||||||||||||||
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