[English] 日本語
Yorodumi
- PDB-7bgy: Cryo-EM Structure of KdpFABC in E2Pi state with MgF4 -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 7bgy
TitleCryo-EM Structure of KdpFABC in E2Pi state with MgF4
Components(Potassium-transporting ATPase ...) x 4
KeywordsMEMBRANE PROTEIN / P-type ATPase / ATP-dependent potassium pump
Function / homology
Function and homology information


P-type K+ transporter / P-type potassium transmembrane transporter activity / potassium:proton antiporter complex / potassium ion-transporting ATPase complex / monoatomic cation transmembrane transport / potassium ion binding / potassium ion transmembrane transport / potassium ion transport / magnesium ion binding / ATP hydrolysis activity ...P-type K+ transporter / P-type potassium transmembrane transporter activity / potassium:proton antiporter complex / potassium ion-transporting ATPase complex / monoatomic cation transmembrane transport / potassium ion binding / potassium ion transmembrane transport / potassium ion transport / magnesium ion binding / ATP hydrolysis activity / ATP binding / plasma membrane
Similarity search - Function
Potassium-transporting ATPase C chain / Potassium-transporting ATPase A chain / K+ transporting P-type ATPase, F subunit / K+-transporting ATPase, c chain / Potassium-transporting ATPase A subunit / F subunit of K+-transporting ATPase (Potass_KdpF) / P-type ATPase, B chain, subfamily IA / E1-E2 ATPase / P-type ATPase, haloacid dehalogenase domain / P-type ATPase, phosphorylation site ...Potassium-transporting ATPase C chain / Potassium-transporting ATPase A chain / K+ transporting P-type ATPase, F subunit / K+-transporting ATPase, c chain / Potassium-transporting ATPase A subunit / F subunit of K+-transporting ATPase (Potass_KdpF) / P-type ATPase, B chain, subfamily IA / E1-E2 ATPase / P-type ATPase, haloacid dehalogenase domain / P-type ATPase, phosphorylation site / P-type ATPase, cytoplasmic domain N / E1-E2 ATPases phosphorylation site. / P-type ATPase, A domain superfamily / P-type ATPase / P-type ATPase, transmembrane domain superfamily / haloacid dehalogenase-like hydrolase / HAD superfamily / HAD-like superfamily
Similarity search - Domain/homology
Chem-9Y0 / TETRAFLUOROMAGNESATE(2-) / Chem-PEV / Potassium-transporting ATPase potassium-binding subunit / Potassium-transporting ATPase ATP-binding subunit / Potassium-transporting ATPase KdpC subunit / Potassium-transporting ATPase KdpF subunit
Similarity search - Component
Biological speciesEscherichia coli K-12 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsSweet, M.E. / Larsen, C. / Pedersen, B.P. / Stokes, D.L.
Funding support United States, Denmark, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01 GM108043 United States
European Research Council (ERC)637372 Denmark
Danish Council for Independent ResearchDFF-8021-00161 Denmark
CitationJournal: Proc Natl Acad Sci U S A / Year: 2021
Title: Structural basis for potassium transport in prokaryotes by KdpFABC.
Authors: Marie E Sweet / Casper Larsen / Xihui Zhang / Michael Schlame / Bjørn P Pedersen / David L Stokes /
Abstract: KdpFABC is an oligomeric K transport complex in prokaryotes that maintains ionic homeostasis under stress conditions. The complex comprises a channel-like subunit (KdpA) from the superfamily of K ...KdpFABC is an oligomeric K transport complex in prokaryotes that maintains ionic homeostasis under stress conditions. The complex comprises a channel-like subunit (KdpA) from the superfamily of K transporters and a pump-like subunit (KdpB) from the superfamily of P-type ATPases. Recent structural work has defined the architecture and generated contradictory hypotheses for the transport mechanism. Here, we use substrate analogs to stabilize four key intermediates in the reaction cycle and determine the corresponding structures by cryogenic electron microscopy. We find that KdpB undergoes conformational changes consistent with other representatives from the P-type superfamily, whereas KdpA, KdpC, and KdpF remain static. We observe a series of spherical densities that we assign as K or water and which define a pathway for K transport. This pathway runs through an intramembrane tunnel in KdpA and delivers ions to sites in the membrane domain of KdpB. Our structures suggest a mechanism where ATP hydrolysis is coupled to K transfer between alternative sites in KdpB, ultimately reaching a low-affinity site where a water-filled pathway allows release of K to the cytoplasm.
History
DepositionJan 9, 2021Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jan 27, 2021Provider: repository / Type: Initial release
Revision 1.1Feb 9, 2022Group: Database references / Category: citation / citation_author / database_2
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _database_2.pdbx_DOI / _database_2.pdbx_database_accession
Revision 1.2Jul 10, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_3d_fitting_list / em_admin / pdbx_initial_refinement_model
Item: _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id ..._em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type / _em_admin.last_update

-
Structure visualization

Movie
  • Deposited structure unit
  • Imaged by Jmol
  • Download
  • Superimposition on EM map
  • EMDB-12184
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: Potassium-transporting ATPase potassium-binding subunit
B: Potassium-transporting ATPase ATP-binding subunit
C: Potassium-transporting ATPase KdpC subunit
D: Potassium-transporting ATPase KdpF subunit
hetero molecules


Theoretical massNumber of molelcules
Total (without water)158,4338
Polymers156,8704
Non-polymers1,5634
Water23413
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration, Purifies as intact complex
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area16270 Å2
ΔGint-153 kcal/mol
Surface area49880 Å2
MethodPISA

-
Components

-
Potassium-transporting ATPase ... , 4 types, 4 molecules ABCD

#1: Protein Potassium-transporting ATPase potassium-binding subunit / ATP phosphohydrolase [potassium-transporting] A chain / Potassium-binding and translocating subunit ...ATP phosphohydrolase [potassium-transporting] A chain / Potassium-binding and translocating subunit A / Potassium-translocating ATPase A chain


Mass: 59247.684 Da / Num. of mol.: 1 / Mutation: Q116R
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: kdpA, b0698, JW0686 / Plasmid: pSD107 / Production host: Escherichia coli (E. coli) / Variant (production host): TK2498 / References: UniProt: P03959
#2: Protein Potassium-transporting ATPase ATP-binding subunit / ATP phosphohydrolase [potassium-transporting] B chain / Potassium-binding and translocating subunit ...ATP phosphohydrolase [potassium-transporting] B chain / Potassium-binding and translocating subunit B / Potassium-translocating ATPase B chain


Mass: 72251.867 Da / Num. of mol.: 1 / Mutation: S162A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: kdpB, b0697, JW0685 / Plasmid: pSD107 / Production host: Escherichia coli (E. coli) / Variant (production host): TK2498 / References: UniProt: P03960, P-type K+ transporter
#3: Protein Potassium-transporting ATPase KdpC subunit / ATP phosphohydrolase [potassium-transporting] C chain / Potassium-binding and translocating subunit ...ATP phosphohydrolase [potassium-transporting] C chain / Potassium-binding and translocating subunit C / Potassium-translocating ATPase C chain


Mass: 22299.225 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: kdpC, b0696, JW0684 / Plasmid: pSD107 / Production host: Escherichia coli (E. coli) / Variant (production host): TK2498 / References: UniProt: P03961
#4: Protein/peptide Potassium-transporting ATPase KdpF subunit / ATP phosphohydrolase [potassium-transporting] F chain / Potassium-binding and translocating subunit ...ATP phosphohydrolase [potassium-transporting] F chain / Potassium-binding and translocating subunit F / Potassium-translocating ATPase F chain


Mass: 3071.714 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: kdpF, b4513, JW0687 / Plasmid: pSD107 / Production host: Escherichia coli (E. coli) / Variant (production host): TK2498 / References: UniProt: P36937

-
Non-polymers , 5 types, 17 molecules

#5: Chemical ChemComp-PEV / (1S)-2-{[(2-AMINOETHOXY)(HYDROXY)PHOSPHORYL]OXY}-1-[(PALMITOYLOXY)METHYL]ETHYL STEARATE / PHOSPHATIDYLETHANOLAMINE / 1-PALMITOYL-2-OLEOYL-SN-GLYCERO-3-PHOSPHOETHANOLAMINE


Mass: 720.012 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C39H78NO8P / Comment: POPE, phospholipid*YM
#6: Chemical ChemComp-9Y0 / (2R)-3-(((2-aminoethoxy)(hydroxy)phosphoryl)oxy)-2-(palmitoyloxy)propyl (E)-octadec-9-enoate


Mass: 717.996 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C39H76NO8P / Feature type: SUBJECT OF INVESTIGATION
#7: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
#8: Chemical ChemComp-MF4 / TETRAFLUOROMAGNESATE(2-) / MAGNESIUMTETRAFLUORIDE


Mass: 100.299 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Formula: F4Mg / Feature type: SUBJECT OF INVESTIGATION
#9: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 13 / Source method: isolated from a natural source / Formula: H2O

-
Details

Has ligand of interestY

-
Experimental details

-
Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

-
Sample preparation

ComponentName: KdpFABC in E2Pi state with MgF4 / Type: COMPLEX
Details: KdpFABC complex solublized in decyl maltopyranoside (DM) detergent.
Entity ID: #1-#4 / Source: RECOMBINANT
Molecular weightValue: 0.16 MDa / Experimental value: NO
Source (natural)Organism: Escherichia coli K-12 (bacteria)
Source (recombinant)Organism: Escherichia coli (E. coli) / Strain: TK2498 / Plasmid: pSD107
Buffer solutionpH: 7.5
Details: A 10x stock solution of MgF4 was added to the protein mixture containing all other buffer components to produce final concentrations 1 mM MgCl2 and 10 mM NaF.
Buffer component
IDConc.NameFormulaBuffer-ID
125 mMtris(hydroxymethyl)aminomethane(HOCH2)3CNH21
2200 mMsodium chlorideNaCl1
35 mMmagnesium chlorideMgCl21
41 mMtris(2-carboxyethyl)phosphineC9H15O6P1
51.5 mg/mLdecyl maltopyranosideC22H42O111
61 mMmagnesium fluorideMgF41
SpecimenConc.: 4.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: DM-solublized KdpFABC complex purified through SEC the same day as grid freezing. Peak fraction diluted to 4.5 mg/mL and complexed with MgF4 solution for 1 hr at room temperature prior to application to grid.
Specimen supportDetails: PELCO easiGlow Glow Discharge Cleaning System used for glow discharge with default settings.
Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K
Details: 3 uL of protein mixture applied to grid. Blot time 4 seconds, blot force 0, no wait before plunging.

-
Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 81000 X / Nominal defocus max: 2200 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 77 K / Temperature (min): 77 K
Image recordingAverage exposure time: 2.7 sec. / Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2391
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV
Image scansSampling size: 5 µm / Width: 5760 / Height: 4092

-
Processing

SoftwareName: PHENIX / Version: dev_3758: / Classification: refinement
EM software
IDNameVersionCategory
2SerialEMimage acquisition
4cryoSPARC2.9.0CTF correction
7UCSF Chimera1.13model fitting
9cryoSPARC2.9.0initial Euler assignment
10cryoSPARC2.9.0final Euler assignment
11cryoSPARC2.9.0classification
12cryoSPARC2.9.03D reconstruction
13PHENIX1.18model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 657658
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 212120 / Details: Non-uniform refinement / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL / Target criteria: Correlation coefficient
Atomic model buildingPDB-ID: 5MRW

5mrw


Accession code: 5MRW / Source name: PDB / Type: experimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00911112
ELECTRON MICROSCOPYf_angle_d0.85515101
ELECTRON MICROSCOPYf_dihedral_angle_d15.6823978
ELECTRON MICROSCOPYf_chiral_restr0.051808
ELECTRON MICROSCOPYf_plane_restr0.0061910

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more