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- PDB-8ps0: Cryo-EM structure of Sodium proton exchanger NhaA with bound card... -

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Basic information

Entry
Database: PDB / ID: 8ps0
TitleCryo-EM structure of Sodium proton exchanger NhaA with bound cardiolipin
ComponentsNa(+)/H(+) antiporter NhaA
KeywordsTRANSPORT PROTEIN / Sodium proton exchanger / Cardiolipin / NhaA
Function / homologyNa+/H+ antiporter NhaA / Na+/H+ antiporter domain superfamily / Na+/H+ antiporter 1 / sodium:proton antiporter activity / regulation of pH / plasma membrane / CARDIOLIPIN / Na(+)/H(+) antiporter NhaA
Function and homology information
Biological speciesEscherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.37 Å
AuthorsGulati, A. / Meier, P. / Kokane, S. / Drew, D.
Funding supportEuropean Union, 1items
OrganizationGrant numberCountry
European Research Council (ERC)CoG-820187European Union
CitationJournal: Nat Commun / Year: 2025
Title: PIP-mediated oligomerization of the endosomal sodium/proton exchanger NHE9.
Authors: Surabhi Kokane / Ashutosh Gulati / Pascal F Meier / Rei Matsuoka / Tanadet Pipatpolkai / Giuseppe Albano / Tin Manh Ho / Lucie Delemotte / Daniel Fuster / David Drew /
Abstract: The strict exchange of Na for H ions across cell membranes is a reaction carried out in almost every cell. Na/H exchangers that perform this task are physiological homodimers, and whilst the ion ...The strict exchange of Na for H ions across cell membranes is a reaction carried out in almost every cell. Na/H exchangers that perform this task are physiological homodimers, and whilst the ion transporting domain is highly conserved, their dimerization differs. The Na/H exchanger NhaA from Escherichia coli has a weak dimerization interface mediated by a β-hairpin domain and with dimer retention dependent on cardiolipin. Similarly, organellar Na/H exchangers NHE6, NHE7 and NHE9 also contain β-hairpin domains and recent analysis of Equus caballus NHE9 indicated PIP lipids could bind at the dimer interface. However, structural validation of the predicted lipid-mediated oligomerization has been lacking. Here, we report cryo-EM structures of E. coli NhaA and E. caballus NHE9 in complex with cardiolipin and phosphatidylinositol-3,5-bisphosphate PI(3,5)P lipids binding at their respective dimer interfaces. We further show how the endosomal specific PI(3,5)P lipid stabilizes the NHE9 homodimer and enhances transport activity. Indeed, we show that NHE9 is active in endosomes, but not at the plasma membrane where the PI(3,5)P lipid is absent. Thus, specific lipids can regulate Na/H exchange activity by stabilizing dimerization in response to either cell specific cues or upon trafficking to their correct membrane location.
History
DepositionJul 13, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Feb 21, 2024Provider: repository / Type: Initial release
Revision 1.1Sep 3, 2025Group: Data collection / Database references / Structure summary
Category: citation / citation_author ...citation / citation_author / em_admin / pdbx_entry_details
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Na(+)/H(+) antiporter NhaA
B: Na(+)/H(+) antiporter NhaA
hetero molecules


Theoretical massNumber of molelcules
Total (without water)89,0015
Polymers84,6092
Non-polymers4,3923
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Na(+)/H(+) antiporter NhaA / Sodium/proton antiporter NhaA


Mass: 42304.422 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli)
Gene: nhaA, nahA, BLM69_005044, BvCmsKKP061_01067, EL79_3853, JNP96_04275, WR15_27505
Production host: Escherichia coli (E. coli) / References: UniProt: Q53YW7
#2: Chemical ChemComp-CDL / CARDIOLIPIN / DIPHOSPHATIDYL GLYCEROL / BIS-(1,2-DIACYL-SN-GLYCERO-3-PHOSPHO)-1',3'-SN-GLYCEROL


Mass: 1464.043 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C81H156O17P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: phospholipid*YM
Has ligand of interestY
Has protein modificationN

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Experimental details

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Experiment

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

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Sample preparation

ComponentName: NhaA dimer / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Escherichia coli (E. coli)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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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 defocus max: 2000 nm / Nominal defocus min: 600 nm
Image recordingElectron dose: 68.11 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.37 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 78917 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 95.47 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00265863
ELECTRON MICROSCOPYf_angle_d0.62587987
ELECTRON MICROSCOPYf_chiral_restr0.0398975
ELECTRON MICROSCOPYf_plane_restr0.0023954
ELECTRON MICROSCOPYf_dihedral_angle_d5.7491868

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