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- PDB-6caa: CryoEM structure of human SLC4A4 sodium-coupled acid-base transpo... -

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Basic information

Entry
Database: PDB / ID: 6caa
TitleCryoEM structure of human SLC4A4 sodium-coupled acid-base transporter NBCe1
ComponentsElectrogenic sodium bicarbonate cotransporter 1
KeywordsTRANSPORT PROTEIN / SLC4A4 / Electrogenic Sodium-Coupled Bicarbonate Cotransporters / NBCe1
Function / homology
Function and homology information


Defective SLC4A4 causes renal tubular acidosis, proximal, with ocular abnormalities and mental retardation (pRTA-OA) / sodium:bicarbonate symporter activity / monoatomic ion homeostasis / Bicarbonate transporters / sodium ion export across plasma membrane / solute:inorganic anion antiporter activity / symporter activity / bicarbonate transport / sodium ion transport / plasma membrane => GO:0005886 ...Defective SLC4A4 causes renal tubular acidosis, proximal, with ocular abnormalities and mental retardation (pRTA-OA) / sodium:bicarbonate symporter activity / monoatomic ion homeostasis / Bicarbonate transporters / sodium ion export across plasma membrane / solute:inorganic anion antiporter activity / symporter activity / bicarbonate transport / sodium ion transport / plasma membrane => GO:0005886 / transmembrane transporter activity / transport across blood-brain barrier / sodium ion transmembrane transport / regulation of membrane potential / positive regulation of glycolytic process / regulation of intracellular pH / transmembrane transport / basolateral plasma membrane / cell surface / extracellular exosome / membrane / identical protein binding / plasma membrane
Similarity search - Function
Sodium bicarbonate cotransporter / Band 3 cytoplasmic domain / Band 3 cytoplasmic domain / Bicarbonate transporter, eukaryotic / Bicarbonate transporter-like, transmembrane domain / HCO3- transporter family / Phosphotransferase/anion transporter
Similarity search - Domain/homology
Electrogenic sodium bicarbonate cotransporter 1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.9 Å
AuthorsHuynh, K.W. / Jiang, J. / Abuladze, N. / Tsirulnikov, K. / Kao, L. / Shao, X. / Newman, D. / Azimov, R. / Pushkin, A. / Zhou, Z.H. / Kurtz, I.
Funding support United States, 11items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)DK077162 United States
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)DK007789 United States
Allan Smidt Charitable Fund United States
Factor Family Foundation United States
Ralph Block Family Foundation United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM071940 United States
National Institutes of Health/National Institute of Dental and Craniofacial Research (NIH/NIDCR)DE025567 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI094386 United States
National Institutes of Health/National Center for Research Resources (NIH/NCRR)1S10RR23057 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1U24GM116792 United States
National Science Foundation (NSF, United States)DBI-1338135 United States
CitationJournal: Nat Commun / Year: 2018
Title: CryoEM structure of the human SLC4A4 sodium-coupled acid-base transporter NBCe1.
Authors: Kevin W Huynh / Jiansen Jiang / Natalia Abuladze / Kirill Tsirulnikov / Liyo Kao / Xuesi Shao / Debra Newman / Rustam Azimov / Alexander Pushkin / Z Hong Zhou / Ira Kurtz /
Abstract: Na-coupled acid-base transporters play essential roles in human biology. Their dysfunction has been linked to cancer, heart, and brain disease. High-resolution structures of mammalian Na-coupled acid- ...Na-coupled acid-base transporters play essential roles in human biology. Their dysfunction has been linked to cancer, heart, and brain disease. High-resolution structures of mammalian Na-coupled acid-base transporters are not available. The sodium-bicarbonate cotransporter NBCe1 functions in multiple organs and its mutations cause blindness, abnormal growth and blood chemistry, migraines, and impaired cognitive function. Here, we have determined the structure of the membrane domain dimer of human NBCe1 at 3.9 Å resolution by cryo electron microscopy. Our atomic model and functional mutagenesis revealed the ion accessibility pathway and the ion coordination site, the latter containing residues involved in human disease-causing mutations. We identified a small number of residues within the ion coordination site whose modification transformed NBCe1 into an anion exchanger. Our data suggest that symporters and exchangers utilize comparable transport machinery and that subtle differences in their substrate-binding regions have very significant effects on their transport mode.
History
DepositionJan 29, 2018Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 7, 2018Provider: repository / Type: Initial release
Revision 1.1Mar 14, 2018Group: Database references / Category: citation
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
Revision 1.2Jul 18, 2018Group: Data collection / Category: em_imaging_optics / Item: _em_imaging_optics.energyfilter_name
Revision 1.3Feb 20, 2019Group: Author supporting evidence / Data collection / Category: em_imaging_optics / pdbx_audit_support
Item: _em_imaging_optics.energyfilter_name / _pdbx_audit_support.funding_organization
Revision 1.4Nov 27, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.5Mar 13, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

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Assembly

Deposited unit
A: Electrogenic sodium bicarbonate cotransporter 1
B: Electrogenic sodium bicarbonate cotransporter 1


Theoretical massNumber of molelcules
Total (without water)232,3452
Polymers232,3452
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration, SDS-PAGE of purified NBCe1
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area2770 Å2
ΔGint-25 kcal/mol
Surface area46820 Å2

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Components

#1: Protein Electrogenic sodium bicarbonate cotransporter 1 / Sodium bicarbonate cotransporter / Na(+)/HCO3(-) cotransporter / Solute carrier family 4 member 4 / kNBC1


Mass: 116172.312 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SLC4A4, NBC, NBC1, NBCE1 / Organ: Kidney / Variant: A / Plasmid: pTT / Cell line (production host): HEK293 / Production host: Homo sapiens (human) / References: UniProt: Q9Y6R1

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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: NBCe1 Dimer / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.260 MDa / Experimental value: YES
Source (natural)Organism: Homo sapiens (human) / Organ: Kidney
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293 / Plasmid: pTT
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMTris-HClTrisC4H12ClNO31
2150 mMsodium chlorideNaClSodium chloride1
SpecimenConc.: 0.4 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Sample was purified in DDM and exchanged into Amphipols (PMAL-C8)
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen 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 FIELDBright-field microscopy / Calibrated magnification: 36764 X / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 24 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 2 / Num. of real images: 3378
EM imaging opticsEnergyfilter name: GIF Quantum LS / Energyfilter upper: 20 eV / Energyfilter lower: 0 eV
Image scansMovie frames/image: 36 / Used frames/image: 2-36

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Processing

SoftwareName: PHENIX / Version: 1.11.1_2575: / Classification: refinement
EM software
IDNameVersionCategory
2Leginonimage acquisition
4CTFFIND4CTF correction
7Coot0.8.7model fitting
9RELION2initial Euler assignment
10RELION2final Euler assignment
12RELION23D reconstruction
13PHENIX1.11.1-2575model refinement
CTF correctionDetails: CTF correction was performed during the 3D refinement in RELION
Type: NONE
Particle selectionNum. of particles selected: 1729419
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 184561 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0097694
ELECTRON MICROSCOPYf_angle_d1.28810456
ELECTRON MICROSCOPYf_dihedral_angle_d6.8294498
ELECTRON MICROSCOPYf_chiral_restr0.0681260
ELECTRON MICROSCOPYf_plane_restr0.0091260

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