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- PDB-9ovr: CryoEM structural insight into the breast cancer pH regulator NBCn1 -

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Basic information

Entry
Database: PDB / ID: 9ovr
TitleCryoEM structural insight into the breast cancer pH regulator NBCn1
ComponentsSodium bicarbonate cotransporter 3
KeywordsPROTEIN TRANSPORT / Transporter / sodium and carbonate cotransporter / pH regulator / breast cancers
Function / homology
Function and homology information


sodium:bicarbonate symporter activity / phagosome acidification / Bicarbonate transporters / monoatomic anion transmembrane transporter activity / auditory receptor cell development / pyrimidine nucleotide biosynthetic process / solute:inorganic anion antiporter activity / stereocilium / bicarbonate transport / purine nucleotide biosynthetic process ...sodium:bicarbonate symporter activity / phagosome acidification / Bicarbonate transporters / monoatomic anion transmembrane transporter activity / auditory receptor cell development / pyrimidine nucleotide biosynthetic process / solute:inorganic anion antiporter activity / stereocilium / bicarbonate transport / purine nucleotide biosynthetic process / RHOD GTPase cycle / RHOF GTPase cycle / RHOJ GTPase cycle / RHOQ GTPase cycle / RHOH GTPase cycle / regulation of intracellular pH / cellular response to growth factor stimulus / transmembrane transport / basolateral plasma membrane / apical plasma membrane / membrane / plasma membrane
Similarity search - Function
Band 3 cytoplasmic domain / Band 3 cytoplasmic domain / Phosphotransferase/anion transporter / Bicarbonate transporter, eukaryotic / Bicarbonate transporter-like, transmembrane domain / HCO3- transporter integral membrane domain
Similarity search - Domain/homology
CARBONATE ION / Sodium bicarbonate cotransporter 3
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsWeiguang, W. / Hristina, R.Z. / Kirill, T. / Hong, Z. / Alexander, P. / Ira, K.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK) United States
CitationJournal: Nat Commun / Year: 2025
Title: CryoEM and computational modeling structural insights into the pH regulator NBCn1.
Authors: Weiguang Wang / Hristina R Zhekova / Kirill Tsirulnikov / B Sridhar Dwadasi / Ehecatl Guzman Aparicio / Rustam Azimov / Natalia Abuladze / Liyo Kao / Dora Acuna / D Peter Tieleman / Z Hong ...Authors: Weiguang Wang / Hristina R Zhekova / Kirill Tsirulnikov / B Sridhar Dwadasi / Ehecatl Guzman Aparicio / Rustam Azimov / Natalia Abuladze / Liyo Kao / Dora Acuna / D Peter Tieleman / Z Hong Zhou / Alexander Pushkin / Ira Kurtz /
Abstract: Breast cancer cells survive despite being exposed to a toxic acidic extracellular environment, by utilizing the NBCn1 transporter. The molecular basis for this phenomenon is unknown, given the lack ...Breast cancer cells survive despite being exposed to a toxic acidic extracellular environment, by utilizing the NBCn1 transporter. The molecular basis for this phenomenon is unknown, given the lack of an NBCn1 atomic structural model. We therefore determined the 3.3 Å cryoEM structure of the human NBCn1 outward facing (OF) conformational state with densities corresponding to the transported ions in the ion coordination site. We further generated NBCn1 inward facing (IF) and intermediate (occluded) structures and characterized the transport cycle and the ion dynamics in the IF and OF states. The results showed that NBCn1 utilizes an elevator-type transport mechanism with a small vertical shift of the ion coordination site between OF and IF conformational states and that the transported ions permeate without significant energy barriers. Functional experiments showed that NBCn1 has an extremely high ion turnover rate (TOR) of ~15,000 s. The unusually high NBCn1 TOR value associated with the small protein structural changes during the OF to IF transitions and the favorable ion permeation energetics provides breast cancer cells with a highly efficient base loading mechanism contributing to their survival advantage.
History
DepositionMay 30, 2025Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 3, 2025Provider: repository / Type: Initial release
Revision 1.0Sep 3, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Sep 3, 2025Data content type: Additional map / Part number: 1 / Data content type: Additional map / Provider: repository / Type: Initial release
Revision 1.0Sep 3, 2025Data content type: FSC / Data content type: FSC / Provider: repository / Type: Initial release
Revision 1.0Sep 3, 2025Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Sep 3, 2025Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Sep 3, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Sep 3, 2025Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.1Mar 18, 2026Group: Data collection / Database references / Category: citation / citation_author / em_admin
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Revision 1.1Mar 18, 2026Data content type: EM metadata / Data content type: EM metadata / EM metadata / Group: Database references / Experimental summary / Data content type: EM metadata / EM metadata / EM metadata / Category: citation / citation_author / em_admin
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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
B: Sodium bicarbonate cotransporter 3
A: Sodium bicarbonate cotransporter 3
hetero molecules


Theoretical massNumber of molelcules
Total (without water)274,95518
Polymers272,4372
Non-polymers2,51816
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein Sodium bicarbonate cotransporter 3 / Electroneutral Na/HCO(3) cotransporter / Sodium bicarbonate cotransporter 2 / Sodium bicarbonate ...Electroneutral Na/HCO(3) cotransporter / Sodium bicarbonate cotransporter 2 / Sodium bicarbonate cotransporter 2b / Bicarbonate transporter / Solute carrier family 4 member 7


Mass: 136218.531 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SLC4A7, BT, NBC2, NBC2B, NBC3, NBCn1, SBC2, SLC4A6 / Production host: Homo sapiens (human) / References: UniProt: Q9Y6M7
#2: Polysaccharide
2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 4 / Source method: obtained synthetically
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#3: Chemical
ChemComp-CO3 / CARBONATE ION


Mass: 60.009 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: CO3 / Feature type: SUBJECT OF INVESTIGATION
#4: Sugar ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C8H15NO6 / Feature type: SUBJECT OF INVESTIGATION
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0
#5: Chemical
ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Na / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY
Has protein modificationY

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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: Transporter branched with NAG and complexed with sodium ions and carbonate.
Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: HEK293T (human)
Buffer solutionpH: 7.4
Buffer component
IDConc.NameFormulaBuffer-ID
1115 mMsodium chlorideNaCl1
220 mMTris aminomethaneTris-HCl1
325 mMsodium bicarbonateNaHCO31
40.01 %LMNGLMNG1
SpecimenConc.: 0.8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
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 / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: OTHER / Nominal magnification: 81000 X / Nominal defocus max: 2400 nm / Nominal defocus min: 1800 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: BASIC
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 95 K / Temperature (min): 79 K
Image recordingAverage exposure time: 2 sec. / Electron dose: 40 e/Å2 / Film or detector model: GATAN K3 BIOCONTINUUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 16274

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Processing

EM software
IDNameVersionCategoryDetails (eV)
1cryoSPARC4.5particle selection
2SerialEM4.2image acquisition
4cryoSPARC4.5CTF correctionPatch CTF
9cryoSPARC4.5initial Euler assignment
10cryoSPARC4.5final Euler assignment
11RELION3.1bclassification
12cryoSPARC4.53D reconstruction
13PHENIX1.19.2_4158model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 2560294
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 267909 / Num. of class averages: 2 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL
RefinementHighest resolution: 3.3 Å
Stereochemistry target values: REAL-SPACE (WEIGHTED MAP SUM AT ATOM CENTERS)
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0039390
ELECTRON MICROSCOPYf_angle_d0.63112744
ELECTRON MICROSCOPYf_dihedral_angle_d12.6321266
ELECTRON MICROSCOPYf_chiral_restr0.0431526
ELECTRON MICROSCOPYf_plane_restr0.0051534

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