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- PDB-8jf1: Human sodium-dependent vitamin C transporter 1 in an apo occluded... -

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Basic information

Entry
Database: PDB / ID: 8jf1
TitleHuman sodium-dependent vitamin C transporter 1 in an apo occluded state
ComponentsSolute carrier family 23 member 1
KeywordsTRANSPORT PROTEIN / Transporter / Membrane protein / Ascorbic acid / Vitamin C / Sodium
Function / homology
Function and homology information


nucleobase transport / L-ascorbate:sodium symporter activity / L-ascorbic acid transmembrane transporter activity / L-ascorbic acid transmembrane transport / nucleobase transmembrane transporter activity / dehydroascorbic acid transmembrane transporter activity / intracellular organelle / sodium ion transmembrane transporter activity / dehydroascorbic acid transport / Vitamin C (ascorbate) metabolism ...nucleobase transport / L-ascorbate:sodium symporter activity / L-ascorbic acid transmembrane transporter activity / L-ascorbic acid transmembrane transport / nucleobase transmembrane transporter activity / dehydroascorbic acid transmembrane transporter activity / intracellular organelle / sodium ion transmembrane transporter activity / dehydroascorbic acid transport / Vitamin C (ascorbate) metabolism / L-ascorbic acid metabolic process / urate transmembrane transporter activity / sodium ion transport / lung development / basal plasma membrane / brain development / response to toxic substance / apical plasma membrane / extracellular exosome / plasma membrane / cytoplasm
Similarity search - Function
Nucleobase cation symporter 2 family / Permease family
Similarity search - Domain/homology
Lauryl Maltose Neopentyl Glycol / CHOLESTEROL / PALMITIC ACID / CHOLESTEROL HEMISUCCINATE / Solute carrier family 23 member 1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.85 Å
AuthorsKobayashi, T.A. / Kusakizako, T. / Nureki, O.
Funding support Japan, 1items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS) Japan
CitationJournal: Nat Commun / Year: 2024
Title: Dimeric transport mechanism of human vitamin C transporter SVCT1.
Authors: Takaaki A Kobayashi / Hiroto Shimada / Fumiya K Sano / Yuzuru Itoh / Sawako Enoki / Yasushi Okada / Tsukasa Kusakizako / Osamu Nureki /
Abstract: Vitamin C plays important roles as a cofactor in many enzymatic reactions and as an antioxidant against oxidative stress. As some mammals including humans cannot synthesize vitamin C de novo from ...Vitamin C plays important roles as a cofactor in many enzymatic reactions and as an antioxidant against oxidative stress. As some mammals including humans cannot synthesize vitamin C de novo from glucose, its uptake from dietary sources is essential, and is mediated by the sodium-dependent vitamin C transporter 1 (SVCT1). Despite its physiological significance in maintaining vitamin C homeostasis, the structural basis of the substrate transport mechanism remained unclear. Here, we report the cryo-EM structures of human SVCT1 in different states at 2.5-3.5 Å resolutions. The binding manner of vitamin C together with two sodium ions reveals the counter ion-dependent substrate recognition mechanism. Furthermore, comparisons of the inward-open and occluded structures support a transport mechanism combining elevator and distinct rotational motions. Our results demonstrate the molecular mechanism of vitamin C transport with its underlying conformational cycle, potentially leading to future industrial and medical applications.
History
DepositionMay 16, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0May 22, 2024Provider: repository / Type: Initial release
Revision 1.1Oct 30, 2024Group: Data collection / Structure summary
Category: em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification
Revision 1.2Dec 4, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
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

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Solute carrier family 23 member 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)69,7149
Polymers64,8611
Non-polymers4,8538
Water362
1
A: Solute carrier family 23 member 1
hetero molecules

A: Solute carrier family 23 member 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)139,42818
Polymers129,7212
Non-polymers9,70716
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation1
Noncrystallographic symmetry (NCS)NCS oper:
IDCodeMatrixVector
1given(1), (1), (1)
2generate(-1), (-1), (1)298.79999, 298.79999

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Components

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Protein / Sugars , 2 types, 2 molecules A

#1: Protein Solute carrier family 23 member 1 / Na(+)/L-ascorbic acid transporter 1 / Sodium-dependent vitamin C transporter 1 / hSVCT1 / Yolk sac ...Na(+)/L-ascorbic acid transporter 1 / Sodium-dependent vitamin C transporter 1 / hSVCT1 / Yolk sac permease-like molecule 3


Mass: 64860.707 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SLC23A1, SVCT1, YSPL3 / Plasmid: pEG BacMam / Cell line (production host): HEK293S GnTI- / Production host: Homo sapiens (human) / References: UniProt: Q9UHI7
#5: 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.: 1 / 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

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Non-polymers , 5 types, 9 molecules

#2: Chemical ChemComp-AV0 / Lauryl Maltose Neopentyl Glycol / 2,2-didecylpropane-1,3-bis-b-D-maltopyranoside / 2-decyl-2-{[(4-O-alpha-D-glucopyranosyl-beta-D-glucopyranosyl)oxy]methyl}dodecyl4-O-alpha-D-glucopyranosyl-beta-D-glucopyranoside


Mass: 1005.188 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C47H88O22 / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical ChemComp-Y01 / CHOLESTEROL HEMISUCCINATE


Mass: 486.726 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C31H50O4 / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical ChemComp-CLR / CHOLESTEROL


Mass: 386.654 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C27H46O / Feature type: SUBJECT OF INVESTIGATION
#6: Chemical ChemComp-PLM / PALMITIC ACID


Mass: 256.424 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C16H32O2 / Feature type: SUBJECT OF INVESTIGATION
#7: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Formula: H2O

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Details

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: Human SVCT1 dimer in an apo occluded state / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
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 magnification: 105000 X / Nominal defocus max: 1600 nm / Nominal defocus min: 800 nm
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 50.9 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 5907
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 25 eV

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Processing

EM software
IDNameVersionCategory
1RELION3.1.1particle selection
2EPU3.9.0image acquisition
4RELION3.1.1CTF correction
7Coot0.9.6model fitting
9RELION3.1.1initial Euler assignment
10RELION3.1.1final Euler assignment
11RELION3.1.1classification
12RELION3.1.13D reconstruction
13Servalcat0.2.115model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 3699304
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 2.85 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 116307 / Symmetry type: POINT
Atomic model buildingPDB-ID: 8JEW

8jew


Accession code: 8JEW / Source name: PDB / Type: experimental model
RefinementResolution: 2.85→2.85 Å / Cor.coef. Fo:Fc: 0.841 / SU B: 9.153 / SU ML: 0.175 / ESU R: 0.226
Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflection
Rwork0.32912 --
obs0.32912 87113 100 %
Solvent computationSolvent model: PARAMETERS FOR MASK CACLULATION
Displacement parametersBiso mean: 70.119 Å2
Refinement stepCycle: 1 / Total: 4221
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
ELECTRON MICROSCOPYr_bond_refined_d0.010.0134336
ELECTRON MICROSCOPYr_bond_other_d0.0020.0174330
ELECTRON MICROSCOPYr_angle_refined_deg1.4891.6385919
ELECTRON MICROSCOPYr_angle_other_deg1.3571.55310025
ELECTRON MICROSCOPYr_dihedral_angle_1_deg3.8665509
ELECTRON MICROSCOPYr_dihedral_angle_2_deg32.82921.192151
ELECTRON MICROSCOPYr_dihedral_angle_3_deg14.28715631
ELECTRON MICROSCOPYr_dihedral_angle_4_deg16.8351516
ELECTRON MICROSCOPYr_chiral_restr0.080.2637
ELECTRON MICROSCOPYr_gen_planes_refined0.0160.024454
ELECTRON MICROSCOPYr_gen_planes_other0.010.02894
ELECTRON MICROSCOPYr_nbd_refined
ELECTRON MICROSCOPYr_nbd_other
ELECTRON MICROSCOPYr_nbtor_refined
ELECTRON MICROSCOPYr_nbtor_other
ELECTRON MICROSCOPYr_xyhbond_nbd_refined
ELECTRON MICROSCOPYr_xyhbond_nbd_other
ELECTRON MICROSCOPYr_metal_ion_refined
ELECTRON MICROSCOPYr_metal_ion_other
ELECTRON MICROSCOPYr_symmetry_vdw_refined
ELECTRON MICROSCOPYr_symmetry_vdw_other
ELECTRON MICROSCOPYr_symmetry_hbond_refined
ELECTRON MICROSCOPYr_symmetry_hbond_other
ELECTRON MICROSCOPYr_symmetry_metal_ion_refined
ELECTRON MICROSCOPYr_symmetry_metal_ion_other
ELECTRON MICROSCOPYr_mcbond_it8.1786.4222048
ELECTRON MICROSCOPYr_mcbond_other8.1596.4172047
ELECTRON MICROSCOPYr_mcangle_it11.5869.6842553
ELECTRON MICROSCOPYr_mcangle_other11.5949.6892554
ELECTRON MICROSCOPYr_scbond_it10.4378.1592288
ELECTRON MICROSCOPYr_scbond_other10.4358.1632289
ELECTRON MICROSCOPYr_scangle_it
ELECTRON MICROSCOPYr_scangle_other15.93711.8073367
ELECTRON MICROSCOPYr_long_range_B_refined23.65818641
ELECTRON MICROSCOPYr_long_range_B_other23.65818642
ELECTRON MICROSCOPYr_rigid_bond_restr
ELECTRON MICROSCOPYr_sphericity_free
ELECTRON MICROSCOPYr_sphericity_bonded
LS refinement shellResolution: 2.8→2.873 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0 0 -
Rwork1.267 6496 -
obs--100 %

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