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- EMDB-45406: Structure of human SLC2A9 transporter -

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Basic information

Entry
Database: EMDB / ID: EMD-45406
TitleStructure of human SLC2A9 transporter
Map dataSharpen final map after refinement
Sample
  • Complex: Codon optimize SLC2A9 gene was synthesize and cloned in 83 nu yeast expressing vector from GenScript. Protein expressed in yeast cells and purified using Ni-affinity followed by Sizing.
    • Protein or peptide: Soluble cytochrome b562,Solute carrier family 2, facilitated glucose transporter member 9
KeywordsTransporter / MEMBRANE PROTEIN
Function / homology
Function and homology information


Defective SLC2A9 causes hypouricemia renal 2 (RHUC2) / fructose transmembrane transporter activity / fructose transmembrane transport / dehydroascorbic acid transport / hexose transmembrane transport / carbohydrate:proton symporter activity / Cellular hexose transport / D-glucose transmembrane transporter activity / D-glucose transmembrane transport / urate transport ...Defective SLC2A9 causes hypouricemia renal 2 (RHUC2) / fructose transmembrane transporter activity / fructose transmembrane transport / dehydroascorbic acid transport / hexose transmembrane transport / carbohydrate:proton symporter activity / Cellular hexose transport / D-glucose transmembrane transporter activity / D-glucose transmembrane transport / urate transport / urate metabolic process / urate transmembrane transporter activity / D-glucose import / transmembrane transporter activity / electron transport chain / basolateral plasma membrane / periplasmic space / electron transfer activity / apical plasma membrane / iron ion binding / heme binding / membrane / plasma membrane
Similarity search - Function
Glucose transporter GLUT / Sugar/inositol transporter / Sugar transport proteins signature 2. / Sugar transport proteins signature 1. / Sugar transporter, conserved site / Major facilitator, sugar transporter-like / Sugar (and other) transporter / Major facilitator superfamily domain / Major facilitator superfamily (MFS) profile. / Cytochrome b562 ...Glucose transporter GLUT / Sugar/inositol transporter / Sugar transport proteins signature 2. / Sugar transport proteins signature 1. / Sugar transporter, conserved site / Major facilitator, sugar transporter-like / Sugar (and other) transporter / Major facilitator superfamily domain / Major facilitator superfamily (MFS) profile. / Cytochrome b562 / Cytochrome b562 / Cytochrome c/b562 / MFS transporter superfamily
Similarity search - Domain/homology
Soluble cytochrome b562 / Solute carrier family 2, facilitated glucose transporter member 9
Similarity search - Component
Biological speciesHomo sapiens (Human) (human) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.37 Å
AuthorsKhandelwal NK / Gupta M / Stroud RM
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS) United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2025
Title: Structural basis of disease mutation and substrate recognition by the human SLC2A9 transporter.
Authors: Nitesh Kumar Khandelwal / Meghna Gupta / Paras Kumar / Sree Ganesh Balasubramani / Ignacia Echeverria / Robert M Stroud /
Abstract: Urate provides ~50% of the reducing potential in human and primate plasma which is key to detoxifying reactive oxygen by-products of cellular metabolism. Urate is the endpoint of purine metabolism in ...Urate provides ~50% of the reducing potential in human and primate plasma which is key to detoxifying reactive oxygen by-products of cellular metabolism. Urate is the endpoint of purine metabolism in primates, and its concentration in plasma is a balance between excretion from kidney and intestine, and subsequent reabsorption in and through cells of kidney proximal tubules to maintain a regulated concentration in plasma. SLC2A9 is the primary transporter that returns urate from the basolateral side of kidney tubule cells back to plasma. A shorter splice variant of SLC2A9 is directed to the apical surface where several transporters recapture urate from the tubule back into cells. Too high a concentration in plasma causes hyperuricemia, is linked to gout, and favors kidney stone formation. To understand the molecular basis of uric acid transport and the role of disease-causing mutations in SLC2A9, we determined structures of human SLC2A9 in its apo form, and its urate-bound form by cryo-EM, at resolution of 3.3 Å and 4.1 Å respectively. Both structures are captured in an inward open conformation. Using the inward-facing structure as a template we modeled the outward-facing conformation to understand the alternating access mechanism. Alternative salt bridge pairs on the cytoplasmic side suggest a mechanism that can balance the energetics of the inward open and outward open states. The location of disease-causing mutants suggests their role in impacting function. Our structures elucidate the molecular basis for urate selectivity and transport and provide a platform for future structure-based drug discovery aimed at reducing plasma urate levels in diseases of hyperuricemia and gout.
History
DepositionJun 18, 2024-
Header (metadata) releaseJan 29, 2025-
Map releaseJan 29, 2025-
UpdateAug 27, 2025-
Current statusAug 27, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45406.png

Downloads & links

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Map

FileDownload / File: emd_45406.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpen final map after refinement
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.82 Å/pix.
x 360 pix.
= 294.804 Å		0.82 Å/pix.
x 360 pix.
= 294.804 Å		0.82 Å/pix.
x 360 pix.
= 294.804 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.8189 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.404
Minimum - Maximum-1.9883963 - 2.7636244
Average (Standard dev.)-0.00036877074 (±0.039325755)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 294.804 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_45406_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Unsharpen final map after refinement

Fileemd_45406_additional_1.map
AnnotationUnsharpen final map after refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map A

Fileemd_45406_half_map_1.map
AnnotationHalf map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B

Fileemd_45406_half_map_2.map
AnnotationHalf map B
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : Codon optimize SLC2A9 gene was synthesize and cloned in 83 nu yea...

EntireName: Codon optimize SLC2A9 gene was synthesize and cloned in 83 nu yeast expressing vector from GenScript. Protein expressed in yeast cells and purified using Ni-affinity followed by Sizing.
Components
  • Complex: Codon optimize SLC2A9 gene was synthesize and cloned in 83 nu yeast expressing vector from GenScript. Protein expressed in yeast cells and purified using Ni-affinity followed by Sizing.
    • Protein or peptide: Soluble cytochrome b562,Solute carrier family 2, facilitated glucose transporter member 9

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Supramolecule #1: Codon optimize SLC2A9 gene was synthesize and cloned in 83 nu yea...

SupramoleculeName: Codon optimize SLC2A9 gene was synthesize and cloned in 83 nu yeast expressing vector from GenScript. Protein expressed in yeast cells and purified using Ni-affinity followed by Sizing.
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (Human) (human)
Molecular weightTheoretical: 69.78906 KDa

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Macromolecule #1: Soluble cytochrome b562,Solute carrier family 2, facilitated gluc...

MacromoleculeName: Soluble cytochrome b562,Solute carrier family 2, facilitated glucose transporter member 9
type: protein_or_peptide / ID: 1
Details: N-terminal Flag tag, HRV3C site followed by BRIL tag N-terminal truncated SlC2A9 isoform 1 (residue 52-540 as per uniprot). C terminal thrombin site followed by 10X his tag,N-terminal Flag ...Details: N-terminal Flag tag, HRV3C site followed by BRIL tag N-terminal truncated SlC2A9 isoform 1 (residue 52-540 as per uniprot). C terminal thrombin site followed by 10X his tag,N-terminal Flag tag, HRV3C site followed by BRIL tag N-terminal truncated SlC2A9 isoform 1 (residue 52-540 as per uniprot). C terminal thrombin site followed by 10X his tag,N-terminal Flag tag, HRV3C site followed by BRIL tag N-terminal truncated SlC2A9 isoform 1 (residue 52-540 as per uniprot). C terminal thrombin site followed by 10X his tag,N-terminal Flag tag, HRV3C site followed by BRIL tag N-terminal truncated SlC2A9 isoform 1 (residue 52-540 as per uniprot). C terminal thrombin site followed by 10X his tag,N-terminal Flag tag, HRV3C site followed by BRIL tag N-terminal truncated SlC2A9 isoform 1 (residue 52-540 as per uniprot). C terminal thrombin site followed by 10X his tag,N-terminal Flag tag, HRV3C site followed by BRIL tag N-terminal truncated SlC2A9 isoform 1 (residue 52-540 as per uniprot). C terminal thrombin site followed by 10X his tag,N-terminal Flag tag, HRV3C site followed by BRIL tag N-terminal truncated SlC2A9 isoform 1 (residue 52-540 as per uniprot). C terminal thrombin site followed by 10X his tag,N-terminal Flag tag, HRV3C site followed by BRIL tag N-terminal truncated SlC2A9 isoform 1 (residue 52-540 as per uniprot). C terminal thrombin site followed by 10X his tag
Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 69.859695 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MASDYKDDDD KGALEVLFQG PSSPMADLED NWETLNDNLK VIEKADNAAQ VKDALTKMRA AALDAQKATP PKLEDKSPDS PEMKDFRHG FDILVGQIDD ALKLANEGKV KEAQAAAEQL KTTRNAYIQK YLSCSLLVAS LAGAFGSSFL YGYNLSVVNA P TPYIKAFY ...String:
MASDYKDDDD KGALEVLFQG PSSPMADLED NWETLNDNLK VIEKADNAAQ VKDALTKMRA AALDAQKATP PKLEDKSPDS PEMKDFRHG FDILVGQIDD ALKLANEGKV KEAQAAAEQL KTTRNAYIQK YLSCSLLVAS LAGAFGSSFL YGYNLSVVNA P TPYIKAFY NESWERRHGR PIDPDTLTLL WSVTVSIFAI GGLVGTLIVK MIGKVLGRKH TLLANNGFAI SAALLMACSL QA GAFEMLI VGRFIMGIDG GVALSVLPMY LSEISPKEIR GSLGQVTAIF ICIGVFTGQL LGLPELLGKE STWPYLFGVI VVP AVVQLL SLPFLPDSPR YLLLEKHNEA RAVKAFQTFL GKADVSQEVE EVLAESRVQR SIRLVSVLEL LRAPYVRWQV VTVI VTMAC YQLCGLNAIW FYTNSIFGKA GIPPAKIPYV TLSTGGIETL AAVFSGLVIE HLGRRPLLIG GFGLMGLFFG TLTIT LTLQ DHAPWVPYLS IVGILAIIAS FCSGPGGIPF ILTGEFFQQS QRPAAFIIAG TVNWLSNFAV GLLFPFIQKS LDTYCF LVF ATICITGAIY LYFVLPETKN RTYAEISQAF SKRNKAYPPE EKIDSAVTDG KINGRPGLVP RGSSAHHHHH HHHHHGA

UniProtKB: Soluble cytochrome b562, Solute carrier family 2, facilitated glucose transporter member 9

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration7 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
300.0 mMNaClSodium Chloride
50.0 mMTris-ClTris

Details: 4 degree cold buffer pH7.5 (300mM NaCl, 50mM Tris and 0.02% GDN)
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Pressure: 0.033 kPa / Details: 30 second glow 30 sec hold
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 10503 / Average exposure time: 2.0 sec. / Average electron dose: 47.7 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: OTHER / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 9590974
CTF correctionType: NONE
Startup modelType of model: NONE / Details: Alphafold2
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.37 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 154457
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
FSC plot (resolution estimation)

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