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- EMDB-16271: Cryo-EM structure of rat SLC22A6 bound to probenecid -

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Basic information

Entry
Database: EMDB / ID: EMD-16271
TitleCryo-EM structure of rat SLC22A6 bound to probenecid
Map data
Sample
  • Complex: OAT1
    • Protein or peptide: Solute carrier family 22 member 6
    • Protein or peptide: Synthetic nanobody (Sybody)
  • Ligand: 4-(dipropylsulfamoyl)benzoic acid
Keywordsmembrane protein / transporter
Function / homology
Function and homology information


Organic anion transport / renal tubular secretion / alpha-ketoglutarate transmembrane transporter activity / alpha-ketoglutarate transport / sodium-independent organic anion transmembrane transporter activity / sodium-independent organic anion transport / metanephric proximal tubule development / prostaglandin transport / prostaglandin transmembrane transporter activity / organic anion transport ...Organic anion transport / renal tubular secretion / alpha-ketoglutarate transmembrane transporter activity / alpha-ketoglutarate transport / sodium-independent organic anion transmembrane transporter activity / sodium-independent organic anion transport / metanephric proximal tubule development / prostaglandin transport / prostaglandin transmembrane transporter activity / organic anion transport / solute:inorganic anion antiporter activity / organic anion transmembrane transporter activity / monoatomic anion transport / antiporter activity / chloride ion binding / xenobiotic transmembrane transporter activity / transmembrane transporter activity / basal plasma membrane / caveola / response to organic cyclic compound / basolateral plasma membrane / protein-containing complex / identical protein binding / plasma membrane
Similarity search - Function
Organic cation transport protein/SVOP / Major facilitator, sugar transporter-like / Sugar (and other) transporter / Major facilitator superfamily domain / Major facilitator superfamily (MFS) profile. / MFS transporter superfamily
Similarity search - Domain/homology
Solute carrier family 22 member 6
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.94 Å
AuthorsParker JL / Kato T / Newstead S
Funding support United Kingdom, 3 items
OrganizationGrant numberCountry
Wellcome Trust215519 United Kingdom
Wellcome Trust219531 United Kingdom
Medical Research Council (MRC, United Kingdom)MR/S021043/1 United Kingdom
CitationJournal: Nat Struct Mol Biol / Year: 2023
Title: Molecular basis for selective uptake and elimination of organic anions in the kidney by OAT1.
Authors: Joanne L Parker / Takafumi Kato / Gabriel Kuteyi / Oleg Sitsel / Simon Newstead /
Abstract: In mammals, the kidney plays an essential role in maintaining blood homeostasis through the selective uptake, retention or elimination of toxins, drugs and metabolites. Organic anion transporters ...In mammals, the kidney plays an essential role in maintaining blood homeostasis through the selective uptake, retention or elimination of toxins, drugs and metabolites. Organic anion transporters (OATs) are responsible for the recognition of metabolites and toxins in the nephron and their eventual urinary excretion. Inhibition of OATs is used therapeutically to improve drug efficacy and reduce nephrotoxicity. The founding member of the renal organic anion transporter family, OAT1 (also known as SLC22A6), uses the export of α-ketoglutarate (α-KG), a key intermediate in the Krebs cycle, to drive selective transport and is allosterically regulated by intracellular chloride. However, the mechanisms linking metabolite cycling, drug transport and intracellular chloride remain obscure. Here, we present cryogenic-electron microscopy structures of OAT1 bound to α-KG, the antiviral tenofovir and clinical inhibitor probenecid, used in the treatment of Gout. Complementary in vivo cellular assays explain the molecular basis for α-KG driven drug elimination and the allosteric regulation of organic anion transport in the kidney by chloride.
History
DepositionDec 6, 2022-
Header (metadata) releaseJul 19, 2023-
Map releaseJul 19, 2023-
UpdateNov 22, 2023-
Current statusNov 22, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_16271.png

Downloads & links

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Map

FileDownload / File: emd_16271.map.gz / Format: CCP4 / Size: 91.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 288 pix.
= 239.616 Å		0.83 Å/pix.
x 288 pix.
= 239.616 Å		0.83 Å/pix.
x 288 pix.
= 239.616 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.832 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.35
Minimum - Maximum-0.82905525 - 1.4224719
Average (Standard dev.)0.0017642981 (±0.041604806)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions288288288
Spacing288288288
CellA=B=C: 239.616 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_16271_msk_1.map
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Half map: #2

Fileemd_16271_half_map_1.map
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Half map: #1

Fileemd_16271_half_map_2.map
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Sample components

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Entire : OAT1

EntireName: OAT1
Components
  • Complex: OAT1
    • Protein or peptide: Solute carrier family 22 member 6
    • Protein or peptide: Synthetic nanobody (Sybody)
  • Ligand: 4-(dipropylsulfamoyl)benzoic acid

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Supramolecule #1: OAT1

SupramoleculeName: OAT1 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Rattus norvegicus (Norway rat)

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Macromolecule #1: Solute carrier family 22 member 6

MacromoleculeName: Solute carrier family 22 member 6 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Molecular weightTheoretical: 60.553559 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MAFNDLLKQV GGVGRFQLIQ VTMVVAPLLL MASHNTLQNF TAAIPPHHCR PPANANLSKD GGLEAWLPLD KQGQPESCLR FTSPQWGPP FYNGTEANGT RVTEPCIDGW VYDNSTFPST IVTEWNLVCS HRAFRQLAQS LYMVGVLLGA MVFGYLADRL G RRKVLILN ...String:
MAFNDLLKQV GGVGRFQLIQ VTMVVAPLLL MASHNTLQNF TAAIPPHHCR PPANANLSKD GGLEAWLPLD KQGQPESCLR FTSPQWGPP FYNGTEANGT RVTEPCIDGW VYDNSTFPST IVTEWNLVCS HRAFRQLAQS LYMVGVLLGA MVFGYLADRL G RRKVLILN YLQTAVSGTC AAYAPNYTVY CVFRLLSGMS LASIAINCMT LNVEWMPIHT RAYVGTLIGY VYSLGQFLLA GI AYAVPHW RHLQLVVSVP FFIAFIYSWF FIESARWYSS SGRLDLTLRA LQRVARINGK QEEGAKLSIE VLRTSLQKEL TLS KGQASA MELLRCPTLR HLFLCLSMLW FATSFAYYGL VMDLQGFGVS MYLIQVIFGA VDLPAKFVCF LVINSMGRRP AQMA SLLLA GICILVNGII PKSHTIIRTS LAVLGKGCLA SSFNCIFLYT GELYPTVIRQ TGLGMGSTMA RVGSIVSPLV SMTAE FYPS MPLFIFGAVP VVASAVTALL PETLGQPLPD TVQDLKSRSR GKQNQQQQEQ QKQMMPLENL YFQ

UniProtKB: Solute carrier family 22 member 6

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Macromolecule #2: Synthetic nanobody (Sybody)

MacromoleculeName: Synthetic nanobody (Sybody) / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 16.079819 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MAGSSSQVQL VESGGGLVQA GGSLRLSCAA SGFPVKTEWM EWYRQAPGKE REWVAAIWSY GSGTRYADSV KGRFTISRDN AKNTVYLQM NSLKPEDTAV YYCLVRVGSW YHGQGTQVTV SAGRAGEQKL ISEEDLNSAV DHHHHHH

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Macromolecule #3: 4-(dipropylsulfamoyl)benzoic acid

MacromoleculeName: 4-(dipropylsulfamoyl)benzoic acid / type: ligand / ID: 3 / Number of copies: 1 / Formula: RTO
Molecular weightTheoretical: 285.359 Da
Chemical component information

ChemComp-RTO:
4-(dipropylsulfamoyl)benzoic acid

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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

BufferpH: 7.5
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Specialist opticsEnergy filter - Name: GIF Bioquantum
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 48.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.94 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.2) / Number images used: 107118
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL
Output model

PDB-8bvt:
Cryo-EM structure of rat SLC22A6 bound to probenecid

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