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- EMDB-34818: The 5-MTHF-bound BRIL-SLC19A1/Fab/Nb ternary complex -

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Basic information

Entry
Database: EMDB / ID: EMD-34818
TitleThe 5-MTHF-bound BRIL-SLC19A1/Fab/Nb ternary complex
Map data
Sample
  • Complex: The 5-MTHF-bound BRIL-SLC19A1/Fab/Nb ternary complex
    • Protein or peptide: BRIL-SLC19A1 chimera
    • Protein or peptide: Anti-BRIL Fab heavy chain
    • Protein or peptide: Anti-Fab nanobody
    • Protein or peptide: Anti-BRIL Fab light chain
  • Ligand: N-[4-({[(6S)-2-AMINO-4-HYDROXY-5-METHYL-5,6,7,8-TETRAHYDROPTERIDIN-6-YL]METHYL}AMINO)BENZOYL]-L-GLUTAMIC ACID
KeywordsSLC19A1 / RFC / transporter / folates / 5-MTHF / TPP / TRANSPORT PROTEIN
Function / homology
Function and homology information


folate:monoatomic anion antiporter activity / methotrexate transport / folic acid transmembrane transporter activity / methotrexate transmembrane transporter activity / folic acid transport / folate transmembrane transport / folate import across plasma membrane / cyclic-GMP-AMP transmembrane transporter activity / cyclic-GMP-AMP transmembrane import across plasma membrane / 2',3'-cyclic GMP-AMP binding ...folate:monoatomic anion antiporter activity / methotrexate transport / folic acid transmembrane transporter activity / methotrexate transmembrane transporter activity / folic acid transport / folate transmembrane transport / folate import across plasma membrane / cyclic-GMP-AMP transmembrane transporter activity / cyclic-GMP-AMP transmembrane import across plasma membrane / 2',3'-cyclic GMP-AMP binding / Metabolism of folate and pterines / positive regulation of cGAS/STING signaling pathway / xenobiotic transmembrane transport / organic anion transport / : / folic acid binding / antiporter activity / folic acid metabolic process / xenobiotic transmembrane transporter activity / transport across blood-brain barrier / brush border membrane / female pregnancy / electron transport chain / response to toxic substance / basolateral plasma membrane / periplasmic space / electron transfer activity / apical plasma membrane / iron ion binding / response to xenobiotic stimulus / heme binding / plasma membrane
Similarity search - Function
Reduced folate transporter SLC19A1 / Reduced folate carrier / Reduced folate carrier / Cytochrome b562 / Cytochrome b562 / MFS transporter superfamily / Cytochrome c/b562
Similarity search - Domain/homology
Soluble cytochrome b562 / Reduced folate transporter
Similarity search - Component
Biological speciesHomo sapiens (human) / Lama glama (llama)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.54 Å
AuthorsZhang Z / Dang Y
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)32171201 China
CitationJournal: Cell Discov / Year: 2022
Title: Molecular mechanism of substrate recognition by folate transporter SLC19A1.
Authors: Yu Dang / Dong Zhou / Xiaojuan Du / Hongtu Zhao / Chia-Hsueh Lee / Jing Yang / Yijie Wang / Changdong Qin / Zhenxi Guo / Zhe Zhang /
Abstract: Folate (vitamin B) is the coenzyme involved in one-carbon transfer biochemical reactions essential for cell survival and proliferation, with its inadequacy causing developmental defects or severe ...Folate (vitamin B) is the coenzyme involved in one-carbon transfer biochemical reactions essential for cell survival and proliferation, with its inadequacy causing developmental defects or severe diseases. Notably, mammalian cells lack the ability to de novo synthesize folate but instead rely on its intake from extracellular sources via specific transporters or receptors, among which SLC19A1 is the ubiquitously expressed one in tissues. However, the mechanism of substrate recognition by SLC19A1 remains unclear. Here we report the cryo-EM structures of human SLC19A1 and its complex with 5-methyltetrahydrofolate at 3.5-3.6 Å resolution and elucidate the critical residues for substrate recognition. In particular, we reveal that two variant residues among SLC19 subfamily members designate the specificity for folate. Moreover, we identify intracellular thiamine pyrophosphate as the favorite coupled substrate for folate transport by SLC19A1. Together, this work establishes the molecular basis of substrate recognition by this central folate transporter.
History
DepositionNov 20, 2022-
Header (metadata) releaseDec 21, 2022-
Map releaseDec 21, 2022-
UpdateOct 16, 2024-
Current statusOct 16, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_34818.png

Downloads & links

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Map

FileDownload / File: emd_34818.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.07 Å/pix.
x 320 pix.
= 342.4 Å		1.07 Å/pix.
x 320 pix.
= 342.4 Å		1.07 Å/pix.
x 320 pix.
= 342.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.07 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.19
Minimum - Maximum-0.0017488354 - 2.1001353
Average (Standard dev.)0.0006511351 (±0.017778859)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 342.40002 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_34818_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_34818_half_map_2.map
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 : The 5-MTHF-bound BRIL-SLC19A1/Fab/Nb ternary complex

EntireName: The 5-MTHF-bound BRIL-SLC19A1/Fab/Nb ternary complex
Components
  • Complex: The 5-MTHF-bound BRIL-SLC19A1/Fab/Nb ternary complex
    • Protein or peptide: BRIL-SLC19A1 chimera
    • Protein or peptide: Anti-BRIL Fab heavy chain
    • Protein or peptide: Anti-Fab nanobody
    • Protein or peptide: Anti-BRIL Fab light chain
  • Ligand: N-[4-({[(6S)-2-AMINO-4-HYDROXY-5-METHYL-5,6,7,8-TETRAHYDROPTERIDIN-6-YL]METHYL}AMINO)BENZOYL]-L-GLUTAMIC ACID

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Supramolecule #1: The 5-MTHF-bound BRIL-SLC19A1/Fab/Nb ternary complex

SupramoleculeName: The 5-MTHF-bound BRIL-SLC19A1/Fab/Nb ternary complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: BRIL-SLC19A1 chimera

MacromoleculeName: BRIL-SLC19A1 chimera / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 75.110344 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: GPSRATADLE DNWETLNDNL KVIEKADNAA QVKDALTKMR AAALDAQKAT PPKLEDKSPD SPEMKDFRHG FDILVGQIDD ALKLANEGK VKEAQAAAEQ LKTTRNAYIQ KYLSWRHLVC YLCFYGFMAQ IRPGESFITP YLLGPDKNFT REQVTNEITP V LSYSYLAV ...String:
GPSRATADLE DNWETLNDNL KVIEKADNAA QVKDALTKMR AAALDAQKAT PPKLEDKSPD SPEMKDFRHG FDILVGQIDD ALKLANEGK VKEAQAAAEQ LKTTRNAYIQ KYLSWRHLVC YLCFYGFMAQ IRPGESFITP YLLGPDKNFT REQVTNEITP V LSYSYLAV LVPVFLLTDY LRYTPVLLLQ GLSFVSVWLL LLLGHSVAHM QLMELFYSVT MAARIAYSSY IFSLVRPARY QR VAGYSRA AVLLGVFTSS VLGQLLVTVG RVSFSTLNYI SLAFLTFSVV LALFLKRPKR SLFFNRDDRG RCETSASELE RMN PGPGGK LGHALRVACG DSVLARMLRE LGDSLRRPQL RLWSLWWVFN SAGYYLVVYY VHILWNEVDP TTNSARVYNG AADA ASTLL GAITSFAAGF VKIRWARWSK LLIAGVTATQ AGLVFLLAHT RHPSSIWLCY AAFVLFRGSY QFLVPIATFQ IASSL SKEL CALVFGVNTF FATIVKTIIT FIVSDVRGLG LPVRKQFQLY SVYFLILSII YFLGAMLDGL RHCQRGHHPR QPPAQG LRS AAEEKAAQAL SVQDKGLGGL QPAQSPPLSP EDSLGAVGPA SLEQRQSDPY LAQAPAPQAA EFLSPVTTPS PCTLCSA QA SGPEAADETC PQLAVHPPGV SKLGLQCLPS DGVQNVNQSN S

UniProtKB: Soluble cytochrome b562, Reduced folate transporter

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Macromolecule #2: Anti-BRIL Fab heavy chain

MacromoleculeName: Anti-BRIL Fab heavy chain / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 27.759207 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGWSLILLFL VAVATRVLSE ISEVQLVESG GGLVQPGGSL RLSCAASGFN VVDFSLHWVR QAPGKGLEWV AYISSSSGST SYADSVKGR FTISADTSKN TAYLQMNSLR AEDTAVYYCA RWGYWPGEPW WKAFDYWGQG TLVTVSSAST KGPSVFPLAP S SKSTSGGT ...String:
MGWSLILLFL VAVATRVLSE ISEVQLVESG GGLVQPGGSL RLSCAASGFN VVDFSLHWVR QAPGKGLEWV AYISSSSGST SYADSVKGR FTISADTSKN TAYLQMNSLR AEDTAVYYCA RWGYWPGEPW WKAFDYWGQG TLVTVSSAST KGPSVFPLAP S SKSTSGGT AALGCLVKDY FPEPVTVSWN SGALTSGVHT FPAVLQSSGL YSLSSVVTVP SSSLGTQTYI CNVNHKPSNT KV DKKVEPK SCDKSNSLEV LFQ

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Macromolecule #3: Anti-Fab nanobody

MacromoleculeName: Anti-Fab nanobody / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Lama glama (llama)
Molecular weightTheoretical: 16.673551 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString:
MKYLLPTAAA GLLLLAAQPA MAGSQVQLQE SGGGLVQPGG SLRLSCAASG RTISRYAMSW FRQAPGKERE FVAVARRSGD GAFYADSVQ GRFTVSRDDA KNTVYLQMNS LKPEDTAVYY CAIDSDTFYS GSYDYWGQGT QVTVSSLEHH HHHH

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Macromolecule #4: Anti-BRIL Fab light chain

MacromoleculeName: Anti-BRIL Fab light chain / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 25.575604 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGWSCIILFL VATATGVHSS DIQMTQSPSS LSASVGDRVT ITCRASQSVS SAVAWYQQKP GKAPKLLIYS ASSLYSGVPS RFSGSRSGT DFTLTISSLQ PEDFATYYCQ QYLYYSLVTF GQGTKVEIKR TVAAPSVFIF PPSDSQLKSG TASVVCLLNN F YPREAKVQ ...String:
MGWSCIILFL VATATGVHSS DIQMTQSPSS LSASVGDRVT ITCRASQSVS SAVAWYQQKP GKAPKLLIYS ASSLYSGVPS RFSGSRSGT DFTLTISSLQ PEDFATYYCQ QYLYYSLVTF GQGTKVEIKR TVAAPSVFIF PPSDSQLKSG TASVVCLLNN F YPREAKVQ WKVDNALQSG NSQESVTEQD SKDSTYSLSS TLTLSKADYE KHKVYACEVT HQGLSSPVTK SFNRGEC

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Macromolecule #5: N-[4-({[(6S)-2-AMINO-4-HYDROXY-5-METHYL-5,6,7,8-TETRAHYDROPTERIDI...

MacromoleculeName: N-[4-({[(6S)-2-AMINO-4-HYDROXY-5-METHYL-5,6,7,8-TETRAHYDROPTERIDIN-6-YL]METHYL}AMINO)BENZOYL]-L-GLUTAMIC ACID
type: ligand / ID: 5 / Number of copies: 1 / Formula: THH
Molecular weightTheoretical: 459.456 Da
Chemical component information

ChemComp-THH:
N-[4-({[(6S)-2-AMINO-4-HYDROXY-5-METHYL-5,6,7,8-TETRAHYDROPTERIDIN-6-YL]METHYL}AMINO)BENZOYL]-L-GLUTAMIC 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: 400
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.7000000000000001 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.54 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 206075
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-8hij:
The 5-MTHF-bound BRIL-SLC19A1/Fab/Nb ternary complex

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