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- EMDB-41862: Structure of Fmoc-Leu-OH bound Sialin -

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Basic information

Entry
Database: EMDB / ID: EMD-41862
TitleStructure of Fmoc-Leu-OH bound Sialin
Map data
Sample
  • Complex: Structure of Fmoc-Leu-OH bound Sialin
    • Protein or peptide: Sialin
  • Ligand: Fluorenylmethyloxycarbonyl chloride
  • Ligand: LEUCINE
KeywordsTransporter / MEMBRANE PROTEIN
Function / homology
Function and homology information


sialic acid:proton symporter activity / D-glucuronate transmembrane transporter activity / Defective SLC17A5 causes Salla disease (SD) and ISSD / Organic anion transport by SLC5/17/25 transporters / sialic acid transmembrane transporter activity / sialic acid transport / carbohydrate:proton symporter activity / Sialic acid metabolism / neurotransmitter loading into synaptic vesicle / monoatomic anion transport ...sialic acid:proton symporter activity / D-glucuronate transmembrane transporter activity / Defective SLC17A5 causes Salla disease (SD) and ISSD / Organic anion transport by SLC5/17/25 transporters / sialic acid transmembrane transporter activity / sialic acid transport / carbohydrate:proton symporter activity / Sialic acid metabolism / neurotransmitter loading into synaptic vesicle / monoatomic anion transport / amino acid transport / monoatomic ion transport / response to bacterium / synaptic vesicle membrane / basolateral plasma membrane / lysosome / apical plasma membrane / lysosomal membrane / glutamatergic synapse / membrane / plasma membrane / cytosol
Similarity search - Function
: / Major facilitator superfamily / Major Facilitator Superfamily / Major facilitator superfamily domain / Major facilitator superfamily (MFS) profile. / MFS transporter superfamily
Similarity search - Domain/homology
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.67 Å
AuthorsSchmiege P / Li X
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)135343 United States
National Institutes of Health/Eunice Kennedy Shriver National Institute of Child Health & Human Development (NIH/NICHD)110229 United States
Welch FoundationI-1957 United States
CitationJournal: Nat Commun / Year: 2024
Title: Structure and inhibition of the human lysosomal transporter Sialin.
Authors: Philip Schmiege / Linda Donnelly / Nadia Elghobashi-Meinhardt / Chia-Hsueh Lee / Xiaochun Li /
Abstract: Sialin, a member of the solute carrier 17 (SLC17) transporter family, is unique in its ability to transport not only sialic acid using a pH-driven mechanism, but also transport mono and diacidic ...Sialin, a member of the solute carrier 17 (SLC17) transporter family, is unique in its ability to transport not only sialic acid using a pH-driven mechanism, but also transport mono and diacidic neurotransmitters, such as glutamate and N-acetylaspartylglutamate (NAAG), into synaptic vesicles via a membrane potential-driven mechanism. While most transporters utilize one of these mechanisms, the structural basis of how Sialin transports substrates using both remains unclear. Here, we present the cryogenic electron-microscopy structures of human Sialin: apo cytosol-open, apo lumen-open, NAAG-bound, and inhibitor-bound. Our structures show that a positively charged cytosol-open vestibule accommodates either NAAG or the Sialin inhibitor Fmoc-Leu-OH, while its luminal cavity potentially binds sialic acid. Moreover, functional analyses along with molecular dynamics simulations identify key residues in binding sialic acid and NAAG. Thus, our findings uncover the essential conformational states in NAAG and sialic acid transport, demonstrating a working model of SLC17 transporters.
History
DepositionSep 7, 2023-
Header (metadata) releaseMay 29, 2024-
Map releaseMay 29, 2024-
UpdateDec 11, 2024-
Current statusDec 11, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_41862.png

Downloads & links

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Map

FileDownload / File: emd_41862.map.gz / Format: CCP4 / Size: 103 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 300 pix.
= 249. Å		0.83 Å/pix.
x 300 pix.
= 249. Å		0.83 Å/pix.
x 300 pix.
= 249. Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.83 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.442
Minimum - Maximum-3.066146 - 4.716381
Average (Standard dev.)0.0026367218 (±0.06121499)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 249.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_41862_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_41862_half_map_2.map
Projections & Slices
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Histogram (log scale)

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Sample components

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Entire : Structure of Fmoc-Leu-OH bound Sialin

EntireName: Structure of Fmoc-Leu-OH bound Sialin
Components
  • Complex: Structure of Fmoc-Leu-OH bound Sialin
    • Protein or peptide: Sialin
  • Ligand: Fluorenylmethyloxycarbonyl chloride
  • Ligand: LEUCINE

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Supramolecule #1: Structure of Fmoc-Leu-OH bound Sialin

SupramoleculeName: Structure of Fmoc-Leu-OH bound Sialin / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Sialin

MacromoleculeName: Sialin / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 55.681137 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MDYKDDDDKR SPVRDLARND GEESTDRTPL LPGAPRAEAA PVCCSARYNL AILAFFGFFI VYALRVNLSV ALVDMVDSNT TLEDNRTSK ACPEHSAPIK VHHNQTGKKY QWDAETQGWI LGSFFYGYII TQIPGGYVAS KIGGKMLLGF GILGTAVLTL F TPIAADLG ...String:
MDYKDDDDKR SPVRDLARND GEESTDRTPL LPGAPRAEAA PVCCSARYNL AILAFFGFFI VYALRVNLSV ALVDMVDSNT TLEDNRTSK ACPEHSAPIK VHHNQTGKKY QWDAETQGWI LGSFFYGYII TQIPGGYVAS KIGGKMLLGF GILGTAVLTL F TPIAADLG VGPLIVLRAL EGLGEGVTFP AMHAMWSSWA PPLERSKLLS ISYAGAQLGT VISLPLSGII CYYMNWTYVF YF FGTIGIF WFLLWIWLVS DTPQKHKRIS HYEKEYILSS LRNQLSSQKS VPWVPILKSL PLWAIVVAHF SYNWTFYTLL TLL PTYMKE ILRFNVQENG FLSSLPYLGS WLCMILSGQA ADNLRAKWNF STLCVRRIFS LIGMIGPAVF LVAAGFIGCD YSLA VAFLT ISTTLGGFCS SGFSINHLDI APSYAGILLG ITNTFATIPG MVGPVIAKSL TPDNTVGEWQ TVFYIAAAIN VFGAI FFTL FAKGEVQNWA LNDHHGHRH

UniProtKB: Sialin

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Macromolecule #2: Fluorenylmethyloxycarbonyl chloride

MacromoleculeName: Fluorenylmethyloxycarbonyl chloride / type: ligand / ID: 2
Details: conjugated to leucine as Fluorenylmethyloxycarbonyl-leucine
Number of copies: 1 / Formula: VP1
Molecular weightTheoretical: 258.7 Da

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Macromolecule #3: LEUCINE

MacromoleculeName: LEUCINE / type: ligand / ID: 3
Details: conjugated to leucine as Fluorenylmethyloxycarbonyl-leucine
Number of copies: 1 / Formula: LEU
Molecular weightTheoretical: 131.173 Da
Chemical component information

ChemComp-LEU:
LEUCINE

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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
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: 2.2 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.67 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 408146
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE

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