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- PDB-8u3e: Structure of Apo Sialin at pH5.0 -

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Basic information

Entry
Database: PDB / ID: 8u3e
TitleStructure of Apo Sialin at pH5.0
ComponentsSialin
KeywordsMEMBRANE PROTEIN / Transporter
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 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 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 / 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)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.19 Å
AuthorsSchmiege, P. / Li, X.
Funding support United States, 3items
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, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 29, 2024Provider: repository / Type: Initial release
Revision 1.1Dec 11, 2024Group: Data collection / Database references / Structure summary
Category: citation / citation_author ...citation / citation_author / em_admin / pdbx_entry_details
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: Sialin


Theoretical massNumber of molelcules
Total (without water)55,6811
Polymers55,6811
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Sialin


Mass: 55681.137 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SLC17A5 / Production host: Homo sapiens (human) / References: UniProt: Q9NRA2
Has protein modificationN

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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: Structure of Apo Sialin at pH5.0 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
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 defocus max: 2200 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameVersionCategory
13PHENIX1.20.1-4487model refinement
14ISOLDE1.6.0model refinement
CTF correctionType: NONE
3D reconstructionResolution: 3.19 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 135283 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0076811
ELECTRON MICROSCOPYf_angle_d0.59512301
ELECTRON MICROSCOPYf_dihedral_angle_d12.8042732
ELECTRON MICROSCOPYf_chiral_restr0.046533
ELECTRON MICROSCOPYf_plane_restr0.003976

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