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- PDB-8qey: Structure of human Asc1/CD98hc heteromeric amino acid transporter -

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Basic information

Entry
Database: PDB / ID: 8qey
TitleStructure of human Asc1/CD98hc heteromeric amino acid transporter
Components
  • 4F2 cell-surface antigen heavy chain
  • Asc-type amino acid transporter 1
KeywordsMEMBRANE PROTEIN / Heteromeric amino acid transporter Transporter APC transporter
Function / homology
Function and homology information


D-serine transmembrane transport / positive regulation of synaptic transmission, glycinergic / D-alanine transmembrane transport / glycine transport / Defective SLC7A7 causes lysinuric protein intolerance (LPI) / neutral L-amino acid secondary active transmembrane transporter activity / apical pole of neuron / aromatic amino acid transmembrane transporter activity / tyrosine transport / L-histidine transport ...D-serine transmembrane transport / positive regulation of synaptic transmission, glycinergic / D-alanine transmembrane transport / glycine transport / Defective SLC7A7 causes lysinuric protein intolerance (LPI) / neutral L-amino acid secondary active transmembrane transporter activity / apical pole of neuron / aromatic amino acid transmembrane transporter activity / tyrosine transport / L-histidine transport / negative regulation of brown fat cell differentiation / amino acid transport complex / L-serine transmembrane transporter activity / L-leucine import across plasma membrane / L-alanine transmembrane transporter activity / L-alanine import across plasma membrane / isoleucine transport / phenylalanine transport / methionine transport / L-amino acid transmembrane transporter activity / valine transport / L-leucine transmembrane transporter activity / calcium:sodium antiporter activity / L-leucine transport / thyroid hormone transport / proline transport / amino acid transmembrane transport / neutral amino acid transport / Amino acid transport across the plasma membrane / neutral L-amino acid transmembrane transporter activity / Tryptophan catabolism / exogenous protein binding / anchoring junction / Basigin interactions / amino acid transport / response to exogenous dsRNA / tryptophan transport / basal plasma membrane / calcium ion transport / double-stranded RNA binding / melanosome / virus receptor activity / basolateral plasma membrane / carbohydrate metabolic process / cadherin binding / symbiont entry into host cell / protein heterodimerization activity / apical plasma membrane / lysosomal membrane / synapse / cell surface / protein homodimerization activity / RNA binding / extracellular exosome / nucleoplasm / membrane / plasma membrane
Similarity search - Function
Solute carrier family 3 member 2, N-terminal domain / 4F2 cell-surface antigen heavy chain / Solute carrier family 3 member 2 N-terminus / : / Amino acid/polyamine transporter I / Amino acid permease / Alpha amylase, catalytic domain / Glycosyl hydrolase, family 13, catalytic domain / Alpha-amylase domain / Glycosyl hydrolase, all-beta / Glycoside hydrolase superfamily
Similarity search - Domain/homology
Amino acid transporter heavy chain SLC3A2 / Asc-type amino acid transporter 1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4 Å
AuthorsMartinez-Molledo, M. / Rullo-Tubau, J. / Errasti-Murugarren, E. / Palacin, M. / Llorca, O.
Funding support Spain, 1items
OrganizationGrant numberCountry
La Caixa FoundationLCF/PR/HR20/52400017 Spain
CitationJournal: Nat Commun / Year: 2024
Title: Structure and mechanisms of transport of human Asc1/CD98hc amino acid transporter.
Authors: Josep Rullo-Tubau / Maria Martinez-Molledo / Paola Bartoccioni / Ignasi Puch-Giner / Ángela Arias / Suwipa Saen-Oon / Camille Stephan-Otto Attolini / Rafael Artuch / Lucía Díaz / Víctor ...Authors: Josep Rullo-Tubau / Maria Martinez-Molledo / Paola Bartoccioni / Ignasi Puch-Giner / Ángela Arias / Suwipa Saen-Oon / Camille Stephan-Otto Attolini / Rafael Artuch / Lucía Díaz / Víctor Guallar / Ekaitz Errasti-Murugarren / Manuel Palacín / Oscar Llorca /
Abstract: Recent cryoEM studies elucidated details of the structural basis for the substrate selectivity and translocation of heteromeric amino acid transporters. However, Asc1/CD98hc is the only neutral ...Recent cryoEM studies elucidated details of the structural basis for the substrate selectivity and translocation of heteromeric amino acid transporters. However, Asc1/CD98hc is the only neutral heteromeric amino acid transporter that can function through facilitated diffusion, and the only one that efficiently transports glycine and D-serine, and thus has a regulatory role in the central nervous system. Here we use cryoEM, ligand-binding simulations, mutagenesis, transport assays, and molecular dynamics to define human Asc1/CD98hc determinants for substrate specificity and gain insights into the mechanisms that govern substrate translocation by exchange and facilitated diffusion. The cryoEM structure of Asc1/CD98hc is determined at 3.4-3.8 Å resolution, revealing an inward-facing semi-occluded conformation. We find that Ser 246 and Tyr 333 are essential for Asc1/CD98hc substrate selectivity and for the exchange and facilitated diffusion modes of transport. Taken together, these results reveal the structural bases for ligand binding and transport features specific to human Asc1.
History
DepositionSep 1, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 10, 2024Provider: repository / Type: Initial release
Revision 1.1Apr 17, 2024Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Oct 16, 2024Group: Data collection / Structure summary
Category: em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Asc-type amino acid transporter 1
B: 4F2 cell-surface antigen heavy chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)127,7807
Polymers124,8942
Non-polymers2,8865
Water00
1


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

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Components

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Protein , 2 types, 2 molecules AB

#1: Protein Asc-type amino acid transporter 1 / Asc-1 / Solute carrier family 7 member 10


Mass: 56837.504 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SLC7A10, ASC1 / Cell line (production host): HEK293-6E / Production host: Homo sapiens (human) / References: UniProt: Q9NS82
#2: Protein 4F2 cell-surface antigen heavy chain / 4F2hc / 4F2 heavy chain antigen / Lymphocyte activation antigen 4F2 large subunit / Solute carrier ...4F2hc / 4F2 heavy chain antigen / Lymphocyte activation antigen 4F2 large subunit / Solute carrier family 3 member 2


Mass: 68056.688 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SLC3A2, MDU1 / Cell line (production host): HEK293-6E / Production host: Homo sapiens (human) / References: UniProt: P08195

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Sugars , 3 types, 4 molecules

#3: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#4: Polysaccharide beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta- ...beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 586.542 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/2,3,2/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5]/1-1-2/a4-b1_b4-c1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{}}}LINUCSPDB-CARE
#6: Sugar ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C8H15NO6
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Non-polymers , 1 types, 1 molecules

#5: Chemical ChemComp-AJP / Digitonin


Mass: 1229.312 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C56H92O29 / Comment: detergent*YM

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Details

Has ligand of interestN
Has protein modificationY

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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: Asc1/CD98hc / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293-6E / Plasmid: pcDNA3.1(+)
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMTris hydrochlorideTris-HCl1
2150 mMSodium chlorideNaCl1
30.1 %Digitonindgt1
SpecimenConc.: 1.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid type: Quantifoil R0.6/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 277.15 K

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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: 2400 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of real images: 31696

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Processing

EM software
IDNameCategory
2EPUimage acquisition
4CTFFINDCTF correction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 139507 / Symmetry type: POINT
RefinementResolution: 4→249.3 Å / Cor.coef. Fo:Fc: 0.82 / SU B: 24.645 / SU ML: 0.27 / ESU R: 0.205
Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflection
Rwork0.41433 --
obs0.41433 506986 100 %
Solvent computationSolvent model: PARAMETERS FOR MASK CACLULATION
Displacement parametersBiso mean: 271.981 Å2
Refinement stepCycle: 1 / Total: 7306
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0047500
ELECTRON MICROSCOPYf_angle_d0.94410228
ELECTRON MICROSCOPYf_dihedral_angle_d13.0931086
ELECTRON MICROSCOPYf_chiral_restr0.0541212
ELECTRON MICROSCOPYf_plane_restr0.0061250
LS refinement shellResolution: 4→4.104 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0 0 -
Rwork0.569 37562 -
obs--100 %

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