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- PDB-8zqo: Human high-affinity choline transporter CHT1 bound to HC-3 under ... -

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Basic information

Entry
Database: PDB / ID: 8zqo
TitleHuman high-affinity choline transporter CHT1 bound to HC-3 under NaCl condition, with sodium and chloride ions coordinated.
ComponentsHigh affinity choline transporter 1
KeywordsSTRUCTURAL PROTEIN / transporter
Function / homology
Function and homology information


choline:sodium symporter activity / Defective SLC5A7 causes distal hereditary motor neuronopathy 7A (HMN7A) / Defective SLC5A7 causes distal hereditary motor neuronopathy 7A (HMN7A) / acetylcholine biosynthetic process / Transport of bile salts and organic acids, metal ions and amine compounds / choline transmembrane transporter activity / Acetylcholine Neurotransmitter Release Cycle / neuromuscular synaptic transmission / choline transport / choline binding ...choline:sodium symporter activity / Defective SLC5A7 causes distal hereditary motor neuronopathy 7A (HMN7A) / Defective SLC5A7 causes distal hereditary motor neuronopathy 7A (HMN7A) / acetylcholine biosynthetic process / Transport of bile salts and organic acids, metal ions and amine compounds / choline transmembrane transporter activity / Acetylcholine Neurotransmitter Release Cycle / neuromuscular synaptic transmission / choline transport / choline binding / synaptic transmission, cholinergic / neurotransmitter transport / neuromuscular junction / transmembrane transport / synaptic vesicle membrane / presynaptic membrane / early endosome membrane / perikaryon / in utero embryonic development / axon / dendrite / synapse / membrane / plasma membrane
Similarity search - Function
: / Sodium/solute symporter / Sodium/glucose symporter superfamily / Sodium:solute symporter family / Sodium:solute symporter family profile.
Similarity search - Domain/homology
Chem-HC6 / High affinity choline transporter 1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.8 Å
AuthorsQiu, Y. / Gao, Y. / Zhao, Y.
Funding support China, 1items
OrganizationGrant numberCountry
Chinese Academy of SciencesXDB37030304 China
CitationJournal: Structure / Year: 2024
Title: Ion coupling and inhibitory mechanisms of the human presynaptic high-affinity choline transporter CHT1.
Authors: Yunlong Qiu / Yiwei Gao / Qinru Bai / Yan Zhao /
Abstract: In cholinergic neurons, choline is the precursor of the excitatory neurotransmitter acetylcholine (ACh), which plays a fundamental role in the brain. The high-affinity choline transporter, CHT1, ...In cholinergic neurons, choline is the precursor of the excitatory neurotransmitter acetylcholine (ACh), which plays a fundamental role in the brain. The high-affinity choline transporter, CHT1, mediates the efficient recycling of choline to facilitate ACh synthesis in the presynapse. Here, we report high-resolution cryoelectron microscopic (cryo-EM) structures of CHT1 in complex with the inhibitors HC-3 and ML352, the substrate choline, and a substrate-free state. Our structures show distinct binding modes of the inhibitors with different chemical structures, revealing their inhibition mechanisms. Additionally, we observed a chloride ion that directly interacts with the substrate choline, thereby stabilizing its binding with CHT1. Two sodium ions, Na2 and Na3, were clearly identified, which we speculate might be involved in substrate binding and conformational transitions, respectively. Our structures provide molecular insights into the coupling mechanism of ion binding with substrate binding and conformational transitions, promoting our understanding of the ion-coupled substrate transport mechanism.
History
DepositionJun 3, 2024Deposition site: PDBJ / Processing site: PDBC
Revision 1.0Dec 4, 2024Provider: repository / Type: Initial release
Revision 1.1Dec 25, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.pdbx_database_id_PubMed / _citation.title / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: High affinity choline transporter 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)64,3226
Polymers63,2391
Non-polymers1,0835
Water181
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 / Sugars , 2 types, 2 molecules A

#1: Protein High affinity choline transporter 1 / Hemicholinium-3-sensitive choline transporter / CHT / Solute carrier family 5 member 7


Mass: 63239.145 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SLC5A7, CHT1 / Cell line (production host): HEK 293-F / Production host: Homo sapiens (human) / References: UniProt: Q9GZV3
#2: 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

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Non-polymers , 4 types, 5 molecules

#3: Chemical ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Na / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical ChemComp-HC6 / (2S,2'S)-2,2'-biphenyl-4,4'-diylbis(2-hydroxy-4,4-dimethylmorpholin-4-ium)


Mass: 414.538 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C24H34N2O4 / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical ChemComp-CL / CHLORIDE ION


Mass: 35.453 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Cl / Feature type: SUBJECT OF INVESTIGATION
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY
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: Human high-affinity choline transporter CHT1 bound to HC-3 under NaCl condition, with sodium and chloride ions coordinated.
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7.5
SpecimenConc.: 10 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: This sample was obtained from the monodispersed peak fractions of the size-exclusion chromatography.
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 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: 2200 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameVersionCategory
1cryoSPARCparticle selection
2EPUimage acquisition
7UCSF Chimera1.15model fitting
9cryoSPARCinitial Euler assignment
10cryoSPARCfinal Euler assignment
11cryoSPARCclassification
12cryoSPARC3D reconstruction
13Coot0.9.6.2-premodel refinement
CTF correctionType: PHASE FLIPPING ONLY
Particle selectionNum. of particles selected: 842831
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 842831 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0034143
ELECTRON MICROSCOPYf_angle_d0.5215665
ELECTRON MICROSCOPYf_dihedral_angle_d3.939584
ELECTRON MICROSCOPYf_chiral_restr0.038660
ELECTRON MICROSCOPYf_plane_restr0.005683

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