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- PDB-8c02: Structure of SLC40/ferroportin in complex with synthetic nanobody... -

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Basic information

Entry
Database: PDB / ID: 8c02
TitleStructure of SLC40/ferroportin in complex with synthetic nanobody Sy3 in occluded conformation
Components
  • Solute carrier family 40 member 1
  • Sybody3
KeywordsTRANSPORT PROTEIN / Iron / SLC40 / Transporter
Function / homology
Function and homology information


spleen trabecula formation / iron ion export across plasma membrane / Defective SLC40A1 causes hemochromatosis 4 (HFE4) (duodenum) / Defective SLC40A1 causes hemochromatosis 4 (HFE4) (macrophages) / Defective CP causes aceruloplasminemia (ACERULOP) / Metal ion SLC transporters / iron ion transmembrane transport / lymphocyte homeostasis / iron ion transmembrane transporter activity / ferrous iron transmembrane transporter activity ...spleen trabecula formation / iron ion export across plasma membrane / Defective SLC40A1 causes hemochromatosis 4 (HFE4) (duodenum) / Defective SLC40A1 causes hemochromatosis 4 (HFE4) (macrophages) / Defective CP causes aceruloplasminemia (ACERULOP) / Metal ion SLC transporters / iron ion transmembrane transport / lymphocyte homeostasis / iron ion transmembrane transporter activity / ferrous iron transmembrane transporter activity / endothelium development / peptide hormone binding / establishment of localization in cell / Iron uptake and transport / multicellular organismal-level iron ion homeostasis / synaptic vesicle / basolateral plasma membrane / intracellular iron ion homeostasis / transcription by RNA polymerase II / apoptotic process / negative regulation of apoptotic process / positive regulation of transcription by RNA polymerase II / nucleoplasm / membrane / identical protein binding / metal ion binding / plasma membrane / cytosol / cytoplasm
Similarity search - Function
Ferroportin-1 / Ferroportin1 (FPN1) / MFS transporter superfamily
Similarity search - Domain/homology
Solute carrier family 40 member 1
Similarity search - Component
Biological speciesHomo sapiens (human)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.09 Å
AuthorsLehmann, E.F. / Liziczai, M. / Drozdzyk, K. / Dutzler, R. / Manatschal, C.
Funding support Switzerland, 1items
OrganizationGrant numberCountry
Swiss National Science Foundation Switzerland
CitationJournal: Elife / Year: 2023
Title: Structures of ferroportin in complex with its specific inhibitor vamifeport.
Authors: Elena Farah Lehmann / Márton Liziczai / Katarzyna Drożdżyk / Patrick Altermatt / Cassiano Langini / Vania Manolova / Hanna Sundstrom / Franz Dürrenberger / Raimund Dutzler / Cristina Manatschal /
Abstract: A central regulatory mechanism of iron homeostasis in humans involves ferroportin (FPN), the sole cellular iron exporter, and the peptide hormone hepcidin, which inhibits Fe transport and induces ...A central regulatory mechanism of iron homeostasis in humans involves ferroportin (FPN), the sole cellular iron exporter, and the peptide hormone hepcidin, which inhibits Fe transport and induces internalization and degradation of FPN. Dysregulation of the FPN/hepcidin axis leads to diverse pathological conditions, and consequently, pharmacological compounds that inhibit FPN-mediated iron transport are of high clinical interest. Here, we describe the cryo-electron microscopy structures of human FPN in complex with synthetic nanobodies and vamifeport (VIT-2763), the first clinical-stage oral FPN inhibitor. Vamifeport competes with hepcidin for FPN binding and is currently in clinical development for β-thalassemia and sickle cell disease. The structures display two distinct conformations of FPN, representing outward-facing and occluded states of the transporter. The vamifeport site is located in the center of the protein, where the overlap with hepcidin interactions underlies the competitive relationship between the two molecules. The introduction of point mutations in the binding pocket of vamifeport reduces its affinity to FPN, emphasizing the relevance of the structural data. Together, our study reveals conformational rearrangements of FPN that are of potential relevance for transport, and it provides initial insight into the pharmacological targeting of this unique iron efflux transporter.
History
DepositionDec 15, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Mar 22, 2023Provider: repository / Type: Initial release
Revision 1.1Mar 29, 2023Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.pdbx_database_id_PubMed ..._citation.journal_volume / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.name

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Solute carrier family 40 member 1
B: Sybody3


Theoretical massNumber of molelcules
Total (without water)79,8802
Polymers79,8802
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, surface plasmon resonance, gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Solute carrier family 40 member 1 / Ferroportin-1 / Iron-regulated transporter 1


Mass: 63536.848 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SLC40A1, FPN1, IREG1, SLC11A3, MSTP079 / Production host: Homo sapiens (human) / References: UniProt: Q9NP59
#2: Antibody Sybody3


Mass: 16343.104 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli)

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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: Complex of ferroportin with synthetic nanobody / Type: COMPLEX
Details: Ferroportin was expressed in human derived HEK cells, whereas the synthetic nanobody was expressed in bacterial cell culture. The two were purified separately and mixed shortly before vitrification
Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 278 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 FIELDBright-field microscopy / Nominal defocus max: 2400 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 66.6 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.20.1_4487: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 4.09 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 139011 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0044338
ELECTRON MICROSCOPYf_angle_d0.8315901
ELECTRON MICROSCOPYf_dihedral_angle_d4.521590
ELECTRON MICROSCOPYf_chiral_restr0.044695
ELECTRON MICROSCOPYf_plane_restr0.007720

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