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- EMDB-8301: Architecture of the Human Mitochondrial Iron-Sulfur Cluster Assem... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-8301 | |||||||||
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Title | Architecture of the Human Mitochondrial Iron-Sulfur Cluster Assembly Machinery: the Complex Formed by the Iron Donor, the Sulfur Donor, and the Scaffold | |||||||||
![]() | Human mitochondrial iron-sulfur cluster assembly machinery: grouping of segments corresponding to one half of the full map (EMD-8293) | |||||||||
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Function / homology | ![]() regulation of ferrochelatase activity / negative regulation of iron ion import across plasma membrane / molybdopterin cofactor metabolic process / Molybdenum cofactor biosynthesis / proprioception / [4Fe-4S] cluster assembly / iron incorporation into metallo-sulfur cluster / positive regulation of lyase activity / positive regulation of succinate dehydrogenase activity / L-cysteine desulfurase complex ...regulation of ferrochelatase activity / negative regulation of iron ion import across plasma membrane / molybdopterin cofactor metabolic process / Molybdenum cofactor biosynthesis / proprioception / [4Fe-4S] cluster assembly / iron incorporation into metallo-sulfur cluster / positive regulation of lyase activity / positive regulation of succinate dehydrogenase activity / L-cysteine desulfurase complex / Mitochondrial iron-sulfur cluster biogenesis / mitochondrial [2Fe-2S] assembly complex / positive regulation of aconitate hydratase activity / positive regulation of mitochondrial electron transport, NADH to ubiquinone / iron chaperone activity / Maturation of TCA enzymes and regulation of TCA cycle / cysteine desulfurase / cysteine desulfurase activity / negative regulation of organ growth / Mo-molybdopterin cofactor biosynthetic process / iron-sulfur cluster assembly complex / Mitochondrial protein import / [2Fe-2S] cluster assembly / oxidative phosphorylation / adult walking behavior / response to iron ion / embryo development ending in birth or egg hatching / iron-sulfur cluster assembly / heme biosynthetic process / negative regulation of multicellular organism growth / organ growth / positive regulation of catalytic activity / muscle cell cellular homeostasis / ferroxidase / negative regulation of release of cytochrome c from mitochondria / iron-sulfur cluster binding / protein autoprocessing / ferroxidase activity / ferric iron binding / mitochondrion organization / ferrous iron binding / 2 iron, 2 sulfur cluster binding / cellular response to hydrogen peroxide / pyridoxal phosphate binding / Maturation of replicase proteins / positive regulation of cell growth / iron ion transport / intracellular iron ion homeostasis / molecular adaptor activity / mitochondrial matrix / iron ion binding / centrosome / positive regulation of cell population proliferation / negative regulation of apoptotic process / protein homodimerization activity / mitochondrion / zinc ion binding / nucleoplasm / nucleus / metal ion binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | single particle reconstruction / negative staining / Resolution: 14.3 Å | |||||||||
![]() | Gakh O / Ranatunga W / Smith DY / Ahlgren EC / Al-Karadaghi S / Thompson JR / Isaya G | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Architecture of the Human Mitochondrial Iron-Sulfur Cluster Assembly Machinery. Authors: Oleksandr Gakh / Wasantha Ranatunga / Douglas Y Smith / Eva-Christina Ahlgren / Salam Al-Karadaghi / James R Thompson / Grazia Isaya / ![]() ![]() Abstract: Fe-S clusters, essential cofactors needed for the activity of many different enzymes, are assembled by conserved protein machineries inside bacteria and mitochondria. As the architecture of the human ...Fe-S clusters, essential cofactors needed for the activity of many different enzymes, are assembled by conserved protein machineries inside bacteria and mitochondria. As the architecture of the human machinery remains undefined, we co-expressed in Escherichia coli the following four proteins involved in the initial step of Fe-S cluster synthesis: FXN (iron donor); [NFS1]·[ISD11] (sulfur donor); and ISCU (scaffold upon which new clusters are assembled). We purified a stable, active complex consisting of all four proteins with 1:1:1:1 stoichiometry. Using negative staining transmission EM and single particle analysis, we obtained a three-dimensional model of the complex with ∼14 Å resolution. Molecular dynamics flexible fitting of protein structures docked into the EM map of the model revealed a [FXN]·[NFS1]·[ISD11]·[ISCU] complex, consistent with the measured 1:1:1:1 stoichiometry of its four components. The complex structure fulfills distance constraints obtained from chemical cross-linking of the complex at multiple recurring interfaces, involving hydrogen bonds, salt bridges, or hydrophobic interactions between conserved residues. The complex consists of a central roughly cubic [FXN]·[ISCU] sub-complex with one symmetric ISCU trimer bound on top of one symmetric FXN trimer at each of its eight vertices. Binding of 12 [NFS1]·[ISD11] sub-complexes to the surface results in a globular macromolecule with a diameter of ∼15 nm and creates 24 Fe-S cluster assembly centers. The organization of each center recapitulates a previously proposed conserved mechanism for sulfur donation from NFS1 to ISCU and reveals, for the first time, a path for iron donation from FXN to ISCU. | |||||||||
History |
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Structure visualization
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 1.9 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 15.8 KB 15.8 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 10.7 KB | Display | ![]() |
Images | ![]() | 85.3 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 400.2 KB | Display | ![]() |
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Full document | ![]() | 399.8 KB | Display | |
Data in XML | ![]() | 9.1 KB | Display | |
Data in CIF | ![]() | 12.1 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 5kz5MC ![]() 8293C C: citing same article ( M: atomic model generated by this map |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | Human mitochondrial iron-sulfur cluster assembly machinery: grouping of segments corresponding to one half of the full map (EMD-8293) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.1 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : NFS1-ISD11-ISCU-FXN
Entire | Name: NFS1-ISD11-ISCU-FXN |
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Components |
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-Supramolecule #1: NFS1-ISD11-ISCU-FXN
Supramolecule | Name: NFS1-ISD11-ISCU-FXN / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: ![]() |
Recombinant expression | Organism: ![]() ![]() |
-Macromolecule #1: Cysteine desulfurase, mitochondrial
Macromolecule | Name: Cysteine desulfurase, mitochondrial / type: protein_or_peptide / ID: 1 / Number of copies: 12 / Enantiomer: LEVO / EC number: cysteine desulfurase |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 43.429648 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: TPLDPRVLDA MLPYLINYYG NPHSRTHAYG WESEAAMERA RQQVASLIGA DPREIIFTSG ATESNNIAIK GVARFYRSRK KHLITTQTE HKCVLDSCRS LEAEGFQVTY LPVQKSGIID LKELEAAIQP DTSLVSVMTV NNEIGVKQPI AEIGRICSSR K VYFHTDAA ...String: TPLDPRVLDA MLPYLINYYG NPHSRTHAYG WESEAAMERA RQQVASLIGA DPREIIFTSG ATESNNIAIK GVARFYRSRK KHLITTQTE HKCVLDSCRS LEAEGFQVTY LPVQKSGIID LKELEAAIQP DTSLVSVMTV NNEIGVKQPI AEIGRICSSR K VYFHTDAA QAVGKIPLDV NDMKIDLMSI SGHKIYGPKG VGAIYIRRRP RVRVEALQSG GGQERGMRSG TVPTPLVVGL GA ACEVAQQ EMEYDHKRIS KLSERLIQNI MKSLPDVVMN GDPKHHYPGC INLSFAYVEG ESLLMALKDV ALSSGSACTS ASL EPSYVL RAIGTDEDLA HSSIRFGIGR FTTEEEVDYT VEKCIQHVKR LREMSPLWEM VQDGIDLKSI KWTQH |
-Macromolecule #2: Frataxin, mitochondrial
Macromolecule | Name: Frataxin, mitochondrial / type: protein_or_peptide / ID: 2 / Number of copies: 12 / Enantiomer: LEVO / EC number: ferroxidase |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 18.849025 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: LRTDIDATCT PRRASSNQRG LNQIWNVKKQ SVYLMNLRKS GTLGHPGSLD ETTYERLAEE TLDSLAEFFE DLADKPYTFE DYDVSFGSG VLTVKLGGDL GTYVINKQTP NKQIWLSSPS SGPKRYDWTG KNWVYSHDGV SLHELLAAEL TKALKTKLDL S SLAYSGKD A |
-Macromolecule #3: Iron-sulfur cluster assembly enzyme ISCU, mitochondrial
Macromolecule | Name: Iron-sulfur cluster assembly enzyme ISCU, mitochondrial type: protein_or_peptide / ID: 3 / Number of copies: 12 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 12.52558 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: GSLDKTSKNV GTGLVGAPAC GDVMKLQIQV DEKGKIVDAR FKTFGCGSAI ASSSLATEWV KGKTVEEALT IKNTDIAKEL CLPPVKLHC SMLAEDAIKA ALADYKLKQE PKKGEAEKK |
-Experimental details
-Structure determination
Method | negative staining |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.3 mg/mL | |||||||||
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Buffer | pH: 8 Component:
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Staining | Type: NEGATIVE / Material: 1% uranyl acetate / Details: 5 and 30 seconds | |||||||||
Grid | Model: Carbon-coated copper grids, EMS / Material: COPPER / Mesh: 400 / Pretreatment - Type: GLOW DISCHARGE / Details: DV-502A vacuum evaporator (Denton Vacuum Inc.) | |||||||||
Details | 50 mM Tris-HCl, pH 8.0, 150 mM NaCl |
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Electron microscopy
Microscope | FEI TECNAI F30 |
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Image recording | Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 466 / Average electron dose: 30.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated defocus max: 3.0 µm / Calibrated defocus min: 0.21 µm / Calibrated magnification: 115000 / Illumination mode: OTHER / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.21 µm / Nominal magnification: 115000 |
Sample stage | Specimen holder model: SIDE ENTRY, EUCENTRIC / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Tecnai F30 / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
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Output model | ![]() PDB-5kz5: |