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- PDB-8rmf: Structure of the core ISC complex under turnover conditions (FDX2... -

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Basic information

Entry
Database: PDB / ID: 8rmf
TitleStructure of the core ISC complex under turnover conditions (FDX2-bound in proximal conformation)
Components
  • Acyl carrier protein
  • Ferredoxin-2, mitochondrial
  • Isoform 1 of Iron-sulfur cluster assembly enzyme ISCU
  • Isoform Mitochondrial of Cysteine desulfurase
  • LYR motif-containing protein 4
KeywordsTRANSFERASE / cysteine desulfurase / FeS biosynthesis / FeS biogenesis / mitochondria / Friedreich's ataxia / frataxin / ferredoxin / FDX2 / iron-sulfur cluster
Function / homology
Function and homology information


Electron transport from NADPH to Ferredoxin / Defective CYP11A1 causes AICSR / negative regulation of iron ion import across plasma membrane / molybdopterin cofactor metabolic process / Molybdenum cofactor biosynthesis / L-cysteine desulfurase complex / iron incorporation into metallo-sulfur cluster / [4Fe-4S] cluster assembly / Mitochondrial iron-sulfur cluster biogenesis / Complex III assembly ...Electron transport from NADPH to Ferredoxin / Defective CYP11A1 causes AICSR / negative regulation of iron ion import across plasma membrane / molybdopterin cofactor metabolic process / Molybdenum cofactor biosynthesis / L-cysteine desulfurase complex / iron incorporation into metallo-sulfur cluster / [4Fe-4S] cluster assembly / Mitochondrial iron-sulfur cluster biogenesis / Complex III assembly / P450-containing electron transport chain / positive regulation of mitochondrial electron transport, NADH to ubiquinone / Maturation of TCA enzymes and regulation of TCA cycle / cysteine desulfurase / cysteine desulfurase activity / Mo-molybdopterin cofactor biosynthetic process / Pregnenolone biosynthesis / mitochondrial [2Fe-2S] assembly complex / iron-sulfur cluster assembly complex / [2Fe-2S] cluster assembly / lipid biosynthetic process / lipid A biosynthetic process / iron-sulfur cluster assembly / acyl binding / acyl carrier activity / iron-sulfur cluster binding / phosphopantetheine binding / Endogenous sterols / electron transport chain / ferrous iron binding / 2 iron, 2 sulfur cluster binding / fatty acid biosynthetic process / pyridoxal phosphate binding / Maturation of replicase proteins / molecular adaptor activity / intracellular iron ion homeostasis / electron transfer activity / nuclear body / mitochondrial matrix / iron ion binding / response to xenobiotic stimulus / lipid binding / centrosome / protein homodimerization activity / mitochondrion / zinc ion binding / nucleoplasm / metal ion binding / nucleus / membrane / cytosol / cytoplasm
Similarity search - Function
LYRM4, LYR domain / : / ISC system FeS cluster assembly, IscU scaffold / Adrenodoxin, iron-sulphur binding site / Adrenodoxin family, iron-sulfur binding region signature. / Cysteine desulfurase IscS / NIF system FeS cluster assembly, NifU, N-terminal / NifU-like N terminal domain / Adrenodoxin / Cysteine desulfurase ...LYRM4, LYR domain / : / ISC system FeS cluster assembly, IscU scaffold / Adrenodoxin, iron-sulphur binding site / Adrenodoxin family, iron-sulfur binding region signature. / Cysteine desulfurase IscS / NIF system FeS cluster assembly, NifU, N-terminal / NifU-like N terminal domain / Adrenodoxin / Cysteine desulfurase / Aminotransferase class-V, pyridoxal-phosphate binding site / Aminotransferases class-V pyridoxal-phosphate attachment site. / Aminotransferase class V domain / Aminotransferase class-V / 2Fe-2S iron-sulfur cluster binding domain / Complex 1 LYR protein domain / Complex 1 protein (LYR family) / Beta-grasp domain superfamily / Acyl carrier protein (ACP) / 2Fe-2S ferredoxin-type iron-sulfur binding domain profile. / 2Fe-2S ferredoxin-type iron-sulfur binding domain / 2Fe-2S ferredoxin-like superfamily / Phosphopantetheine attachment site / Phosphopantetheine attachment site. / Phosphopantetheine attachment site / ACP-like superfamily / Carrier protein (CP) domain profile. / Phosphopantetheine binding ACP domain / Pyridoxal phosphate-dependent transferase, small domain / Pyridoxal phosphate-dependent transferase, major domain / Pyridoxal phosphate-dependent transferase
Similarity search - Domain/homology
Chem-8Q1 / : / FE2/S2 (INORGANIC) CLUSTER / Acyl carrier protein / Ferredoxin-2, mitochondrial / Iron-sulfur cluster assembly enzyme ISCU / LYR motif-containing protein 4 / Cysteine desulfurase
Similarity search - Component
Biological speciesHomo sapiens (human)
Escherichia coli BL21 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.33 Å
AuthorsSteinhilper, R. / Murphy, B.J.
Funding support Germany, 1items
OrganizationGrant numberCountry
Max Planck Society Germany
CitationJournal: Nat Commun / Year: 2024
Title: Two-stage binding of mitochondrial ferredoxin-2 to the core iron-sulfur cluster assembly complex.
Authors: Ralf Steinhilper / Linda Boß / Sven-A Freibert / Vinzent Schulz / Nils Krapoth / Susann Kaltwasser / Roland Lill / Bonnie J Murphy /
Abstract: Iron-sulfur (FeS) protein biogenesis in eukaryotes begins with the de novo assembly of [2Fe-2S] clusters by the mitochondrial core iron-sulfur cluster assembly (ISC) complex. This complex comprises ...Iron-sulfur (FeS) protein biogenesis in eukaryotes begins with the de novo assembly of [2Fe-2S] clusters by the mitochondrial core iron-sulfur cluster assembly (ISC) complex. This complex comprises the scaffold protein ISCU2, the cysteine desulfurase subcomplex NFS1-ISD11-ACP1, the allosteric activator frataxin (FXN) and the electron donor ferredoxin-2 (FDX2). The structural interaction of FDX2 with the complex remains unclear. Here, we present cryo-EM structures of the human FDX2-bound core ISC complex showing that FDX2 and FXN compete for overlapping binding sites. FDX2 binds in either a 'distal' conformation, where its helix F interacts electrostatically with an arginine patch of NFS1, or a 'proximal' conformation, where this interaction tightens and the FDX2-specific C terminus binds to NFS1, facilitating the movement of the [2Fe-2S] cluster of FDX2 closer to the ISCU2 FeS cluster assembly site for rapid electron transfer. Structure-based mutational studies verify the contact areas of FDX2 within the core ISC complex.
History
DepositionJan 5, 2024Deposition site: PDBE / Processing site: PDBE
Revision 1.0Dec 18, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Isoform Mitochondrial of Cysteine desulfurase
B: LYR motif-containing protein 4
C: Acyl carrier protein
D: Isoform 1 of Iron-sulfur cluster assembly enzyme ISCU
I: Ferredoxin-2, mitochondrial
E: Isoform Mitochondrial of Cysteine desulfurase
F: LYR motif-containing protein 4
H: Isoform 1 of Iron-sulfur cluster assembly enzyme ISCU
G: Acyl carrier protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)180,70814
Polymers179,3399
Non-polymers1,3695
Water4,558253
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Protein , 5 types, 9 molecules AEBFCGDHI

#1: Protein Isoform Mitochondrial of Cysteine desulfurase


Mass: 45165.566 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: NFS1, NIFS, HUSSY-08 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q9Y697, cysteine desulfurase
#2: Protein LYR motif-containing protein 4


Mass: 13502.373 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: LYRM4, C6orf149, ISD11, CGI-203 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q9HD34
#3: Protein Acyl carrier protein / ACP / Cytosolic-activating factor / CAF / Fatty acid synthase acyl carrier protein


Mass: 8645.460 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P0A6A8
#4: Protein Isoform 1 of Iron-sulfur cluster assembly enzyme ISCU / NifU-like N-terminal domain-containing protein / NifU-like protein


Mass: 15684.119 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ISCU, NIFUN / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q9H1K1
#5: Protein Ferredoxin-2, mitochondrial / Adrenodoxin-like protein / Ferredoxin-1-like protein


Mass: 13344.046 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: FDX2, FDX1L / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q6P4F2

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Non-polymers , 4 types, 258 molecules

#6: Chemical ChemComp-8Q1 / S-[2-({N-[(2R)-2-hydroxy-3,3-dimethyl-4-(phosphonooxy)butanoyl]-beta-alanyl}amino)ethyl] dodecanethioate / S-dodecanoyl-4'-phosphopantetheine


Mass: 540.651 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C23H45N2O8PS / Feature type: SUBJECT OF INVESTIGATION
#7: Chemical ChemComp-FE2 / FE (II) ION


Mass: 55.845 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Fe / Feature type: SUBJECT OF INVESTIGATION
#8: Chemical ChemComp-FES / FE2/S2 (INORGANIC) CLUSTER


Mass: 175.820 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Fe2S2 / Feature type: SUBJECT OF INVESTIGATION
#9: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 253 / 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 core ISC complex (NFS1-ISD11-ACP1-ISCU2-FXN/FDX2) under turnover conditions
Type: COMPLEX / Entity ID: #1-#5 / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R0.6/1
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS 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
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 80 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)

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Processing

EM software
IDNameVersionCategory
2EPU3.1image acquisition
3EPU3.3image acquisition
8Coot0.9model fitting
10PHENIX1.20.1model refinement
14cryoSPARC4.1.23D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 2.33 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 195329 / Symmetry type: POINT
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
16NZU

6nzu

16NZU1PDBexperimental model
22Y5C

2y5c

12Y5C2PDBexperimental model

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