FDX2

FDX2 is a mitochondrial [2Fe-2S] ferredoxin dedicated to iron-sulfur cluster biogenesis. It accepts electrons from NADPH via ferredoxin reductase FDXR and donates them to the NFS1/ISD11/ACP cysteine desulfurase complex to drive [2Fe-2S] cluster assembly on ISCU2, and subsequently catalyzes reductive [2Fe-2S] cluster fusion on the ISCA1-ISCA2 scaffold (with IBA57) to generate [4Fe-4S] clusters for mitochondrial apoproteins; its paralog FDX1 cannot substitute in these reactions, and the distinct substrate specificity is encoded by small conserved sequence motifs (PMID:20547883, PMID:32817474, PMID:36280795). FDX2 and frataxin compete for the same binding site on NFS1, and their stoichiometric balance governs Fe/S cluster output such that excess FDX2 inhibits frataxin-stimulated desulfurase activity and partial FDX2 reduction ameliorates frataxin-deficient phenotypes (PMID:41372402). Loss-of-function mutations in FDX2 cause a mitochondrial disorder with impaired respiratory chain complex activities, mitochondrial iron accumulation, and myopathy (PMID:24281368, PMID:41372147).

1. 2010 High

   The first functional dichotomy between the two human ferredoxins was established: FDX2 is essential for Fe/S cluster and heme A biosynthesis while FDX1 is specific for steroidogenesis, resolving a long-standing question about why mammals retain two mitochondrial ferredoxins.

   Evidence RNAi depletion of FDX1 and FDX2 in human cells with measurement of multiple mitochondrial pathways

   PMID:20547883

   Open questions at the time

   • No reconstituted biochemical system to show direct electron transfer to Fe/S assembly machinery
   • Heme A involvement of FDX2 was later reassigned to FDX1
   • Molecular basis of substrate selectivity between the two paralogs unknown
2. 2013 Medium

   A human loss-of-function mutation confirmed FDX2's physiological essentiality for Fe/S-dependent mitochondrial enzymes and established FDX2 deficiency as a cause of mitochondrial disease.

   Evidence Exome sequencing identifying c.1A>T start-codon mutation; respiratory chain enzyme assays in patient muscle

   PMID:24281368

   Open questions at the time

   • Single family; no rescue experiment to confirm causality
   • Degree of residual FDX2 protein expression not quantified precisely
   • No structure-function insight from this mutation
3. 2017 High

   The physical interaction between FDX2 and the NFS1/ISD11/ACP cysteine desulfurase complex was mapped at atomic resolution, revealing that FDX2 binds more tightly than FDX1 and supports faster Fe/S cluster assembly on ISCU.

   Evidence NMR binding-interface mapping, isothermal titration calorimetry, and in vitro Fe/S cluster assembly on ISCU

   PMID:28001042

   Open questions at the time

   • Binding interface of FDX2 with downstream late-acting ISC components not characterized
   • Physiological relevance of the affinity difference between FDX1 and FDX2 for the desulfurase complex not tested in cells
4. 2020 High

   FDX2 was shown to be uniquely required for the late step of [4Fe-4S] cluster maturation, catalyzing reductive [2Fe-2S] cluster fusion on the ISCA1-ISCA2 scaffold in an IBA57-dependent manner — a reaction no other cellular reductant could support.

   Evidence Full in vitro reconstitution of [4Fe-4S] aconitase maturation with all physiological ISC components

   PMID:32817474

   Open questions at the time

   • Structural basis of FDX2 recognition by the ISCA1-ISCA2 scaffold not determined
   • Whether FDX2 transfers one or two electrons per fusion event unclear
5. 2022 High

   The molecular determinants encoding ferredoxin substrate specificity were identified: small conserved sequence motifs can transfer FDX2's Fe/S assembly role to FDX1 and vice versa, definitively separating the two paralogs' functions.

   Evidence Motif-swapping mutagenesis combined with RNAi knockdown and in vitro electron transfer assays

   PMID:36280795

   Open questions at the time

   • Crystal or cryo-EM structure of the FDX2–target complex showing how these motifs mediate recognition unavailable
   • Whether the motifs affect binding affinity, electron transfer rate, or both was not quantitatively dissected
6. 2024 High

   The pathogenic P144L mutation was shown to impair FDXR-dependent electron delivery to FDX2 by disrupting protein-protein recognition at FDX2's C-terminal tail without perturbing the [2Fe-2S] cluster, revealing a functional role for the C-terminus in the electron transfer relay.

   Evidence NMR structural/dynamic comparison of WT vs. P144L FDX2; in vitro electron transfer assays with FDXR

   PMID:39467201

   Open questions at the time

   • Quantitative impact of P144L on in-cell Fe/S cluster levels not measured
   • Whether other disease mutations similarly target the FDXR interface unknown
7. 2024 Medium

   Conditional FDX2 knockout in cancer cells revealed that global Fe/S protein loss triggers Fe²⁺ overload, DNA damage, and p53 activation, with cell fate (senescence, apoptosis, or ferroptosis) determined by p53 status and lipid homeostasis.

   Evidence Conditional KO in ovarian cancer cells; Fe²⁺ quantification; DNA damage markers; ferroptosis sensitivity assays

   PMID:39151727

   Open questions at the time

   • Single cancer cell type; generalizability to other tissues not established
   • Whether ferroptosis sensitivity is a direct consequence of Fe/S loss or secondary iron overload not distinguished
8. 2025 High

   A forward genetic screen uncovered that FDX2 and frataxin compete for the same binding site on NFS1, establishing that their stoichiometric balance directly regulates Fe/S cluster output and providing a mechanistic basis for therapeutic reduction of FDX2 in Friedreich's ataxia.

   Evidence Genome-scale C. elegans screen; in vitro NFS1 activity titrations; mammalian cell Fe/S assays; mouse Friedreich's ataxia model with FDX2 heterozygous reduction

   PMID:41372402

   Open questions at the time

   • Structural basis of competitive binding at the shared NFS1 site not resolved at atomic level
   • Long-term safety and efficacy of FDX2 reduction in mammalian Friedreich's ataxia models not determined
   • Whether FDX2-frataxin competition is regulated by post-translational modifications unknown
9. 2025 Medium

   Paramagnetic NMR combined with DFT calculations resolved the electronic structure of FDX2's [2Fe-2S]²⁺ cluster, revealing inequivalent iron centers and spin-density transfer through C-H···S-Fe³⁺ interactions.

   Evidence Paramagnetic NMR with full binding-loop assignment; DFT quantum chemical calculations

   PMID:40121555

   Open questions at the time

   • No functional mutagenesis to test how perturbing these electronic features affects electron transfer
   • Whether these electronic properties differ between FDX2 and FDX1 and account for specificity not examined

• Key unresolved questions include the atomic-resolution structure of FDX2 in complex with NFS1 or ISCA1-ISCA2, the mechanism by which FDX2-frataxin competition is physiologically regulated, and whether FDX2 reduction is a viable therapeutic strategy for Friedreich's ataxia in long-term preclinical models.
• No high-resolution structure of FDX2 bound to any of its physiological partner complexes
• Regulatory mechanisms controlling FDX2 expression or stability in response to Fe/S demand unknown
• Therapeutic window for FDX2 reduction in Friedreich's ataxia not defined