Affinage

GLRX5

Glutaredoxin-related protein 5, mitochondrial · UniProt Q86SX6

Length
157 aa
Mass
16.6 kDa
Annotated
2026-06-10
25 papers in source corpus 15 papers cited in narrative 15 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 6/6 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

GLRX5 is a mitochondrial matrix monothiol glutaredoxin that serves as a central hub of the iron-sulfur (Fe/S) cluster assembly pathway, required for maturation of all cellular Fe/S proteins regardless of cluster type or subcellular location (PMID:11950925, PMID:23615440, PMID:40074084). It coordinates a bridging [2Fe-2S] cluster through its active-site Cys67 and two glutathione molecules, transitioning from a monomeric apoprotein to a cluster-bound oligomer, with glutathione contacts dictating cluster chirality (PMID:21029046, PMID:29264659). GLRX5 receives pre-assembled clusters from the scaffold protein ISCU/Isu1, a transfer facilitated by the mitochondrial Hsp70 chaperone Ssq1, which binds GLRX5 at a site distinct from the scaffold and positions the two proteins for rapid cluster hand-off (PMID:23615440); reconstitution in isolated mitochondria places GLRX5 upstream of the Isa1/Isa2 machinery and shows it is needed both for intramitochondrial Fe/S synthesis and for the exported intermediate that supports cytoplasmic Fe/S assembly (PMID:40074084). Downstream, GLRX5 forms cluster-bridged heterodimers with BOLA1 and BOLA3 that diverge functionally: BOLA1-GLRX5 binds a reduced Rieske-type cluster and cannot donate it (a non-trafficking, redox role), whereas BOLA3-GLRX5 carries an oxidized ferredoxin-type cluster competent for trafficking (PMID:28483642, PMID:32542995). Through this activity GLRX5 enables maturation of Fe/S enzymes including IRP1, mitochondrial aconitase, ferrochelatase and succinate dehydrogenase, lipoylation of the pyruvate dehydrogenase and α-ketoglutarate dehydrogenase complexes, and heme biosynthesis via ferrochelatase and ALAS2 (PMID:26100117, PMID:30660387). Loss-of-function GLRX5 mutations in humans cause sideroblastic anemia and variant nonketotic hyperglycinemia, with patient defects in lipoylation and PDH activity corrected by wild-type GLRX5 (PMID:24334290, PMID:30660387).

Mechanistic history

Synthesis pass · year-by-year structured walk · 14 steps
  1. 1999 High

    Established the founding function of the GLRX5 ortholog as a monothiol glutaredoxin protecting cells against oxidative protein damage, before any Fe/S role was known.

    Evidence Genetic deletion of yeast grx5 with protein carbonyl assays and oxidant sensitivity screens

    PMID:10567543

    Open questions at the time
    • Did not identify a molecular substrate or distinguish a direct redox role from an indirect consequence of metabolic dysfunction
    • No connection to Fe/S biology yet
  2. 2002 High

    Localized the protein to the mitochondrial matrix and placed it in the Fe/S assembly machinery, recasting the oxidative phenotype as a downstream consequence of failed Fe/S biogenesis.

    Evidence Subcellular fractionation, suppressor-overexpression epistasis (SSQ1, ISA2), Fe/S enzyme assays and iron measurement in yeast null mutants

    PMID:11950925

    Open questions at the time
    • Did not define the biochemical step catalyzed within the pathway
    • Mechanism of iron accumulation not resolved
  3. 2002 High

    Mapped the residues essential for both Fe/S assembly and oxidant resistance, identifying the active-site cysteine and the glutathione cleft as functionally required.

    Evidence Site-directed mutagenesis with 3D modeling and multiple phenotypic readouts in yeast

    PMID:12138088

    Open questions at the time
    • Structural model not experimentally determined at the time
    • Did not establish whether the active-site cysteine binds a cluster directly
  4. 2003 High

    Defined the redox chemistry of the active site, showing a GSH-dependent thiol reductase cycle and quantifying its sluggish kinetics relative to dithiol glutaredoxins.

    Evidence Purified wild-type and cysteine-mutant proteins, redox potential and pKa measurements, in vitro glutathionylation/reduction assays

    PMID:12730244

    Open questions at the time
    • Did not reconcile the slow reductase activity with the essential Fe/S role
    • Physiological substrate of the reductase activity not identified
  5. 2006 Medium

    Demonstrated functional conservation by showing human GLRX5 targeted to yeast mitochondria fully rescues Fe/S defects, validating the yeast model for human biology.

    Evidence Heterologous complementation of yeast grx5 null with human GLRX5 plus localization

    PMID:16566929

    Open questions at the time
    • Cross-species rescue does not prove identical human partner interactions
    • Single lab
  6. 2010 Medium

    Established direct in vivo interaction with the Isa scaffold proteins and linked GLRX5 to mtDNA integrity, beginning to define its protein partnerships within the pathway.

    Evidence Bimolecular fluorescence complementation, multi-copy suppressor screen and Fe/S enzyme assays in fission yeast

    PMID:20085751

    Open questions at the time
    • BiFC indicates proximity but not direct stable complex stoichiometry
    • Directionality of cluster flow between Grx5 and Isa not resolved
  7. 2010 High

    Provided the structural basis for cluster coordination, showing a [2Fe-2S] cluster bridged by Cys67 from two protomers and two glutathione thiols, with an apo-monomer/holo-oligomer transition.

    Evidence X-ray crystallography of human GLRX5 with biophysical and enzymatic validation

    PMID:21029046

    Open questions at the time
    • Static structure does not capture cluster transfer dynamics
    • Relationship between the crystallographic tetramer and physiological transfer complexes unclear
  8. 2013 High

    Defined the Hsp70 chaperone Ssq1 as the platform that juxtaposes scaffold and GLRX5 to enable rapid cluster transfer, and established GLRX5 as required for maturation of all cellular Fe/S proteins.

    Evidence In vivo and in vitro interaction, ATPase, and cluster transfer assays with genetic depletion in yeast

    PMID:23615440

    Open questions at the time
    • Did not resolve how cluster is released to specific downstream acceptors
    • Human ortholog (HSPA9) interaction inferred by homology, not directly tested here
  9. 2013 High

    Linked GLRX5 to human disease and to lipoylation, showing patient mutations cause variant nonketotic hyperglycinemia via defective lipoylation, with rescue by wild-type GLRX5.

    Evidence Patient cell biochemistry, transfection rescue, and enzyme activity assays

    PMID:24334290

    Open questions at the time
    • Connection between Fe/S defect and lipoylation loss is indirect via Fe/S-dependent lipoyl synthase
    • Did not quantify cluster occupancy in patient cells
  10. 2016 High

    Separated GLRX5 function into cluster-binding and cluster-transfer steps through human KO cells and structure-function mutants, mapping discrete defects to downstream Fe/S targets and lipoylation.

    Evidence CRISPR GLRX5 knockout K562 cells with K101Q/L148S mutant rescue and multiple Fe/S enzyme and lipoylation readouts

    PMID:26100117

    Open questions at the time
    • Did not identify the direct acceptor proteins for each transfer step
    • Mechanism by which L148S impairs transfer not structurally resolved
  11. 2017 High

    Identified divergent BOLA1 and BOLA3 heterocomplexes with distinct cluster types, revealing two functionally specialized GLRX5-containing complexes.

    Evidence UV/vis, CD, EPR, NMR spectroscopy and protein-protein docking on reconstituted human complexes

    PMID:28483642

    Open questions at the time
    • Functional roles assigned by cluster type were not yet tested by transfer assays
    • In vivo abundance of each complex not measured
  12. 2017 Medium

    Showed that glutathione in the binding pocket controls cluster chirality and that thiol substitutions permit transfer to apo ferredoxin, refining the chemistry of cluster handling.

    Evidence In vitro reconstitution with natural and non-natural amino acid substitutions, CD spectroscopy, transfer kinetics to apo ferredoxin

    PMID:29264659

    Open questions at the time
    • Single lab in vitro reconstitution
    • Functional consequence of altered chirality in cells not established
  13. 2020 Medium

    Distinguished the BOLA1-GLRX5 and BOLA3-GLRX5 complexes functionally, showing BOLA1-GLRX5 accepts but cannot donate cluster (a non-trafficking/redox role) versus the donating BOLA3-GLRX5 complex.

    Evidence CD spectroscopy cluster transfer kinetics with defined donor/acceptor proteins

    PMID:32542995

    Open questions at the time
    • In vitro inability to donate does not exclude a partner-dependent donation in cells
    • Physiological redox substrate of BOLA1-GLRX5 unidentified
  14. 2025 High

    Positioned GLRX5 upstream of the Isa1/Isa2 machinery as the central hub for both mitochondrial and exported cytoplasmic Fe/S assembly using organellar reconstitution.

    Evidence In vitro import of purified Grx5 precursor into isolated yeast mitochondria with Fe/S synthesis assays and genetic depletion of Grx5/Isa1/Isa2

    PMID:40074084

    Open questions at the time
    • Molecular identity of the exported (Fe-S)int intermediate not defined
    • How GLRX5 partitions cluster between mitochondrial maturation and export not resolved

Open questions

Synthesis pass · forward-looking unresolved questions
  • The physiological redox substrate of the GLRX5 thiol reductase activity and the precise molecular nature of the exported Fe/S intermediate it generates remain unresolved.
  • No identified in vivo target of GLRX5 glutaredoxin reductase activity distinct from its Fe/S role
  • Chemical identity and protein carrier of the cytoplasmic (Fe-S)int intermediate unknown
  • Determinants directing cluster to BOLA1 vs BOLA3 complexes in cells not defined

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0140104 molecular carrier activity 3 GO:0016491 oxidoreductase activity 2 GO:0140098 catalytic activity, acting on RNA 2
Localization
GO:0005739 mitochondrion 2
Pathway
R-HSA-1852241 Organelle biogenesis and maintenance 3 R-HSA-1430728 Metabolism 2 R-HSA-1643685 Disease 2
Partners
BOLA1BOLA3HSPA9ISCA1ISCA2ISCU
Complex memberships
BOLA1-GLRX5 [2Fe-2S]-bridged heterodimerBOLA3-GLRX5 [2Fe-2S]-bridged heterodimer

Evidence

Reading pass · 15 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
1999 Yeast Grx5 (ortholog of human GLRX5) is a monothiol glutaredoxin required to protect against constitutive oxidative protein damage; grx5 null mutants show increased total protein carbonyl content and specific oxidation of proteins such as transketolase, and are highly sensitive to menadione and hydrogen peroxide. Genetic deletion (grx5 null mutant), protein carbonyl assay, growth sensitivity assays, synthetic lethality with grx2 and grx3/grx4 double mutants Molecular and cellular biology High 10567543
2002 Yeast Grx5 localizes to the mitochondrial matrix (mature form lacks the first 29 amino acids of the translation product) and is required for iron-sulfur (Fe/S) cluster assembly; absence of Grx5 causes iron accumulation and inactivation of Fe/S-dependent enzymes. Overexpression of SSQ1 and ISA2 (Fe/S assembly genes) suppresses grx5 phenotypes, placing Grx5 in the mitochondrial Fe/S assembly machinery. Subcellular fractionation/localization, genetic epistasis (suppressor overexpression), enzyme activity assays, iron measurement in null mutants Molecular biology of the cell High 11950925
2002 Structure-function analysis of yeast Grx5 identifies Cys60 and Gly61 as essential for function (Fe/S cluster assembly and oxidant resistance), while Cys117 is not essential; Gly115/Gly116 are important for glutathione cleft formation; Phe50 is required for proper thioredoxin-fold beta-sheet structure. Site-directed mutagenesis of conserved residues, 3D modeling, phenotypic assays (oxidant sensitivity, respiratory growth, amino acid auxotrophy, iron accumulation) The Journal of biological chemistry High 12138088
2003 Yeast Grx5 has a redox potential of -175 mV; the conserved Cys60 (pKa 5.0) forms a transient mixed disulfide with glutathione (GSSG), which then promotes a decrease in Cys117 pKa (8.2 → lower) triggering an intramolecular disulfide bond between Cys60 and Cys117; the disulfide is reduced by GSH ~20× more slowly than E. coli Grx1; Grx5 efficiently reduces glutathiolated substrate proteins, consistent with a thiol reductase function in mitochondria. Purification of wild-type and C60S/C117S mutant proteins, redox potential measurement, iodoacetamide titration at different pH, in vitro glutathionylation/reduction assays The Journal of biological chemistry High 12730244
2006 Human GLRX5 (hGRX5) contains a mitochondrial targeting sequence and, when expressed in the mitochondrial matrix of yeast grx5 null mutants, fully rescues the Fe/S cluster assembly defects, demonstrating functional conservation of human GLRX5 in mitochondrial Fe/S biogenesis. Heterologous complementation in yeast grx5 null mutant, mitochondrial targeting/localization, phenotypic rescue assays FEBS letters Medium 16566929
2010 Fission yeast Grx5 interacts in vivo with Fe/S scaffold proteins Isa1 and Isa2 in mitochondria (demonstrated by bimolecular fluorescence complementation); multi-copy overexpression of isa1+ or isa2+ (but not isu1+ or ssc1+) suppresses grx5 null growth defects and partially restores Fe/S enzyme activities; Grx5 also supports mitochondrial DNA integrity. Bimolecular fluorescence complementation (BiFC) for in vivo protein interaction, multi-copy suppressor screen, Fe/S enzyme activity assays, mtDNA quantification Biochemical and biophysical research communications Medium 20085751
2011 Crystal structure of human GLRX5 bound to two [2Fe-2S] clusters and four GSH molecules reveals: (1) tetrameric organization with clusters buried in the interior; (2) each [2Fe-2S] cluster coordinated by the N-terminal active-site Cys67 thiols from two protomers and two cysteine thiols from two GSH molecules; (3) the apoprotein is monomeric while the holo form is tetrameric; (4) glutathionylation of Cys67 occurs in the absence of cluster, potentially protecting it; (5) apo-GLRX5 reduces glutathione mixed disulfides ~100× more slowly than GLRX2 but is active as a glutathione-dependent electron donor for ribonucleotide reductase. X-ray crystallography, gel-filtration chromatography, analytical ultracentrifugation, mass spectrometry, in vitro enzymatic assay The Biochemical journal High 21029046
2013 Mitochondrial Hsp70 chaperone Ssq1 interacts with Grx5 at a binding site distinct from that of scaffold protein Isu1; Grx5 binding is most pronounced for the ADP-bound form of Ssq1; Grx5 binding does not stimulate Ssq1 ATPase activity; the proximity of Isu1 and Grx5 on Ssq1 facilitates rapid Fe/S cluster transfer from Isu1 to Grx5; Grx5 and its bound Fe/S cluster are required for maturation of all cellular Fe/S proteins regardless of cluster type or subcellular localization. In vivo and in vitro interaction assays, ATPase activity measurement, Fe/S cluster transfer assays, genetic depletion studies Molecular biology of the cell High 23615440
2013 Human GLRX5 mutations cause variant nonketotic hyperglycinemia with deficient lipoylation of mitochondrial proteins and reduced pyruvate dehydrogenase activity; transfection with wild-type GLRX5 corrects the biochemical deficiency in patient cells, establishing GLRX5 as required for lipoylation (and thus indirectly for glycine cleavage enzyme function). Patient cell biochemistry, transfection rescue experiments, enzyme activity assays (lipoylation, PDH, glycine cleavage), genetic sequencing Brain : a journal of neurology High 24334290
2016 Using GLRX5 knockout K562 cells, the K101Q mutation (preventing Fe/S binding to GLRX5) and the L148S mutation (interfering with Fe/S transfer from GLRX5 to downstream targets IRP1, mitochondrial aconitase, and ferrochelatase) define distinct functional domains; GLRX5 is required for lipoylation of pyruvate dehydrogenase complex and α-ketoglutarate dehydrogenase complex components. CRISPR/genetic knockout of GLRX5 in K562 cells, mutant transfection, Fe/S enzyme activity assays (IRP1, m-aconitase, ferrochelatase, succinate dehydrogenase), lipoylation western blot Journal of cellular biochemistry High 26100117
2017 Human mitochondrial BOLA1 and BOLA3 each form [2Fe-2S] cluster-bridged dimeric heterocomplexes with GRX5; BOLA1-GRX5 coordinates a reduced Rieske-type [2Fe-2S]1+ cluster while BOLA3-GRX5 coordinates an oxidized ferredoxin-like [2Fe-2S]2+ cluster; BOLA1-GRX5 is preferentially formed over BOLA3-GRX5 due to higher cluster binding affinity. UV/vis spectroscopy, CD spectroscopy, EPR spectroscopy, NMR spectroscopy, computational protein-protein docking (experimentally-driven structural models) Biochimica et biophysica acta. General subjects High 28483642
2017 Glutathione residues in the GRX5 binding pocket provide ionic and hydrogen-bonding contacts critical for cluster chirality; Cys67 of GRX5 coordinates the [2Fe-2S] cluster through glutathione; substitution of glutathione analogs or other thiols (DTT, L-cysteine) allows cluster reconstitution and transfer to apo ferredoxin 1 at comparable rates, but alters CD spectra reflecting perturbations in local cluster chirality. In vitro chemical reconstitution with natural and non-natural amino acid substitutions, CD spectroscopy, cluster transfer kinetics to apo ferredoxin Journal of biological inorganic chemistry Medium 29264659
2019 GLRX5 mutations (Cys67Tyr and Met128Lys) in a sideroblastic anemia patient impair both ferrochelatase activity (without porphyrin accumulation) and ALAS2 activity (possibly via defective succinyl-CoA biogenesis); structural analysis confirms Cys67 coordinates the [2Fe-2S] cluster and Met128 is implicated in partner protein interactions; GLRX5 loss also causes oxidative stress (reduced glutathione, decreased aconitase activity), mtDNA damage, and decreased respiratory chain complex I and IV activities. Patient-derived lymphoblastoid and CD34+ cell biochemistry, 3D structure analysis, enzyme activity assays (ferrochelatase, ALAS2, aconitase, complexes I/IV), oxidative stress markers, mtDNA quantification Molecular genetics and metabolism Medium 30660387
2020 [2Fe-2S]-bridged BOLA1-GLRX5 heterodimeric complex can accept cluster from ISCU or [2Fe-2S](GS)4 but not from ISCA1 or ISCA2; the holo BOLA1-GLRX5 complex is incapable of donating cluster to apo protein acceptors, providing experimental evidence for a non-trafficking (likely redox) role distinct from the cluster-donating BOLA3-GLRX5 complex. CD spectroscopy-based cluster transfer kinetics, in vitro reconstitution with defined donor/acceptor proteins The FEBS journal Medium 32542995
2025 Purified Grx5 precursor protein, imported into isolated Grx5-depleted yeast mitochondria, rescues both intra-mitochondrial Fe/S cluster synthesis and generation of the exported Fe/S intermediate (Fe-S)int required for cytoplasmic Fe/S assembly; mitochondria lacking Isa1 or Isa2 can still synthesize [2Fe-2S] but not [4Fe-4S] clusters and can still support cytoplasmic Fe/S assembly, placing Grx5 upstream of Isa1/Isa2 as a central hub for both mitochondrial and cytoplasmic Fe/S cluster biogenesis. Isolated mitochondria and cytoplasm from S. cerevisiae, in vitro import of purified Grx5 precursor, Fe/S cluster synthesis assays in isolated organelles, genetic depletion of Grx5/Isa1/Isa2 The Journal of biological chemistry High 40074084

Source papers

Stage 0 corpus · 25 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2002 Grx5 is a mitochondrial glutaredoxin required for the activity of iron/sulfur enzymes. Molecular biology of the cell 379 11950925
1999 Grx5 glutaredoxin plays a central role in protection against protein oxidative damage in Saccharomyces cerevisiae. Molecular and cellular biology 246 10567543
2013 Variant non ketotic hyperglycinemia is caused by mutations in LIAS, BOLA3 and the novel gene GLRX5. Brain : a journal of neurology 170 24334290
2003 Biochemical characterization of yeast mitochondrial Grx5 monothiol glutaredoxin. The Journal of biological chemistry 109 12730244
2013 The mitochondrial Hsp70 chaperone Ssq1 facilitates Fe/S cluster transfer from Isu1 to Grx5 by complex formation. Molecular biology of the cell 105 23615440
2011 The crystal structure of human GLRX5: iron-sulfur cluster co-ordination, tetrameric assembly and monomer activity. The Biochemical journal 102 21029046
2006 Prokaryotic and eukaryotic monothiol glutaredoxins are able to perform the functions of Grx5 in the biogenesis of Fe/S clusters in yeast mitochondria. FEBS letters 63 16566929
2002 Structure-function analysis of yeast Grx5 monothiol glutaredoxin defines essential amino acids for the function of the protein. The Journal of biological chemistry 63 12138088
2010 Monothiol glutaredoxin Grx5 interacts with Fe-S scaffold proteins Isa1 and Isa2 and supports Fe-S assembly and DNA integrity in mitochondria of fission yeast. Biochemical and biophysical research communications 53 20085751
2017 Structural insights into the molecular function of human [2Fe-2S] BOLA1-GRX5 and [2Fe-2S] BOLA3-GRX5 complexes. Biochimica et biophysica acta. General subjects 44 28483642
2016 Functional Analysis of GLRX5 Mutants Reveals Distinct Functionalities of GLRX5 Protein. Journal of cellular biochemistry 36 26100117
2019 GLRX5 mutations impair heme biosynthetic enzymes ALA synthase 2 and ferrochelatase in Human congenital sideroblastic anemia. Molecular genetics and metabolism 25 30660387
2004 Predictive reconstruction of the mitochondrial iron-sulfur cluster assembly metabolism. II. Role of glutaredoxin Grx5. Proteins 25 15382238
2020 A novel lncRNA BADLNCR1 inhibits bovine adipogenesis by repressing GLRX5 expression. Journal of cellular and molecular medicine 19 32449295
2024 Salvia miltiorrhiza Bge. processed with porcine cardiac blood inhibited GLRX5-mediated ferroptosis alleviating cerebral ischemia-reperfusion injury. Phytomedicine : international journal of phytotherapy and phytopharmacology 12 38677272
2023 Exosomes from hypoxic pretreated ADSCs attenuate ultraviolet light-induced skin injury via GLRX5 delivery and ferroptosis inhibition. Photochemical & photobiological sciences : Official journal of the European Photochemistry Association and the European Society for Photobiology 11 38100056
2021 HACE1, GLRX5, and ELP2 gene variant cause spastic paraplegies. Acta neurologica Belgica 9 33813722
2025 Mitochondrial glutaredoxin Grx5 functions as a central hub for cellular iron-sulfur cluster assembly. The Journal of biological chemistry 8 40074084
2021 Case Report: A Variant Non-ketotic Hyperglycinemia With GLRX5 Mutations: Manifestation of Deficiency of Activities of the Respiratory Chain Enzymes. Frontiers in genetics 8 34054912
2021 GLRX5-associated [Fe-S] cluster biogenesis disorder: further characterisation of the neurological phenotype and long-term outcome. Orphanet journal of rare diseases 7 34732213
2017 Investigation of glutathione-derived electrostatic and hydrogen-bonding interactions and their role in defining Grx5 [2Fe-2S] cluster optical spectra and transfer chemistry. Journal of biological inorganic chemistry : JBIC : a publication of the Society of Biological Inorganic Chemistry 7 29264659
2020 Cluster exchange reactivity of [2Fe-2S]-bridged heterodimeric BOLA1-GLRX5. The FEBS journal 6 32542995
2024 Case report: Unveiling genetic and phenotypic variability in Nonketotic hyperglycinemia: an atypical early onset case associated with a novel GLRX5 variant. Frontiers in genetics 1 39323869
2009 Cloning, overproduction, purification, crystallization and preliminary X-ray diffraction analysis of yeast glutaredoxin Grx5. Acta crystallographica. Section F, Structural biology and crystallization communications 1 19478456
2025 Infant With a Severe Form of GLRX5-Related Atypical Hyperglycinemia Exhibiting Novel Cardiac and Neurologic Disease Manifestations at Autopsy. Pediatric and developmental pathology : the official journal of the Society for Pediatric Pathology and the Paediatric Pathology Society 0 40415601

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