Superoxide-Driven Aconitase FE-S Center Cycling

1997 ◽  
Vol 17 (1) ◽  
pp. 33-42 ◽  
Author(s):  
Paul R. Gardner
Keyword(s):  
Mammalian Cells ◽  
Oxidant Stress ◽  
Iron Sulfur Cluster ◽  
Cycle Enzyme ◽  
Reactive Iron ◽  
Iron Sulfur ◽  
Iron Sulfur Center ◽  
Cyclical Process ◽  
Electron Carriers ◽  
Sulfur Containing

O−2 produced by the autoxidation of respiratory chain electron carriers, and other cellular reductants, inactivates bacterial and mammalian iron-sulfur-containing (de)hydratases including the citric acid cycle enzyme aconitase. Release of the solvent-exposed iron atom and oxidation of the [4Fe-4S]2+ cluster accompanies loss of catalytic activity. Rapid reactivation is achieved by iron-sulfur cluster reduction and Fe2+ insertion. Inactivation-reactivation is a dynamic and cyclical process which modulates aconitase and (de)hydratase activities in Escherichia coli and mammalian cells. The balance of inactive and active aconitase provides a sensitive measure of the changes in steady-statO−2 levels occuring in living cells and mitochondria under stress conditions. Aconitases are also inactivated by other oxidants including O2, H2O2, NO., and ONOO− which are associated with inflammation, hyperoxia and other pathophysiological conditions. Loss of aconitase activity during oxidant stress may impair energy production, and the liberation of reactive iron may further enhance oxidative damage. Iron-sulfur center cycling may also serve adaptive functions by modulating gene expression or by signaling metabolic quiescence.

scholarly journals Recombination of 2Fe-2S ferredoxins reveals differences in the inheritance of thermostability and midpoint potential

2020 ◽  
Author(s):  
Ian J. Campbell ◽  
Dimithree Kahanda ◽  
Joshua T. Atkinson ◽  
Othneil N. Sparks ◽  
Jinyoung Kim ◽  
...  
Keyword(s):  
Electron Transfer ◽  
Physical Properties ◽  
Sulfite Reductase ◽  
Amino Acid Substitutions ◽  
Iron Sulfur Cluster ◽  
Midpoint Potential ◽  
Electron Transfer Pathway ◽  
Iron Sulfur ◽  
Wide Range ◽  
Electron Carriers

ABSTRACTHomologous recombination can be used to create enzymes that exhibit distinct activities and stabilities from proteins in nature, allowing researchers to overcome component limitations in synthetic biology. To investigate how recombination affects the physical properties of an oxidoreductase that transfers electrons, we created ferredoxin (Fd) chimeras by recombining distantly-related cyanobacterial and cyanomyophage Fds that present similar midpoint potentials but distinct thermostabilities. Fd chimeras having a wide range of amino acid substitutions retained the ability to coordinate an iron-sulfur cluster, although their thermostabilities varied with the fraction of residues inherited from each parent. The midpoint potentials of chimeric Fds also varied. However, all of the synthetic Fds exhibited midpoint potentials outside of the parental protein range. Each of the chimeric Fds could also be used to build synthetic pathways that support electron transfer between Fd-NADP reductase and sulfite reductase in Escherichia coli, although the chimeric Fds varied in the expression required to support similar levels of cellular electron transfer. These results show how recombination can be used to rapidly diversify the physical properties of protein electron carriers and reveal differences in the inheritance of thermostability and electrochemical properties. Furthermore, they illustrate how electron transfer efficiencies of chimeric Fds can be rapidly evaluated using a synthetic electron transfer pathway.

scholarly journals Catalytic Intermediate Crystal Structures of Cysteine Desulfurase from the Archaeon Thermococcus onnurineus NA1

Archaea ◽  
2017 ◽  
Vol 2017 ◽  
pp. 1-11 ◽  
Author(s):  
Thien-Hoang Ho ◽  
Kim-Hung Huynh ◽  
Diem Quynh Nguyen ◽  
Hyunjae Park ◽  
Kyoungho Jung ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Elemental Sulfur ◽  
Catalytic Mechanism ◽  
Cysteine Desulfurase ◽  
Thermococcus Onnurineus Na1 ◽  
Iron Sulfur Cluster ◽  
Terminal Electron Acceptor ◽  
Iron Sulfur ◽  
Angle Rotation ◽  
Sulfur Containing

Thermococcus onnurineus NA1 is an anaerobic archaeon usually found in a deep-sea hydrothermal vent area, which can use elemental sulfur (S0) as a terminal electron acceptor for energy. Sulfur, essential to many biomolecules such as sulfur-containing amino acids and cofactors including iron-sulfur cluster, is usually mobilized from cysteine by the pyridoxal 5′-phosphate- (PLP-) dependent enzyme of cysteine desulfurase (CDS). We determined the crystal structures of CDS from Thermococcus onnurineus NA1 (ToCDS), which include native internal aldimine (NAT), gem-diamine (GD) with alanine, internal aldimine structure with existing alanine (IAA), and internal aldimine with persulfide-bound Cys356 (PSF) structures. The catalytic intermediate structures showed the dihedral angle rotation of Schiff-base linkage relative to the PLP pyridine ring. The ToCDS structures were compared with bacterial CDS structures, which will help us to understand the role and catalytic mechanism of ToCDS in the archaeon Thermococcus onnurineus NA1.

scholarly journals Loss of Mitochondrial Localization of Human FANCG Causes Defective FANCJ Helicase

2020 ◽  
Vol 40 (23) ◽  
Author(s):  
Jagadeesh Chandra Bose K ◽  
Bishwajit Singh Kapoor ◽  
Kamal Mandal ◽  
Shubhrima Ghosh ◽  
Raveendra B. Mokhamatam ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Fanconi Anemia ◽  
Protein G ◽  
Repair Pathway ◽  
Mitochondrial Localization ◽  
Iron Sulfur Cluster ◽  
Iron Sulfur ◽  
Damage Repair ◽  
N Terminus ◽  
Sulfur Containing

ABSTRACT Fanconi anemia (FA) is a unique DNA damage repair pathway. To date, 22 genes have been identified that are associated with the FA pathway. A defect in any of those genes causes genomic instability, and the patients bearing the mutation become susceptible to cancer. In our earlier work, we identified that Fanconi anemia protein G (FANCG) protects the mitochondria from oxidative stress. In this report, we have identified eight patients having a mutation (C.65G>C), which converts arginine at position 22 to proline (p.Arg22Pro) in the N terminus of FANCG. The mutant protein, hFANCGR22P, is able to repair the DNA and able to retain the monoubiquitination of FANCD2 in the FANCGR22P/FGR22P cell. However, it lost mitochondrial localization and failed to protect mitochondria from oxidative stress. Mitochondrial instability in the FANCGR22P cell causes the transcriptional downregulation of mitochondrial iron-sulfur cluster biogenesis protein frataxin (FXN) and the resulting iron deficiency of FA protein FANCJ, an iron-sulfur-containing helicase involved in DNA repair.

scholarly journals Mitochondrial Dynamics: Functional Link with Apoptosis

2012 ◽  
Vol 2012 ◽  
pp. 1-10 ◽  
Author(s):  
Hidenori Otera ◽  
Katsuyoshi Mihara
Keyword(s):  
Mammalian Cells ◽  
Molecular Mechanisms ◽  
Current Knowledge ◽  
Mitochondrial Dynamics ◽  
Functional Link ◽  
Iron Sulfur Cluster ◽  
Atp Production ◽  
Iron Sulfur ◽  
Cellular Apoptosis

Mitochondria participate in a variety of physiologic processes, such as ATP production, lipid metabolism, iron-sulfur cluster biogenesis, and calcium buffering. The morphology of mitochondria changes dynamically due to their frequent fusion and division in response to cellular conditions, and these dynamics are an important constituent of apoptosis. The discovery of large GTPase family proteins that regulate mitochondrial dynamics, together with novel insights into the role of mitochondrial fusion and fission in apoptosis, has provided important clues to understanding the molecular mechanisms of cellular apoptosis. In this paper, we briefly summarize current knowledge of the role of mitochondrial dynamics in apoptosis and cell pathophysiology in mammalian cells.

scholarly journals Mouse Knock-out of IOP1 Protein Reveals Its Essential Role in Mammalian Cytosolic Iron-Sulfur Protein Biogenesis

2011 ◽  
Vol 286 (18) ◽  
pp. 15797-15805 ◽  
Author(s):  
Daisheng Song ◽  
Frank S. Lee
Keyword(s):  
Mammalian Cells ◽  
Essential Role ◽  
Cluster Assembly ◽  
Iron Sulfur Cluster ◽  
Knock Out ◽  
Adult Mice ◽  
Iron Sulfur ◽  
Sulfur Protein ◽  
Iron Sulfur Protein ◽  
Cellular Compartments

Iron-sulfur proteins play an essential role in a variety of biologic processes and exist in multiple cellular compartments. The biogenesis of these proteins has been the subject of extensive investigation, and particular focus has been placed on the pathways that assemble iron-sulfur clusters in the different cellular compartments. Iron-only hydrogenase-like protein 1 (IOP1; also known as nuclear prelamin A recognition factor like protein, or NARFL) is a human protein that is homologous to Nar1, a protein in Saccharomyces cerevisiae that, in turn, is an essential component of the cytosolic iron-sulfur protein assembly pathway in yeast. Previous siRNA-induced knockdown studies using mammalian cells point to a similar role for IOP1 in mammals. In the present studies, we pursued this further by knocking out Iop1 in Mus musculus. We find that Iop1 knock-out results in embryonic lethality before embryonic day 10.5. Acute, inducible global knock-out of Iop1 in adult mice results in lethality and significantly diminished activity of cytosolic aconitase, an iron-sulfur protein, in liver extracts. Inducible knock-out of Iop1 in mouse embryonic fibroblasts results in diminished activity of cytosolic but not mitochondrial aconitase and loss of cell viability. Therefore, just as with knock-out of Nar1 in yeast, we find that knock-out of Iop1/Narfl in mice results in lethality and defective cytosolic iron-sulfur cluster assembly. The findings demonstrate an essential role for IOP1 in this pathway.

scholarly journals SLC25A39 is necessary for mitochondrial glutathione import in mammalian cells

2021 ◽  
Author(s):  
Ying Wang ◽  
Frederick S Yen ◽  
Xiphias Ge Zhu ◽  
Rebecca C Timson ◽  
Ross Weber ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Redox Homeostasis ◽  
Protein Abundance ◽  
Iron Sulfur Cluster ◽  
Oxidative Reactions ◽  
Iron Sulfur ◽  
Molecular Machinery ◽  
Mitochondrial Glutathione ◽  
Proliferation In Vitro

Glutathione (GSH) is a small molecule thiol abundantly present in all eukaryotes with key roles in oxidative metabolism. Mitochondria, as the major site of oxidative reactions, must maintain sufficient levels of GSH to perform protective and biosynthetic functions. GSH is exclusively synthesized in the cytosol, yet the molecular machinery involved in mitochondrial GSH import remain elusive. Here, using organellar proteomics and metabolomics approaches, we identify SLC25A39, a mitochondrial membrane carrier of unknown function, to regulate GSH transport into mitochondria. SLC25A39 loss reduces mitochondrial GSH import and abundance without impacting whole cell GSH levels. Cells lacking both SLC25A39 and its paralog SLC25A40 exhibit defects in the activity and stability of iron-sulfur cluster containing proteins. Moreover, mitochondrial GSH import is necessary for cell proliferation in vitro and red blood cell development in mice. Remarkably, the heterologous expression of an engineered bifunctional bacterial GSH biosynthetic enzyme (GshF) in mitochondria enabled mitochondrial GSH production and ameliorated the metabolic and proliferative defects caused by its depletion. Finally, GSH availability negatively regulates SLC25A39 protein abundance, coupling redox homeostasis to mitochondrial GSH import in mammalian cells. Our work identifies SLC25A39 as an essential and regulated component of the mitochondrial GSH import machinery.

scholarly journals Mutations in the iron-sulfur cluster ligands of the human ferrochelatase lead to erythropoietic protoporphyria

Blood ◽  
2000 ◽  
Vol 96 (4) ◽  
pp. 1545-1549 ◽  
Author(s):  
Xiaoye Schneider-Yin ◽  
Laurent Gouya ◽  
Morna Dorsey ◽  
Urszula Rüfenacht ◽  
Jean-Charles Deybach ◽  
...  
Keyword(s):  
Enzymatic Activity ◽  
Point Mutations ◽  
Erythropoietic Protoporphyria ◽  
Iron Sulfur Cluster ◽  
Iron Sulfur ◽  
Iron Sulfur Center ◽  
Physiologic Role ◽  
Skin Photosensitivity ◽  
New Mutations

Abstract Ferrochelatase (FECH; EC 4.99.1.1) catalyzes the terminal step of the heme biosynthetic pathway. Defects in the human FECHgene may lead to erythropoietic protoporphyria (EPP), a rare inherited disorder characterized by diminished FECH activity with protoporphyrin overproduction and subsequent skin photosensitivity and in rare cases liver failure. Inheritance of EPP appeared to be autosomal dominant with possible modulation by low expression of the wild-type FECH allele. Animal FECHs have been demonstrated to be [2Fe-2S] cluster-containing proteins. Although enzymatic activity and stability of the protein appear to be dependent on the presence of the [2Fe-2S] cluster, the physiologic role of the iron-sulfur center remains to be unequivocally established. Three of the 4 [2Fe-2S] cluster-coordinating cysteines (ie, C403, C406, and C411 in the human enzyme) are located within the C-terminal domain. In this study 5 new mutations are identified in patients with EPP. Three of the point mutations, in 3 patients, resulted in FECH variants with 2 of the [2Fe-2S] cluster cysteines substituted with tyrosine, serine, and glycine (ie, C406Y, C406S, and C411G) and with undetectable enzymatic activity. Further, one of the patients exhibited a triple point mutation (T1224→A, C1225→T, and T1231→G) leading to the N408K/P409S/C411G variant. This finding is entirely novel and has not been reported in EPP. The mutations of the codons for 2 of the [2Fe-2S] cluster ligands in patients with EPP supports the importance of the iron-sulfur center for the proper functioning of mammalian FECH and, in at least humans, its absence has a direct clinical impact.

scholarly journals Mutations in the iron-sulfur cluster ligands of the human ferrochelatase lead to erythropoietic protoporphyria

Blood ◽  
2000 ◽  
Vol 96 (4) ◽  
pp. 1545-1549
Author(s):  
Xiaoye Schneider-Yin ◽  
Laurent Gouya ◽  
Morna Dorsey ◽  
Urszula Rüfenacht ◽  
Jean-Charles Deybach ◽  
...  
Keyword(s):  
Enzymatic Activity ◽  
Point Mutations ◽  
Erythropoietic Protoporphyria ◽  
Iron Sulfur Cluster ◽  
Iron Sulfur ◽  
Iron Sulfur Center ◽  
Physiologic Role ◽  
Skin Photosensitivity ◽  
New Mutations

Ferrochelatase (FECH; EC 4.99.1.1) catalyzes the terminal step of the heme biosynthetic pathway. Defects in the human FECHgene may lead to erythropoietic protoporphyria (EPP), a rare inherited disorder characterized by diminished FECH activity with protoporphyrin overproduction and subsequent skin photosensitivity and in rare cases liver failure. Inheritance of EPP appeared to be autosomal dominant with possible modulation by low expression of the wild-type FECH allele. Animal FECHs have been demonstrated to be [2Fe-2S] cluster-containing proteins. Although enzymatic activity and stability of the protein appear to be dependent on the presence of the [2Fe-2S] cluster, the physiologic role of the iron-sulfur center remains to be unequivocally established. Three of the 4 [2Fe-2S] cluster-coordinating cysteines (ie, C403, C406, and C411 in the human enzyme) are located within the C-terminal domain. In this study 5 new mutations are identified in patients with EPP. Three of the point mutations, in 3 patients, resulted in FECH variants with 2 of the [2Fe-2S] cluster cysteines substituted with tyrosine, serine, and glycine (ie, C406Y, C406S, and C411G) and with undetectable enzymatic activity. Further, one of the patients exhibited a triple point mutation (T1224→A, C1225→T, and T1231→G) leading to the N408K/P409S/C411G variant. This finding is entirely novel and has not been reported in EPP. The mutations of the codons for 2 of the [2Fe-2S] cluster ligands in patients with EPP supports the importance of the iron-sulfur center for the proper functioning of mammalian FECH and, in at least humans, its absence has a direct clinical impact.

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