The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform
Arne Holmgren - One of the best experts on this subject based on the ideXlab platform.
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modulation of thiol dependent redox system by metal ions via thioredoxin and Glutaredoxin systems
Metallomics, 2018Co-Authors: Yanfang Ouyang, Yi Peng, Arne HolmgrenAbstract:The thioredoxin and Glutaredoxin systems possess a variety of biological activities in mammalian cells, including the defense against oxidative stress, regulation of DNA synthesis, the cell cycle and the mediation of apoptosis. The thioredoxin system, comprised of NADPH, thioredoxin reductase (TrxR) and thioredoxin (Trx), exerts its activities via a disulfide–dithiol exchange reaction. Mammalian TrxRs are selenoproteins; the thiols and selenols in the active site of these enzymes confer the thioredoxin system to work as soft bases, which have a high affinity with soft acids, including numerous metal ions. In this review we focus on recent advances in the modulation of thioredoxin and Glutaredoxin systems by metal ion soft acids. Numerous clinical metal-containing drugs, such as platinum- and gold-containing compounds, show inhibitory effects on the thioredoxin system, providing strategies to develop novel anti-cancer drugs. Moreover, inhibition of the Trx system by soft acids, such as mercury-, chromium- and arsenic-containing compounds cause changes in the cellular redox state and contribute to their cell toxicity. In addition, metal ions are also involved in the regulation of the Glutaredoxin system. Iron ions participate in regulating Grx2 activity via iron–sulfur cluster formation. Moreover, Grx5 in mitochondria contains a 2Fe–2S cluster stabilized by GSH, which can mediate cellular iron metabolism. Collectively, these results demonstrate that metal ions are major players in regulating the Trx and Grx systems-mediated cellular redox processes and thus, provide an opportunity to understand the functions of metal ions in thiol metabolism dysfunction-related diseases.
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cellular redox systems impact the aggregation of cu zn superoxide dismutase linked to familial amyotrophic lateral sclerosis
Journal of Biological Chemistry, 2016Co-Authors: Cristina Alvarezzaldiernas, Yujuan Zheng, Hongqian Yang, Juan Blasi, Carles Solsona, Arne HolmgrenAbstract:Protein misfolding is implicated in neurodegenerative diseases such as ALS, where mutations of superoxide dismutase 1 (SOD1) account for about 20% of the inherited mutations. Human SOD1 (hSOD1) contains four cysteines, including Cys(57) and Cys(146), which have been linked to protein stability and folding via forming a disulfide bond, and Cys(6) and Cys(111) as free thiols. But the roles of the cellular oxidation-reduction (redox) environment in SOD1 folding and aggregation are not well understood. Here we explore the effects of cellular redox systems on the aggregation of hSOD1 proteins. We found that the known hSOD1 mutations G93A and A4V increased the capability of the thioredoxin and Glutaredoxin systems to reduce hSOD1 compared with wild-type hSOD1. Treatment with inhibitors of these redox systems resulted in an increase of hSOD1 aggregates in the cytoplasm of cells transfected with mutants but not in cells transfected with wild-type hSOD1 or those containing a secondary C111G mutation. This aggregation may be coupled to changes in the redox state of the G93A and A4V mutants upon mild oxidative stress. These results strongly suggest that the thioredoxin and Glutaredoxin systems are the key regulators for hSOD1 aggregation and may play critical roles in the pathogenesis of ALS.
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an unusual mode of iron sulfur cluster coordination in a teleost Glutaredoxin
Biochemical and Biophysical Research Communications, 2013Co-Authors: Lars Brautigam, Arne Holmgren, C. Johansson, Carsten Berndt, Bastian Kubsch, Michael A Mcdonough, Eckhard BillAbstract:Abstract Glutaredoxins that contain a Cys-X-X-Cys active site motif are glutathione-dependent thiol-disulfide oxidoreductases. Vertebrate Glutaredoxin 2 is characterized by two extra cysteines that form an intra-molecular disulfide bridge. Zebrafish Glutaredoxin 2 contains four additional cysteines that are conserved within the infraclass of bony fish (teleosts). Here, we present a biochemical and biophysical characterization of zebrafish Glutaredoxin 2, focusing on iron–sulfur-cluster coordination. The coordination of [2Fe2S]2+-clusters in monomers of this protein was revealed by both absorption and Mossbauer spectroscopy as well as size exclusion chromatography. All other holo-Glutaredoxins represent [FeS]-cluster bridged dimers using two molecules of non-covalently bound glutathione and the N-terminal active site cysteines as ligands. These cysteine residues were not required for [FeS]-cluster coordination in zebrafish Glutaredoxin 2. A crystal structure of the teleost protein revealed high structural similarity to its human homologue. The two vertebrate-specific cysteines as well as two of the teleost-specific cysteines are positioned within a radius of 7 A near the C-terminus suggesting a potential role in [FeS]-cluster coordination. Indeed, mutated proteins lacking these teleost-specific cysteines lost the ability to bind the cofactor. Hence, the apparent mode of [FeS]-cluster coordination in zebrafish Glutaredoxin 2 could be different from all yet described [FeS]-Glutaredoxins.
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glutathione and Glutaredoxin act as a backup of human thioredoxin reductase 1 to reduce thioredoxin 1 preventing cell death by aurothioglucose
Journal of Biological Chemistry, 2012Co-Authors: Huihui Zhang, Arne HolmgrenAbstract:Thioredoxin reductase 1 (TrxR1) in cytosol is the only known reductant of oxidized thioredoxin 1 (Trx1) in vivo so far. We and others found that aurothioglucose (ATG), a well known active-site inhibitor of TrxR1, inhibited TrxR1 activity in HeLa cell cytosol but had no effect on the viability of the cells. Using a redox Western blot analysis, no change was observed in redox state of Trx1, which was mainly fully reduced with five sulfhydryl groups. In contrast, auranofin killed cells and oxidized Trx1, also targeting mitochondrial TrxR2 and Trx2. Combining ATG with ebselen gave a strong synergistic effect, leading to Trx1 oxidation, reactive oxygen species accumulation, and cell death. We hypothesized that there should exist a backup system to reduce Trx1 when only TrxR1 activity was lost. Our results showed that physiological concentrations of glutathione, NADPH, and glutathione reductase reduced Trx1 in vitro and that the reaction was strongly stimulated by Glutaredoxin1. Simultaneous depletion of TrxR activity by ATG and glutathione by buthionine sulfoximine led to overoxidation of Trx1 and loss of HeLa cell viability. In conclusion, the Glutaredoxin system and glutathione have a backup role to keep Trx1 reduced in cells with loss of TrxR1 activity. Monitoring the redox state of Trx1 shows that cell death occurs when Trx1 is oxidized, followed by general protein oxidation catalyzed by the disulfide form of thioredoxin.
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vertebrate specific Glutaredoxin is essential for brain development
Proceedings of the National Academy of Sciences of the United States of America, 2011Co-Authors: Lars Brautigam, Christopher Horst Lillig, Arne Holmgren, Jose R Godoy, Lena Dorothee Schutte, Timour Prozorovski, Manuela Gellert, Giselbert Hauptmann, Carsten BerndtAbstract:Cellular functions and survival are dependent on a tightly controlled redox potential. Currently, an increasing amount of data supports the concept of local changes in the redox environment and specific redox signaling events controlling cell function. Specific protein thiol groups are the major targets of redox signaling and regulation. Thioredoxins and Glutaredoxins catalyze reversible thiol-disulfide exchange reactions and are primary regulators of the protein thiol redox state. Here, we demonstrate that embryonic brain development depends on the enzymatic activity of Glutaredoxin 2. Zebrafish with silenced expression of Glutaredoxin 2 lost virtually all types of neurons by apoptotic cell death and the ability to develop an axonal scaffold. As demonstrated in zebrafish and in a human cellular model for neuronal differentiation, Glutaredoxin 2 controls axonal outgrowth via thiol redox regulation of collapsin response mediator protein 2, a central component of the semaphorin pathway. This study provides an example of a specific thiol redox regulation essential for vertebrate embryonic development.
Carsten Berndt - One of the best experts on this subject based on the ideXlab platform.
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an unusual mode of iron sulfur cluster coordination in a teleost Glutaredoxin
Biochemical and Biophysical Research Communications, 2013Co-Authors: Lars Brautigam, Arne Holmgren, C. Johansson, Carsten Berndt, Bastian Kubsch, Michael A Mcdonough, Eckhard BillAbstract:Abstract Glutaredoxins that contain a Cys-X-X-Cys active site motif are glutathione-dependent thiol-disulfide oxidoreductases. Vertebrate Glutaredoxin 2 is characterized by two extra cysteines that form an intra-molecular disulfide bridge. Zebrafish Glutaredoxin 2 contains four additional cysteines that are conserved within the infraclass of bony fish (teleosts). Here, we present a biochemical and biophysical characterization of zebrafish Glutaredoxin 2, focusing on iron–sulfur-cluster coordination. The coordination of [2Fe2S]2+-clusters in monomers of this protein was revealed by both absorption and Mossbauer spectroscopy as well as size exclusion chromatography. All other holo-Glutaredoxins represent [FeS]-cluster bridged dimers using two molecules of non-covalently bound glutathione and the N-terminal active site cysteines as ligands. These cysteine residues were not required for [FeS]-cluster coordination in zebrafish Glutaredoxin 2. A crystal structure of the teleost protein revealed high structural similarity to its human homologue. The two vertebrate-specific cysteines as well as two of the teleost-specific cysteines are positioned within a radius of 7 A near the C-terminus suggesting a potential role in [FeS]-cluster coordination. Indeed, mutated proteins lacking these teleost-specific cysteines lost the ability to bind the cofactor. Hence, the apparent mode of [FeS]-cluster coordination in zebrafish Glutaredoxin 2 could be different from all yet described [FeS]-Glutaredoxins.
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crucial function of vertebrate Glutaredoxin 3 picot in iron homeostasis and hemoglobin maturation
Molecular Biology of the Cell, 2013Co-Authors: Petra Haunhorst, Lars Brautigam, Ulrich Muhlenhoff, Roland Lill, Bastian Hoffmann, Oliver Stehling, Evamaria Hanschmann, Carsten BerndtAbstract:The mechanisms by which eukaryotic cells handle and distribute the essential micronutrient iron within the cytosol and other cellular compartments are only beginning to emerge. The yeast monothiol multidomain Glutaredoxins (Grx) 3 and 4 are essential for both transcriptional iron regulation and intracellular iron distribution. Despite the fact that the mechanisms of iron metabolism differ drastically in fungi and higher eukaryotes, the Glutaredoxins are conserved, yet their precise function in vertebrates has remained elusive. Here we demonstrate a crucial role of the vertebrate-specific monothiol multidomain Grx3 (PICOT) in cellular iron homeostasis. During zebrafish embryonic development, depletion of Grx3 severely impairs the maturation of hemoglobin, the major iron-consuming process. Silencing of human Grx3 expression in HeLa cells decreases the activities of several cytosolic Fe/S proteins, for example, iron-regulatory protein 1, a major component of posttranscriptional iron regulation. As a consequence, Grx3-depleted cells show decreased levels of ferritin and increased levels of transferrin receptor, features characteristic of cellular iron starvation. Apparently, Grx3-deficient cells are unable to efficiently use iron, despite unimpaired cellular iron uptake. These data suggest an evolutionarily conserved role of cytosolic monothiol multidomain Glutaredoxins in cellular iron metabolism pathways, including the biogenesis of Fe/S proteins and hemoglobin maturation.
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vertebrate specific Glutaredoxin is essential for brain development
Proceedings of the National Academy of Sciences of the United States of America, 2011Co-Authors: Lars Brautigam, Christopher Horst Lillig, Arne Holmgren, Jose R Godoy, Lena Dorothee Schutte, Timour Prozorovski, Manuela Gellert, Giselbert Hauptmann, Carsten BerndtAbstract:Cellular functions and survival are dependent on a tightly controlled redox potential. Currently, an increasing amount of data supports the concept of local changes in the redox environment and specific redox signaling events controlling cell function. Specific protein thiol groups are the major targets of redox signaling and regulation. Thioredoxins and Glutaredoxins catalyze reversible thiol-disulfide exchange reactions and are primary regulators of the protein thiol redox state. Here, we demonstrate that embryonic brain development depends on the enzymatic activity of Glutaredoxin 2. Zebrafish with silenced expression of Glutaredoxin 2 lost virtually all types of neurons by apoptotic cell death and the ability to develop an axonal scaffold. As demonstrated in zebrafish and in a human cellular model for neuronal differentiation, Glutaredoxin 2 controls axonal outgrowth via thiol redox regulation of collapsin response mediator protein 2, a central component of the semaphorin pathway. This study provides an example of a specific thiol redox regulation essential for vertebrate embryonic development.
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characterization of the human monothiol Glutaredoxin 3 picot as iron sulfur protein
Biochemical and Biophysical Research Communications, 2010Co-Authors: Petra Haunhorst, Carsten Berndt, Susanne Eitner, Jose R Godoy, Christopher Horst LilligAbstract:Mammalian Glutaredoxin 3 (Grx3/PICOT) is an essential protein involved in the regulation of signal transduction, for instance during immune cell activation and development of cardiac hypertrophy, presumably in response to redox signals. This function requires the sensing of such stresses by a hitherto unknown mechanism. Here, we characterized Grx3/PICOT as iron–sulfur protein. The protein binds two bridging [2Fe–2S] clusters in a homodimeric complex with the active site cysteinyl residues of its two monothiol Glutaredoxin domains and glutathione bound non-covalently to the Grx domains. Co-immunoprecipitation of 55-iron with Grx3/PICOT from Jurkat cells suggested the presence of these cofactors under physiological conditions. The [2Fe–2S] 2+
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thiol redox control via thioredoxin and Glutaredoxin systems
Biochemical Society Transactions, 2005Co-Authors: Arne Holmgren, C. Johansson, Carsten Berndt, Maria Lonn, Christoph Hudemann, Christopher Horst LilligAbstract:The Trx (thioredoxin) and Grx (Glutaredoxin) systems control cellular redox potential, keeping a reducing thiol-rich intracellular state, which on generation of reactive oxygen species signals through thiol redox control mechanisms. Here, we give a brief overview of the human Trx and Grx systems. The main part focuses on our current knowledge about mitochondrial Grx2, which facilitates mitochondrial redox homoeostasis during oxidative stress-induced apoptosis.
Christopher Horst Lillig - One of the best experts on this subject based on the ideXlab platform.
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iron sulfur Glutaredoxin 2 protects oligodendrocytes against damage induced by nitric oxide release from activated microglia
Glia, 2017Co-Authors: Klaudia Lepka, Christopher Horst Lillig, Eckhard Bill, Katrin Volbracht, Reiner Schneider, Natalia Rios, Thomas Hildebrandt, Jens Ingwersen, Timur Prozorovski, Jack Van HorssenAbstract:Demyelinated brain lesions, a hallmark of autoimmune neuroinflammatory diseases like multiple sclerosis, result from oligodendroglial cell damage. Activated microglia are considered a major source of nitric oxide and subsequent peroxynitrite-mediated damage of myelin. Here, we provide biochemical and biophysical evidence that the oxidoreductase Glutaredoxin 2 inhibits peroxynitrite formation by transforming nitric oxide into dinitrosyl-diglutathionyl-iron-complexes. Glutaredoxin 2 levels influence both survival rates of primary oligodendrocyte progenitor cells and preservation of myelin structure in cerebellar organotypic slice cultures challenged with activated microglia or nitric oxide donors. Of note, Glutaredoxin 2-mediated protection is not linked to its enzymatic activity as oxidoreductase, but to the disassembly of its uniquely coordinated iron-sulfur cluster using glutathione as non-protein ligand. The protective effect of Glutaredoxin 2 is connected to decreased protein carbonylation and nitration. In line, brain lesions of mice suffering from experimental autoimmune encephalomyelitis, an animal model of multiple sclerosis, show decreased Glutaredoxin 2 expression and increased nitrotyrosine formation indicating that this type of protection is missing in the inflamed central nervous system. Our findings link inorganic biochemistry to neuroinflammation and identify Glutaredoxin 2 as a protective factor against neuroinflammation-mediated myelin damage. Thus, improved availability of glutathione-coordinated iron-sulfur clusters emerges as a potential therapeutic approach in inflammatory demyelination.
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vertebrate specific Glutaredoxin is essential for brain development
Proceedings of the National Academy of Sciences of the United States of America, 2011Co-Authors: Lars Brautigam, Christopher Horst Lillig, Arne Holmgren, Jose R Godoy, Lena Dorothee Schutte, Timour Prozorovski, Manuela Gellert, Giselbert Hauptmann, Carsten BerndtAbstract:Cellular functions and survival are dependent on a tightly controlled redox potential. Currently, an increasing amount of data supports the concept of local changes in the redox environment and specific redox signaling events controlling cell function. Specific protein thiol groups are the major targets of redox signaling and regulation. Thioredoxins and Glutaredoxins catalyze reversible thiol-disulfide exchange reactions and are primary regulators of the protein thiol redox state. Here, we demonstrate that embryonic brain development depends on the enzymatic activity of Glutaredoxin 2. Zebrafish with silenced expression of Glutaredoxin 2 lost virtually all types of neurons by apoptotic cell death and the ability to develop an axonal scaffold. As demonstrated in zebrafish and in a human cellular model for neuronal differentiation, Glutaredoxin 2 controls axonal outgrowth via thiol redox regulation of collapsin response mediator protein 2, a central component of the semaphorin pathway. This study provides an example of a specific thiol redox regulation essential for vertebrate embryonic development.
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characterization of the human monothiol Glutaredoxin 3 picot as iron sulfur protein
Biochemical and Biophysical Research Communications, 2010Co-Authors: Petra Haunhorst, Carsten Berndt, Susanne Eitner, Jose R Godoy, Christopher Horst LilligAbstract:Mammalian Glutaredoxin 3 (Grx3/PICOT) is an essential protein involved in the regulation of signal transduction, for instance during immune cell activation and development of cardiac hypertrophy, presumably in response to redox signals. This function requires the sensing of such stresses by a hitherto unknown mechanism. Here, we characterized Grx3/PICOT as iron–sulfur protein. The protein binds two bridging [2Fe–2S] clusters in a homodimeric complex with the active site cysteinyl residues of its two monothiol Glutaredoxin domains and glutathione bound non-covalently to the Grx domains. Co-immunoprecipitation of 55-iron with Grx3/PICOT from Jurkat cells suggested the presence of these cofactors under physiological conditions. The [2Fe–2S] 2+
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thioredoxin and related molecules from biology to health and disease
Antioxidants & Redox Signaling, 2007Co-Authors: Christopher Horst Lillig, Arne HolmgrenAbstract:Thioredoxin and Glutaredoxin systems in mammalian cells utilize thiol and selenol groups to maintain a reducing intracellular redox state acting as antioxidants and reducing agents in redox signaling with oxidizing reactive oxygen species. During the last decade, the functional roles of thioredoxin in particular have continued to expand, also including novel functions such as a secreted growth factor or a chemokine for immune cells. The role of thioredoxin and Glutaredoxin in antioxidant defense and the role of thioredoxin in controlling recruitment of inflammatory cells offer potential use in clinical therapy. The fundamental differences between bacterial and mammalian thioredoxin reductases offer new principles for treatment of infections. Clinical drugs already in use target the active site selenol in thioredoxin reductases, inducing cell death in tumor cells. Thioredoxin and binding proteins (ASK1 and TBP2) appear to control apoptosis or metabolic states such as carbohydrate and lipid metabolism relat...
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thiol redox control via thioredoxin and Glutaredoxin systems
Biochemical Society Transactions, 2005Co-Authors: Arne Holmgren, C. Johansson, Carsten Berndt, Maria Lonn, Christoph Hudemann, Christopher Horst LilligAbstract:The Trx (thioredoxin) and Grx (Glutaredoxin) systems control cellular redox potential, keeping a reducing thiol-rich intracellular state, which on generation of reactive oxygen species signals through thiol redox control mechanisms. Here, we give a brief overview of the human Trx and Grx systems. The main part focuses on our current knowledge about mitochondrial Grx2, which facilitates mitochondrial redox homoeostasis during oxidative stress-induced apoptosis.
Chris M Grant - One of the best experts on this subject based on the ideXlab platform.
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non reciprocal regulation of the redox state of the glutathione Glutaredoxin and thioredoxin systems
EMBO Reports, 2003Co-Authors: Eleanor W Trotter, Chris M GrantAbstract:Our studies in yeast show that there is an essential requirement for either an active thioredoxin or an active glutathione (GSH)–Glutaredoxin system for cell viability. Glutathione reductase (Glr1) and thioredoxin reductase (Trr1) are key regulatory enzymes that determine the redox state of the GSH–Glutaredoxin and thioredoxin systems, respectively. Here we show that Trr1 is required during normal cell growth, whereas there is no apparent requirement for Glr1. Analysis of the redox state of thioredoxins and Glutaredoxins in glr1 and trr1 mutants reveals that thioredoxins are maintained independently of the glutathione system. In contrast, there is a strong correlation between the redox state of Glutaredoxins and the oxidation state of the GSSG/2GSH redox couple. We suggest that independent redox regulation of thioredoxins enables cells to survive in conditions under which the GSH–Glutaredoxin system is oxidized.
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regulation of protein s thiolation by Glutaredoxin 5 in the yeast saccharomyces cerevisiae
Journal of Biological Chemistry, 2002Co-Authors: Daniel Shenton, Gabriel G Perrone, Kathryn A Quinn, Ian W Dawes, Chris M GrantAbstract:The irreversible oxidation of cysteine residues can be prevented by protein S-thiolation, a process by which protein -SH groups form mixed disulfides with low molecular weight thiols such as glutathione. We report here that this protein modification is not a simple response to the cellular redox state, since different oxidants lead to different patterns of protein S-thiolation. SDS-polyacrylamide gel electrophoresis shows that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is the major target for modification following treatment with hydroperoxides (hydrogen peroxide or tert-butylhydroperoxide), whereas this enzyme is unaffected following cellular exposure to the thiol oxidant diamide. Further evidence that protein S-thiolation is tightly regulated in response to oxidative stress is provided by the finding that the Tdh3 GAPDH isoenzyme, and not the Tdh2 isoenzyme, is S-thiolated following exposure to H(2)O(2) in vivo, whereas both GAPDH isoenzymes are S-thiolated when H(2)O(2) is added to cell-free extracts. This indicates that cellular factors are likely to be responsible for the difference in GAPDH S-thiolation observed in vivo rather than intrinsic structural differences between the GAPDH isoenzymes. To begin to search for factors that can regulate the S-thiolation process, we investigated the role of the Glutaredoxin family of oxidoreductases. We provide the first evidence that protein dethiolation in vivo is regulated by a monothiol-Glutaredoxin rather than the classical Glutaredoxins, which contain two active site cysteine residues. In particular, Glutaredoxin 5 is required for efficient dethiolation of the Tdh3 GAPDH isoenzyme.
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role of the glutathione Glutaredoxin and thioredoxin systems in yeast growth and response to stress conditions
Molecular Microbiology, 2001Co-Authors: Chris M GrantAbstract:Sulphydryl groups (-SH) play a remarkably broad range of roles in the cell, and the redox status of cysteine residues can affect both the structure and the function of numerous enzymes, receptors and transcription factors. The intracellular milieu is usually a reducing environment as a result of high concentrations of the low-molecular-weight thiol glutathione (GSH). However, reactive oxygen species (ROS), which are the products of normal aerobic metabolism, as well as naturally occurring free radical-generating compounds, can alter this redox balance. A number of cellular factors have been implicated in the regulation of redox homeostasis, including the glutathione/Glutaredoxin and thioredoxin systems. Glutaredoxins and thioredoxins are ubiquitous small heat-stable oxidoreductases that have proposed functions in many cellular processes, including deoxyribonucleotide synthesis, repair of oxidatively damaged proteins, protein folding and sulphur metabolism. This review describes recent findings in the lower eukaryote Saccharomyces cerevisiae that are leading to a better understanding of their role in redox homeostasis in eukaryotic cell metabolism.
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the yeast saccharomyces cerevisiae contains two Glutaredoxin genes that are required for protection against reactive oxygen species
Molecular Biology of the Cell, 1998Co-Authors: Sandra Luikenhuis, Ian W Dawes, Gabriel G Perrone, Chris M GrantAbstract:Glutaredoxins are small heat-stable proteins that act as glutathione-dependent disulfide oxidoreductases. Two genes, designatedGRX1 and GRX2, which share 40–52% identity and 61–76% similarity with ...
Lars Brautigam - One of the best experts on this subject based on the ideXlab platform.
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Glutaredoxin regulates vascular development by reversible glutathionylation of sirtuin 1
Proceedings of the National Academy of Sciences of the United States of America, 2013Co-Authors: Lars Brautigam, Timour Prozorovski, Lasse Jensen, Gereon Poschmann, Staffan Nystrom, Sarah Bannenberg, Kristian Dreij, Klaudia Lepka, Sergio J Montano, Orhan AktasAbstract:Embryonic development depends on complex and precisely orchestrated signaling pathways including specific reduction/oxidation cascades. Oxidoreductases of the thioredoxin family are key players conveying redox signals through reversible posttranslational modifications of protein thiols. The importance of this protein family during embryogenesis has recently been exemplified for Glutaredoxin 2, a vertebrate-specific glutathione-disulfide oxidoreductase with a critical role for embryonic brain development. Here, we discovered an essential function of Glutaredoxin 2 during vascular development. Confocal microscopy and time-lapse studies based on two-photon microscopy revealed that morpholino-based knockdown of Glutaredoxin 2 in zebrafish, a model organism to study vertebrate embryogenesis, resulted in a delayed and disordered blood vessel network. We were able to show that formation of a functional vascular system requires Glutaredoxin 2-dependent reversible S-glutathionylation of the NAD(+)-dependent protein deacetylase sirtuin 1. Using mass spectrometry, we identified a cysteine residue in the conserved catalytic region of sirtuin 1 as target for Glutaredoxin 2-specific deglutathionylation. Thereby, Glutaredoxin 2-mediated redox regulation controls enzymatic activity of sirtuin 1, a mechanism we found to be conserved between zebrafish and humans. These results link S-glutathionylation to vertebrate development and successful embryonic angiogenesis.
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an unusual mode of iron sulfur cluster coordination in a teleost Glutaredoxin
Biochemical and Biophysical Research Communications, 2013Co-Authors: Lars Brautigam, Arne Holmgren, C. Johansson, Carsten Berndt, Bastian Kubsch, Michael A Mcdonough, Eckhard BillAbstract:Abstract Glutaredoxins that contain a Cys-X-X-Cys active site motif are glutathione-dependent thiol-disulfide oxidoreductases. Vertebrate Glutaredoxin 2 is characterized by two extra cysteines that form an intra-molecular disulfide bridge. Zebrafish Glutaredoxin 2 contains four additional cysteines that are conserved within the infraclass of bony fish (teleosts). Here, we present a biochemical and biophysical characterization of zebrafish Glutaredoxin 2, focusing on iron–sulfur-cluster coordination. The coordination of [2Fe2S]2+-clusters in monomers of this protein was revealed by both absorption and Mossbauer spectroscopy as well as size exclusion chromatography. All other holo-Glutaredoxins represent [FeS]-cluster bridged dimers using two molecules of non-covalently bound glutathione and the N-terminal active site cysteines as ligands. These cysteine residues were not required for [FeS]-cluster coordination in zebrafish Glutaredoxin 2. A crystal structure of the teleost protein revealed high structural similarity to its human homologue. The two vertebrate-specific cysteines as well as two of the teleost-specific cysteines are positioned within a radius of 7 A near the C-terminus suggesting a potential role in [FeS]-cluster coordination. Indeed, mutated proteins lacking these teleost-specific cysteines lost the ability to bind the cofactor. Hence, the apparent mode of [FeS]-cluster coordination in zebrafish Glutaredoxin 2 could be different from all yet described [FeS]-Glutaredoxins.
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crucial function of vertebrate Glutaredoxin 3 picot in iron homeostasis and hemoglobin maturation
Molecular Biology of the Cell, 2013Co-Authors: Petra Haunhorst, Lars Brautigam, Ulrich Muhlenhoff, Roland Lill, Bastian Hoffmann, Oliver Stehling, Evamaria Hanschmann, Carsten BerndtAbstract:The mechanisms by which eukaryotic cells handle and distribute the essential micronutrient iron within the cytosol and other cellular compartments are only beginning to emerge. The yeast monothiol multidomain Glutaredoxins (Grx) 3 and 4 are essential for both transcriptional iron regulation and intracellular iron distribution. Despite the fact that the mechanisms of iron metabolism differ drastically in fungi and higher eukaryotes, the Glutaredoxins are conserved, yet their precise function in vertebrates has remained elusive. Here we demonstrate a crucial role of the vertebrate-specific monothiol multidomain Grx3 (PICOT) in cellular iron homeostasis. During zebrafish embryonic development, depletion of Grx3 severely impairs the maturation of hemoglobin, the major iron-consuming process. Silencing of human Grx3 expression in HeLa cells decreases the activities of several cytosolic Fe/S proteins, for example, iron-regulatory protein 1, a major component of posttranscriptional iron regulation. As a consequence, Grx3-depleted cells show decreased levels of ferritin and increased levels of transferrin receptor, features characteristic of cellular iron starvation. Apparently, Grx3-deficient cells are unable to efficiently use iron, despite unimpaired cellular iron uptake. These data suggest an evolutionarily conserved role of cytosolic monothiol multidomain Glutaredoxins in cellular iron metabolism pathways, including the biogenesis of Fe/S proteins and hemoglobin maturation.
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vertebrate specific Glutaredoxin is essential for brain development
Proceedings of the National Academy of Sciences of the United States of America, 2011Co-Authors: Lars Brautigam, Christopher Horst Lillig, Arne Holmgren, Jose R Godoy, Lena Dorothee Schutte, Timour Prozorovski, Manuela Gellert, Giselbert Hauptmann, Carsten BerndtAbstract:Cellular functions and survival are dependent on a tightly controlled redox potential. Currently, an increasing amount of data supports the concept of local changes in the redox environment and specific redox signaling events controlling cell function. Specific protein thiol groups are the major targets of redox signaling and regulation. Thioredoxins and Glutaredoxins catalyze reversible thiol-disulfide exchange reactions and are primary regulators of the protein thiol redox state. Here, we demonstrate that embryonic brain development depends on the enzymatic activity of Glutaredoxin 2. Zebrafish with silenced expression of Glutaredoxin 2 lost virtually all types of neurons by apoptotic cell death and the ability to develop an axonal scaffold. As demonstrated in zebrafish and in a human cellular model for neuronal differentiation, Glutaredoxin 2 controls axonal outgrowth via thiol redox regulation of collapsin response mediator protein 2, a central component of the semaphorin pathway. This study provides an example of a specific thiol redox regulation essential for vertebrate embryonic development.