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Arne Holmgren - One of the best experts on this subject based on the ideXlab platform.
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Thioredoxins and glutaredoxins as facilitators of protein Folding.
Biochimica et biophysica acta, 2008Co-Authors: Carsten Berndt, Christopher Horst Lillig, Arne HolmgrenAbstract:Thiol-disulfide oxidoreductase systems of bacterial cytoplasm and eukaryotic cytosol favor reducing conditions and protein thiol groups, while bacterial periplasm and eukaryotic endoplasmatic reticulum provide oxidizing conditions and a machinery for disulfide bond formation in the secretory pathway. Oxidoreductases of the Thioredoxin Fold superfamily catalyze steps in oxidative protein Folding via protein-protein interactions and covalent catalysis to act as chaperones and isomerases of disulfides to generate a native Fold. The active site dithiol/disulfide of Thioredoxin Fold proteins is CXXC where variations of the residues inside the disulfide ring are known to increase the redox potential like in protein disulfide isomerases. In the catalytic mechanism Thioredoxin Fold proteins bind to target proteins through conserved backbone-backbone hydrogen bonds and induce conformational changes of the target disulfide followed by nucleophilic attack by the N-terminally located low pK(a) Cys residue. This generates a mixed disulfide covalent bond which subsequently is resolved by attack from the C-terminally located Cys residue. This review will focus on two members of the Thioredoxin superfamily of proteins known to be crucial for maintaining a reduced intracellular redox state, Thioredoxin and glutaredoxin, and their potential functions as facilitators and regulators of protein Folding and chaperone activity.
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characterization of human glutaredoxin 2 as iron sulfur protein a possible role as redox sensor
Proceedings of the National Academy of Sciences of the United States of America, 2005Co-Authors: Christopher Horst Lillig, Carsten Berndt, Maria Lönn, Olivia Vergnolle, Christoph Hudemann, Eckhard Bill, Arne HolmgrenAbstract:Human mitochondrial glutaredoxin 2 (Grx2) is a glutathione-dependent oxidoreductase (active site: Cys-Ser-Tyr-Cys) that facilitates the maintenance of mitochondrial redox homeostasis upon induction of apoptosis by oxidative stress. Here, we have characterized Grx2 as an iron–sulfur center-containing member of the Thioredoxin Fold protein family. Mossbauer spectroscopy revealed the presence of a four cysteine-coordinated nonoxidizable [2Fe-2S]2+ cluster that bridges two Grx2 molecules via two structural Cys residues to form dimeric holo Grx2. Coimmunoprecipitation of radiolabeled iron with Grx2 from human cell lines indicated the presence of the cluster in vivo. The [2Fe-2S]-bridged dimer was enzymatically inactive, but degradation of the cluster and the resulting monomerization of Grx2 activated the protein. Slow degradation under aerobic conditions was prevented by the presence of glutathione, whereas glutathione disulfide as well as one-electron oxidants or reductants promoted monomerization of Grx2. We propose that the iron–sulfur cluster serves as a redox sensor for the activation of Grx2 during conditions of oxidative stress when free radicals are formed and the glutathione pool becomes oxidized.
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Short interfering RNA-mediated silencing of glutaredoxin 2 increases the sensitivity of HeLa cells toward doxorubicin and phenylarsine oxide.
Proceedings of the National Academy of Sciences of the United States of America, 2004Co-Authors: Christopher Horst Lillig, Maria Lönn, Mari Enoksson, Aristi P. Fernandes, Arne HolmgrenAbstract:Glutaredoxin (Grx) belongs to the Thioredoxin Fold superfamily and catalyzes glutathione-dependent oxidoreductions. The recently discovered mitochondrial and nuclear Grx (Grx2) differs from the more abundant cytosolic Grx (Grx1) by its higher affinity toward S-glutathionylated proteins and by being a substrate for Thioredoxin reductase. Here, we have successfully established a method to silence the expression of Grx2 in HeLa cells by using short interfering RNA to study its role in the cell. Cells with levels of Grx2
Antonio C. Bianco - One of the best experts on this subject based on the ideXlab platform.
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the deiodinase trio and thyroid hormone signaling
Methods of Molecular Biology, 2018Co-Authors: Antonio C. Bianco, Rodrigo R Da ConceicaoAbstract:Thyroid hormone signaling is customized in a time and cell-specific manner by the deiodinases, homodimeric Thioredoxin Fold containing selenoproteins. This ensures adequate T3 action in developing tissues, healthy adults and many disease states. D2 activates thyroid hormone by converting the pro-hormone T4 to T3, the biologically active thyroid hormone. D2 expression is tightly regulated by transcriptional mechanisms triggered by endogenous as well as environmental cues. There is also an on/off switch mechanism that controls D2 activity that is triggered by catalysis and functions via D2 ubiquitination/deubiquitination. D3 terminates thyroid hormone action by inactivation of both T4 and T3 molecules. Deiodinases play a role in thyroid hormone homeostasis, development, growth and metabolic control by affecting the intracellular levels of T3 and thus gene expression on a cell-specific basis. In many cases, tight control of these pathways by T3 is achieved with coordinated reciprocal changes in D2-mediated thyroid hormone activation D3-mediated thyroid hormone inactivation.
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The type II deiodinase is retrotranslocated to the cytoplasm and proteasomes via p97/Atx3 complex.
Molecular endocrinology (Baltimore Md.), 2013Co-Authors: Rafael Arrojo E Drigo, Balazs Gereben, Péter Egri, Antonio C. BiancoAbstract:The type II iodothyronine deiodinase (D2) is a type I endoplasmic reticulum (ER)-resident Thioredoxin Fold-containing selenoprotein that activates thyroid hormone. D2 is inactivated by ER-associated ubiquitination and can be reactivated by two ubiquitin-specific peptidase-class D2-interacting deubiquitinases (DUBs). Here, we used D2-expressing cell models to define that D2 ubiquitination (UbD2) occurs via K48-linked ubiquitin chains and that exposure to its natural substrate, T4, accelerates UbD2 formation and retrotranslocation to the cytoplasm via interaction with the p97-ATPase complex. D2 retrotranslocation also includes deubiquitination by the p97-associated DUB Ataxin-3 (Atx3). Inhibiting Atx3 with eeyarestatin-I did not affect D2:p97 binding but decreased UbD2 retrotranslocation and caused ER accumulation of high-molecular weight UbD2 bands possibly by interfering with the D2-ubiquitin-specific peptidases binding. Once in the cytosol, D2 is delivered to the proteasomes as evidenced by coprecipitati...
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Activation and inactivation of thyroid hormone by deiodinases: local action with general consequences.
Cellular and Molecular Life Sciences, 2007Co-Authors: Balazs Gereben, Aniko Zeold, Monica Dentice, Domenico Salvatore, Antonio C. BiancoAbstract:The thyroid hormone plays a fundamental role in the development, growth, and metabolic homeostasis in all vertebrates by affecting the expression of different sets of genes. A group of Thioredoxin Fold-containing selenoproteins known as deiodinases control thyroid hormone action by activating or inactivating the precursor molecule thyroxine that is secreted by the thyroid gland. These pathways ensure regulation of the availability of the biologically active molecule T3, which occurs in a time-and tissue-specific fashion. In addition, because cells and plasma are in equilibrium and deiodination affects central thyroid hormone regulation, these local deiodinase-mediated events can also affect systemic thyroid hormone economy, such as in the case of non-thyroidal illness. Heightened interest in the field has been generated following the discovery that the deiodinases can be a component in both the Sonic hedgehog signaling pathway and the TGR-5 signaling cascade, a G-protein-coupled receptor for bile acids. These new mechanisms involved in deiodinase regulation indicate that local thyroid hormone activation and inactivation play a much broader role than previously thought.
Balazs Gereben - One of the best experts on this subject based on the ideXlab platform.
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The type II deiodinase is retrotranslocated to the cytoplasm and proteasomes via p97/Atx3 complex.
Molecular endocrinology (Baltimore Md.), 2013Co-Authors: Rafael Arrojo E Drigo, Balazs Gereben, Péter Egri, Antonio C. BiancoAbstract:The type II iodothyronine deiodinase (D2) is a type I endoplasmic reticulum (ER)-resident Thioredoxin Fold-containing selenoprotein that activates thyroid hormone. D2 is inactivated by ER-associated ubiquitination and can be reactivated by two ubiquitin-specific peptidase-class D2-interacting deubiquitinases (DUBs). Here, we used D2-expressing cell models to define that D2 ubiquitination (UbD2) occurs via K48-linked ubiquitin chains and that exposure to its natural substrate, T4, accelerates UbD2 formation and retrotranslocation to the cytoplasm via interaction with the p97-ATPase complex. D2 retrotranslocation also includes deubiquitination by the p97-associated DUB Ataxin-3 (Atx3). Inhibiting Atx3 with eeyarestatin-I did not affect D2:p97 binding but decreased UbD2 retrotranslocation and caused ER accumulation of high-molecular weight UbD2 bands possibly by interfering with the D2-ubiquitin-specific peptidases binding. Once in the cytosol, D2 is delivered to the proteasomes as evidenced by coprecipitati...
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Activation and inactivation of thyroid hormone by deiodinases: local action with general consequences.
Cellular and Molecular Life Sciences, 2007Co-Authors: Balazs Gereben, Aniko Zeold, Monica Dentice, Domenico Salvatore, Antonio C. BiancoAbstract:The thyroid hormone plays a fundamental role in the development, growth, and metabolic homeostasis in all vertebrates by affecting the expression of different sets of genes. A group of Thioredoxin Fold-containing selenoproteins known as deiodinases control thyroid hormone action by activating or inactivating the precursor molecule thyroxine that is secreted by the thyroid gland. These pathways ensure regulation of the availability of the biologically active molecule T3, which occurs in a time-and tissue-specific fashion. In addition, because cells and plasma are in equilibrium and deiodination affects central thyroid hormone regulation, these local deiodinase-mediated events can also affect systemic thyroid hormone economy, such as in the case of non-thyroidal illness. Heightened interest in the field has been generated following the discovery that the deiodinases can be a component in both the Sonic hedgehog signaling pathway and the TGR-5 signaling cascade, a G-protein-coupled receptor for bile acids. These new mechanisms involved in deiodinase regulation indicate that local thyroid hormone activation and inactivation play a much broader role than previously thought.
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the iodothyronine selenodeiodinases are Thioredoxin Fold family proteins containing a glycoside hydrolase clan gh a like structure
Journal of Biological Chemistry, 2003Co-Authors: Isabelle Callebaut, Balazs Gereben, Cyntia Curciomorelli, Jeanpaul Mornon, Christoph Buettner, Stephen A Huang, Bertrand Castro, Tatiana L Fonseca, John W Harney, Reed P LarsenAbstract:The three iodothyronine selenodeiodinases catalyze the initiation and termination of thyroid hormone effects in vertebrates. Structural analyses of these proteins have been hindered by their integral membrane nature and the inefficient eukaryotic-specific pathway for selenoprotein synthesis. Hydrophobic cluster analysis used in combination with Position-specific Iterated BLAST reveals that their extramembrane portion belongs to the Thioredoxin-Fold superfamily for which experimental structure information exists. Moreover, a large deiodinase region imbedded in the Thioredoxin Fold shares strong similarities with the active site of iduronidase, a member of the clan GH-A-Fold of glycoside hydrolases. This model can explain a number of results from previous mutagenesis analyses and permits new verifiable insights into the structural and functional properties of these enzymes.
Christopher Horst Lillig - One of the best experts on this subject based on the ideXlab platform.
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Molecular architecture of Streptococcus pneumoniae surface Thioredoxin-Fold lipoproteins crucial for extracellular oxidative stress resistance and maintenance of virulence.
EMBO molecular medicine, 2013Co-Authors: Malek Saleh, Christopher Horst Lillig, Sergio G. Bartual, Mohammed R. Abdullah, Inga Jensch, Tauseef M. Asmat, Lothar Petruschka, Thomas Pribyl, Manuela Gellert, Haike AntelmannAbstract:The respiratory pathogen Streptococcus pneumoniae has evolved efficient mechanisms to resist oxidative stress conditions and to displace other bacteria in the nasopharynx. Here we characterize at physiological, functional and structural levels two novel surface-exposed Thioredoxin-family lipoproteins, Etrx1 and Etrx2. The impact of both Etrx proteins and their redox partner methionine sulfoxide reductase SpMsrAB2 on pneumococcal pathogenesis was assessed in mouse virulence studies and phagocytosis assays. The results demonstrate that loss of function of either both Etrx proteins or SpMsrAB2 dramatically attenuated pneumococcal virulence in the acute mouse pneumonia model and that Etrx proteins compensate each other. The deficiency of Etrx proteins or SpMsrAB2 further enhanced bacterial uptake by macrophages, and accelerated pneumococcal killing by H2O2 or free methionine sulfoxides (MetSO). Moreover, the absence of both Etrx redox pathways provokes an accumulation of oxidized SpMsrAB2 in vivo. Taken together our results reveal insights into the role of two extracellular electron pathways required for reduction of SpMsrAB2 and surface-exposed MetSO. Identification of this system and its target proteins paves the way for the design of novel antimicrobials.
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14:Thioredoxins and Glutaredoxins. Functions and Metal Ion Interactions
Metallothioneins and Related Chelators, 2009Co-Authors: Christopher Horst Lillig, Carsten BerndtAbstract:Thioredoxins and glutaredoxins represent the major cellular systems for the reduction of protein disulfides and protein de-glutathionylation, respectively. These two systems are involved in many aspects of human health, for instance as electron donors of metabolic enzymes and by controlling and maintaining the cellular redox state. The members of this protein family are characterized by a common structural motif, the Thioredoxin Fold. This basic architecture consists of a central four-stranded β-sheet surrounded by three α-helices. During the past few years accumulating evidence suggests a close relationship between these redoxins, most of all the glutaredoxins, and the cellular iron pool. Today we know that the Thioredoxin Fold cannot only be utilized for specific protein-protein interactions but also for interactions with metals, for instance iron-sulfur centers. Within this chapter, we summarize these recent findings and discuss the potential physiological implications of these metal interactions.
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Thioredoxins and glutaredoxins as facilitators of protein Folding.
Biochimica et biophysica acta, 2008Co-Authors: Carsten Berndt, Christopher Horst Lillig, Arne HolmgrenAbstract:Thiol-disulfide oxidoreductase systems of bacterial cytoplasm and eukaryotic cytosol favor reducing conditions and protein thiol groups, while bacterial periplasm and eukaryotic endoplasmatic reticulum provide oxidizing conditions and a machinery for disulfide bond formation in the secretory pathway. Oxidoreductases of the Thioredoxin Fold superfamily catalyze steps in oxidative protein Folding via protein-protein interactions and covalent catalysis to act as chaperones and isomerases of disulfides to generate a native Fold. The active site dithiol/disulfide of Thioredoxin Fold proteins is CXXC where variations of the residues inside the disulfide ring are known to increase the redox potential like in protein disulfide isomerases. In the catalytic mechanism Thioredoxin Fold proteins bind to target proteins through conserved backbone-backbone hydrogen bonds and induce conformational changes of the target disulfide followed by nucleophilic attack by the N-terminally located low pK(a) Cys residue. This generates a mixed disulfide covalent bond which subsequently is resolved by attack from the C-terminally located Cys residue. This review will focus on two members of the Thioredoxin superfamily of proteins known to be crucial for maintaining a reduced intracellular redox state, Thioredoxin and glutaredoxin, and their potential functions as facilitators and regulators of protein Folding and chaperone activity.
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characterization of human glutaredoxin 2 as iron sulfur protein a possible role as redox sensor
Proceedings of the National Academy of Sciences of the United States of America, 2005Co-Authors: Christopher Horst Lillig, Carsten Berndt, Maria Lönn, Olivia Vergnolle, Christoph Hudemann, Eckhard Bill, Arne HolmgrenAbstract:Human mitochondrial glutaredoxin 2 (Grx2) is a glutathione-dependent oxidoreductase (active site: Cys-Ser-Tyr-Cys) that facilitates the maintenance of mitochondrial redox homeostasis upon induction of apoptosis by oxidative stress. Here, we have characterized Grx2 as an iron–sulfur center-containing member of the Thioredoxin Fold protein family. Mossbauer spectroscopy revealed the presence of a four cysteine-coordinated nonoxidizable [2Fe-2S]2+ cluster that bridges two Grx2 molecules via two structural Cys residues to form dimeric holo Grx2. Coimmunoprecipitation of radiolabeled iron with Grx2 from human cell lines indicated the presence of the cluster in vivo. The [2Fe-2S]-bridged dimer was enzymatically inactive, but degradation of the cluster and the resulting monomerization of Grx2 activated the protein. Slow degradation under aerobic conditions was prevented by the presence of glutathione, whereas glutathione disulfide as well as one-electron oxidants or reductants promoted monomerization of Grx2. We propose that the iron–sulfur cluster serves as a redox sensor for the activation of Grx2 during conditions of oxidative stress when free radicals are formed and the glutathione pool becomes oxidized.
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Short interfering RNA-mediated silencing of glutaredoxin 2 increases the sensitivity of HeLa cells toward doxorubicin and phenylarsine oxide.
Proceedings of the National Academy of Sciences of the United States of America, 2004Co-Authors: Christopher Horst Lillig, Maria Lönn, Mari Enoksson, Aristi P. Fernandes, Arne HolmgrenAbstract:Glutaredoxin (Grx) belongs to the Thioredoxin Fold superfamily and catalyzes glutathione-dependent oxidoreductions. The recently discovered mitochondrial and nuclear Grx (Grx2) differs from the more abundant cytosolic Grx (Grx1) by its higher affinity toward S-glutathionylated proteins and by being a substrate for Thioredoxin reductase. Here, we have successfully established a method to silence the expression of Grx2 in HeLa cells by using short interfering RNA to study its role in the cell. Cells with levels of Grx2
Javier Santos - One of the best experts on this subject based on the ideXlab platform.
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recognition between a short unstructured peptide and a partially Folded fragment leads to the Thioredoxin Fold sharing native like dynamics
Proteins, 2012Co-Authors: Andres Binolfi, Javier Santos, José M. Delfino, Claudio O. Fernández, Mauricio P. SicaAbstract:Thioredoxins (TRXs) constitute attractive α/β scafFolds for investigating molecular recognition. The interaction between the recombinant fragment spanning the sequence 1–93 of full-length TRX (TRX1-93) and the synthetic peptide comprising residues 94–108 (TRX94-108), plus a C-terminal tyrosine tag (the numbering scheme used in entry pdb 2TRX is used throughout the article, two complementary moieties of E. coli TRX, brings about the consolidation of a native-like complex. Despite its reduced thermodynamic stability, this complex is able to acquire fine structural features remarkably similar to those characteristic of full-length TRX, namely, hydrodynamic behavior, assessed by diffusion-ordered spectroscopy (DOSY)-NMR; the pattern of secondary structure, as revealed by three-bond HNHα coupling constants and secondary shifts for Hα/CO/Cα/Cβ; native-like tertiary structural signatures revealed by near-UV circular dichroism (CD) spectroscopy. The complex exhibits a relaxation behavior compatible with that expected for a native-like structure. However, heteronuclear nuclear Overhauser effect (NOE)s reveal an enhanced dynamics for the complex by comparison with full-length TRX. Furthermore, higher R2 values for residues 43–50 and 74–89 would likely result from an exchange process modulated by the peptide at the interface region. The slow kinetics of the consolidation reaction was followed by CD and real-time NMR. Equilibrium titration experiments by NMR yield a KD value of 1.4 ± 1.0 μM and a second low-affinity (>150 μM) binding event in the vicinity of the active site. Molecular dynamics simulations of both the isolated fragment TRX1-93 and the complex suggest the destabilization of α2 and α3 helical elements and the persistence of β-structure in the absence of TRX94-108. Altogether, structural and dynamic evidence presented herein points to the key role played by the C-terminal helix in establishing the overall Fold. This critical switch module endows reduced TRX with the ability to act as a cooperative Folding unit. Proteins 2012;. © 2012 Wiley Periodicals, Inc.
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Recognition between a short unstructured peptide and a partially Folded fragment leads to the Thioredoxin Fold sharing native‐like dynamics
Proteins, 2012Co-Authors: Andres Binolfi, José M. Delfino, Claudio O. Fernández, Mauricio P. Sica, Javier SantosAbstract:Thioredoxins (TRXs) constitute attractive α/β scafFolds for investigating molecular recognition. The interaction between the recombinant fragment spanning the sequence 1–93 of full-length TRX (TRX1-93) and the synthetic peptide comprising residues 94–108 (TRX94-108), plus a C-terminal tyrosine tag (the numbering scheme used in entry pdb 2TRX is used throughout the article, two complementary moieties of E. coli TRX, brings about the consolidation of a native-like complex. Despite its reduced thermodynamic stability, this complex is able to acquire fine structural features remarkably similar to those characteristic of full-length TRX, namely, hydrodynamic behavior, assessed by diffusion-ordered spectroscopy (DOSY)-NMR; the pattern of secondary structure, as revealed by three-bond HNHα coupling constants and secondary shifts for Hα/CO/Cα/Cβ; native-like tertiary structural signatures revealed by near-UV circular dichroism (CD) spectroscopy. The complex exhibits a relaxation behavior compatible with that expected for a native-like structure. However, heteronuclear nuclear Overhauser effect (NOE)s reveal an enhanced dynamics for the complex by comparison with full-length TRX. Furthermore, higher R2 values for residues 43–50 and 74–89 would likely result from an exchange process modulated by the peptide at the interface region. The slow kinetics of the consolidation reaction was followed by CD and real-time NMR. Equilibrium titration experiments by NMR yield a KD value of 1.4 ± 1.0 μM and a second low-affinity (>150 μM) binding event in the vicinity of the active site. Molecular dynamics simulations of both the isolated fragment TRX1-93 and the complex suggest the destabilization of α2 and α3 helical elements and the persistence of β-structure in the absence of TRX94-108. Altogether, structural and dynamic evidence presented herein points to the key role played by the C-terminal helix in establishing the overall Fold. This critical switch module endows reduced TRX with the ability to act as a cooperative Folding unit. Proteins 2012;. © 2012 Wiley Periodicals, Inc.
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Consolidation of the Thioredoxin Fold by peptide recognition: interaction between E. coli Thioredoxin fragments 1-93 and 94-108.
Biochemistry, 2007Co-Authors: Javier Santos, Cristina Marino-buslje, Claudia Kleinman, Mario R. Ermácora, José M. DelfinoAbstract:Escherichia coli Thioredoxin (TRX) catalyzes redox reactions via the reversible oxidation of the conserved active center WCGPC. TRX is a monomeric α/β protein with a Fold characterized by a central...