Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
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Schistosoma mansoni, a trematode parasite, which causes schistosomiasis and affects more than 200 million people worldwide, lives in an aerobic environment and therefore needs an effective redox mechanism for surviving reactive oxygen species from its host. Although, the host has two different redox systems: glutaredoxin and thioredoxin, the parasite has only one unique multifunctional enzyme, thioredoxin glutathione reductase (TGR) involving a fusion of two proteins, glutaredoxin (Grx) and thioredoxin reductase (TR), for performing all the redox activities. This dependence of S. mansoni on a single protein, TGR, for its protection from oxidative stress, makes it a promising drug target. Here, we describe a suitably validated, homology model for S. mansoni TGR (SmTGR), developed using both TR and Grx templates, functionally complete in the dimeric form with cofactors NADP(H) and FAD. Comparative analysis of substrate and inhibitor binding pockets of our model with crystal structures of parent TR as well as with that of glutathione reductase (GR), which is an essential component of the Grx system, appears to provide greater insight into the functioning of TGR. This also augments recent observations reported on the basis of X-ray structure data on SmTGR monomer lacking the C-terminal selenocysteine tail.
J Mol Graph Model 2009 Feb
PMID:Comparative modeling of thioredoxin glutathione reductase from Schistosoma mansoni: a multifunctional target for antischistosomal therapy. 1907 May 22

Peroxiredoxins are ubiquitous enzymes which protect cells against oxidative stress. The first step of catalysis is common to all peroxiredoxins and results in oxidation of a conserved peroxidatic cysteine residue to sulfenic acid. This forms an intermolecular disulfide bridge in the case of 2-Cys peroxiredoxins, which is a substrate for the thioredoxin system. 1-Cys Prx's contain a peroxidatic cysteine but do not contain a second conserved cysteine residue, and hence the identity of the in vivo reduction system has been unclear. Here, we show that the yeast mitochondrial 1-Cys Prx1 is reactivated by glutathionylation of the catalytic cysteine residue and subsequent reduction by thioredoxin reductase (Trr2) coupled with glutathione (GSH). This novel mechanism does not require the usual thioredoxin (Trx3) redox partner of Trr2 for antioxidant activity, although in vitro assays show that the Trr2/Trx3 and Trr2/GSH systems exhibit similar capacities for supporting Prx1 catalysis. Our data also indicate that mitochondria are a main target of cadmium-induced oxidative stress and that Prx1 is particularly required to protect against mitochondrial oxidation. This study demonstrates a physiological reaction mechanism for 1-Cys peroxiredoxins and reveals a new role in protection against mitochondrial heavy metal toxicity.
Mol Cell Biol 2009 Jun
PMID:Antioxidant activity of the yeast mitochondrial one-Cys peroxiredoxin is dependent on thioredoxin reductase and glutathione in vivo. 1933 53

The indolequinone ES936 {5-methoxy-1,2-dimethyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione} was previously developed in our lab as an antitumor agent against pancreatic cancer. The objective of this study was to identify indolequinones with improved potency against pancreatic cancer and to define their mechanisms of action. Pancreatic cancer cell lines PANC-1, MIA PaCa-2, and BxPC-3 were used in in vitro assays [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) and clonogenic assays]; indolequinones displayed potent cytotoxicity against all three cell lines, and two specific classes of indolequinone were particularly potent agents. These indolequinones induced caspase-dependent apoptosis but no redox cycling or oxidative stress in MIA PaCa-2 and BxPC-3 cells. Selected indolequinones were also screened against the NCI-60 cell line panel and were found to be particularly effective against colon, renal, and melanoma cancer cells. A potential target of these indolequinones was identified as thioredoxin reductase. Indolequinones were found to be potent inhibitors of thioredoxin reductase activity both in pancreatic cancer cells and in cell-free systems. The mechanism of action of the indolequinones was shown to involve metabolic reduction, loss of a leaving group to generate a reactive electrophile resulting in alkylation of the selenocysteine residue in the active site of thioredoxin reductase. In vivo efficacy of the indolequinones was also tested in the MIA PaCa-2 pancreatic tumor xenograft in nude mice, and lead indolequinones demonstrated high efficacy and low toxicity. Inhibition of thioredoxin reductase represents a potential novel target in pancreatic cancer and may provide a biomarker of effect of lead indolequinones in this type of cancer.
Mol Pharmacol 2009 Jul
PMID:Potent activity of indolequinones against human pancreatic cancer: identification of thioredoxin reductase as a potential target. 1936 12

Thioredoxin and thioredoxin reductase from the psychrophilic eubacterium Pseudoalteromonas haloplanktis were obtained as recombinant His-tagged proteins (rPhTrx and rPhTrxR, respectively). rPhTrxR is organised as a homodimeric flavoenzyme, whereas rPhTrx is a small monomeric protein, both containing a functional disulfide bridge. However, three additional cysteines are present as free thiols in purified rPhTrxR. When individually tested in specific assays, rPhTrxR and rPhTrx display a full activity at low temperatures, an indispensable requirement for cold-adapted proteins. In particular, rPhTrxR catalyses the NADPH dependent reduction of DTNB and rPhTrx provokes the insulin precipitation in the presence of DTT. The analysis of the effect of temperature on these reactions indicates that rPhTrxR is more cold-adapted than rPhTrx, having a higher psychrophilicity. The combined activity of rPhTrxR and rPhTrx, tested in a reconstituted assay containing NADPH as electrons donor and human insulin as the thioredoxin substrate, demonstrates a direct functional interaction between the purified recombinant components of the thioredoxin system of P. haloplanktis. Furthermore, the NADPH-dependent reduction of rPhTrx catalysed by rPhTrxR is fully reversible and allows the determination of its redox potential, whose value is in the range of other bacterial and archaeal thioredoxins. The analysis of the thermostability of rPhTrxR points to its discrete heat resistance. However, rPhTrx is much more heat resistant, with a half inactivation time of about 4 h at 95 degrees C. This exceptional heat resistance for a psychrophilic protein is significantly decreased by the reduction of the disulfide bridge of rPhTrx. Functionality, thermodependence and thermostability of the P. haloplanktis thioredoxin system point to the relevance of this key mechanism for the preservation of the reduced state of cytoplasmic proteins even in a cold-adapted source.
Mol Biosyst 2009 May
PMID:Differential cold-adaptation among protein components of the thioredoxin system in the psychrophilic eubacterium Pseudoalteromonas haloplanktis TAC 125. 1938 66

Infections with the microaerophilic parasite Trichomonas vaginalis are treated with the 5-nitroimidazole drug metronidazole, which is also in use against Entamoeba histolytica, Giardia intestinalis and microaerophilic/anaerobic bacteria. Here we report that in T. vaginalis the flavin enzyme thioredoxin reductase displays nitroreductase activity with nitroimidazoles, including metronidazole, and with the nitrofuran drug furazolidone. Reactive metabolites of metronidazole and other nitroimidazoles form covalent adducts with several proteins that are known or assumed to be associated with thioredoxin-mediated redox regulation, including thioredoxin reductase itself, ribonucleotide reductase, thioredoxin peroxidase and cytosolic malate dehydrogenase. Disulphide reducing activity of thioredoxin reductase was greatly diminished in extracts of metronidazole-treated cells and intracellular non-protein thiol levels were sharply decreased. We generated a highly metronidazole-resistant cell line that displayed only minimal thioredoxin reductase activity, not due to diminished expression of the enzyme but due to the lack of its FAD cofactor. Reduction of free flavins, readily observed in metronidazole-susceptible cells, was also absent in the resistant cells. On the other hand, iron-depleted T. vaginalis cells, expressing only minimal amounts of PFOR and hydrogenosomal malate dehydrogenase, remained fully susceptible to metronidazole. Thus, taken together, our data suggest a flavin-based mechanism of metronidazole activation and thereby challenge the current model of hydrogenosomal activation of nitroimidazole drugs.
Mol Microbiol 2009 Apr
PMID:Trichomonas vaginalis: metronidazole and other nitroimidazole drugs are reduced by the flavin enzyme thioredoxin reductase and disrupt the cellular redox system. Implications for nitroimidazole toxicity and resistance. 1941 1

The expression of the genes encoding the ferredoxin-thioredoxin system including the ferredoxin-thioredoxin reductase (FTR) genes ftrC and ftrV and the four different thioredoxin genes trxA (m-type; slr0623), trxB (x-type; slr1139), trxC (sll1057) and trxQ (y-type; slr0233) of the cyanobacterium Synechocystis sp. PCC 6803 has been studied according to changes in the photosynthetic conditions. Experiments of light-dark transition indicate that the expression of all these genes except trxQ decreases in the dark in the absence of glucose in the growth medium. The use of two electron transport inhibitors, 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB), reveals a differential effect on thioredoxin genes expression being trxC and trxQ almost unaffected, whereas trxA, trxB, and the ftr genes are down-regulated. In the presence of glucose, DCMU does not affect gene expression but DBMIB still does. Analysis of the single TrxB or TrxQ and the double TrxB TrxQ Synechocystis mutant strains reveal different functions for each of these thioredoxins under different growth conditions. Finally, a Synechocystis strain was generated containing a mutated version of TrxB (TrxBC34S), which was used to identify the potential in-vivo targets of this thioredoxin by a proteomic analysis.
Mol Plant 2009 Mar
PMID:Photosynthetic regulation of the cyanobacterium Synechocystis sp. PCC 6803 thioredoxin system and functional analysis of TrxB (Trx x) and TrxQ (Trx y) thioredoxins. 1982 13

The stress-associated protein SAP12 belongs to the stress-associated protein (SAP) family with 14 members in Arabidopsis thaliana. SAP12 contains two AN1 zinc fingers and was identified in diagonal 2D redox SDS-PAGE as a protein undergoing major redox-dependent conformational changes. Its transcript was strongly induced under cold and salt stress in a time-dependent manner similar to SAP10, with high levels after 6 h and decreasing levels after 24 and 48 h. The transcript regulation resembled those of the stress marker peroxiredoxin PrxIID at 24 and 48 h. Recombinant SAP12 protein showed redox-dependent changes in quaternary structure as visualized by altered electrophoretic mobility in non-reducing SDS polyacrylamide gel electrophoresis. The oxidized oligomer was reduced by high dithiothreitol concentrations, and also by E. coli thioredoxin TrxA with low dithiothreitol (DTT) concentrations or NADPH plus NADPH-dependent thioredoxin reductase. From Western blots, the SAP12 protein amount was estimated to be in the range of 0.5 ng mug(-1) leaf protein. SAP12 protein decreased under salt and cold stress. These data suggest a redox state-linked function of SAP12 in plant cells particularly under cold and salt stress.
Mol Plant 2009 Mar
PMID:Redox-dependent regulation of the stress-induced zinc-finger protein SAP12 in Arabidopsis thaliana. 1982 20

Thioredoxins (Trx) are ubiquitous proteins that participate in thiol disulfide reactions via two active site cysteine residues, allowing Trx to reduce disulfide bonds in target proteins. Recent progress in proteome analysis has resulted in identification of a wide range of potential target proteins for Trx, indicating that Trx plays a key role in several aspects of cell metabolism. In contrast to other organisms, plants contain multiple forms of Trx that are classified based on their primary structures and sub-cellular localization. The reduction of cytosolic and mitochondrial types of Trx is dependent on NADPH and catalyzed by NADPH-dependent thioredoxin reductase (NTR). In barley, two isoforms each of Trx and NTR have been identified and investigated using proteomics, gene expression, and structural studies. This review outlines the diverse roles suggested for cytosolic/mitochondrial-type Trx systems in cereal seeds and summarizes the current knowledge of the barley system including recent data on function, regulation, interactions, and structure. Directions for future research are discussed.
Mol Plant 2009 May
PMID:From proteomics to structural studies of cytosolic/mitochondrial-type thioredoxin systems in barley seeds. 1982 23

NADPH thioredoxin reductase C (NTRC) is a chloroplast enzyme able to conjugate NADPH thioredoxin reductase (NTR) and thioredoxin (TRX) activities for the efficient reduction of 2-Cys peroxiredoxin (2-Cys PRX). Because NADPH can be produced in chloroplasts during darkness, NTRC plays a key role for plant peroxide detoxification during the night. Here, it is shown that the quaternary structure of NTRC is highly dependent on its redox status. In vitro, most of the enzyme adopted an oligomeric state that disaggregated in dimers upon addition of NADPH, NADH, or DTT. Gel filtration and Western blot analysis of protein extracts from Arabidopsis chloroplast stroma showed that native NTRC forms aggregates, which are sensitive to NADPH and DTT, suggesting that the aggregation state might be a significant aspect of NTRC activity in vivo. Moreover, the enzyme is localized in clusters in Arabidopsis chloroplasts. NTRC triple and double mutants, A164G-V182E-R183F and A164G-R183F, replacing key residues of NADPH binding site, showed reduced activity but were still able to dimerize though with an increase in intermediary forms. Based on these results, we propose that the catalytically active form of NTRC is the dimer, which formation is induced by NADPH.
Mol Plant 2009 May
PMID:The quaternary structure of NADPH thioredoxin reductase C is redox-sensitive. 1982 29

Diatoms are unicellular algae of great ecological importance. So far, very little is known about the regulation of carbon fixation in these algae; however, there are strong indications that in diatom plastids, the ferredoxin/thioredoxin system might play a minor role in redox regulation of the photosynthetic reactions compared to land plants. Until now, it is unknown whether there are fewer or other target enzymes of thioredoxins in diatoms. Only a single potential target enzyme for thioredoxin, the plastidic fructose-1,6-bisphosphatase, has yet been identified. Nevertheless, during the annotation of the genome of the diatom Phaeodactylum tricornutum, we identified several genes encoding different thioredoxins. Utilizing in vivo expression of GFP:presequence fusion proteins in P. tricornutum, we were able to show that these thioredoxins are targeted either into plastids, mitochondria, or remain in the cytosol. Surprisingly, two of the three usually cytosolic thioredoxin h proteins are apparently plastid associated and, together with a thioredoxin reductase, putatively located in the periplastidic compartment. This is one of the few indications for so far unknown enzymatic reactions in the space between the two pairs of diatom plastid envelope membranes.
Mol Plant 2009 May
PMID:The presence and localization of thioredoxins in diatoms, unicellular algae of secondary endosymbiotic origin. 1982 30


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