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- Chasapis, Christos T., et al.
(författare)
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Implications of the mitochondrial interactome of mammalian thioredoxin 2 for normal cellular function and disease
- 2019
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Ingår i: Free Radical Biology & Medicine. - : Elsevier. - 0891-5849 .- 1873-4596. ; 137, s. 59-73
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Tidskriftsartikel (refereegranskat)abstract
- Multiple thioredoxin isoforms exist in all living cells. To explore the possible functions of mammalian mitochondrial thioredoxin 2 (Trx2), an interactome of mouse Trx2 was initially created using (i) a monothiol mouse Trx2 species for capturing protein partners from different organs and (ii) yeast two hybrid screens on human liver and rat brain cDNA libraries. The resulting interactome consisted of 195 proteins (Trx2 included) plus the mitochondrial 16S RNA. 48 of these proteins were classified as mitochondrial (MitoCarta2.0 human inventory). In a second step, the mouse interactome was combined with the current four-membered mitochondrial sub-network of human Trx2 (BioGRID) to give a 53-membered human Trx2 mitochondrial interactome (52 interactor proteins plus the mitochondrial 16S RNA). Although thioredoxins are thiol-employing disulfide oxidoreductases, approximately half of the detected interactions were not due to covalent disulfide bonds. This finding reinstates the extended role of thioredoxins as moderators of protein function by specific non-covalent, protein-protein interactions. Analysis of the mitochondrial interactome suggested that human Trx2 was involved potentially in mitochondrial integrity, formation of iron sulfur clusters, detoxification of aldehydes, mitoribosome assembly and protein synthesis, protein folding, ADP ribosylation, amino acid and lipid metabolism, glycolysis, the TCA cycle and the electron transport chain. The oxidoreductase functions of Trx2 were verified by its detected interactions with mitochondrial peroxiredoxins and methionine sulfoxide reductase. Parkinsons disease, triosephosphate isomerase deficiency, combined oxidative phosphorylation deficiency, and lactate dehydrogenase b deficiency are some of the diseases where the proposed mitochondrial network of Trx2 may be implicated.
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| 2. |
- Fernandes, Aristi Potamitou, et al.
(författare)
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A Novel Monothiol Glutaredoxin (Grx4) from Escherichia coli Can Serve as a Substrate for Thioredoxin Reductase
- 2005
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 280:26, s. 24544-24552
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Tidskriftsartikel (refereegranskat)abstract
- Glutaredoxins are ubiquitous proteins that catalyze the reduction of disulfides via reduced glutathione (GSH). Escherichia coli has three glutaredoxins (Grx1, Grx2, and Grx3), all containing the classic dithiol active site CPYC. We report the cloning, expression, and characterization of a novel monothiol E. coli glutaredoxin, which we name glutaredoxin 4 (Grx4). The protein consists of 115 amino acids (12.7 kDa), has a monothiol (CGFS) potential active site and shows high sequence homology to the other monothiol glutaredoxins and especially to yeast Grx5. Experiments with gene knock-out techniques showed that the reading frame encoding Grx4 was essential. Grx4 was inactive as a GSH-disulfide oxidoreductase in a standard glutaredoxin assay with GSH and hydroxyethyl disulfide in a complete system with NADPH and glutathione reductase. An engineered CGFC active site mutant did not gain activity either. Grx4 in reduced form contained three thiols, and treatment with oxidized GSH resulted in glutathionylation and formation of a disulfide. Remarkably, this disulfide of Grx4 was a direct substrate for NADPH and E. coli thioredoxin reductase, whereas the mixed disulfide was reduced by Grx1. Reduced Grx4 showed the potential to transfer electrons to oxidized E. coli Grx1 and Grx3. Grx4 is highly abundant (750–2000 ng/mg of total soluble protein), as determined by a specific enzyme-link immunosorbent assay, and most likely regulated by guanosine 3′,5′-tetraphosphate upon entry to stationary phase. Grx4 was highly elevated upon iron depletion, suggesting an iron-related function for the protein.
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| 3. |
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| 5. |
- Lu, Jun, et al.
(författare)
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Inhibition of Bacterial Thioredoxin Reductase: An Antibiotic Mechanism Targetting Bacteria Lacking Glutathione
- 2013
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Ingår i: The FASEB Journal. - : Wiley. - 0892-6638 .- 1530-6860. ; 27:4, s. 1394-1403
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Tidskriftsartikel (refereegranskat)abstract
- Increasing antibiotic resistance makes the identification of new antibacterial principles an urgent task. The thioredoxin system including thioredoxinreductase (TrxR), thioredoxin (Trx), and NADPH plays critical roles in cellular DNA synthesis and defense against oxidative stress. Notably, TrxR is very different in structure and mechanism in mammals and bacteria. Ebselen [2-phenyl-1,2 benzisoselenazol-3(2H)-one], a well-known antioxidant and a substrate for mammalian TrxR and Trx, is rapidly bacteriocidal for methicillin-resistant Staphylococcus aureus by an unknown mechanism. We have discovered that ebselen is a competitive inhibitor of Escherichia coli TrxR with a K-i of 0.52 +/- 0.13 mu M, through reaction with the active site dithiol of the enzyme. Bacteria lacking glutathione (GSH) and glutaredoxin, in which TrxR and Trx are essential for DNA synthesis, were particularly sensitive to ebselen. In growth-inhibited E. coli strains, Trx1 and Trx2 were oxidized, demonstrating that electron transfer via thioredoxin was blocked. Ebselen and its sulfur analog ebsulfur were bactericidal for GSH-negative pathogens. Ebsulfur inhibited a clinically isolated Helicobacter pylori strain with a minimum inhibitory concentration value as low as 0.39 mu g/ml. These results demonstrate that bacterial Trx and TrxR are viable antibacterial drug targets using benzisoselenazol and benzisothiazol derivates.-Lu, J., Vlamis-Gardikas, A., Kandasamy, K., Zhao, R., Gustafsson, T. N., Engstrand, L., Hoffner, S., Engman, L., Holmgren, A. Inhibition of bacterial thioredoxin reductase: an antibiotic mechanism targeting bacteria lacking glutathione.
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| 6. |
- Panagopoulos, Ioannis, et al.
(författare)
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Characterization of the native CREB3L2 transcription factor and the FUS/CREB3L2 chimera
- 2007
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Ingår i: Genes, Chromosomes and Cancer. - : Wiley. - 1045-2257 .- 1098-2264. ; 46:2, s. 181-191
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Tidskriftsartikel (refereegranskat)abstract
- CREB3L2 was first identified as the 3'-partner of FUS in a fusion gene that seems to be specific for low grade fibromyxoid sarcoma. In silico analyses suggest that the predicted CREB3L2 protein is a member of the CREB3 family of transcription factors, with its bZIP domain being highly similar to that in CREB3L1, CREB3L3, CREB3L4, CREB3, and Drosophila Bbf-2. In the present study, the authors assessed various cellular outcomes after transfection of NIH3T3 and HEK-293 cells with constructs containing full-length and truncated versions of CREB3L2 and FUS/CREB3L2. Northern blot of CREB3L2 mRNA revealed a 7.4 kbp band that contains 0.4 kbp and 5.5 kbp untranslated 5' and 3' regions, respectively. CREB3L2 constructs containing the first 120 amino acids (aa) showed the highest transcriptional activation. Much stronger transcriptional activation was consistently seen for the FUS/CREB3L2 constructs than for the corresponding CREB3L2 constructs. Transcriptional activity was achieved through the box-B element, ATF6 and CRE binding sites, as well as the GRP78 promoter. Proteins encoded by full-length CREB3L2 and FUS/CREB3L2 were localized to reticular structures of the cytoplasm, whereas the corresponding, truncated proteins lacking the transmembrane domain and the carboxy-terminal part of CREB3L2 resided within the nucleus. The results of the present study show that CREB3L2 is not only structurally, but also functionally very similar to CREB3L1. Thus, studies regarding the pathways influenced by wild-type CREB3L2 should provide valuable clues to the pathogenetic significance of the FUS/CREB3L2 chimera in low grade fibromyxoid sarcoma.
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| 7. |
- Papadopoulos, Evangelos, et al.
(författare)
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Solution structure and biophysical properties of MqsA, a Zn-containing antitoxin from Escherichia coli
- 2012
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Ingår i: Biochimica et Biophysica Acta - Proteins and Proteomics. - : Elsevier BV. - 1570-9639 .- 1878-1454. ; 1824:12, s. 1401-1408
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Tidskriftsartikel (refereegranskat)abstract
- The gene ygiT (mqsA) of Escherichia coli encodes MqsA, the antitoxin of the motility quorum sensing regulator (MqsR). Both proteins are considered to form a DNA binding complex and to be involved in the formation of biofilms and persisters. We have determined the three-dimensional solution structure of MqsA by high-resolution NMR. The protein comprises a well-defined N-terminal domain with a Zn finger motif usually found in eukaryotes, and a defined C-terminal domain with a typical prokaryotic DNA binding helix-turn-helix motif. The two well-defined domains of MqsA have almost identical structure in solution and in the two published crystal structures of dimeric MqsA bound to either MqsR or DNA. However, the connection of the two domains with a flexible linker yields a large variety of possible conformations in solution, which is not reflected in the crystal structures. MqsA binds Zn with all four cysteines, a stoichiometry of 1:1 and a femtomolar affinity (K-a >= 10(17) M-1 at 23 degrees C, pH 7.0).
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