Thioredoxin, Trx

Fig. 2.2 Simplified scheme of oxidant/antioxidant regulation ofNF-KB activation. Different stimuli, leading to an increase of ROS generation inside the ceU, activate the phosphorylation of IkB inhibitory protein and the subsequent proteolysis. Thioredoxin (Trx) may reduce activated NF-kB proteins facilitating nuclear translocation.Qnce released from IkB, the NF-kB complex translocates into the nucleus and the binding to DNA domain in the promoters and enhancers of genes such as TNF-a, IL-1, proliferation and chemotactic factors, adhesion molecule. Some of these genes, in turn, may further induce NF-kB activation, leading to a vicious circle if the regulatory cellular system escapes from...

Mammalian thioredoxin reductases are a family of selenium-containing pyridine nucleotide-disulfide oxidoreductases. These enzymes catalyze NADPH-dependent reduction of the redox protein thioredoxin (Trx), which contains a redox-active disulfide and dithiol group and by itself may function as an efficient cytosolic antioxidant [77]. One of the functions of Trx/ thioredoxin reductase system is the NADPH-catalyzed reduction of protein disulfide [78] ... 

Yoon, B.I., Hirahayashi, Y, Kaneko, T., Kodama, Y, Kano, J., Yodoi, J., Kim, D.Y and Inoue, T. (2001) Transgene expression of thioredoxin (TRX-ADF) protects against 2,3,7,8-tetrachlorodihenzo-p-dioxrn (TCDD)-induced hematotoxicity. Archives of Environmental Contamination and Toxicology, 41, 232-236. 

In neural cells, the redox status is controlled by the thioredoxin (Trx) and glutathione (GSH) systems that scavenge harmful intracellular ROS. Thioredoxins are antioxidants that serve as a general protein disulphide oxidoreductase (Saitoh et al., 1998). They interact with a broad range of proteins by a redox mechanism based on the reversible oxidation of 2 cysteine thiol groups to a disulphide, accompanied by the transfer of 2 electrons and 2 protons. These proteins maintain their reduced state through the thioredoxin system, which consists of NADPH, thioredoxin reductase (TR), and thioredoxin (Trx) (Williams, Jr. et al., 2000 Saitoh et al., 1998). The thioredoxin system is a system inducible by oxidative stress that reduces the disulfide bond in proteins (Fig. 7.4). It is a major cellular redox system that maintains cysteine residues in the reduced state in numerous proteins. 

Another form of displayed peptide library is the FliTrx Random Peptide Display Library (Invitrogen, Paisley, UK), which uses the bacterial flagellum to display random peptide libraries on the E. coli cell surface (14). This library was constructed in the pFliTrx vector, which positions the random peptides in a flagellin (Fli) thioredoxin (Trx) fusion protein. Biopanning with bacteria works surprisingly well in our experience (15) and screening on nitrocellulose is similar to the Smith method. 

Fig. 4. Possible roles of glutathione, glutaredoxin (Grx) and thioredoxin (Trx) in type I iodothyronine deiodination.

TrxRs are homodimeric flavoproteins [80] that catalyze the NADPH-dependent reduction of thioredoxin (Trx), a ubiquitous 12 kDa protein that is the major protein disulfide reductase in cells [81], and belongs to the pyridine nucleotide-disulfide oxidoreductase family [82]. Each monomer includes an FAD prosthetic group, a NADPH binding site and an active site containing a redox-active selenol group. Electrons are transferred from NADPH via FAD to the active-site selenol of TrxR, which then reduces the substrate Trx [83]. The crystal structure of TrxR is shown in Fig. 13 [84],... 

FIGURE 2. Three-dimensional structure of conserved residues and the substrate GDP in the active site of E. coli protein R1 (A) (Eriksson et al., 1997), and proposed reaction mechanism for RNR (B), A turnover is initiated by long-range RTF between Tyrl22 in R2 and Cys439 in Rl, and completed by a reversal of this RTP (thin arrows). Oxidised Rl is reduced by the thioredoxin (Trx), or glutaredoxin system. 

FTR plays the central role in light regulation of the activity of enzymes involved in oxygenic photosynthesis. The light signal is converted into reducing equivalents in the form of reduced 2Fe Fd by photosystem I and FTR catalyzes the reduction of disulfides in thioredoxin (Trx) / and m using the... 

Figure 1 Reactions involved in the glycine reduction pathway hy Clostridia. Pa, Pb, and Pc denote the components of the glycine reduction complex where Pb is specific for the substrate (e.g. glycine, betaine, sarcosine). The reduction of oxidized protein Pa occurs via a redox chain involving thioredoxin reductase (TR) and thioredoxin (TRX). The involvement of ketene as an intermediate has not yet been proven...

Thioredoxin (TRX) is a pleiotropic polypeptide also known as T-cell leukemia-derived factor (Tagaya et al., 1989 Yodoi and Uchiyama, 1992). It has two redox-reactive cysteine residues in the reactive center (Cys-32 and Cys-35) (Holmgren, 1972, 1985, 1989 Buchanan et al., 1994). Reductases are used to donate electrons from NADPH to facilitate the reduction oxidation reaction between the dithiol or disulhde forms of TRX (Luthman and Holmgren, 1982 Watson et al., 2004). 

A set of hybrid proteins containing the Ap40 peptide domain and dimeric glucose dehydrogenase (GDH) or thioredoxin (Trx) have been constructed in present work. Expression and purification of the proteins was accomplished using prokaryotic system based on Escherichia coli. Details of the construction and expression of hybrid proteins are subject of special report and will be presented elsewhere. Hen egg lysozyme was fibrilized as described in our recent work. 

After publishing the facile synthesis of tran -cyclooctene derivatives from the readily accessible cfr-cyclooctene (Royzen et al., 2008), Fox and colleagues extended this work to develop a fast bioconjugation strategy based upon the IVED-DA cycloadditions (Blackman et al., 2008). Maleimide functional frans-cyclooctene was coupled to thioredoxin (after reduction to produce the solvent exposed cysteine) via standard Michael addition chemistry. The now trans-cyclooctene functional thioredoxin (Trx-ene) could then be further modified extremely efficientiy with an electron-deficient tetrazine (3,6-di-(2-pyidyl)-i-tetrazine) shown in the left side of Figure 2.15. The overall strategy is depicted in Scheme 2.5. 

Scheme 2.5 Modification of thioredoxin (Trx) via the IVED-DA cycloaddition (Blackman, Royzen and Fox, 2008). (Reproduced with permission from A.J. Inglis and C. Barner-Kowollik, Ultra rapid approaches to mild macromolecular conjugation, Macromolecular Rapid Communications, 2010, 31, 14, 1247-1266. Wiley-VCH Verlag GmbH Co. KGaA.)...

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