Peroxidases of the peroxiredoxin (Prx) family contain a Cys residue that is preceded by a conserved sequence in the NH₂-terminal region. A new type of mammalian Prx, designated PrxV, has now been identified as the result of a data base search with this conserved Cys-containing sequence. The 162-amino acid PrxV shares only ∼10% sequence identity with previously identified mammalian Prx enzymes and contains Cys residues at positions 73 and 152 in addition to that (Cys⁴⁸) corresponding to the conserved Cys. Analysis of mutant human PrxV proteins in which each of these three Cys residues was individually replaced with serine suggested that the sulfhydryl group of Cys⁴⁸ is the site of oxidation by peroxides and that oxidized Cys⁴⁸ reacts with the sulfhydryl group of Cys¹⁵² to form an intramolecular disulfide linkage. The oxidized intermediate of PrxV is thus distinct from those of other Prx enzymes, which form either an intermolecular disulfide or a sulfenic acid intermediate. The disulfide formed by PrxV is reduced by thioredoxin but not by glutaredoxin or glutathione. Thus, PrxV mutants lacking Cys⁴⁸ or Cys¹⁵² showed no detectable thioredoxin-dependent peroxidase activity, whereas mutation of Cys⁷³ had no effect on activity. Immunoblot analysis revealed that PrxV is widely expressed in rat tissues and cultured mammalian cells and is localized intracellularly to cytosol, mitochondria, and peroxisomes. The peroxidase function of PrxV in vivo was demonstrated by the observations that transient expression of the wild-type protein, but not that of the Cys⁴⁸ mutant, in NIH 3T3 cells inhibited H₂O₂ accumulation and activation of c-Jun NH₂-terminal kinase induced by tumor necrosis factor-α.

Peroxiredoxins (Prxs) are a family of antioxidant proteins that reduce peroxide levels by using reducing agents such as thioredoxin. These proteins were characterized to have a number of cellular functions, including cell proliferation and differentiation and protection of specific proteins from oxidative damage. However, the physiological roles of the peroxiredoxins have not been determined. To clarify the physiological relevance of this protein type, we generated a mouse model deficient in Prx II, which is abundantly expressed in all types of cells. The Prx II^–/– mice were healthy in appearance and fertile. However, they had splenomegaly caused by the congestion of red pulp with hemosiderin accumulation. Heinz bodies were detected in their peripheral blood, and morphologically abnormal cells were elevated in the dense red blood cell (RBC) fractions, which contained markedly higher levels of reactive oxygen species (ROS). The Prx II^–/– mice had significantly decreased hematocrit levels, but increased reticulocyte counts and erythropoietin levels, indicative of a compensatory action to maintain hematologic homeostasis in the mice. In addition, a labeling experiment with the thiol-modifying reagent biotinylated iodoacetamide (BIAM) in Prx II^–/– mice revealed that a variety of RBC proteins were highly oxidized. Our results suggest that Prx II^–/– mice have hemolytic anemia and that Prx II plays a major role in protecting RBCs from oxidative stress in mice.

Peroxiredoxins (Prxs) play an important role in regulating cellular differentiation and proliferation in several types of mammalian cells. One mechanism for this action involves modulation of hydrogen peroxide (H₂O₂)-mediated cellular responses. This report examines the expression of Prx I and Prx II in thyroid cells and their roles in eliminating H₂O₂ produced in response to thyrotropin (TSH). Prx I and Prx II are constitutively expressed in FRTL-5 thyroid cells. Prx I expression, but not Prx II expression, is stimulated by exposure to TSH and H₂O₂. In addition, methimazole induces a high level of Prx I mRNA and protein in these cells. Overexpression of Prx I and Prx II enhances the elimination of H₂O₂ produced by TSH in FRTL-5 cells. Treatment with 500 μm H₂O₂ causes apoptosis in FRTL-5 cells as evidenced by standard assays of apoptosis (i.e. terminal deoxynucleotidyl transferase deoxyuridine triphosphate-biotin nick end labeling, BAX expression, and poly(ADP-ribose) polymerase cleavage. Overexpression of Prx I and Prx II reduces the amount of H₂O₂-induced apoptosis measured by these assays. These results suggest that Prx I and Prx II are involved in the removal of H₂O₂ in thyroid cells and can protect these cells from undergoing apoptosis. These proteins are likely to be involved in the normal physiological response to TSH-induced production of H₂O₂ in thyroid cells.

Antioxidant enzymes as redox-based biomarkers: a brief review Antioxidant enzyme;Cysteine oxidation;Disease;ROS;Redox proteomics; The field of redox proteomics focuses to a large extent on analyzing cysteine oxidation in proteins under different experimental conditions and states of diseases. The identification and localization of oxidized cysteines within the cellular milieu is critical for understanding the redox regulation of proteins under physiological and pathophysiological conditions, and it will in turn provide important information that are potentially useful for the development of novel strategies in the treatment and prevention of diseases associated with oxidative stress. Antioxidant enzymes that catalyze oxidation/reduction processes are able to serve as redox biomarkers in various human diseases, and they are key regulators controlling the redox state of functional proteins. Redox regulators with antioxidant properties related to active mediators, cellular organelles, and the surrounding environments are all connected within a network and are involved in diseases related to redox imbalance including cancer, ischemia/reperfusion injury, neurodegenerative diseases, as well as normal aging. In this review, we will briefly look at the selected aspects of oxidative thiol modification in antioxidant enzymes and thiol oxidation in proteins affected by redox control of antioxidant enzymes and their relation to disease. [BMB Reports 2015; 48(4): 200-208]

Mammalian 2-Cys peroxiredoxin II (Prx II) is a cellular peroxidase that eliminates endogenous H₂O₂. The involvement of Prx II in the regulation of lipopolysaccharide (LPS) signaling is poorly understood. In this report, we show that LPS induces substantially enhanced inflammatory events, which include the signaling molecules nuclear factor κB and mitogen-activated protein kinase (MAPK), in Prx II–deficient macrophages. This effect of LPS was mediated by the robust up-regulation of the reactive oxygen species (ROS)–generating nicotinamide adenine dinucleotide phosphate (NADPH) oxidases and the phosphorylation of p47^phox. Furthermore, challenge with LPS induced greater sensitivity to LPS-induced lethal shock in Prx II–deficient mice than in wild-type mice. Intravenous injection of Prx II–deficient mice with the adenovirus-encoding Prx II gene significantly rescued mice from LPS-induced lethal shock as compared with the injection of a control virus. The administration of catalase mimicked the reversal effects of Prx II on LPS-induced inflammatory responses in Prx II–deficient cells, which suggests that intracellular H₂O₂ is attributable, at least in part, to the enhanced sensitivity to LPS. These results indicate that Prx II is an essential negative regulator of LPS-induced inflammatory signaling through modulation of ROS synthesis via NADPH oxidase activities and, therefore, is crucial for the prevention of excessive host responses to microbial products.