Inactivation of Glutathione Peroxidase by Nitric Oxide

Abstract

S-nitro-N-acetyl-DL-penicillamine (SNAP), a nitric oxide (NO) donor, inactivated bovine glutathione peroxidase (GPx) in a dose-and time-dependent manner. The IC 50 of SNAP for GPx was 2 M at 1 h of incubation and was 20% of the IC 50 for another thiol enzyme, glyceraldehyde-3-phosphate dehydrogenase, in which a specific cysteine residue is known to be nitrosylated. Incubation of the inactivated GPx with 5 mM dithiothreitol within 1 h restored about 50% of activity of the start of the SNAP incubation. A longer exposure to NO donors, however, irreversibly inactivated the enzyme. The similarity of the inactivation with SNAP and reactivation with dithiothreitol of GPx to that of glyceraldehyde-3-phosphate dehydrogenase, suggested that NO released from SNAP modified a cysteine-like essential residue on GPx. When U937 cells were incubated with 100 M SNAP for 1 h, a significant decrease in GPx activity was observed although the change was less dramatic than that with the purified enzyme, and intracellular peroxide levels increased as judged by flow cytometric analysis using a peroxide-sensitive dye. Other major antioxidative enzymes, copper/zinc superoxide dismutase, manganese superoxide dismutase, and catalase, were not affected by SNAP, which suggested that the increased accumulation of peroxides in SNAP-treated cells was due to inhibition of GPx activity by NO. Moreover, stimulation with lipopolysaccharide significantly decreased intracellular GPx activity in RAW 264.7 cells, and this effect was blocked by NO synthase inhibitor N -methyl-L-arginine. This indicated that GPx was also inactivated by endogenous NO. This mechanism may at least in part explain the cytotoxic effects of NO on cells and NOinduced apoptotic cell death.

A mosquito 2-Cys peroxiredoxin protects against nitrosative and oxidative stresses associated with malaria parasite infection

“…Insects seem to lack GPx [47] and GR [12,48] and instead substitute a thioredoxin system for similar chemical reactions [12,48]. High levels of NO synthesis in insects, such as we have observed in A. stephensi, may have been a strong selective pressure for this trait because NO inactivates GPx in a dose-dependent manner [49].…”

“…The peroxidatic cysteine of 1-Cys, 2-Cys, and BCP-like Prx proteins is conserved in the predicted AsPrx-4783 and corresponds to C 49 (Fig. 1).…”

“…1), which are required to activate the peroxidatic cysteine. Specifically, T 46 hydrogen bonds to C 49 , whereas R 125 interacts with the peroxidatic Cys to lower the pK a of the ionized form of thiol [30,31].…”

Oxidative and nitrosative stress induces peroxiredoxins in pancreatic beta cells

“…In addition, inactivation of glutathione peroxidase by nitric oxide has been observed [65]. The bacterial peroxiredoxin AhpC (alkylhydroperoxide reductase subunit C) has been shown to protect cells against reactive nitrogen intermediates, such as nitric oxide [25].…”

Endotoxemia, pentose cycle, and the oxidant/antioxidant balance in the hepatic sinusoid

“…Nitric oxide production is minimal in resting sinusoidal cells but it becomes substantial after the induction of inducible nitric oxide synthase (iNOS) by LPS, bacteria, or cytokines [20,44]. Nitric oxide, when present in high concentrations, may directly alter cellular proteins, including NADPH oxidase [45], peroxidases [46], catalase [47], and glucose metabolic enzymes, thereby modulating their activity [48][49][50]. Nitric oxide synthesis is dependent on glutathione [51], whereas nitric oxide depletes GSH and activates the HMS [48].…”