Different consequences of reactions with hydrogen peroxide and t-butyl hydroperoxide in the hyperoxidative inactivation of rat peroxiredoxin-4

Abstract

Eukaryotic typical 2-Cys type peroxiredoxin (Prx) is inactivated by hyperoxidation of the peroxidatic cysteine to a sulphinic acid in a catalytic cycle-dependent manner. This inactivation process has been well documented for cytosolic isoforms of Prx. However, such a hyperoxidative inactivation has not fully been investigated in Prx-4, a secretable endoplasmic reticulum-resident isoform, in spite of being a typical 2-Cys type, and details of this process are reported herein. As has been observed in many peroxiredoxins, the peroxidase activity of Prx-4 was almost completely inhibited in the reaction with t-butyl hydroperoxide. On the other hand, when H(2)O(2) was used as the substrate, the peroxidase activity significantly remained after oxidative damage. In spite of these different consequences, mass spectrometric analyses indicated that both reactions resulted in the same oxidative damage, i.e. sulphinic acid formation at the peroxidatic cysteine, suggesting that another cysteine in the active site confers the peroxidase activity. As suggested by the analyses using cysteine-substituted mutants sulphinic acid formation at the peroxidatic cysteine may play a role in the development of the possible alternative mechanism, thereby sustaining the peroxidase activity that prefers H(2)O(2).

MrSkn7 Controls Sporulation, Cell Wall Integrity, Autolysis, and Virulence in Metarhizium robertsii

“…3A and B). Alternatively, it could be due to the fact that t-butyl-hydrogen peroxide but not H 2 O 2 inhibits cell peroxidase activity (46). The exact mechanism remains to be determined; however, this observation is in contrast to the null mutants of A. fumigatus and A. nidulans yeasts that are sensitive to both oxidants (5).…”

The role of peroxiredoxins in cancer

“…In addition, it has an important chaperone function, operating by means of a versatile mechanism that allows it to switch from redox-dependent and reversibly convertible, disulfide-linked homodimers to higher-order multimers, which enables the interaction with binding partners, including stress-responsive kinases, membrane proteins and immune modulators (110). As a chaperone protein, it cooperates with the protein disulfide isomerase (PDI), a key foldase and chaperone at the ER level, mediating the oxidative folding of various ER proteins (111).…”

Physiological and pathological views of peroxiredoxin 4

“…This altered form of PRDX4 shows a substrate preference that differs from that of an intact fully active enzyme. The altered form displayed higher activity toward hydrogen peroxide compared to t-butyl hydroperoxide while the intact acts more actively on t-butyl hydroperoxide [29].…”

“…This inactivation process is more prominent with higher concentrations of peroxide as the substrate. In the case of PRDX4, however, activity was observed even after hyperoxidation of the catalytic Cys [29]. It appears that the enzyme still functions, albeit with altered kinetic properties, probably by an alternative catalytic mechanism in which the resolving Cys residue directly reacts with peroxide and interacts with Trx.…”

“…Alternatively, the Cys-SOH can further react with peroxide and is "hyperoxidized" or "overoxidized" into cysteine sulfinic acid, Cys-SO 2 H [28,29]. Thus, the interchain disulfide formation competes with hyperoxidation for the Cys-SOH.…”