Deciphering the in vivo redox behavior of human peroxiredoxins I and II by expressing in budding yeast

Free Radic Biol Med. 2019 Dec;145:321-329. doi: 10.1016/j.freeradbiomed.2019.09.034. Epub 2019 Sep 30.

Abstract

Peroxiredoxins (Prxs), scavenge cellular peroxides by forming recyclable disulfides but under high oxidative stress, hyperoxidation of their active-site Cys residue results in loss of their peroxidase activity. Saccharomyces cerevisiae deficient in human Prx (hPrx) orthologue TSA1 show growth defects under oxidative stress. They can be complemented with hPRXI but not by hPRXII, but it is not clear how the disulfide and hyperoxidation states of the hPrx vary in yeast under oxidative stress. To understand this, we used oxidative-stress sensitive tsa1tsa2Δ yeast strain to express hPRXI or hPRXII. We found that hPrxI in yeast exists as a mixture of disulfide-linked dimer and reduced monomer but becomes hyperoxidized upon elevated oxidative stress as analyzed under denaturing conditions (SDS-PAGE). In contrast, hPrxII was present predominantly as the disulfide in unstressed cells and readily converted to its hyperoxidized, peroxidase-inactive form even with mild oxidative stress. Interestingly, we found that plant extracts containing polyphenol antioxidants provided further protection against the growth defects of the tsa1tsa2Δ strain expressing hPrx and preserved the peroxidase-active forms of the Prxs. The extracts also helped to protect against hyperoxidation of hPrxs in HeLa cells. Based on these findings we can conclude that resistance to oxidative stress of yeast cells expressing individual hPrxs requires the hPrx to be maintained in a redox state that permits redox cycling and peroxidase activity. Peroxidase activity decreases as the hPrx becomes hyperoxidized and the limited protection by hPrxII compared with hPrxI can be explained by its greater sensitivity to hyperoxidation.

Keywords: Antioxidants; Hyperoxidation; Polyphenol; Reactive oxygen species; Tsa1.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antioxidants / metabolism
  • Catalytic Domain / genetics
  • Cysteine / metabolism
  • Disulfides / metabolism
  • HeLa Cells
  • Homeodomain Proteins / genetics*
  • Homeodomain Proteins / metabolism
  • Humans
  • Hydrogen Peroxide / metabolism
  • Oxidation-Reduction / drug effects
  • Oxidative Stress / genetics*
  • Peroxidases / genetics*
  • Peroxidases / metabolism
  • Peroxides / metabolism
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins / genetics*
  • Saccharomyces cerevisiae Proteins / metabolism

Substances

  • Antioxidants
  • Disulfides
  • Homeodomain Proteins
  • PRRX1 protein, human
  • PRRX2 protein, human
  • Peroxides
  • Saccharomyces cerevisiae Proteins
  • Hydrogen Peroxide
  • Peroxidases
  • Tsa1 protein, S cerevisiae
  • Cysteine