Tsa1 is the dominant peroxide scavenger and a source of H2O2-dependent GSSG production in yeast

Abstract

AbstractHydrogen peroxide (H2O2) is an important biological molecule, functioning both as a second messenger in cell signaling and, especially at higher concentrations, as a cause of cell damage. Cells harbor multiple enzymes that have peroxide reducing activityin vitro. However, the contribution of each of these enzymes towards peroxide scavengingin vivois less clear. Therefore, to directly investigatein vivoperoxide scavenging, we used the genetically encoded peroxide sensors, roGFP2-Tsa2ΔCRand HyPer7, to systematically screen the peroxide scavenging capacity of yeast thiol and heme peroxidase mutants. We show that the 2-Cys peroxiredoxin Tsa1 alone is responsible for almost all exogenous H2O2andtert-butyl hydroperoxide scavenging. The two catalases and cytochromecperoxidase only produce observable scavenging defects at higher H2O2concentrations when these three heme peroxidases are deleted in combination. We also analyzed the reduction of Tsa1in vitro, revealing that the enzyme is efficiently reduced by thioredoxin 1 with a rate constant of 2.8×106M−1s−1. When thioredoxins are oxidized, Tsa1 can become an important source of H2O2-dependent cytosolic glutathione disulfide production in yeast. Our findings clarify the importance of the various thiol and heme peroxidases for peroxide removal and suggest that most thiol peroxidases have alternative or specialized functions in specific subcellular compartments.