A comparison of Prx- and OxyR-based H2O2 probes expressed in S. cerevisiae

Genetically encoded fluorescent H₂O₂ probes continue to advance the field of redox biology. Here, we compare the previously established peroxiredoxin-based H₂O₂ probe roGFP2-Tsa2ΔC_R with the newly described OxyR-based H₂O₂ probe HyPer7, using yeast as the model system. Although not as sensitive as roGFP2-Tsa2ΔC_R, HyPer7 is much improved relative to earlier HyPer versions, most notably by ratiometric pH stability. The most striking difference between the two probes is the dynamics of intracellular probe reduction. HyPer7 is rapidly reduced, predominantly by the thioredoxin system, whereas roGFP2-Tsa2ΔC_R is reduced more slowly, predominantly by the glutathione system. We discuss the pros and cons of each probe and suggest that future side-by-side measurements with both probes may provide information on the relative activity of the two major cellular reducing systems.