# Biochemical and Functional Profiling of Thioredoxin-Dependent Cytosolic GPX-like Proteins in Euglena gracilis

## TL;DR

This study investigates how the organism Euglena gracilis manages oxidative stress using thioredoxin-dependent enzymes instead of typical antioxidant systems.

## Contribution

The study reveals the biochemical and functional roles of thioredoxin-dependent cytosolic GPX-like proteins in ROS homeostasis in Euglena.

## Key findings

- EgGPXLs use thioredoxin to reduce H2O2 and t-BOOH with measurable peroxidase activity.
- Silencing EgGPXL genes did not cause critical effects under normal or high-light conditions.
- EgGPXL suppression rescued cells from the negative effects of EgPrx1/EgPrx4 silencing.

## Abstract

Unlike plants and animals, the phytoflagellate Euglena gracilis lacks catalase and contains a non-selenocysteine glutathione peroxidase-like protein (EgGPXL), two peroxiredoxins (EgPrx1 and EgPrx4), and one ascorbate peroxidase in the cytosol to maintain reactive oxygen species (ROS) homeostasis. In the present study, the full-length cDNA of three cytosolic EgGPXLs was obtained and further characterized biochemically and functionally. These EgGPXLs used thioredoxin instead of glutathione as an electron donor to reduce the levels of H2O2 and t-BOOH. The specific peroxidase activities of these enzymes for H2O2 and t-BOOH were 1.3 to 4.9 and 0.79 to 3.5 µmol/min/mg protein, respectively. Cytosolic EgGPXLs and EgPrx1/EgPrx4 were silenced simultaneously to investigate the synergistic effects of these genes on the physiological function of E. gracilis. The suppression of cytosolic EgGPXL genes was unable to induce any critical phenomena in Euglena under normal (100 μmol photons m−2 s−1) and high-light conditions (350 μmol photons m−2 s−1) at both autotrophic and heterotrophic states. Unexpectedly, the suppression of EgGPXL genes was able to rescue the EgPrx1/EgPrx4-silenced cell line from a critical situation. This study explored the potential resilience of Euglena to ROS, even with restriction of the cytosolic antioxidant system, indicating the involvement of some compensatory mechanisms.