Optimizing the trapping and identification of sulfenylated proteins in the green microalga Chlamydomonas reinhardtii

In photosynthetic organisms, chloroplast is one of the main sources of reactive oxygen species (ROS). The most common ROS produced during photosynthesis are superoxide anion (O2-), singlet oxygen (1O2) and hydrogen peroxide (H2O2) [1]. In this study we mainly focused on H2O2 which can also act as a signalling molecule by oxidizing cysteine residues of proteins into their sulfenylated form. The aim of the project is to identify these proteins in order to establish their role during oxidative stress in the green microalga C. reinhardtii. We optimized a protocol for trapping sulfenylated proteins from [2] by using a benzothiazine-based chemoselective probe (BTD) [3] specific to recognize -SOH. We succeeded to label sulfenylated proteins after addition of H2O2. However, we are still in the process to optimize the workflow for modified peptides identification by mass spectrometry analyses. Preliminary results on wild type cells have shown that some photosynthetic proteins are susceptible to H2O2 modification, such us the light-harvesting complexes (LHC) of photosysthem II, LHCII3, LHCB3 and LHCII-1.3 important to capture light during photosynthesis. Our analyses will provide information on the Chlamydomonas sulfenome, and will be compared to the other post translational modifications already identified on the Chlamydomonas proteome, such as glutathionylation and nitrosylation.
Funded by FNRS-FWO EOS Project 30829584

[1] Asada (2006). Plant Physiol. 141 (2): 391-396
[2] Huang et al. (2019). PNAS. 116 (42): 21256-21261
[3] Gupta et al. (2017) J. Am. Chem. Soc. 139 (159): 5588–5595