Sequential enrichment of phosphorylation on reversibly oxidized proteoforms – part 2

Post-translational modifications (PTMs) are necessary for the regulation of critical processes including metabolism, signaling, and overall homeostasis. While proteins PTMs have been largely investigated independently in bottom-up proteomics methodologies, examination into how different PTMs interact, or crosstalk, will reveal a more complete understanding of the reciprocity of signaling cascades across numerous pathways. Combinatorial reversible thiol oxidation and phosphorylation in eukaryotes is largely recognized, but rigorous approaches for experimental verification are underdeveloped and must be advanced to begin meaningful definition of crosstalk in targeted pathway and systems biology research. Herein, we applied protein-level enrichment of reversibly oxidized proteoforms in Chlamydomonas reinhardtii with subsequent phosphopeptide analysis to determine the extent of phosphorylation in the redox thiol proteome.