Chemical profiling of live cell temporal redox dynamics in a photoautotrophic cyanobacterium

We have developed a quantitative chemical probe approach for live cell labeling of proteins that are sensitive to redox modifications. We utilize this in vivo strategy coupled to mass spectrometry-based proteomics to identify 176 proteins undergoing ~5-10 fold dynamic redox change in response to nutrient limitation and subsequent replenishment in the photoautotrophic cyanobacterium, Synechococcus sp. PCC 7002. This method enables the identification of redox changes in as little as 30 seconds after nutrient perturbation, and oscillations in reduction and oxidation for 60 minutes following the perturbation. The redox changes were validated by demonstrating that protein abundances did not change per global proteomic analyses. Peptides identified by MS for global and probe-labeled samples were required to be at least six amino acids in length having a mass spectra generating function score of <=1E-10, which corresponds to an FDR of <1%. Additionally, only peptides unique in identifying a single protein were utilized to estimate protein abundances, and proteins represented by <2 unique peptides were removed. This resulted in the identification of 176 redox probe labeled protein identifications, and 808 protein identifications in the global data.