Dynamic protein phosphorylation in Streptococcus pyogenes during growth, stationary phase and starvation

Abstract Background Phosphorylation of proteins at serine, threonine, and tyrosine residues plays an important role in physiological processes of bacteria such as cell cycle, metabolism, virulence, dormancy, sporulation, and stationary phase functions. Streptococcus pyogenes possesses a single known transmembrane serine/threonine kinase belonging to the class of penicillin-binding protein and serine/threonine-associated (PASTA) kinases. To gain initial insights into the targets and dynamics of protein phosphorylation in S. pyogenes serotype M49 strain 591, we performed a proteomics and phosphoproteomics workflow using cultures from different growth conditions, stationary phase and starvation. Results The quantitative analysis of dynamic phosphorylation, which included a subset of 463 out of 815 identified phosphorylation sites, revealed two main types of phosphorylation events, distinguished by the growth phase in which they predominantly occur and their preference for either threonine or serine. A small group of phosphorylation events occurred almost exclusively at threonine residues of proteins related to the cell cycle and was enhanced in growing cells. Many of these phosphorylation sites are highly conserved targets of PASTA kinases in streptococci. The majority of phosphorylation events occurred during stationary phase or starvation, preferentially at serine residues. These phosphorylations may be important for regulatory processes in stationary phase or for persister cell formation, but their function and the kinases responsible for their formation need to be elucidated in further analyses. Moreover, our data indicate that the vast majority of proteins can be phosphorylated, but identification of their phosphopeptides depends on the sensitivity of the proteomic methods used. Conclusions PASTA kinase-dependent cell cycle regulation processes found in related bacteria are conserved in S. pyogenes , but account for only a small part of all phosphorylation events. Most phosphorylation events take place during stationary growth phase and starvation. This phenomenon has also been described for some other bacteria and may therefore be a general feature of bacterial protein phosphorylation.