The proteome and phosphoproteome of Streptococcus pyogenes M49 was investigated at different growth phases of cultures grown in either rich medium (THY), chemically defined medium without carbon source, or chemically defined medium containing 1% fructose. Of the 815 phosphosites identified, 463 were included in a label-free quantitative analysis of dynamic protein phosphorylation. A small group of phosphorylation events, almost exclusively at threonine residues, was strongest during growth and decreased during stationary phase. These included phosphorylation sites of the PASTA kinase SP-STK and its putative substrates suggesting that the PASTA kinase-dependent processes regulating the cell cycle in related bacteria are also conserved in S. pyogenes. Most phosphorylation events occurred preferentially at serine residues in the stationary growth phase and under starvation conditions. The elucidation of the physiological significance and the responsible kinases of these phosphorylations require further investigations. In addition to phosphorylation events at S/T/Y residues, phosphoglycerylation of lysine (PGK) occurred frequently among the enriched phosphopeptides.