The enzyme activity of sortase A is regulated by phosphorylation in Staphylococcus aureus .

In many Gram-positive bacteria, the transpeptidase enzyme sortase A (SrtA) anchors surface proteins to cell wall and plays a critical role in the bacterial pathogenesis. Here, we show that in Staphylococcus aureus , an important human pathogen, the SrtA is phosphorylated by serine/threonine protein kinase Stk1. S. aureus SrtA can also be phosphorylated by small-molecule phosphodonor acetyl phosphate (AcP) in vitro . We determined that various amino acid residues of S. aureus SrtA are subject to phosphorylation, primarily on its catalytic site residue cysteine-184 in the context of a bacterial cell lysate. Both Stk1 and AcP-mediated phosphorylation inhibited the enzyme activity of SrtA in vitro . Consequently, deletion of gene (i.e. stp1 ) encoding serine/threonine phosphatase Stp1, the corresponding phosphatase of Stk1, caused an increase in the phosphorylation level of SrtA. The stp1 deletion mutant mimicked the phenotypic traits of srtA deletion mutant (i.e. attenuated growth where either hemoglobin or heme as a sole iron source and reduced liver infections in a mouse model of systemic infection). Importantly, the phenotypic defects of the stp1 deletion mutant can be alleviated by overexpressing srtA . Taken together, our finding suggests that phosphorylation plays an important role in modulating the activity of SrtA in S. aureus.