Use of molecular homology model to identify inhibitors of Staphylococcus pseudintermedius sortase A

Conventional antibiotic-based therapies for the treatment of Staphylococcus pseudintermedius infections have become less reliable in recent years due to the increased prevalence of methicillin resistance and multidrug resistance. Therefore, development of alternate therapeutic strategies is a high priority. Treatments based on sortase inhibition hold potential to address this shortcoming. Sortase A (SrtA) is a transpeptidase, commonly produced by Gram positive bacteria, that interacts with proteins baring a C-terminal Leu-Pro-X-Thr-Gly (LPXTG) motif, anchoring them on the peptidoglycan cell wall. SrtA substrates include numerous virulence factors that may overcome the host immune response including protein A, fibrinogen binding proteins and fibronectin binding proteins. In the present study, the srtA gene from S. pseudintermedius was identified and its conservation and level of transcription determined in isolates representing major clonal complexes. The gene was synthesized and then expressed in Escherichia coli and the recombinant enzyme’s activity measured using a synthetic substrate. A three dimensional homology model of SrtA was generated and used in virtual screening libraries of chemicals for potential inhibitors. Four compounds that showed 50% or greater inhibition of SrtA activity were identified. A thermal shift assay confirmed binding of one inhibitor to the active site of SrtA. SrtA inhibitor was active in preventing protein A anchoring in bacterial cultures. The results confirm the function of the srtA encoded protein and indicate that SrtA inhibition has potential for treatment of S. pseudintermedius infections