The increasing antimicrobial resistance of Staphylococcus aureus represents one of the greatest challenges in modern medicine and has driven the development of novel therapeutic strategies targeting bacterial virulence factors. One of the promising targets is serine hydrolase FphI, for which no specific small-molecule inhibitors have been reported to date.
In this master's thesis, a focused library of carbamoyl fluorides was synthesized and evaluated for their inhibitory activity against FphI. Biochemical evaluation identified several highly potent inhibitors with nanomolar activity. Kinetic studies confirmed their two-step covalent mechanism of inhibition, while mass spectrometry and X-ray crystallography verified covalent modification of the catalytic Ser94 residue and revealed the binding mode of the inhibitors within the enzyme active site. Molecular modelling and molecular dynamics simulations demonstrated that the formation of a covalent bond between the carbamoyl fluoride warhead and the catalytic serine residue is essential for effective inhibition, whereas non-covalent interactions alone are insufficient to ensure stable ligand binding. The selected compounds did not exhibit significant antibacterial activity.
The results of this master's thesis describe the first covalent inhibitors of FphI and provide important insights into its inhibition mechanism and biological role. The developed compounds represent promising starting points for the further development of selective FphI inhibitors and chemical probes for investigating the biological role of this enzyme, but not for development of novel antibiotics against Staphylococcus aureus.
|
