| Staphylococcus aureus is the leading cause of hospital-acquired infections in the United States and many strains have become resistant to available antibiotics, raising the need for new drugs. This dissertation focuses on structural studies by nuclear magnetic resonance (NMR) spectroscopy and inhibitor discovery of a new target enzyme called sortase A (SrtA), a transpeptidase in Gram-positive bacteria that anchors surface proteins to the cell wall peptidylglycan. SrtA is highly conserved and critical for achieving bacterial virulence. Sortase inhibitors may function as anti-infective agents in Gram-positive bacteria that do not affect microbial viability outside the host or induce selective pressures that lead to drug resistance. The process by which SrtA anchors a polypeptide to the peptidylglycan has been extensively studied. However, the exact mechanism through which SrtA recognizes its substrates remained unclear. Detailed structural studies of SrtA-substrate complexes have been hindered because of the low affinity of these interactions. This problem was overcome by determining the solution structure of a covalent SrtA-peptide complex that mimics the first thioacyl intermediate of catalysis. For the first time, this complex structure, supplemented by biochemical studies, has revealed how the sorting signal is recognized. Additionally, dynamics data have shed light onto how sortase enzymes anchor proteins containing conserved LPXTG sorting signals. By using high-throughput screening, several compounds that inhibit SrtA with high potency have been identified. A structure activity relationship (SAR) analysis and molecular docking have been performed on these lead compounds in order to optimize their potency. |
