Sortase Enzyme Mediated Surface Protein Display in Gram-positive Bacteria: Mechanistic Studies and Inhibitor Development
Staphylococcus aureus is a leading cause of hospital-acquired infections in the United States. The rapid emergence of multidrug-resistant strains has created an urgent need for new antibiotics. S. aureus and other Gram-positive pathogens use sortase enzymes to display surface virulence factors to promote infections. Compounds that inhibit sortase might therefore function as potent anti-infective agents. This dissertation focuses on the discovery of sortase A (SrtA) inhibitors, as well as more fundamental studies of the mechanism of catalysis. Both structural (NMR) and in silico approaches were used for inhibitor development, which resulted in the discovery of a very potent pyridazinone class compound that effectively inhibits S. aureus SrtA at nanomolar concentrations. This compound and its related derivatives also inhibit Bacillus anthracis SrtA, but they are not toxic to human cells. Therefore, they are promising leads for further development into a therapeutic. To gain insights into the sortase catalytic mechanism, I also determined the 3D structure of the B. anthracis SrtA bound to a substrate analog using NMR methods. The structure reveals a novel N-terminal extension that regulates substrate entry, and a substrate-induced disorder to order transition of one of the active site loops. Ultimately, this new structural information may be useful in guiding the design of pan-sortase inhibitors that can be used to treat a broad spectrum of bacterial infections.