Antibiotic resistance is becoming an increasing threat to global health. This fellowship aims to produce inhibitor libraries that target enzymes associated with bacterial survival and ultimately antibiotic resistance. Nucleoside natural products have shown biological activity towards bacterial cell wall synthesis, making them excellent scaffolds for inhibitor development. However, the complexity of these natural products makes synthesis, controlling selectivity, and minimizing toxicity a challenge. The family of enzymes that are targeted in this proposal are monotopic phosphoglycosyl transferases (PGTs). These PGTs are enzymes that catalyze the initial step in the synthesis of glycoconjugates that are linked with bacterial virulence. Once a set of inhibitors are synthesized, they will be tested with biochemical assays with various monotopic PGTs to establish their IC50 and Kd values. However, a major limitation with antibiotic development is the inability of some inhibitors to traverse the membrane and accumulate in Gram-negative bacteria. For that reason, the inhibitors will be tested for cellular uptake in Gram-negative bacteria and quantified using liquid chromatography coupled with mass spectroscopy (LC-MS). Lastly, chemoenzymatic syntheses of a UDP-sugars, will be performed and used in combination with biochemical assays to aid in the characterization of a monotopic PGT. The culmination of the work proposed in this fellowship will contribute to the development of effective inhibitors targeting monotopic PGTs, which will provide tools to study glycoconjugate biosynthesis and ultimately aid in the fight against antibiotic resistance.
Phosphoglycosyl transferases (PGTs) are responsible for catalyzing the synthesis of complex glycan products associated with bacterial virulence. PGTs are involved in the initial step in the bacterial glycosylation biosynthesis by transferring a phosphate moiety from a uridine diphosphate-sugar (UDP-sugar) to membrane bound undecaprenol phosphate, thus making nucleoside-based small molecules excellent scaffolds for antibiotic applications. This study aims to design and synthesize effective uridine-based inhibitors to target monotopic PGTs.