Design and synthesis of nucleoside-based small molecules to inhibit phosphoglycosyl transferases

项目摘要/摘要 抗生素耐药性正日益成为对全球健康的威胁。这一奖学金的目的是生产 以与细菌生存和最终抗生素耐药性相关的酶为靶标的抑制剂库。 核苷天然产物对细菌细胞壁的合成显示出生物活性，使它们 用于缓蚀剂开发的极佳支架。然而，这些天然产品的复杂性使得 合成，控制选择性，并将毒性降至最低是一个挑战。作为靶点的一系列酶 这个建议是单位磷酸糖基转移酶(PGTS)。这些PGT是催化 与细菌毒力有关的糖偶联物合成的第一步。一旦一组抑制剂被 合成后，将用不同的单位PGT进行生化检测，以确定它们的IC50和 Kd值。然而，抗生素开发的一个主要限制是某些抑制剂不能通过 并在革兰氏阴性菌体内蓄积。出于这个原因，将对这些抑制剂进行细胞测试 革兰氏阴性菌摄取的液-质联用定量研究 (LC-MS)。最后，将进行UDP-糖的化学酶合成，并与 生物化学分析，以帮助确定单一性PGT的特征。年提出的工作的高潮 这项研究将有助于开发针对单位PGTS的有效抑制剂，这将 提供研究糖共轭生物合成的工具，并最终帮助对抗抗生素耐药性。

Project Summary/Abstract 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.