Disrupting Dogma: Investigating LPS Biosynthesis Inhibition as an Alternative Mechanism of Action of Aminoglycoside Antibiotics

项目摘要 由于许多革兰氏阴性细菌物种表现出抗菌药物耐药性， 新的抑制剂和现有抑制剂的优化是必要的，以使有效的治疗， 疾病。我们实验室和其他实验室的最新研究成果表明，氨基糖苷类药物具有最小的 影响蛋白质合成，事实上，它们在大肠杆菌中有效地与庚糖基转移酶I（HepI）结合。这 这是一个重要的发现，因为它可能会使这类抗生素得到显着的重新设计， 副作用较少的药物，因为HepI和核糖体结合位点的大小非常不同， 在电荷上有很大的差异（HepI带正电荷，而核糖体带负电荷）。这 一项提案将推动重新设计氨基糖苷类抗生素的努力，以增强HepI的结合， 与其他细胞靶点结合，可能导致耳毒性和肾毒性等副作用。 我们的研究将解决三个假设：（1）氨基糖苷类药物结合到其他细胞靶点， 核糖体包括庚糖基转移酶，（2）理解HepI-氨基糖苷相互作用 将使结构修饰和杀菌活性的优化成为可能，以及（3）计算方法 可以促进氨基糖苷类药物的重新设计。迄今为止，重新设计氨基糖苷类以更有效地结合 核糖体未能导致更有效的药物，这可能是因为作用机制涉及 其他酶如HepI。这项工作有望加强药物发现的努力，同时也提供培训， 我实验室的学生和卫斯理大学的两个高级生物化学课程的学生，在21世纪的世纪药物发现 方法.

Project Summary With numerous Gram-negative bacterial species demonstrating antimicrobial drug resistance, the identification of new inhibitors and the optimization of existing inhibitors is necessary to enable an effective treatment of illnesses. Recent research efforts in our lab and others have demonstrated that aminoglycosides have minimal impact on protein synthesis and in fact they potently bind to heptosytransferase I (HepI) in Escherichia coli. This is an important finding, because it may allow for this class of antibiotics to be dramatically redesigned to be better drugs with fewer side effects, because the HepI and ribosome binding sites have very different sizes and they have dramatic differences in charges (HepI is positively charged, while the ribosome is negatively charged). This proposal will advance efforts to redesign aminoglycoside antibiotics to enhance HepI binding and to reduce binding to other cellular targets that can lead to side effects like oto- and nephrotoxicity. Our investigation will address three hypotheses: (1) that aminoglycosides bind to other cellular targets beyond the ribosome including heptosyltransferase enzymes, (2) understanding the HepI-aminoglycoside interactions will enable structural modification and optimization of bactericidal activity, and (3) that computational methods can enhance aminoglycoside redesign. To date, efforts to redesign aminoglycosides for more potent binding to the ribosome has failed to lead to more potent drugs, and this is likely because the mechanism of action involves other enzymes like HepI. This work promises to enhance drug discovery efforts while also providing training for students in my lab and in two upper-level biochemistry courses at Wesleyan in 21st century drug discovery methods.