Discovery and characterization of new bacterial cell wall targets and inhibitors to treat resistant infections

PROJECT SUMMARY Bacterial cells are surrounded by multilayered cell envelopes that are required for their growth and play crucial roles in their ability to cause disease. The cytoplasmic membrane forms the foundational layer of the envelope and maintaining the appropriate membrane lipid composition is critical for cell viability. Beyond the cytoplasmic membrane is the peptidoglycan cell wall, which forms a mesh-like framework necessary for bacteria to withstand fluctuations in turgor pressure. In Gram-negative bacteria, the cell wall is thin and relatively fragile, but is surrounded by a second membrane called the outer membrane that provides an additional layer of protection. The outer membrane serves as a major barrier to entry of antibiotics. Depending on the defect, damage to any of the bacterial cell envelope layers can be lethal or can potentiate the effects of cell wall-targeting antibiotics. Understanding the molecular mechanisms underlying cell envelope biogenesis is therefore important for developing new strategies to treat resistant infections. This proposal has four aims. The first will investigate glycopeptide antibiotics, which inhibit synthesis of cell wall peptidoglycan. We have found that some glycopeptides inhibit the initiation step of peptidoglycan polymerization, whereas others inhibit its elongation. We will elucidate the molecular basis for these different mechanisms to lay the groundwork for rational design of new analogs. The next three aims will focus on uncovering mechanisms for how the individual layers of the bacterial cell envelope are assembled and how the different processes are coordinated. Aim 2 will focus on mechanisms of peptidoglycan acetylation and will explore the role of PG acetylation in regulating cell wall hydrolases. Aim 3 will investigate how cell wall hydrolase activity is coordinated with LPS transport to ensure that the peptidoglycan layer expands as the outer membrane expands. Aim 4 will investigate how lipoteichoic acid assembly is coordinated with phospholipid recycling to maintain membrane homeostasis. The work in Aims 2-4 will identify new vulnerabilities that can be targeted to overcome drug-resistant infections.