A healthy cell wall is indispensable for survival of bacteria. The building blocks of the bacterial peptidoglycan, the major constituent of cell wall, are assembled within the cytoplasm, then they are transported to the surface of the cytoplasmic membrane. The final steps of cell wall assembly take place on the cytoplasmic membrane and they involve the interplay among the activities of a set of enzymes referred to as penicillin-binding proteins (PBPs) and lytic transglcosylases. PBPs catalyze the final steps of cell wall maturation. The roles of lytic transglycosylases are less well understood. They are involved in recycling of cell wall, as well as in restructuring of the cell wall for incorporation of macromolecular systems such as various secretion systems, uptake systems and pilus that requires localized degradation/alteration of the cell wall. Investigations of the activities of lytic transglycosylases are disclosed in this grant application.
Three Specific Aims are offered.
Specific Aim 1 is to clone the genes, to express and to purify the proteins and to perform studies on the reactions of the six Escherichia coli lytic transglycosylases. Synthetic substrates that closely mimic the polymeric substrates for these enzymes are to be developed for detailed study of the microscopic steps of the reactions of these enzymes.
Specific Aim 2 describes the detailed studies of inhibition of lytic transglycosylases. These studies will prepare molecules as both mechanistic probes and as potential antibiotics.
Specific Aim 3 will deal with a key step in cell wall maturation catalyzed by Peptidoglycan Undecaprenyl Releasing Enzyme. It is expected that the outcome of these studies will shed definitive light on the biochemical mechanisms of lytic transglycosylases and pave the way for discoveries of novel inhibitors as potential future antibiotics.

Public Health Relevance

Bacteria require a healthy cell wall for survival. Processes that lead to the formation of cell wall are complex and are targets for antibiotics. This grant application proposes to investigate cell wall and its formation and degradation. The knowledge from the proposed study will provide greater understanding of how bacterial cell wall exists and functions and how potential antibiotic molecules can be generated that target the cell wall.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Synthetic and Biological Chemistry B Study Section (SBCB)
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Fabian, Miles
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University of Notre Dame
Schools of Arts and Sciences
Notre Dame
United States
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Lee, Mijoon; Batuecas, María T; Tomoshige, Shusuke et al. (2018) Exolytic and endolytic turnover of peptidoglycan by lytic transglycosylase Slt of Pseudomonas aeruginosa. Proc Natl Acad Sci U S A 115:4393-4398
Tomoshige, Shusuke; Dik, David A; Akabane-Nakata, Masaaki et al. (2018) Total Syntheses of Bulgecins A, B, and C and Their Bactericidal Potentiation of the ?-Lactam Antibiotics. ACS Infect Dis 4:860-867
Byun, Byungjin; Mahasenan, Kiran V; Dik, David A et al. (2018) Mechanism of the Escherichia coli MltE lytic transglycosylase, the cell-wall-penetrating enzyme for Type VI secretion system assembly. Sci Rep 8:4110
Dik, David A; Fisher, Jed F; Mobashery, Shahriar (2018) Cell-Wall Recycling of the Gram-Negative Bacteria and the Nexus to Antibiotic Resistance. Chem Rev 118:5952-5984
Light, Samuel H; Cahoon, Laty A; Mahasenan, Kiran V et al. (2017) Transferase Versus Hydrolase: The Role of Conformational Flexibility in Reaction Specificity. Structure 25:295-304
Lee, Mijoon; Hesek, Dusan; Dik, David A et al. (2017) From Genome to Proteome to Elucidation of Reactions for All Eleven Known Lytic Transglycosylases from Pseudomonas aeruginosa. Angew Chem Int Ed Engl 56:2735-2739
Dik, David A; Marous, Daniel R; Fisher, Jed F et al. (2017) Lytic transglycosylases: concinnity in concision of the bacterial cell wall. Crit Rev Biochem Mol Biol 52:503-542
Acebrón, Iván; Mahasenan, Kiran V; De Benedetti, Stefania et al. (2017) Catalytic Cycle of the N-Acetylglucosaminidase NagZ from Pseudomonas aeruginosa. J Am Chem Soc 139:6795-6798
Lee, Mijoon; Hesek, Dusan; Lastochkin, Elena et al. (2017) Deciphering the Nature of Enzymatic Modifications of Bacterial Cell Walls. Chembiochem 18:1696-1702
Dik, David A; Domínguez-Gil, Teresa; Lee, Mijoon et al. (2017) Muropeptide Binding and the X-ray Structure of the Effector Domain of the Transcriptional Regulator AmpR of Pseudomonas aeruginosa. J Am Chem Soc 139:1448-1451

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