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; Dhar, Supurna; De Benedetti, Stefania et al. (2016) Muropeptides in Pseudomonas aeruginosa and their Role as Elicitors of β-Lactam-Antibiotic Resistance. Angew Chem Int Ed Engl 55:6882-6
Sandalova, Tatyana; Lee, Mijoon; Henriques-Normark, Birgitta et al. (2016) The crystal structure of the major pneumococcal autolysin LytA in complex with a large peptidoglycan fragment reveals the pivotal role of glycans for lytic activity. Mol Microbiol 101:954-67
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Lee, Mijoon; Hesek, Dusan; Blázquez, Blas et al. (2015) Catalytic spectrum of the penicillin-binding protein 4 of Pseudomonas aeruginosa, a nexus for the induction of β-lactam antibiotic resistance. J Am Chem Soc 137:190-200
Rico-Lastres, Palma; Díez-Martínez, Roberto; Iglesias-Bexiga, Manuel et al. (2015) Substrate recognition and catalysis by LytB, a pneumococcal peptidoglycan hydrolase involved in virulence. Sci Rep 5:16198
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Artola-Recolons, Cecilia; Lee, Mijoon; Bernardo-García, Noelia et al. (2014) Structure and cell wall cleavage by modular lytic transglycosylase MltC of Escherichia coli. ACS Chem Biol 9:2058-66
Martínez-Caballero, Siseth; Lee, Mijoon; Artola-Recolons, Cecilia et al. (2013) Reaction products and the X-ray structure of AmpDh2, a virulence determinant of Pseudomonas aeruginosa. J Am Chem Soc 135:10318-21
Zhang, Weilie; Lee, Mijoon; Hesek, Dusan et al. (2013) Reactions of the three AmpD enzymes of Pseudomonas aeruginosa. J Am Chem Soc 135:4950-3
Johnson, Jarrod W; Fisher, Jed F; Mobashery, Shahriar (2013) Bacterial cell-wall recycling. Ann N Y Acad Sci 1277:54-75

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