Solids NMR Characterization of Bacterial Cell Walls

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This project is designed to continue the development of solids state NMR as a tool for characterization of bacterial peptidoglycan, specifically the peptidoglycan of B. subtilis. Both whole cells and cell walls of bacteria grown in defined media containing 13C and 15N -labeled amino acids will be examined by solids state NMR for the identification and quantitative analysis of cell wall substructures. Gas chromatograph-mass spectroscopy techniques will be developed to complement the NMR data and will allow complete analyses in the presence of label scrambling. The bacterial cell wall is vital for the existence of these organisms in the environment and its metabolism represents a major focus for attack by antibiotics. Recent NMR experiments aimed at developing a new analytical technique for determination of cell-wall cross-linking were carried out on the bacterium A. viridans. This bacterium transports both labeled alanine and lysine readily and incorporates them into the cell wall with no scrambling. NMR can then be used to measure directly the cross-link density in the bacterial cell wall. An important step in the development of solids state NMR as a tool for the characterization of peptidoglycan substructures in bacterial cell walls is an extension of the technique to bacteria which are more commonly studied, e.g. B. Subtilis.***//