UCLA

The peptidoglycan (PG) cell wall is the ultimate determinant of bacterial cell morphology and the target of many different antibiotics. Its construction is a complex process involving a large number of enzymatic reactions spanning multiple subcellular compartments. Herein, the spatial and temporal organization of cell wall synthesis is examined in the gram-negative bacterium Caulobacter crescentus. Mutants of MreB, a morphogenetic protein and homolog of eukaryotic actin, possess cell shape abnormalities and cell wall defects. It was observed that MreB forms dynamic localization patterns similar to PG biosynthetic enzymes and insertion patterns. It is believed that cell wall breakdown and requisite insertion is controlled by a complex of enzymes directed by morphogenetic proteins. This dissertation describes interactions within this complex that occur between morphogenetic and PG synthetic enzymes located in the cytosol, membrane and periplasm. We discovered and confirmed protein-protein interactions that may be relevant as targets of rational drug discovery initiatives. We translated a critical protein-protein interaction into an assay amenable for high-throughput screening (HTS) of small molecule inhibitors. After careful optimization and validation we screened more than 77,000 unique molecules that yielded three lead compounds for further investigation. This research presented shows how new biological insights into an essential microbial pathway can potentially lead to new methods and targets for antibiotic drug discovery.