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Please use this identifier to cite or link to this item: http://arks.princeton.edu/ark:/88435/dsp015x21th651
 Title: GENETIC ANALYSIS OF CELL ENVELOPE BIOGENESIS IN ESCHERICHIA COLI Authors: Sutterlin, Holly Ann Advisors: Silhavy, Thomas J Contributors: Molecular Biology Department Keywords: cell envelopeEscherichia colilipopolysaccharideouter membrane Subjects: Molecular biologyMicrobiologyGenetics Issue Date: 2014 Publisher: Princeton, NJ : Princeton University Abstract: The Escherichia coli cell envelope is a multi-layered structure that serves as a selective permeability barrier, permitting influx of nutrients, preventing entry of potentially toxic molecules, and allowing efflux of waste products. The cell envelope consists of an inner and outer membrane that delimit the aqueous periplasm, which houses the peptidoglycan cell wall. The lipids, proteins, glycan chains, and peptides that make up the cell envelope structures are all transported and assembled by discrete pathways that are tightly coordinated with one another. In this thesis, we focus on understanding the assembly and function of the LptDE subcomplex responsible for assembling the glycolipid lipopolysaccharide in the outer leaflet of the outer membrane. Through the use of bacterial genetics and biochemistry, we have isolated and characterized lptD mutants that affect assembly and/or function of LptD. Our work provides insight into the robust interaction between LptD and LptE, which is necessary for proper assembly and function of the subcomplex. We also shed light on how LptD interacts with LptA to complete the Lpt transenvelope bridge as well as how it interacts with LPS, suggesting a model for LPS transport and assembly. In addition to learning about the Lpt pathway, lptD mutants reported here have contributed to our knowledge of other cell envelope processes as well. We present results that show BamD can bind to substrate, through the use of an LptD mutant substrate. Using a different lptD mutation, we show that accumulation of phosphatidic acid in the outer membrane alters the permeability barrier and increases resistance to vancomycin. Finally, through the analysis of a dominant mutation in the retrograde phospholipid transport pathway that was originally identified as a suppressor of an lptD mutation, we demonstrate that the outer membrane physically contributes to cell integrity to prevent lysis when the local peptidoglycan structure is weakened. Taken together, the results presented here illustrate that cell envelope biogenesis is a highly complex process that involves an elegant interplay between all of the pathways that contribute to its assembly. URI: http://arks.princeton.edu/ark:/88435/dsp015x21th651 Alternate format: The Mudd Manuscript Library retains one bound copy of each dissertation. Search for these copies in the library's main catalog Type of Material: Academic dissertations (Ph.D.) Language: en Appears in Collections: Molecular Biology

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