Date of Award
Spring 2014
Document Type
Restricted Thesis
Terms of Use
© 2014 Ted Goh. All rights reserved. Access to this work is restricted to users within the Swarthmore College network and may only be used for non-commercial, educational, and research purposes. Sharing with users outside of the Swarthmore College network is expressly prohibited. For all other uses, including reproduction and distribution, please contact the copyright holder.
Degree Name
Bachelor of Arts
Department
Biology Department
First Advisor
Amy Cheng Vollmer
Abstract
The PhoQ/PhoP signaling system in Escherichia coli senses low magenesium, low pH, and the presence of cationic microbial peptides in the periplasm and responds to these stresses by regulating genes important for growth in these conditions, as well as genes important for virulence in pathogenic strains. The sensor kinase PhoQ modulates the phosphorylation state of its cognate response regulator PhoP, thus activating or deactivating its ability for gene regulation. MgrB is a 47 amino acid protein that has been found to repress PhoQ kinase activity by negative feedback, thus decreasing PhoP-regulated transcription; however, its repressive mechanism remains largely unclear. Unfortunately, due to its small size (5 kDa) and hydrophobic surfaces in the inner membrane environment, protein purification and common structural prediction approaches like x-ray crystallography are difficult to perform. In this study, in order to identify critical residues for its structure and function, alanine scanning site-directed mutagenesis was performed on the periplasmic domain of MgrB (25-47), previously predicted to be important for direct PhoQ interaction. Transcriptional reporter assays measuring the modulation of PhoP-regulated transcription and fluorescence measurements of GFP-MgrB were performed in order the assess the effect of each mutation on MgrB activity. Interestingly, in agreement with previous studies, two cysteines were found to be crucial and likely to form intra- or intermolecular disulfide bonds. In addition, several models have been proposed to characterize various facets of the MgrB-PhoQ interaction.
Recommended Citation
Goh, Ted , '14, "Identification of critical residues in the periplasmic domain of MgrB in the PhoQ/PhoP signaling system of Escherichia coli" (2014). Senior Theses, Projects, and Awards. 131.
https://works.swarthmore.edu/theses/131