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"Involvement of the YihE protein kinase in the bacterial stress response"

by
Angella M. Dorsey-Oresto
Microbiology and Molecular Genetics Program
B.A. 2004, Drew University, Madison, NJ



Thesis Advisor: Karl Drlica, Ph.D.
Professor
Department of Microbiology & Molecular Genetics
Public Health Research Institute

Wednesday, July 28, 2010
1:00 p.m., ICPH Auditorium


Abstract

Resistance to antimicrobials is an increasing problem that is not being solved by educational efforts or the discovery of new agents. To prolong the efficacy of currently available antimicrobials, experiments were designed to identify key components of bacterial stress response pathways that might serve as potential targets for antimicrobial-enhancing agents. The present work represents an initial characterization of a newly identified stress response gene, yihE. Loss of functional yihE resulted in hyperlethal susceptibility of Escherichia coli to a variety of stressors including nalidixic acid, kanamycin, mitomycin C, hydrogen peroxide, and UV-irradiation. The hyperlethal activity of these stressors was reversed by co-deletion of the mazEF toxin-antitoxin module. When YihE was expressed and purified it was found to be a eukaryotic-like Ser/Thr protein kinase capable of both autophosphorylation and transphosphorylation of protein substrates such as myelin basic protein. The kinase activity of YihE plays a central role in protecting E. coli from a variety of lethal stressors, since mutations in important active-site residues abolished the ability of wild-type yihE to complement a yihE-deficient mutant in trans. It is hypothesized that the YihE protein kinase protects cells during stress by decreasing MazF toxicity, although the effect may be indirect, because YihE did not phosphorylate purified MazE or MazF in vitro. The natural substrate of YihE is still unknown. yihE-mediated hyperlethality during stress probably resulted from decreased accumulation of reactive oxygen species, since the effect of loss of functional yihE was abolished by treatment with sub-inhibitory concentrations of bipyridyl and thiourea, compounds that interfere with stress-mediated accumulation of hydroxyl radical.


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