Analysis of the Impact of pH on Nitric Oxide Detoxification in Escherichia coli.

Abstract

Nitric oxide (NO•) is an important antimicrobial defense in innate immunity, and the virulence of many pathogens relies on NO• detoxification. Invading pathogens are engulfed by immune cells and subjected to acidic, nitrosative, nutritional, and oxidative stresses. Previous research has suggested an interaction between acidic stress and NO• tolerance which alludes to the possibility for each system to affect the other system’s response. An increased understanding of this interaction could lead to better strategies to impair NO• defenses and reduce the virulence of pathogens. This thesis explores the impact of pH on NO• detoxification in Escherichia coli. The conditions tested required development of a novel method for conducting NO• detoxification assays, which was validated here by ensuring the [NO•] profile was the same under both pH conditions without cells. Using NO• detoxification assays, no significant difference in NO• clearance times for wild-type (WT) E. coli was observed between neutral (7.5) and acidic pH (4.5). Mutants lacking important acid resistance genes were then assayed to assess their involvement in NO• detoxification under acidic conditions. Of all the mutants tested, the imp4213 mutant was suggested to have an impaired NO• detoxification when subjected to acidic stress compared to neutral conditions.