A Kinetic Platform to Determine the Fate of Hydrogen Peroxide in Escherichia coli

Hydrogen peroxide (H(2)O(2)) is used by phagocytic cells of the innate immune response to kill engulfed bacteria. H(2)O(2) diffuses freely into bacteria, where it can wreak havoc on sensitive biomolecules if it is not rapidly detoxified. Accordingly, bacteria have evolved numerous systems to defend themselves against H(2)O(2), and the importance of these systems to pathogenesis has been substantiated by the many bacteria that require them to establish or sustain infections. The kinetic competition for H(2)O(2) within bacteria is complex, which suggests that quantitative models will improve interpretation and prediction of network behavior. To date, such models have been of limited scope, and this inspired us to construct a quantitative, systems-level model of H(2)O(2) detoxification in Escherichia coli that includes detoxification enzymes, H(2)O(2)-dependent transcriptional regulation, enzyme degradation, the Fenton reaction and damage caused by •OH, oxidation of biomolecules by H(2)O(2), and repair processes. After using an iterative computational and experimental procedure to train the model, we leveraged it to predict how H(2)O(2) detoxification would change in response to an environmental perturbation that pathogens encounter within host phagosomes, carbon source deprivation, which leads to translational inhibition and limited availability of NADH. We found that the model accurately predicted that NADH depletion would delay clearance at low H(2)O(2) concentrations and that detoxification at higher concentrations would resemble that of carbon-replete conditions. These results suggest that protein synthesis during bolus H(2)O(2) stress does not affect clearance dynamics and that access to catabolites only matters at low H(2)O(2) concentrations. We anticipate that this model will serve as a computational tool for the quantitative exploration and dissection of oxidative stress in bacteria, and that the model and methods used to develop it will provide important templates for the generation of comparable models for other bacterial species.