Metabolite analysis of Escherichia coli in response to changes in oxygen levels

• Main Author: Yasid, Nur Adeela
• Other Authors: Williamson, Michael Paul
• Published: University of Sheffield 2013
• Subjects: 579.3
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.589192

E. coli is a versatile bacterium that has three metabolic modes: aerobic, anaerobic and fermentation. The major environmental factor that controls the switching between these metabolic modes is oxygen availability. E. coli in the intestinal tract is completely anaerobic but certain parts of the intestinal tract close to the gut are microaerobic. The change from anaerobic to aerobic or microaerobic is very rapid and leads to major metabolic change. The changes have been characterized by using NMR. A quenching technique has been developed by using ethanol/NaCl to cool the cells very rapidly after removal from the chemostat to inactivate the metabolic activity. The measurement error from most sources is small with acceptable error from extraction of intracellular metabolites. The technique is being used to studying the transition from anaerobic to aerobic at different levels of oxygen. Changes in the major intracellular and extracellular metabolites have been measured, where the metabolic activity inside the cells showed not much change. The metabolic profile of pyruvate that goes up and down in response to the switch led to the study of movement of pyruvate inside the cells. Tagging of the pykF gene with GFP and 3xFLAG tag (in particular) indicated that the localisation of the PykF protein changes from cytoplasm to cell membrane. This explained why the concentration of pyruvate is low inside the cells, suggesting that the pyruvate is transported out directly when oxygen is introduced.