Effect of fadR gene knockout on the metabolism of Escherichia coli based on analyses of protein expressions, enzyme activities and intracellular metabolite concentrations

Abstract

The metabolic effect of fadR gene knockout on Escherichia coli was investigated based on comparative analyses of protein expressions, enzyme activities and intracellular metabolite concentrations between the parent strain Escherichia coli BW25113 and its fadR knockout mutant JWK1176. The E. coli strains were grown in a minimal medium using glucose as the carbon source. Two-dimensional gel electrophoresis and MALDI-TOF mass spectrometry were used to detect and identify the proteins expressed. Enzyme activities and intracellular metabolite concentrations were also measured. Results showed that the fadR knockout E. coli reduced acetate excretion by 27.3%, accompanied by 8.5% enhancement in biomass yield. Expressions of proteins involved in glucose transport, energy metabolism and some of amino acid biosynthesis pathways were upregulated, whereas those for fatty acids biosynthesis, such as AccB and FabD were downregulated. Besides, UspA and DnaK, the universal stress and heat shock proteins, were induced in response to fadR knockout. There was 3.7- and 1.9-fold increase in the enzyme activities of isocitrate lyase and malate synthase, respectively, in the fadR mutant compared to the parent. Intracellular concentrations of acetyl coenzyme A, pyruvate and phosphoenolpyruvate deceased, whereas the concentrations of isocitrate, α-ketoglutarate, malate, oxaloacetate and aspartate increased. These results suggest that the fadR knockout E. coli activates the glyoxylate shunt and enhances the capacity of energy metabolism and biosynthesis, which leads to the reduction of acetate excretion and improvement of the biomass yield.