Enhanced Phenyllactic Acid Production in Escherichia coli Via Oxygen Limitation and Shikimate Pathway Gene Expression

Abstract

3-Phenyllactic acid (PhLA) is useful as a start-up material in the pharmaceutical and biorefinery industries. To enhance the production of PhLA from glucose using recombinant Escherichia coli, the effects of glucose concentration and oxygen limitation on PhLA production are assessed in a fed-batch system using dissolved oxygen (DO)-stat method. The highest titer of PhLA (7.3 g L^-1 ) is observed with a high concentration of glucose and under oxygen-limited conditions (DO = 0 ppm). Under oxygen limitation, cell growth and the formation of acetate and l-phenylalanine (Phe) by-products after 72 h of cultivation are reduced by 30%, 70%, and 81%, respectively, as compared to that under high DO conditions (DO = 5 ppm). Gene expression levels are compared between low and high DO conditions by quantitative polymerase chain reaction (qPCR) analysis. Several genes in the glycolysis (gapA and pykA), pentose phosphate (tktA), and early shikimate pathways for PhLA biosynthesis (aroF, aroG, and aroH) are upregulated under oxygen limitation. The results suggest that oxygen limitation affects metabolism in the shikimate pathway at both metabolic and transcriptional levels and that controlling the DO level is critical for enhanced production of a variety of aromatic compounds through the shikimate pathway.

Keywords: Escherichia coli; gene expression; oxygen limitation; phenyllactic acid; shikimate.