Metabolic engineering of Escherichia coli for the enhanced production of l-tyrosine.

Abstract

Despite wide applications of l-tyrosine in the market, microbial overproduction of l-tyrosine has been a great challenge due to the complex gene regulations involved in its biosynthetic pathway. To this end, effects of knocking out tyrR on the l-tyrosine production were further explored during the strain development. Also, blocking cellular uptake of l-tyrosine by knocking out tyrosine transporters was examined with respect to l-tyrosine production. Using feedback-resistant aroG and tyrA genes (aroGfbr and tyrAfbr hereafter) as initial overexpression targets, which encode 3-deoxy-7-phosphoheptulonate synthase and chorismate mutase or prephenate dehydrogenase, respectively, various combinations of genes were subsequently overexpressed in the Escherichia coli wild-type and tyrR knockout strain, and their effects on the l-tyrosine production were examined. Co-overexpression of aroGfbr , aroL and tyrC, a gene from Zymomonas mobilis functionally similar to tyrAfbr , but insensitive to l-tyrosine, led to the greatest l-tyrosine production regardless of the strains and plasmid constructs examined in this study. The strain BTY2.13 overexpressing the abovementioned three genes together with the removal of the l-tyrosine-specific transporter (tyrP) produced 43.14 g/L of l-tyrosine by fed-batch fermentation using the exponential feeding followed by DO-stat feeding method. This outcome suggested that the tyrR gene knockout was not mandatory for the l-tyrosine overproduction, but the production performance of strains having tyrR appeared to be highly affected by vector systems and feeding methods. With an optimal vector system and a feeding method, tyrP knockout appeared to be more effective in enhancing the l-tyrosine than tyrR knockout.