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 (aroG(fbr) and tyrA(fbr) 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 aroG(fbr), aroL and tyrC, a gene from Zymomonas mobilis functionally similar to tyrA(fbr), 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 43.14g/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.