P114: Enhancing glucose transport in an Escherichia coli strain devoid of the Phosphoenolpyruvate: sugar phosphotransferase system for shikimic acid production

Shikimic acid (SA) is a precursor in chemical synthesis of oseltamivir phosphate, an antiviral drug prescribed in the treatment of several types of influenza including A, B, H5N1 and A/H1N1. SA production by fermentation processes from glucose using recombinant microorganisms is starting to supply commercial demand for this compound. In Escherichia coli, Phosphoenolpyruvate (PEP): sugar phosphotransferase system (PTS) inactivation (PTS¯) increases PEP availability; however µ decreases substantially as compared to a wild type strain. A PTS¯ derivative strain from E. coli JM101 designated as PB12, which is a mutant capable of growing in glucose due to overexpression of genes coding for galactose permease system (galP) and glucokinase (glk), was used as genetic background to obtain a derivative namely PB12.SA22 (PTS¯ glc⁺ aroK¯ aroL¯ aroG^fbr tktA aroB aroE) which produces 7.1 g/L of SA in 0.5 L batch cultures grown in mineral broth supplemented with 25 g/L of glucose and 15 g/L of yeast extract. In this work, the replacement of native promoter region of galP gene with inducible hybrid strong promoter trc in strain PB12.SA22 was evaluated. [¹⁴C]-glucose initial uptake rate was 276.6% higher than in PB12.SA22 strain. µ, q_s and volumetric productivity were improved in 24.1%, 75.8% and 66.7% respectively as well. When grown in 100 g/L of glucose the evaluated strain was able to consume 28% more glucose than PB12.SA22 and produced 12.5 g/L of SA. These results show that contending with glucose transport capacity constraint in PB12.SA22 strain allows improving SA production process.