Recruiting alternative glucose utilization pathways for improving succinate production.

Abstract

The phosphoenolpyruvate (PEP): carbohydrate phosphotransferase system (PTS) of Escherichia coli was usually inactivated to increase PEP supply for succinate production. However, cell growth and glucose utilization rate decreased significantly with PTS inactivation. In this work, two glucose transport proteins and two glucokinases (Glk) from E. coli and Zymomonas mobilis were recruited in PTS(-) strains, and their impacts on glucose utilization and succinate production were compared. All PTS(-) strains recruiting Z. mobilis glucose facilitator Glf had higher glucose utilization rates than PTS(-) strains using E. coli galactose permease (GalP), which was suggested to be caused by higher glucose transport velocity and lower energetic cost of Glf. The highest rate obtained by combinatorial modulation of glf and glk ( E. coli ) (2.13 g/L•h) was 81 % higher than the wild-type E. coli and 30 % higher than the highest rate obtained by combinatorial modulation of galP and glk ( E. coli ). On the other hand, although glucokinase activities increased after replacing E. coli Glk with isoenzyme of Z. mobilis, glucose utilization rate decreased to 0.58 g/L•h, which was assumed due to tight regulation of Z. mobilis Glk by energy status of the cells. For succinate production, using GalP led to a 20 % increase in succinate productivity, while recruiting Glf led to a 41 % increase. These efficient alternative glucose utilization pathways obtained in this work can also be used for production of many other PEP-derived chemicals, such as malate, fumarate, and aromatic compounds.