Metabolic evolution of non-transgenic Escherichia coli SZ420 for enhanced homoethanol fermentation from xylose

Efficient utilization of pentose sugars (xylose and arabinose) is an essential requirement for economically viable ethanol production from cellulosic biomass. The desirable pentose-fermenting ethanologenic biocatalysts are the native microorganisms or the engineered derivatives without recruited exogenous gene(s). We have used a metabolic evolution (adaptive selection) approach to improve a non-transgenic homoethanol Escherichia coli SZ420 (ldhA pflB ackA frdBC pdhR::pflBp6-aceEF-lpd) for xylose fermentation. An improved mutant, E. coli KC01, was evolved through a 3 month metabolic evolution process. This evolved mutant increased pyruvate dehydrogenase activity by 100%, cell growth rate (h(-1)) by 23%, volumetric ethanol productivity by 65% and ethanol tolerance by 200%. These improvements enabled KC01 to complete 50 g xylose l(-1) fermentations with an ethanol titer of 23 g l(-1) and a yield of 90%. The improved cell growth and ethanol production of KC01 are likely attributed to its three fold increased ethanol tolerance.