Development of new ethanologenic Escherichia coli strains for fermentationof lignocellulosic biomass

Two new ethanologenic strains (FBR4 and FBR5) of Escherichia coli were cons tructed and used to ferment corn fiber hydrolysate. The strains carry the p lasmid pLOI297, which contains the genes from Zymomonas mobilis necessary f or efficiently converting pyruvate into ethanol. Both strains selectively m aintained the plasmid when grown anaerobically. Each culture was serially t ransferred 10 times in anaerobic culture with sugar-limited medium containi ng xylose, but no selective antibiotic. An average of 93 and 95% of the FBR 4 and FBR5 cells, respectively, maintained pLOI297 in anaerobic culture. Th e fermentation performances of the repeatedly transferred cultures were com pared with those of cultures freshly revived from stock in pH-controlled ba tch fermentations with 10% (w/v) xylose. Fermentation results were similar fur all the cultures. Fermentations were completed within 60 h and ethanol yields were 86-92% of theoretical. Maximal ethanol concentrations were 3.9- 4.2% (w/v). The strains were also tested for their ability to ferment corn fiber hydrolysate, which contained 8.5% (w/v) total sugars (2.0% arabinose, 2.8% glucose, and 3.7% xylose). E. coli FBR5 produced more ethanol than FB R4 from the corn fiber hydrolysate. E. coli FBR5 fermented all but 0.4% (w/ v) of the available sugar, whereas strain FBR4 left 1.6% unconsumed. The fe rmentation with FBR5 was completed within 55 h and yielded 0.46 g of ethano l/g of available sugar, 90% of the maximum obtainable.