Metabolic engineering of yeasts for production of bulk fermentation products from xylose: ethanol and lactic acid

Kluyveromyces marxianus, a yeast naturally assimilating but not fermenting xylose, was genetically engineered to produce ethanol from xylose efficiently. Genetic engineering included replacing the natural xylose utilization pathway via xylose reductase and xylitol dehydrogenase to xylulose by a fungal xylose isomerase converting xylose directly to xylulose. Furthermore xylulokinase was overexpressed to improve efficiency of xylose to ethanol fermentation. The resulting strain produced 37 g/L ethanol with a yield of 0.4 g/g xylose used and an ethanol production rate of 0.94 g/L*h in shake flask fermentation tests. The original strain produced xylitol very efficiently in these conditions but did not produce any ethanol. The above strain was further engineered to produce lactic acid from xylose by expressing the L-lactate dehydrogenase (LDH) from Lactobacillus helveticus. The resulting strain produced lactate from xylose in shake flask test conditions with 77% yield and 1 g/L*h productivity. Lactobacillus helveticus LDH was also introduced to Pichia stipitis, a yeast naturally fermenting xylose. Lactate was the main product from xylose in shake flask test and was produced with about 60% yield and 0.4 g/L*h productivity.