Simultaneous utilization of glucose and galactose to produce enantiopure (2R,3R)-butanediol by an engineered Saccharomyces cerevisiae strain

Due to increasing concerns on sustainability, intensive research has been devoted to microbial production of fuels and chemicals from renewable feedstocks such as lignocellulose biomass and marine macroalgae. One such example is the biological production of 2,3-butanediol (BDO), an important chemical with extensive industrial applications. Although many native hosts such as Klebsiella and Enterobacter species can accumulate BDO to high levels, they are pathogenic and synthesize a mixture of BDO stereoisomers, preventing their commercial applications. Here we report the engineering of an industrially friendly host, Saccharomyces cerevisiae, to produce BDO at high titer and yield. By inactivation of pyruvate decarboxylases (PDCs) followed by overexpression of MTH1 and adaptive evolution, the resultant yeast grew on glucose as the sole carbon source with ethanol production completely eliminated. Moreover, the pdc- strain consumed glucose and galactose simultaneously, which to our knowledge is unprecedented in S. cerevisiae strains. Subsequent introduction of a BDO biosynthetic pathway consisting of the cytosolic acetolactate synthase (cytoILV2), Bacillus subtilis acetolactate decarboxylase (BsAlsD), and the butanediol dehydrogenase (BDH1) resulted in the production of enantiopure (2R,3R)-butanediol (R-BDO). In shake flask fermentation, a yield over 70% of the theoretical value was achieved. Using fed-batch fermentation, more than 100 g/L R-BDO was synthesized from a mixture of glucose and galactose, two major components in red algae. The high titer and yield of the enantiopure R-BDO produced as well as the ability to co-ferment glucose and galactose make our engineered yeast strain a superior host for cost-effective production of bio-based BDO from renewable resources.