Engineering xylose fermentation in an industrial yeast: continuous cultivation as a tool for selecting improved strains

Production of second-generation ethanol from lignocellulosic residues should be fueling the energy matrix in the near future. Lignocellulosic biomass has received considerable attention as an alternativ e rene w able resource to w ard reducing the demand for fossil energy sources, contributing to a future sustainable bio-based economy. Fermentation of lignocellulosic h y droly sates poses many scientific and technological challenges as the drawback of Saccharom y ces cere visiae's inability in fermenting pentose sugars (deriv ed from hemicellulose). To o v ercome the inability of S. cerevisiae to ferment xylose and increase yeast robustness in the presence of inhibitory compound-containing media, the industrial S. cerevisiae strain SA-1 was engineered using CRISPR-Cas9 with the oxidoreductive xylose pathway from Scheffersomyces stipitis (encoded by XYL1 , XYL2 , and XYL3 ). The engineered strain was then cultivated in a xylose-limited chemostat under increasing dilution rates (for 64 da y s) to impro v e its xylose consumption kinetics under aerobic conditions. T he e v olv ed strain (DPY06) and its parental strain (SA-1 XR/XDH) w ere e v aluated under microaerobic in a hemicellulosic h y droly sate-based medium. DPY06 exhibited 35% higher volumetric ethanol productivity compared to its parental strain.