(766b) Combinatorial and Evolutionary Engineering of Pentose Sugar Catabolism in Saccharomyces Cerevisiae for Efficient Biofuels Production

Biofuels production from lignocellulosic biomass can be both sustainable and economical when all available carbon sources are completely utilized. Pentose sugars, such as xylose and arabinose, constitute significant portion of lignocellulosic biomass hydrolysates. However, these sugars are poorly utilized by the yeast Saccharomyces cerevisiae despite decades of research. Here, we discuss the advantages of both combinatorial and evolutionary engineering approaches for the improvement of pentose sugar utilization. Specifically, we discuss the directed evolution of a xylose isomerase-based pathway to achieve a 61 fold improvement in growth rate and 8 fold improvement in ethanol production from xylose. We next discuss combinatorial and evolutionary engineering efforts to establish functional pathways. In particular, key catabolic enzymes were either randomly mutated from a previously reported enzyme or newly obtained from a pentose sugar utilizing fungi. We show that alternative pentose sugar catabolic pathways in S. cerevisiae can perform better compared with previously reported pentose sugar pathways. Moreover, key design principles for catabolic enzyme pathways can be extracted from these studies. These strains can be further improved through adaptive or evolutionary engineering efforts. Finally, these studies describe an effective combinatorial engineering strategy to develop efficient heterologous pathway in S. cerevisiae.