Improving xylose consumption in Rhodotorula toruloides through heterologous expression of xylose reductase and xylulokinase

The oleaginous yeast Rhodotorula toruloides is a promising host for sustainable bioproduction due to its capacity to naturally utilize xylose present in lignocellulosic biomass, an abundant and renewable resource. However, its xylose consumption pathway is still not completely understood. To better understand the potential limitations in xylose utilization in R. toruloides, heterologous xylose reductase from Scheffersomyces stipitis, together with the native and heterologous xylulokinases from three different microorganisms (Scheffersomyces stipitis, Candida intermedia, and Escherichia coli) were overexpressed solely and in combination. The overexpression of xylulokinases showed more significant improvements in terms of xylose consumption rate compared to the single overexpression of xylose reductase. When the heterologous xylulokinase from Escherichia coli was overexpressed, the specific xylose consumption rate was improved by 66% and the maximum specific growth rate by 30% compared to the parental strain. The xylose specific consumption rate increased by 146% and the maximum specific growth rate increased by 118% when heterologous genes for xylose reductase and xylulokinase from E. coli were overexpressed together. These results suggest that the low expression of xylulokinase in R. toruloides, which has been reported previously, could limit its sugar consumption, while supporting higher lipid accumulation in this yeast.