7-05: Metabolic engineering of oleaginous bacteria for lignocellulose biomass-based production of biofuel

Lignocellulosic biomass feedstocks composed of cellulose, hemicellulose, and lignin are the most abundant renewable resources in the world. Both hexoses from cellulose and pentoses including D-xylose and L-arabinose from hemicellulose are released during the hydrolysis. Other than the sugars, the chemicals such as furfural and hydroxymethylfurfural (HMF), which strongly inhibit the growth and fermentation of microorganisms, are also formed in the hydrolysates. Thus, development of the strains with the capability of full utilization of sugars including pentose and tolerance to the inhibitors will be the critical issue for production of fuels from lignocellulose. In this study, the high lipid producing oleaginous strain, Rhodococcus opacus PD630 (DSMZ 44193) was metabolically engineered for utilization of the renewable resources. The metabolic pathways of D-xylose and L-arabinose were engineered into the strain by heterologous expression of the genes involved in pentose utilization. The recombinants were able to utilize pentose effectively, while accumulating lipids to more than 60% of cell dry weight (CDW) under nitrogen-limiting conditions. The lipid can be used as feedstock to be further processed into “drop-in” fuel.  In order to remove the inhibitors from the hydrolysates, the gene clusters responsible for furfural and HMF degradation from Bradyrhizobium japonicum USDA 110 were introduced into the substrate broadened hosts. The resulting recombinants could utilize furfural and HMF as sole carbon sources and production of lipids by the recombinants is being studied. Our study generates the robust strains for lipid production and also sheds the light on the detoxification of the lignocellulosic hydrolysates in situ.