Abstract
BackgroundProduction of biofuels and green chemicals by microbes is currently of great interest due to the increasingly limited reserves of fossil fuels. Biodiesel, especially fatty acid ethyl esters (FAEEs), is considered as an attractive alternative because of its similarity with petrodiesel and compatibility with existing infrastructures. Cost-efficient bio-production of FAEEs requires a highly lipogenic production host that is suitable for large-scale fermentation. As a non-model oleaginous yeast that can be cultured to an extremely high cell density and accumulate over 70% cell mass as lipids, Rhodotorula toruloides represents an attractive host for FAEEs production.ResultsWe first constructed the FAEE biosynthetic pathways in R. toruloides by introducing various wax ester synthase genes from different sources, and the bifunctional wax ester synthase/acyl-CoA-diacyglycerol acyltransferase (WS/DGAT) gene from Acinetobacter baylyi was successfully expressed, leading to a production of 826 mg/L FAEEs through shake-flask cultivation. We then mutated this bifunctional enzyme to abolish the DGAT activity, and further improved the titer to 1.02 g/L. Finally, to elevate the performance of Δku70-AbWS* in a bioreactor, both batch and fed-batch cultivation strategies were performed. The FAEEs titer, productivity and yield were 4.03 g/L, 69.5 mg/L/h and 57.9 mg/g (mg FAEEs/g glucose) under batch cultivation, and 9.97 g/L, 90.6 mg/L/h, and 86.1 mg/g under fed-batch cultivation. It is worth mentioning that most of the produced FAEEs were secreted out of the cell, which should greatly reduce the cost of downstream processing.ConclusionWe achieved the highest FAEEs production in yeast with a final titer of 9.97 g/L and demonstrated that the engineered R. toruloides has the potential to serve as a platform strain for efficient production of fatty acid-derived molecules.
Highlights
Production of biofuels and green chemicals by microbes is currently of great interest due to the increasingly limited reserves of fossil fuels
Additional file 1: Table S1 summarizes the application of these enzymes in S. cerevisiae and their specific enzyme activities for fatty acid ethyl esters (FAEEs) production
While we have demonstrated that R. toruloides has great potential for large-scale production of biodiesels during this study, a major bottleneck to implement an economically feasible process is the additional cost associated with providing exogenous ethanol
Summary
Production of biofuels and green chemicals by microbes is currently of great interest due to the increasingly limited reserves of fossil fuels. As one of the most promising alternative energy sources, biodiesel has high energy density, and is compatible with current infrastructure [2] It showed several advantages over petrodiesel, such as higher lubricity, and lower tailpipe emissions [3, 4]. Commercial strategies for biodiesel production mainly use plant oils or animal fats as feedstock to produce FAMEs through chemical transesterification [6, 7]. These strategies compete for lands and materials required for food production and cause environmental problems [2, 8]. It is desirable to use renewable plant biomass as a feedstock to produce FAEEs by microbial fermentation [9]
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