| Summary: | Abstract Energy shortage and environmental concern urgently require establishing the feasible bio-refinery process from various feedstocks. The methylotrophic yeast Ogataea polymorpha is thermo-tolerant and can utilize various carbon sources, such as glucose, xylose and methanol, which makes it a promising host for bio-manufacturing. Here, we explored the capacity of O. polymorpha for overproduction of free fatty acids (FFAs) from multiple substrates. The engineered yeast produced 674 mg/L FFA from 20 g/L glucose in shake flask and could sequentially utilize the mixture of glucose and xylose. However, the FFA producing strain failed to survive in sole methanol and supplementing co-substrate xylose promoted methanol metabolism. A synergistic utilization of xylose and methanol was observed in the FFA producing strain. Finally, a mixture of glucose, xylose and methanol was evaluated for FFA production (1.2 g/L). This study showed that O. polymorpha is an ideal host for chemical production from various carbon sources. Graphical Abstract
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