Functional overexpression of genes involved in erythritol synthesis in the yeast Yarrowia lipolytica

Aleksandra M. Mirończuk, Anna Biegalska, Adam Dobrowolski

Erythritol, a four-carbon polyol synthesized by microorganisms as an osmoprotectant, is a natural sweetener produced on an industrial scale for decades. Despite the fact that the yeast Yarrowia lipolytica has been reported since the 1970s as an erythritol producer, the metabolic pathway of this polyol has never been characterized. It was shown that erythritol synthesis in yeast occurs via the pentose phosphate pathway (PPP). The oleaginous yeast Y. lipolytica is a good host for converting inexpensive glycerol into a value-added product such as erythritol. Glycerol is a renewable feedstock which is produced on a large scale as a waste product by many branches of industry. In this study, we functionally overexpressed four genes involved in the pentose phosphate pathway (PPP): gene YALI0E06479g encoding transketolase (TKL1), gene YALI0F15587g encoding transaldolase (TAL1), gene YALI0E22649g encoding glucose-6-phosphate dehydrogenase (ZWF1), and gene YALI0B15598g encoding 6-phosphogluconate dehydrogenase (GND1). Here, we show that the crucial gene for erythritol synthesis in Y. lipolytica is transketolase. Overexpression of this gene results in a twofold improvement in erythritol synthesis during a shake-flask experiment (58 g/L). Moreover, overexpression of TKL1 allows for efficient production of erythritol independently from the supplied dissolved oxygen. Fermentation conducted in a 5-L bioreactor at low agitation results in almost 70% higher titer of erythritol over the control strain. This work presents the importance of the PPP in erythritol synthesis and the feasibility for economic production of erythritol from glycerol by the yeast Y. lipolytica.