Enhancing erythritol productivity in Yarrowia lipolytica using metabolic engineering

Abstract

Erythritol (1,2,3,4-butanetetrol) is a four-carbon sugar alcohol with sweetening properties that is used by the agrofood industry as a food additive. In this study, we demonstrated that metabolic engineering can be used to improve the production of erythritol from glycerol in the yeast Yarrowia lipolytica. The best results were obtained using a mutant that overexpressed GUT1 and TKL1, which encode a glycerol kinase and a transketolase, respectively, and in which EYK1, which encodes erythrulose kinase, was disrupted; the latter enzyme is involved in an early step of erythritol catabolism. In this strain, erythritol productivity was 75% higher than in the wild type; furthermore, the culturing time needed to achieve maximum concentration was reduced by 40%. An additional advantage is that the strain was unable to consume the erythritol it had created, further increasing the process's efficiency. The erythritol productivity values we obtained here are among the highest reported thus far.

Homology‐independent genome integration enables rapid library construction for enzyme expression and pathway optimization in Yarrowia lipolytica

“…Yarrowia lipolytica , an unconventional and oleaginous yeast, has attracted an increasing attention in recent years (Darvishi, Ariana, Marella, & Borodina, ; Hussain et al, ; Markham and Alper, ; Xie, ) and has been exploited to produce a variety of products, such as fatty acid derivatives (Tai & Stephanopoulos, ; Xue et al, ), succinic acid (Cui et al, ; C. Gao et al, ), erythritol (Carly et al, ; Mirończuk, Biegalska, & Dobrowolski, ), and lipids (Qiao, Wasylenko, Zhou, Xu, & Stephanopoulos, ). Y. lipolytica is also an excellent host for the production of secondary metabolites such as α‐farnesene (Yang, Nambou, Wei, & Hua, ), lycopene (Matthäus, Ketelhot, Gatter, & Barth, ), and β‐carotene (S. Gao et al, ; Larroude et al, ), because it contains many precursors and cofactors.…”

Erythritol production by yeasts: a snapshot of current knowledge

“…Moreover, cells grown on raw glycerol produced more erythritol as compared with pure glycerol, indicating that the impurities might have a beneficial effect on erythritol production (Kobayashi, Iwata, Mizushima, Ogihara, & Kasumi, ). These results demonstrated raw glycerol to be both a cheap and an efficient alternative to glucose, and it was used successfully in a large number of studies (Carly, Vandermies, et al, ; Mirończuk, Dobrowolski, et al, ; Tomaszewska, Rywińska, & Rymowicz, ).…”

“…The central role of transketolase and ER in erythritol synthesis was further emphasized by the fact that both enzymes have a higher activity level under high osmotic pressure (Tomaszewska, Rakicka, Rymowicz, & Rywińska, ; Yang et al, ). A correlation was also found between transketolase activity and erythritol production, and transketolase overexpression was found to increase both erythritol yield and productivity (Carly et al, ; Sawada, Taki, Yamakawa, & Seki, ). A similar correlation between ER activity and erythritol production was found in Y. lipolytica (Janek et al, ).…”

“…Genetic engineering gave promising results, but more information would be needed for further improvement. Although enzymes from the pentose phosphate pathway are usually well characterized, and ER genes had been identified in several organisms, the gene coding for erythrose-4-phopshate phosphatase has still not been identified in yeast (Carly, Vandermies, et al, 2017b;Deng, Han, Liu, Jia, & Zhou, 2012;Lee, Kim, et al, 2003a).…”

Sugar Alcohols and Organic Acids Synthesis in Yarrowia lipolytica: Where Are We?

“…Other strategies of metabolic engineering consisted of overexpressing key genes of the PPP, namely TKL1 (YALI0E06479g) encoding transketolase, gene TAL1 (YALI0F15587g) encoding transaldolase (Figure 2). In Y. lipolytica strain Po1d, overexpression of TKL1 led to a 19% and 17% increased EOL titer and productivity (43 g/L, 0.04 g/(g DCW •h), respectively) [106]. A similar strategy in the strain MK1, led to 51% and 47% increases (51 g/L, 0.53 g/(L•h), respectively) [107].…”