Biotechnology Bulletin ›› 2023, Vol. 39 ›› Issue (8): 137-147.doi: 10.13560/j.cnki.biotech.bull.1985.2023-0138

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Modification of Pichia pastoris for Erythritol Production by Metabolic Engineering

ZHAO Si-jia1(), WANG Xiao-lu2, SUN Ji-lu1, TIAN Jian2(), ZHANG Jie2()   

  1. 1. College of Food Science and Technology, Hebei Agricultural University, Baoding 071000
    2. State Key Laboratory of Animal Nutrition, Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193
  • Received:2023-02-19 Online:2023-08-26 Published:2023-09-05
  • Contact: TIAN Jian, ZHANG Jie E-mail:sijia_zhao@163.com;tianjian@caas.cn;zhangjie09@caas.cn

Abstract:

The aim of this study is to construct an erythritol-producing strain using Pichia pastoris as the chassis cell. An erythritol-producing P. pastoris strain was developed by regulating the expression of phosphofructokinase gene(pfk)in glycolysis pathway, knocking out the genes associated with the production of by-products arabitol and ribitol, and overexpressing 4-phosphate erythrose phosphorylase, erythrose reductase and sugar alcohol phosphatase genes derived from different organisms. Next, we investigated the effects of overexpressions of erythrose reductase and two key enzymes transketolase(TKL)and ribulose-phosphate epimerase(RPE)involved in pentose phosphate pathway on the erythritol production. The results showed that strain C8 harboring pyp1 gene derived from Saccharomyces cerevisiae and yidA gene derived from Escherichia coli had the ability to produce erythritol. The erythritol production of the C8 strain in shake-flask fermentation was 30 mg/L. Furthermore, the overexpression of tkl and rpe genes enhanced the erythritol production of C10 strain by about 40-fold. The erythritol production of C10 reached 1.2 and 10.6 g/L in the shake-flask and high-cell-density fermentations, respectively. Further the overexpression of erythrose reductase did not cause the erythritol production increased, while caused the production of by-products increased. In this study, the erythritol synthetic pathway was successfully constructed in P. pastoris for the first time, which laid a foundation for engineering P. pastoris for efficient production of erythritol and other high-value compounds.

Key words: erythritol, Pichia pastoris, metabolic engineering, fermentation