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Efficient production of 2′-fucosyllactose in Pichia pastoris through metabolic engineering and constructing an orthogonal energy supply system

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Název: Efficient production of 2′-fucosyllactose in Pichia pastoris through metabolic engineering and constructing an orthogonal energy supply system
Autoři: Yi Li, Xiang Wang, Kaidi Chen, Zhoukang Zhuang, Hongting Tang, Tao Yu, Wenbing Cao
Zdroj: Synth Syst Biotechnol
Synthetic and Systems Biotechnology, Vol 10, Iss 3, Pp 807-815 (2025)
Informace o vydavateli: Elsevier BV, 2025.
Rok vydání: 2025
Témata: Microbial cell factory, QH301-705.5, Original Research Article, Biology (General), Cofactor engineering, Orthogonal energy module, TP248.13-248.65, 2′-FL biosynthesis, Biotechnology
Popis: 2'-fucosyllactose (2'-FL) holds significant role in the infants' nutrition. While microbial production of 2'-FL has predominantly utilized Escherichia coli and Saccharomyces cerevisiae, the potential of Pichia pastoris, renowned for its robust NADPH regeneration capability, remains underexplored. Herein, we systematically engineered the metabolism of P. pastoris to develop an efficient 2'-FL-producing cell factory. We first constructed the de novo biosynthesis pathway for 2'-FL in P. pastoris, achieving an initial titer of 0.143 g/L. By optimizing enzyme selection and solubility of α-1,2-fucosyltransferase (FutC), 2'-FL production was enhanced by nearly ten folds. Subsequently, engineering NADPH supply further increased the 2'-FL production by 170 %. Furthermore, we enhanced energy supply by incorporating an orthogonal energy module based on the methanol dissimilation pathway and increasing GTP availability, resulting in a 32 % improvement in 2'-FL production. Finally, through the optimization of fermentation condition, we realized the production titer of 2'-FL to 3.50 g/L in shake-flask, representing the highest titer in P. pastoris. These findings highlight the potential of P. pastoris as a chassis to produce chemicals by providing abundant NADPH and utilizing methanol as co-substrate to supply sufficient energy.
Druh dokumentu: Article
Other literature type
Jazyk: English
ISSN: 2405-805X
DOI: 10.1016/j.synbio.2025.04.002
Přístupová URL adresa: https://pubmed.ncbi.nlm.nih.gov/40297761
https://doaj.org/article/18da426add6947e38835bd8f22cee215
Rights: CC BY NC ND
Přístupové číslo: edsair.doi.dedup.....ad91c04eaa91e74798439893182b8caf
Databáze: OpenAIRE
Popis
Abstrakt:2'-fucosyllactose (2'-FL) holds significant role in the infants' nutrition. While microbial production of 2'-FL has predominantly utilized Escherichia coli and Saccharomyces cerevisiae, the potential of Pichia pastoris, renowned for its robust NADPH regeneration capability, remains underexplored. Herein, we systematically engineered the metabolism of P. pastoris to develop an efficient 2'-FL-producing cell factory. We first constructed the de novo biosynthesis pathway for 2'-FL in P. pastoris, achieving an initial titer of 0.143 g/L. By optimizing enzyme selection and solubility of α-1,2-fucosyltransferase (FutC), 2'-FL production was enhanced by nearly ten folds. Subsequently, engineering NADPH supply further increased the 2'-FL production by 170 %. Furthermore, we enhanced energy supply by incorporating an orthogonal energy module based on the methanol dissimilation pathway and increasing GTP availability, resulting in a 32 % improvement in 2'-FL production. Finally, through the optimization of fermentation condition, we realized the production titer of 2'-FL to 3.50 g/L in shake-flask, representing the highest titer in P. pastoris. These findings highlight the potential of P. pastoris as a chassis to produce chemicals by providing abundant NADPH and utilizing methanol as co-substrate to supply sufficient energy.
ISSN:2405805X
DOI:10.1016/j.synbio.2025.04.002