Metabolic engineering of Komagataella phaffii for enhanced 3-hydroxypropionic acid (3-HP) production from methanol DOI Creative Commons
Sílvia Àvila-Cabré, Joan Albiol, Pau Ferrer

и другие.

Journal of Biological Engineering, Год журнала: 2025, Номер 19(1)

Опубликована: Фев. 20, 2025

Bioconversion of methanol derived from CO2 reduction into value-added chemicals provides a unique approach for mitigating global warming and reducing fossil fuels dependence. Production 3-hydroxypropionic acid (3-HP), key building block the development biobased products such as acrylates 1,3-propanediol, has been successfully achieved using sole carbon energy source in methylotrophic yeast Komagataella phaffii (syn. Pichia pastoris). However, challenges remain meeting commercially relevant concentrations, yields productivities 3-HP, prompting further strain optimization. In present study, we have combined metabolic engineering strategies aiming at increasing precursors supply redirecting flux towards 3-HP production. A combinatorial strategy targeting export was applied to original producing K. harboring synthetic β-alanine pathway mutated NADP-dependent formate dehydrogenase Pseudomonas sp. 101 (PseFDH(V9)). To do so, several genes encoding enzymes catalyzing reactions immediately upstream were overexpressed enhance availability. only overexpression pyruvate carboxylase PYC2 gene significantly increased yield on biomass (YP/X) small-scale cultivations. Co-overexpression lactate permeases ESBP6 JEN1 led 55% improvement titer product deep-well plate cultures compared reference strain, mostly due Esbp6 activity, proving its effectiveness transporter. Deletion native FDH1 did not increase entering assimilatory pathway. Instead, knockout strains showed severe growth defects toxic intermediates accumulation. Co-expression PseFDH(V9) these failed compensate loss FDH. The combining PYC2, ESBP6, tested fed-batch pH 5, achieving concentration 27.0 g l- 1, with 0.19 g- volumetric productivity 0.56 1 h- feeding phase These results represent 42% final over 20% 3-HP-producing strain. Furthermore, bioreactor-scale cultivations 3.5 revealed robustness overexpressing monocarboxylate transporters. Our point out potential transporters efficiently drive phaffii, leading higher titers, yields, productivities, even lower conditions.

Язык: Английский

Metabolic engineering of Komagataella phaffii for enhanced 3-hydroxypropionic acid (3-HP) production from methanol DOI Creative Commons
Sílvia Àvila-Cabré, Joan Albiol, Pau Ferrer

и другие.

Journal of Biological Engineering, Год журнала: 2025, Номер 19(1)

Опубликована: Фев. 20, 2025

Bioconversion of methanol derived from CO2 reduction into value-added chemicals provides a unique approach for mitigating global warming and reducing fossil fuels dependence. Production 3-hydroxypropionic acid (3-HP), key building block the development biobased products such as acrylates 1,3-propanediol, has been successfully achieved using sole carbon energy source in methylotrophic yeast Komagataella phaffii (syn. Pichia pastoris). However, challenges remain meeting commercially relevant concentrations, yields productivities 3-HP, prompting further strain optimization. In present study, we have combined metabolic engineering strategies aiming at increasing precursors supply redirecting flux towards 3-HP production. A combinatorial strategy targeting export was applied to original producing K. harboring synthetic β-alanine pathway mutated NADP-dependent formate dehydrogenase Pseudomonas sp. 101 (PseFDH(V9)). To do so, several genes encoding enzymes catalyzing reactions immediately upstream were overexpressed enhance availability. only overexpression pyruvate carboxylase PYC2 gene significantly increased yield on biomass (YP/X) small-scale cultivations. Co-overexpression lactate permeases ESBP6 JEN1 led 55% improvement titer product deep-well plate cultures compared reference strain, mostly due Esbp6 activity, proving its effectiveness transporter. Deletion native FDH1 did not increase entering assimilatory pathway. Instead, knockout strains showed severe growth defects toxic intermediates accumulation. Co-expression PseFDH(V9) these failed compensate loss FDH. The combining PYC2, ESBP6, tested fed-batch pH 5, achieving concentration 27.0 g l- 1, with 0.19 g- volumetric productivity 0.56 1 h- feeding phase These results represent 42% final over 20% 3-HP-producing strain. Furthermore, bioreactor-scale cultivations 3.5 revealed robustness overexpressing monocarboxylate transporters. Our point out potential transporters efficiently drive phaffii, leading higher titers, yields, productivities, even lower conditions.

Язык: Английский

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