Enhancing precursor supply and modulating metabolism to achieve high-level production of β-farnesene in Yarrowia lipolytica

Bi, Haoran; Xu, Chenchen; Bao, Yufei; Zhang, Changwei; Wang, Kai; Zhang, Yang; Wang, Meng; Chen, Biqiang; Fang, Yunming; Tan, Tianwei

doi:10.1016/j.biortech.2023.129171

Cited by 18 publications

(13 citation statements)

References 46 publications

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“…In contrast, the introduction of phosphoketolase bypass, which has been shown to be largely beneficial for enhancing the production of metabolites downstream of acetyl-CoA in various studies (15–18, 20, 21), led to decreased production, yield, and content of beta-carotene in our study. In three studies that have utilized the same set of genes for the phosphoketolase bypass as we did, the authors have observed positive impact on the production of acetate (17) and lipids (15), and no impact on the production of alpha-humulene (19).…”

Section: Resultscontrasting

confidence: 78%

“…Regarding the modifications involving the carnitine shuttle in Y. lipolytica , previous studies done on the W29 derived strains have reported positive effects on the production of lipids (14), beta-farnesene (20), and trans-nerolidol (21). However, Guo and coworkers (19) have reported no effect of ScCAT2 gene overexpression on alpha-humulene production.…”

Section: Resultsmentioning

confidence: 99%

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Impact of central carbon metabolism bypasses on the production of beta-carotene inYarrowia lipolytica

Markuš,

Soldat,

Magdevska

et al. 2023

Preprint

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Yarrowia lipolyticais an oleaginous yeast with ever growing popularity in the metabolic engineering circles. It is well known for its ability to accommodate a high carbon flux through acetyl-CoA and is being extensively studied for production of chemicals derived from it.We investigated the effects of modifying the upstream metabolism leading to acetyl-CoA on beta-carotene production, including its titer, yield, and content. We examined the pyruvate and the phosphoketolase bypass, both of which are stoichiometrically favorable for the production of acetyl-CoA and beta-carotene. Additionally, we examined a set of genes involved in the carnitine shuttle. We constructed a set of parental strains derived from theY. lipolyticaYB-392 wild-type strain, each with a different capacity for beta-carotene production, and introduced genes for the metabolic bypasses in each of the constructed parental strains. Subsequently, we subjected these constructed strains to a series of fermentation experiments.We discovered that altering the upstream metabolism in most cases led to a decrease in performance for production of beta-carotene. Most notably, a set of genes used for the pyruvate bypass (YlPDC2,YlALD5, andYlACS1) and the phosphoketolase bypass (LmXPKandCkPTA) resulted in the reduction of more than 30%.Our findings contribute to our understanding ofY. lipolytica’s metabolic capacity and suggest that production of beta-carotene is most likely not limited solely by the acetyl-CoA supply. We also highlight a complex nature of engineeringY. lipolytica, as most of the results from studies using a different strain background did not align with our findings.

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Section: Resultscontrasting

confidence: 78%

Section: Resultsmentioning

confidence: 99%

Impact of central carbon metabolism bypasses on the production of beta-carotene inYarrowia lipolytica

Markuš,

Soldat,

Magdevska

et al. 2023

Preprint

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“…In addition, other researchers have also synthesized β-farnesene using Y. lipolytica as the host cell. Bi et al achieved a fermentation titer of 28.9 g/L β-farnesene in Y. lipolytica by modulating the regeneration of NAD(P)H, the supply of precursors and ATP [53]. Liu et al achieved a high yield of β-farnesene (31.9 g/L) using waste edible oil as a carbon source by genetically engineering the lipid synthesis pathway and combining β-farnesene synthase engineering [54].…”

Section: Microbial Synthesis Of Sesquiterpenesmentioning

confidence: 99%

Biosynthesis Progress of High-Energy-Density Liquid Fuels Derived from Terpenes

Liu,

Lin,

Han

et al. 2024

Microorganisms

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High-energy-density liquid fuels (HED fuels) are essential for volume-limited aerospace vehicles and could serve as energetic additives for conventional fuels. Terpene-derived HED biofuel is an important research field for green fuel synthesis. The direct extraction of terpenes from natural plants is environmentally unfriendly and costly. Designing efficient synthetic pathways in microorganisms to achieve high yields of terpenes shows great potential for the application of terpene-derived fuels. This review provides an overview of the current research progress of terpene-derived HED fuels, surveying terpene fuel properties and the current status of biosynthesis. Additionally, we systematically summarize the engineering strategies for biosynthesizing terpenes, including mining and engineering terpene synthases, optimizing metabolic pathways and cell-level optimization, such as the subcellular localization of terpene synthesis and adaptive evolution. This article will be helpful in providing insight into better developing terpene-derived HED fuels.

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“…However, the successful implementation of heterologous pathways and perturbation of native microbial metabolism often requires extensive optimization to ensure economically viable production titers. Metabolic engineering allows rewiring of cellular metabolism to increase product titers by enhancing precursor supply 3 , 4 , tackling co-factor limitations 5 – 7 , interrupting competitive pathways 8 , 9 , preventing buildup of toxic intermediates 10 and improving heterologous pathway flux 11 . Over the past years, several tools and strategies have been developed to facilitate yield optimization, including chassis strain improvement 12 – 14 , directed mutagenesis 15 – 17 , synthetic compartmentalization 18 , 19 and pathway gene expression optimization 20 .…”

Section: Introductionmentioning

confidence: 99%

Combinatorial optimization of gene expression through recombinase-mediated promoter and terminator shuffling in yeast

Cautereels,

Smets,

Bircham

et al. 2024

Nat Commun

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Microbes are increasingly employed as cell factories to produce biomolecules. This often involves the expression of complex heterologous biosynthesis pathways in host strains. Achieving maximal product yields and avoiding build-up of (toxic) intermediates requires balanced expression of every pathway gene. However, despite progress in metabolic modeling, the optimization of gene expression still heavily relies on trial-and-error. Here, we report an approach for in vivo, multiplexed Gene Expression Modification by LoxPsym-Cre Recombination (GEMbLeR). GEMbLeR exploits orthogonal LoxPsym sites to independently shuffle promoter and terminator modules at distinct genomic loci. This approach facilitates creation of large strain libraries, in which expression of every pathway gene ranges over 120-fold and each strain harbors a unique expression profile. When applied to the biosynthetic pathway of astaxanthin, an industrially relevant antioxidant, a single round of GEMbLeR improved pathway flux and doubled production titers. Together, this shows that GEMbLeR allows rapid and efficient gene expression optimization in heterologous biosynthetic pathways, offering possibilities for enhancing the performance of microbial cell factories.

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Enhancing precursor supply and modulating metabolism to achieve high-level production of β-farnesene in Yarrowia lipolytica

Cited by 18 publications

References 46 publications

Impact of central carbon metabolism bypasses on the production of beta-carotene inYarrowia lipolytica

Impact of central carbon metabolism bypasses on the production of beta-carotene inYarrowia lipolytica

Biosynthesis Progress of High-Energy-Density Liquid Fuels Derived from Terpenes

Combinatorial optimization of gene expression through recombinase-mediated promoter and terminator shuffling in yeast

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