APPLIED AND ENVIRONMENTAL MICROBIOLOGY
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2024
·Vol. 91
·期 1
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IF 3.9
Yuan-Ru Xiong,Yuan-Chun Fang,Min He,Ke-Jing Li,Lei Qi,Yang Sui,Ke Zhang,Xue-Chang Wu,Liang Meng,Ou Li,Dao-Qiong Zheng
This study explored the genomic alterations in Yarrowia lipolytica, a key yeast in industrial biotechnology, under both spontaneous and mutagen-induced conditions. Our findings reveal that spontaneous mutations occur at a rate of approximately 4 × 10−10 events per base pair per cell division, primarily manifesting as single-nucleotide variations (SNVs) and small insertions and deletions (InDels). Notably, C-to-T/G-to-A transitions and C-to-A/G-to-T transversions dominate the spontaneous SNVs, while 1 bp deletions, likely resulting from template slippage, are the most frequent InDels. Furthermore, chromosomal aneuploidy and rearrangements occur, albeit at a lower frequency. Exposure to ultraviolet (UV) light, methylmethane sulfonate (MMS), and Zeocin significantly enhances the rates of SNVs and alters their mutational spectra in distinct patterns. Notably, Zeocin-induced SNVs are predominantly T-to-A and T-to-G substitutions, often occurring within the 5′-TGT*−3′ motif (* denotes the mutated base). Additionally, Zeocin exhibits a higher potency in stimulating InDels compared to UV and MMS. Translesion DNA synthesis is implicated as the primary mechanism behind most Zeocin-induced SNVs and some InDels, whereas non-homologous end joining serves as the main pathway for Zeocin-mediated InDels. Intriguingly, the study identifies the gene YALI1_E21053g, encoding a protein kinase, as negatively associated with Zeocin resistance. Overall, our results not only deepened our knowledge about the genome evolution in Y. lipolytica but also provided reference to develop innovative strategies to harness its genetic potential.
研究领域:
微生物学
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遗传学
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分子生物学
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高影响因子
Results in Engineering
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2025
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IF 7.9
Wei Kuang,Yuxiang Hong,Zimu Hu,Peng Xu
Liquid C1 feedstock, including methanol, formaldehyde and formate can be easily manufactured from greenhouse gases CO 2 and CH 4 . Yarrowia lipolytica , isolated from marine ecosystem, is a non-conventional yeast which is capable of assimilating multiple range of hydrophobic compounds including alcohols, lipids, hydrocarbons, and volatile fatty acids. This yeast has been extensively engineered as a benchmark host to produce lipids, resveratrol, carotenoids, squalene and other natural products. Connecting C1 feedstock pathways to these important metabolites holds promise to further reduce costs and improve sustainability. Herein we tested the growth fitness of Y. lipolytica when exposed to various one-carbon (C1) liquid feedstocks (methanol, formaldehyde, and formate). We firstly examined the cell growth kinetics with an automatic Cell Growth Analyzer and fitted the data with a Monod-type growth model. We obtained the inhibition constants of the three liquid C1s from the growth fitness dose-response curves. We found that the lagging phase duration is exponentially correlated with the toxicity of the tested C1 solvents. Notably, Y. lipolytica exhibited tolerance to methanol (IC 50 871 mM) and formate (IC 50 42.6 mM), but showed sensitivity to formaldehyde (IC 50 3.8 mM). We further demonstrated that glucose and formate co-substrate could improve cell growth and lipid production. This work broadens Y. lipolytica ’s substrate scope and facilitates sustainable and cost-efficient production of fuels and chemicals from C1 feedstocks.
研究领域:
微生物学
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生物工程
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环境科学
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JOURNAL OF CHROMATOGRAPHY A
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2017
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IF 3.981
Chen Zhao,Yan Yang,Linfang Wei,Qinghua Guo,Hao Fang,Shaolin Chen,Qiang Hua
Yarrowia lipolytica is an oleaginous yeast with promise in producing terpenoids such as lycopene . Though methods for analyzing primary metabolic intermediates have been established, further work is needed to better analyze nucleotides and coenzymes . Here, we presented an optimized method for the separation of nucleotides and coenzymes in Y. lipolytica using the capillary electrophoresis. The separation of twelve metabolites including four coenzymes, five nucleotides and three nucleosides was achieved within 32 min using a voltage of 15 kV and 70 mM sodium carbonate/hydrogencarbonate buffer with 1.0% β -CD at pH 10. The results show that the concentrations of adenosine triphosphate and nicotinamide adenine dinucleotide phosphate changed significantly between lycopene producing strain and the control, indicating that these two metabolites may be closely related with lycopene production. The optimized method provides a useful approach for future metabolic analysis of fermentation process as well as industrial strain improvement.
研究领域:
生物化学
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代谢工程
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PubMed
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JOURNAL OF CHEMICAL TECHNOLOGY AND BIOTECHNOLOGY
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2011
·Vol. 87
·期 4
·553-558
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IF 1.818
Xiao-Feng Wang,Xu-Guang Shen,Yong-Chuan Sun,He-Yun Zhao,Li Xu,Yun Liu,Yun-Jun Yan
BACKGROUND: Yarrowia lipolytica lipase LIP2 (YlLIP2) is an important industrial enzyme that has many potential applications. Although it has been successfully expressed in Pichia pastoris under the control of the AOX1 promoter ( pAOX1 ), there have been many efforts to develop new alternative promoters to pAOX1 in order to avoid using methanol in the fermentation. Investigation of YlLIP2 production in P. pastoris using the formaldehyde dehydrogenase 1 promoter ( pFLD1 ) is especially attractive, since little is known about its application in methanol-free culture strategies. RESULTS: Three fed-batch cultivations were performed to investigate the production of YlLIP2 in a pFLD1 -based system. When methanol was used as the fed-batch feeding substrate, the maximum YlLIP2 activity obtained in a 10-L bioreactor was 30 000 U mL −1 after 143 h of culture, whereas the maximum YlLIP2 activity was further increased to 35 000 U mL −1 by adopting a co-induction strategy with methanol and methylamine as a mixed fed-batch substrate. Furthermore, the maximum YlLIP2 activity reached 13 000 U mL −1 after 80 h of cultivation in a methanol-free culture. CONCLUSION: The expression levels of YlLIP2 in the pFLD1 -based system were comparable with those in a pAOX1 -based system. The results suggest that pFLD1 is an attractive alternative to pAOX1 , and may make it feasible to induce high yields of protein expression. Copyright © 2011 Society of Chemical Industry
研究领域:
生物技术
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分子生物学
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高影响因子
BIOCHEMICAL ENGINEERING JOURNAL
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2015
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IF 2.467
Qin Xu,Wei-Yi Tao,He Huang,Shuang Li
An NADPH-dependent carbonyl reductase (YlCR2) from Yarrowia lipolytica was discovered by genome mining, overexpressed in Escherichia coli BL21 and purified to homogeneity. To efficiently synthesize ethyl ( S )-4-chloro-3-hydroxybutanoate (( S )-CHBE) (99%, e.e), the highly stereoselective bioreduction of ethyl 4-chloro-3-oxobutanoate (COBE) into ( S )-CHBE with the recombinant E. coli BL21/pETYlCR2 was successfully demonstrated in an n -butyl acetate-water biphasic system (1:1, v/v) with NADPH self-regeneration by substrate-coupled system using sorbitol or mannitol as co-substrate. The optimum reaction condition for the biotransformation of COBE in the biphasic system were 3000 mM COBE, (1.2 or 1.3 mmol/mmol COBE) mannitol or sorbitol, 0.2 mM or 0.4 mM NADP + , 0.12 or 0.14 g (wet weight)/ml cell dosage, pH 5.0, 30 °C; ( S )-CHBE with yield of 90% and e.e of 99% was obtained after 10 h reaction. Furthermore, 3000 mM COBE could also be completely biotransformed after 20 h without addition of expensive cofactor NADP + . Significantly, E. coli BL21/pETYlCR2 shows the high potential in the industrial production of ( S )-CHBE.
研究领域:
生物化学
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合成生物学
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酶工程
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DOI
高影响因子
BioDesign Research
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2025
·
IF 4.7
Yuxiang Hong,Yang Gu,Dewei Lin,Zizhao Wu,Wenhao Chen,Tianjian Lu,Pornpatsorn Lertphadungkit,Jingbo Ma,Haili Wang,Bo Zhou,Gil Bar-Sela,Idan Cohen,Peng Xu
Cannabis sativa has long been a cornerstone of both medicinal and cultural practices, with its therapeutic use spanning over 2,700 years. Central to its therapeutic effects are cannabinoids, which interact with the endocannabinoid system to influence various physiological processes such as anxiety, pain, and inflammation. Despite its benefits, cannabinoid production faces challenges and scarcity from plant extraction. This work leverages Yarrowia lipolytica as a platform for cannabinoid biosynthesis. By optimizing precursor supply, engineering biomolecular condensate-like dual prenyltransferase expression and expanding endogenous metabolism with a noncanonical polyketide synthase, we achieved the de novo biosynthesis of various cannabinoid and its analogs. Our engineered Y. lipolytica produced ∼3.5 mg/L of cannabigerolic acid, 18.8 mg/L of orsellinic acid, and 0.5 mg/L of cannabigerorcinic acid. Additionally, CBGA titer reached 15.7 mg/L with olivetolic acid supplement. This work demonstrates Y. lipolytica 's versatility as a promising host for cannabinoid and its analogs production, which opens avenues for further research and medicinal applications.
研究领域:
合成生物学
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代谢工程
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植物化学
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生物技术
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DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
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2026
·
IF 6.7
Guiping Guo,Huibin Chen,Hailin Wang,Guilin Zhang,Xiaodan Fu,Man Yuen Lee,Yun-Chung Leung,Ningyang Li,Zhen Liu
Crocetin, a bioactive carotenoid derivative from saffron, exhibits antioxidant, anti-inflammatory, and metabolic regulatory properties. Its low natural abundance and limited extraction production hinder large-scale production. Microbial synthesis offers a promising alternative but is constrained by low carotenoid cleavage dioxygenase (CCD2) expression. Here, we engineered a zeaxanthin-producing Yarrowia lipolytica strain to enhance CsCCD2 expression via fermentation temperature optimization, SUMOstar fusion, subcellular localization, and precursor supply improvement. The engineered strain achieved a crocetin titer of 196.52 mg/L in shake flasks and 1735.73 mg/L in 10 L fed-batch fermentation, setting a new microbial production record─16-fold higher than previous reports. This work provides a foundation for efficient, large-scale microbial crocetin production and its industrial application.
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PubMed
高影响因子
Biomolecules
·
2024
·Vol. 14
·期 6
·712
·
IF 4.8
Gehad G. Ibrahim,Madhavi Perera,Saadiah A. Abdulmalek,Jinyong Yan,Yunjun Yan
Resveratrol, a phenylpropanoid compound, exhibits diverse pharmacological properties, making it a valuable candidate for health and disease management. However, the demand for resveratrol exceeds the capacity of plant extraction methods, necessitating alternative production strategies. Microbial synthesis offers several advantages over plant-based approaches and presents a promising alternative.Yarrowia lipolyticastands out among microbial hosts due to its safe nature, abundant acetyl-CoA and malonyl-CoA availability, and robust pentose phosphate pathway. This study aimed to engineerY. lipolyticafor resveratrol production. The resveratrol biosynthetic pathway was integrated intoY. lipolyticaby adding genes encoding tyrosine ammonia lyase fromRhodotorula glutinis, 4-coumarate CoA ligase fromNicotiana tabacum, and stilbene synthase fromVitis vinifera. This resulted in the production of 14.3 mg/L resveratrol. A combination of endogenous and exogenous malonyl-CoA biosynthetic modules was introduced to enhance malonyl-CoA availability. This included genes encoding acetyl-CoA carboxylase 2 fromArabidopsis thaliana, malonyl-CoA synthase, and a malonate transporter protein fromBradyrhizobium diazoefficiens. These strategies increased resveratrol production to 51.8 mg/L. The further optimization of fermentation conditions and the utilization of sucrose as an effective carbon source in YP media enhanced the resveratrol concentration to 141 mg/L in flask fermentation. By combining these strategies, we achieved a titer of 400 mg/L resveratrol in a controlled fed-batch bioreactor. These findings demonstrate the efficacy ofY. lipolyticaas a platform for the de novo production of resveratrol and highlight the importance of metabolic engineering, enhancing malonyl-CoA availability, and media optimization for improved resveratrol production.
研究领域:
生物技术
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代谢工程
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微生物学
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药学
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生物化学
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产品关联
DOI
PubMed
高影响因子
Biotechnology for Biofuels
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2020
·Vol. 13
·期 1
·1-13
·
IF 4.815
Zhang Jin-Lai,Bai Qiu-Yan,Peng Yang-Zi,Fan Jie,Jin Cong-Cong,Cao Ying-Xiu,Yuan Ying-Jin
Background Lupeol exhibits novel physiological and pharmacological activities, such as anticancer and immunity-enhancing activities. However, cytotoxicity remains a challenge for triterpenoid overproduction in microbial cell factories. As lipophilic and relatively small molecular compounds, triterpenes are generally secreted into the extracellular space. The effect of increasing triterpene efflux on the synthesis capacity remains unknown.Results In this study, we developed a strategy to enhance triterpene efflux through manipulation of lipid components in Y. lipolytica by overexpressing the enzyme Δ9-fatty acid desaturase (OLE1) and disturbing phosphatidic acid phosphatase (PAH1) and diacylglycerol kinase (DGK1). By this strategy combined with two-phase fermentation, the highest lupeol production reported to date was achieved, where the titer in the organic phase reached 381.67 mg/L and the total production was 411.72 mg/L in shake flasks, exhibiting a 33.20-fold improvement over the initial strain. Lipid manipulation led to a twofold increase in the unsaturated fatty acid (UFA) content, up to 61–73%, and an exceptionally elongated cell morphology, which might have been caused by enhanced membrane phospholipid biosynthesis flux. Both phenotypes accelerated the export of toxic products to the extracellular space and ultimately stimulated the capacity for triterpenoid synthesis, which was proven by the 5.11-fold higher ratio of extra/intracellular lupeol concentrations, 2.79-fold higher biomass accumulation and 2.56-fold higher lupeol productivity per unit OD in the modified strains. This strategy was also highly efficient for the biosynthesis of other triterpenes and sesquiterpenes, including α-amyrin, β-amyrin, longifolene, longipinene and longicyclene.Conclusion sIn conclusion, we successfully created a high-yield lupeol-producing strain via lipid manipulation. We demonstrated that the enhancement of lupeol efflux and synthesis capacity was induced by the increased UFA content and elongated cell morphology. Our study provides a novel strategy to promote the biosynthesis of valuable but toxic products in microbial cell factories.
研究领域:
生物技术
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微生物学
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代谢工程
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药理学
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DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
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2023
·
IF 6.1
Qian-Qian Peng,Qi Guo,Cheng Chen,Ping Song,Yue-Tong Wang,Xiao-Jun Ji,Chao Ye,Tian-Qiong Shi
Patchoulol is an important sesquiterpene alcohol with a strong and lasting odor, which has led to prominent applications in perfumes and cosmetics. In this study, systematic metabolic engineering strategies were adopted to create an efficient yeast cell factory for patchoulol overproduction. First, a baseline strain was constructed by selecting a highly active patchoulol synthase. Subsequently, the mevalonate precursor pool was expanded to boost patchoulol synthesis. Moreover, a method for downregulating squalene synthesis based on Cu2+-repressible promoter was optimized, which significantly improved the patchoulol titer by 100.9% to 124 mg/L. In addition, a protein fusion strategy resulted in a final titer of 235 mg/L in shake flasks. Finally, 2.864 g/L patchoulol could be produced in a 5 L bioreactor, representing a remarkable 1684-fold increase compared to the baseline strain. To our knowledge, this is the highest patchoulol titer reported so far.
研究领域:
生物技术
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代谢工程
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合成生物学
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产品关联
DOI
PubMed
高影响因子
BIORESOURCE TECHNOLOGY
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2026
·
IF 9
Jianglong Li,Xin Li,Kai Linghu,Niping Yang,Zhengxuan Shao,Jingwen Zhou,Haitong Hou,Weizhu Zeng
Retinol (Vitamin A) is a micronutrient that is essential for human health. This study has engineered the oleaginous yeast Yarrowia lipolytica for high-level de novo retinol biosynthesis. The precursor flux based on a previously established high-lycopene strain was further strengthened by overexpressing GGPP synthase and the β-carotene synthesis pathway, which achieved 822.7 mg·L −1 β-carotene. The retinol pathway was subsequently established by introducing β-carotene dioxygenase (Blh) and retinol dehydrogenase (RDH12). The catalytic activity of Blh was significantly improved using directed evolution in order to address the excessively low catalytic efficiency of the key rate-limiting enzymes. Furthermore, retinol production was improved using co-factor engineering to increase the availability of NADPH and knocking out the oxidase gene to maintain the cellular redox balance. Finally, the retinol titer reached 8.92 g·L −1 using butylated hydroxytoluene (BHT) as an antioxidant in a 5-L fed-batch fermentation process, which represents the highest reported microbial titer, establishing Y. lipolytica as a potent platform for sustainable vitamin A production.
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PubMed
高影响因子
Synthetic and Systems Biotechnology
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2025
·
IF 4.4
Mingyu Dai,Qingyi Cao,Shuanglong Jia,Jiang Chen,Hui-Ying Xu,Bang-Ce Ye,Wei-Bing Liu,Ying Zhou
β-Elemene, a pharmacologically active sesquiterpene derived from the traditional Chinese medicinal herb Curcuma wenyujin, exhibits broad-spectrum antitumor and anti-inflammatory activities. Currently, its commercial production relies heavily on plant extraction, a process associated with high costs and environmental burdens, limiting industrial scalability. To address these challenges, we developed a metabolically engineered Yarrowia lipolytica strain as an efficient microbial cell factory for de novo Germacrene A biosynthesis, the direct precursor of β-elemene. Through combinatorial optimization strategies—including (1) mevalonate (MVA) pathway enhancement, (2) copy number amplification of germacrene A synthase (GAS), (3) β-oxidation pathway reinforcement, and (4) introduction of the isopentenyl utilization pathway (IUP)—we significantly improved β-elemene production. The engineered strain achieved a titer of 3.08 ± 0.05 g/L in shake-flask cultures, with a yield of 51.27 ± 0.75 mg/g glucose, representing a 3.5-fold increase over the parental strain. Our work highlights the potential of Y. lipolytica as a sustainable and scalable platform for high-value sesquiterpene production, offering a viable alternative to plant-derived extraction.
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