Studies on Upgradation of Waste Fish Oil to Lipid-Rich Yeast Biomass in Yarrowia lipolytica Batch Cultures.
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SUBMITTER: Fabiszewska AU
PROVIDER: S-EPMC7922964 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundLimonene is an important biologically active natural product widely used in the food, cosmetic, nutraceutical and pharmaceutical industries. However, the low abundance of limonene in plants renders their isolation from plant sources non-economically viable. Therefore, engineering microbes into microbial factories for producing limonene is fast becoming an attractive alternative approach that can overcome the aforementioned bottleneck to meet the needs of industries and make limonene production more sustainable and environmentally friendly.ResultsIn this proof-of-principle study, the oleaginous yeast Yarrowia lipolytica was successfully engineered to produce both d-limonene and l-limonene by introducing the heterologous d-limonene synthase from Citrus limon and l-limonene synthase from Mentha spicata, respectively. However, only 0.124 mg/L d-limonene and 0.126 mg/L l-limonene were produced. To improve the limonene production by the engineered yeast Y. lipolytica strain, ten genes involved in the mevalonate-dependent isoprenoid pathway were overexpressed individually to investigate their effects on limonene titer. Hydroxymethylglutaryl-CoA reductase (HMGR) was found to be the key rate-limiting enzyme in the mevalonate (MVA) pathway for the improving limonene synthesis in Y. lipolytica. Through the overexpression of HMGR gene, the titers of d-limonene and l-limonene were increased to 0.256 mg/L and 0.316 mg/L, respectively. Subsequently, the fermentation conditions were optimized to maximize limonene production by the engineered Y. lipolytica strains from glucose, and the final titers of d-limonene and l-limonene were improved to 2.369 mg/L and 2.471 mg/L, respectively. Furthermore, fed-batch fermentation of the engineered strains Po1g KdHR and Po1g KlHR was used to enhance limonene production in shake flasks and the titers achieved for d-limonene and l-limonene were 11.705 mg/L (0.443 mg/g) and 11.088 mg/L (0.385 mg/g), respectively. Finally, the potential of using waste cooking oil as a carbon source for limonene biosynthesis from the engineered Y. lipolytica strains was investigated. We showed that d-limonene and l-limonene were successfully produced at the respective titers of 2.514 mg/L and 2.723 mg/L under the optimal cultivation condition, where 70% of waste cooking oil was added as the carbon source, representing a 20-fold increase in limonene titer compared to that before strain and fermentation optimization.ConclusionsThis study represents the first report on the development of a new and efficient process to convert waste cooking oil into d-limonene and l-limonene by exploiting metabolically engineered Y. lipolytica strains for fermentation. The results obtained in this study lay the foundation for more future applications of Y. lipolytica in converting waste cooking oil into various industrially valuable products.
| S-EPMC6781337 | biostudies-literature
Project description:Time-course transcriptomic profilling of the oleaginous yeast Yarrowia lipolytica, during a controlled fed-batch. A nitrogen limitation was applied during the course of the fed-batch to initiate de novo biolipid synthesis.
2011-05-15 | GSE29046 | GEO
Project description:Backgroundβ-Farnesene is a sesquiterpene with versatile industrial applications. The production of β-farnesene from waste lipid feedstock is an attractive method for sustainable production and recycling waste oil. Yarrowia lipolytica is an unconventional oleaginous yeast, which can use lipid feedstock and has great potential to synthesize acetyl-CoA-derived chemicals.ResultsIn this study, we engineered Y. lipolytica to produce β-farnesene from lipid feedstock. To direct the flux of acetyl-CoA, which is generated from lipid β-oxidation, to β-farnesene synthesis, the mevalonate synthesis pathway was compartmentalized into peroxisomes. β-Farnesene production was then engineered by the protein engineering of β-farnesene synthase and pathway engineering. The regulation of lipid metabolism by enhancing β-oxidation and eliminating intracellular lipid synthesis was further performed to improve the β-farnesene synthesis. As a result, the final β-farnesene production with bio-engineering reached 35.2 g/L and 31.9 g/L using oleic acid and waste cooking oil, respectively, which are the highest β-farnesene titers reported in Y. lipolytica.ConclusionsThis study demonstrates that engineered Y. lipolytica could realize the sustainable production of value-added acetyl-CoA-derived chemicals from waste lipid feedstock.
| S-EPMC9528160 | biostudies-literature
Project description:The natural plant product bisabolene serves as a precursor for the production of a wide range of industrially relevant chemicals. However, the low abundance of bisabolene in plants renders its isolation from plant sources non-economically viable. Therefore, creation of microbial cell factories for bisabolene production supported by synthetic biology and metabolic engineering strategies presents a more competitive and environmentally sustainable method for industrial production of bisabolene. In this proof-of-principle study, for the first time, we engineered the oleaginous yeast Yarrowia lipolytica to produce α-bisabolene, β-bisabolene and γ-bisabolene through heterologous expression of the α-bisabolene synthase from Abies grandis, the β-bisabolene synthase gene from Zingiber officinale and the γ-bisabolene synthase gene from Helianthus annuus respectively. Subsequently, two metabolic engineering approaches, including overexpression of the endogenous mevalonate pathway genes and introduction of heterologous multidrug efflux transporters, were employed in order to improve bisabolene production. Furthermore, the fermentation conditions were optimized to maximize bisabolene production by the engineered Y. lipolytica strains from glucose. Finally, we explored the potential of the engineered Y. lipolytica strains for bisabolene production from the waste cooking oil. To our knowledge, this is the first report of bisabolene production in Y. lipolytica using metabolic engineering strategies. These findings provide valuable insights into the engineering of Y. lipolytica for a higher-level production of bisabolene and its utilization in converting waste cooking oil into various industrially valuable products.
| S-EPMC8601197 | biostudies-literature
Project description:The global limitation of fossil fuels impels scientists to search for new energy sources. A good alternative is biodiesel produced from crop plants. However, its production requires huge quantities of farmland, fertilizers and fresh water, which is in conflict with the human demand for water for consumption and land for food production. Thus, production of single cell oil (SCO) by oleaginous microorganisms remains the best solution for the coming years. Whereas most microorganisms require fresh water for proper cell metabolism, in this study we demonstrate that the unconventional yeast Yarrowia lipolytica is able to produce huge quantities of fatty acid in seawater-based medium. Here we shown that Y. lipolytica is able to produce fatty acids in medium based on seawater and crude glycerol as the main carbon source, which allows for low-cost production of SCO, is beneficial for industrial application and is ecologically friendly.
| S-EPMC6431633 | biostudies-literature
Project description:Time-course transcriptomic profilling of the oleaginous yeast Yarrowia lipolytica, during a controlled fed-batch. A nitrogen limitation was applied during the course of the fed-batch to initiate de novo biolipid synthesis. Dual staining, one replicate per slide. On each slide, a time point sample and a reference sample are spotted. Reference samples were obtained by pooling the RNAs from the time-course samples.
2011-05-15 | E-GEOD-29046 | biostudies-arrayexpress
Project description:Oleaginous yeasts are fast emerging as a possible feedstock for biodiesel production. Yarrowia lipolytica, a model oleaginous yeast is known to utilize a variety of hydrophobic substrates for lipid accumulation including waste cooking oil (WCO). Approaches to increase lipid content in this yeast include metabolic engineering which requires manipulation of multiple genes in the lipid biosynthesis pathway. A classical and cost-effective approach, namely, random chemical mutagenesis on the yeast can lead to increased production of biodiesel as is explored here.In this study, chemical mutagenesis using the alkylating agent, N- methyl-N'-nitro-N-nitrosoguanidine (MNNG) as well as an additional treatment with cerulenin, a fatty acid synthase inhibitor generated 800 mutants of Y. lipolytica NCIM 3589 (761 MNNG treated and 39 MNNG + cerulenin treated). A three-stage screening using Sudan Black B plate technique, Nile red fluorimetry and total lipid extraction using solvent was performed, which enabled selection of ten high lipid yielding mutants. Time course studies of all the ten mutants were further undertaken in terms of biomass, lipid yield and lipid content to select three stable mutants (YlB6, YlC7 and YlE1) capable of growing and accumulating lipid on WCO, with lipid contents of 55, 60 and 67% as compared to 45% for the wild type. The mutants demonstrated increased volumetric lipid productivities (0.062, 0.044 and 0.041 g L-1 h-1) as compared to the wild type (0.033 g L-1 h-1). The fatty acid profile of the three mutants consisted of a high content of C16 and C18 saturated and monounsaturated fatty acids and was found to be suitable for biodiesel production. The fuel properties, namely, density, kinematic viscosity, total acid number, iodine value of the three mutants were evaluated and found to lie within the limits specified by internationally accepted standards. Additionally, it was noted that the mutants demonstrated better cetane numbers and higher heating values than the wild type strain.The chemical mutagenesis strategy adopted in this study resulted in the successful isolation of three stable high SCO yielding mutants. The mutants, namely, YlB6, YlC7 and YlE1 exhibited a 1.22, 1.33 and 1.49-fold increase in lipid contents when grown on 100 g L-1 waste cooking oil than the parental yeast strain. The fatty acid methyl ester (FAME) profiles of all the three mutants was determined to be suitable for biodiesel suggesting their potential applicability while simultaneously addressing the management of waste cooking oil.
| S-EPMC5655982 | biostudies-literature
Project description:We previously developed a fermentation protocol for lipid accumulation in the oleaginous yeast Y. lipolytica. This process was used to perform transcriptomic time-course analyses to explore gene expression in Y. lipolytica during the transition from biomass production to lipid accumulation. In this experiment, a biomass concentration of 54.6 g(CDW)/l, with 0.18 g/g(CDW) lipid was obtained in ca. 32 h, with low citric acid production. A transcriptomic profiling was performed on 11 samples throughout the fermentation. Through statistical analyses, 569 genes were highlighted as differentially expressed at one point during the time course of the experiment. These genes were classified into 9 clusters, according to their expression profiles. The combination of macroscopic and transcriptomic profiles highlighted 4 major steps in the culture: (i) a growth phase, (ii) a transition phase, (iii) an early lipid accumulation phase, characterized by an increase in nitrogen metabolism, together with strong repression of protein production and activity; (iv) a late lipid accumulation phase, characterized by the rerouting of carbon fluxes within cells. This study explores the potential of Y. lipolytica as an alternative oil producer, by identifying, at the transcriptomic level, the genes potentially involved in the metabolism of oleaginous species.
| S-EPMC3222671 | biostudies-literature
Project description: Not available
| S-EPMC10540775 | biostudies-literature
Project description:The yeast strain Yarrowia lipolytica IPS 21 was tested for its ability to degrade potentially toxic chrome-tanned leather shavings (CTLS) in a liquid environment. Biological and chemical parameters were monitored during a 48-h period of biotransformation of the protein-rich waste. CTLS was added at a concentration of 0.1-4% (wt/wt) to a modified YPG medium (15 g L-1 yeast extract and 5 g L-1 NaCl). Biodegradation and bioconversion were performed in a one-step process. It was found that the higher degradation rate depended on the activity of the proteases and the pH of the medium, but not on the initial inoculum ratio and the activity of the dehydrogenase. The highest efficiency of the process was obtained for 4% (wt/wt) CTLS on day 2 (degradation rate 58-67%, biomass production 2.11-2.20 g L-1, protease activity 312 U mg-1 protein, and pH 9.20). Our results showed that total chromium was probably not transported across the cytoplasmic membrane of Y. lipolytica IPS21 and that chromium (III) was not oxidized to chromium (VI). The phytotoxicity of selected amino acid supernatants [2.5% (vol/vol)] was tested after the bioconversion process. It was found that the supernatants had a stimulating effect on the plants tested. The root elongation was 29-28% higher than that of the reference samples. This result makes Y. lipolytica IPS21 a potential candidate for safely converting potentially toxic protein-rich wastes into valuable products without enzyme isolation, e.g., amino acid fertilizers. IMPORTANCE Enzyme technologies have the greatest practical relevance to environmental trends. Overcoming the barrier of the high cost of carbon substrates used for biotransformation is the main challenge of these methods. The huge potential of the use of extracellular proteases of Yarrowia species or amino acids in various industries indicates the need for the extension of basic research on waste as a carbon source for this yeast. The experiments demonstrated that it is possible to use Y. lipolytica IPS21 for bioconversion of chrome-tanned leather shavings (CTLS) in a single-step process and to produce high-value amino acid supernatant without having an isolated enzyme. In our study, we show the effect of 2.5% (vol/vol) CTLS supernatant obtained from Y. lipolytica IPS21 on the elongation of the root system of selected plants and provide information on the effect of environmental factors on the efficiency of the bioconversion and the migration of chromium.
| S-EPMC10581069 | biostudies-literature