Oleaginous yeasts have been increasingly explored for production of chemicals and fuels via metabolic engineering. Particularly, there is a growing interest in using oleaginous yeasts for the synthesis of lipid-related products due to their high lipogenesis capability, robustness, and ability to utilize a variety of substrates. Most of the metabolic engineering studies in oleaginous yeasts focused on Yarrowia that already has plenty of genetic engineering tools. However, recent advances in systems biology and synthetic biology have provided new strategies and tools to engineer those oleaginous yeasts that have naturally high lipid accumulation but lack genetic tools, such as Rhodosporidium, Trichosporon, and Lipomyces. This review highlights recent accomplishments in metabolic engineering of oleaginous yeasts and recent advances in the development of genetic engineering tools in oleaginous yeasts within the last 3 years.

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Metabolic Engineering of Oleaginous Yeasts for Production of Fuels and Chemicals.

Lipomyces starkeyi: Its current status as a potential oil producer

Oils and oleochemicals produced by microbial cells offer an attractive alternative to petroleum and food-crop derived oils for the production of transport fuel and oleochemicals An emerging candidate for industrial single cell oil production is the oleaginous yeast Lipomyces starkeyi This yeast is capable of accumulating storage lipids to concentrations greater than 60% of the dry cell weight From the perspective of industrial biotechnology L starkeyi is an excellent chassis for single-cell oil and oleochemical production as it can use a wide variety of carbon and nitrogen sources as feedstock The strain has been used to produce lipids from hexose and pentose sugars derived from cellulosic hydrolysates as well as crude glycerol and even sewage sludge L starkeyi also produces glucanhydrolases that have a variety of industrial applications and displays potential to be employed for bioremediation Despite its excellent properties for biotechnology applications, adoption of L starkeyi as an industrial chassis has been hindered by the difficulty of genetically manipulating the strain This review will highlight the industrial potential of L starkeyi as a chassis for the production of lipids, oleochemicals and other biochemicals Additionally, we consider progress and challenges in engineering this organism for industrial applications

Lipomyces starkeyi: an emerging cell factory for production of lipids, oleochemicals and biotechnology applications.

Enhancing microbial lipids yield for biodiesel production by oleaginous yeast Lipomyces starkeyi fermentation: A review

Microbial oil triacylglycerol (TAG) from the renewable feedstock attract much attention. The oleaginous yeast Yarrowia lipolytica has become the most studied for lipid biosynthesis. Fatty acid desaturases catalyze the introduction of a double bond into fatty-acid hydrocarbon chains to produce unsaturated fatty acids. Desaturases are known to enhance lipid accumulation. In this study, we have achieved a significant increase in lipid production and increase the unsaturated fatty acids content in Y. lipolytica. By comparing the expression of the native genes of △-9 stearoyl-CoA desaturase (SCD) and △12 desaturase (△12D), and an exogenous △15 desaturase (△15D) from flax in the strain with deleted peroxisomal biogenesis factor 10 (PEX10) and overexpressed diacylglyceride acyl-transferase (DGA1), we found that the strain with overexpressed △15 desaturase accumulated 30.7% lipid. Simultaneously, we explored the effect of two copies of desaturase genes (12D-SCD, 15D-SCD, 12D-15D) on lipid production, and found co-expression of △12D and △15D accumulated 42.6% lipid. The lipid content was further increased by 56.3% through the deletion of the multifunctional enzyme (MFE1) and the overexpression of acetyl-CoA carboxylase (ACC1). Finally, the lipid productivity of 50 g/L and maximal lipid content of 77.8% DCW are obtained using a 5-L stirred-tank bioreactor during the stationary phase in the engineered YL-10. Our result demonstrated that the △12 and △15 desaturases play an important role in lipid production in Y. lipolytica and provides an effective strategy for biodiesel development.

Overexpression of △12, △15-Desaturases for Enhanced Lipids Synthesis in Yarrowia lipolytica.

Long chain polyunsaturated fatty acids (LCPUFA) are known to play an important role in human health and nutrition. Considering the limitation of LCPUFA sources, it is necessary to search new avenues for their production. Oleaginous yeasts are an attractive target for harvesting single cell oil, mainly because of the ease of cultivation with cheaper raw material. Lipomyces starkeyi is one such oleaginous yeast, which can accumulate oil to the extent of 60 % of its biomass and where genetic transformation can be achieved. In our earlier work, Δ15 desaturase gene (AEP37840) from flax was transformed into L. starkeyi. In the present work, we report optimization of medium for the production of ω-3 enriched oil from this transformed yeast. A basic medium containing 20 g/l glucose as a carbon source and 10 g/l yeast extract as a nitrogen source was used during fermentation. At regular time intervals, glucose was fed to maintain high C:N ratio (65:10) during fermentation. Under the most favorable conditions, dry biomass and total lipid content were 18 and 7.29 g/l, respectively. Prior to genetic transformation, L. starkeyi contained 56.03 mg/l DHA along with 71.4 mg/l EPA and 42.2 mg/l ALA. Genetic engineering of this yeast resulted in a strain that produced 1080 mg/l DHA (17.4 %) along with 74.28 mg/l EPA and 126.72 mg/l ALA possibly through modification of PUFA biosynthetic pathway. To the best of our knowledge, this is a first report of DHA enrichment and opens up avenues for LCPUFA production through L. starkeyi.

Production of polyunsaturated fatty acids in recombinant Lipomyces starkeyi through submerged fermentation

Long-chain polyunsaturated fatty acids (LCPUFA) i.e., eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have established role in the prevention of several diseases like atherosclerosis, cardiovascular disorders, and arthritis. However, LCPUFA sources are limited for availability and acceptability problems of fish and flax oil. Microbial sources of n-3 PUFA are of merit being cheaper, contamination free and vegan. We here report fungi that can convert ∝-linolenic acid (ALA) from flax seed meal (FSM) into EPA and DHA in dose-dependent manner and therefore serve as valuable LCPUFA sources. Mucor sp. and R. oligosporous cultivated on FSM supplemented medium accumulated 34 and 36% lipids of dry biomass. Wherein, Mucor sp produced 6.01mg DHA per gram of lipid at 10% FSM at 28°C, while Rhizopus oligosporus produced 14.32 mg of DHA/gm of lipid at 3% FSM at 20°C. These novel yet exploitable sources of LCPUFA are valuable to ensure EPA/DHA for human supplementation.

Bioconversion of alpha-linolenic acid into long chain polyunsaturated fatty acids by oleaginous fungi

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Production of polyunsaturated fatty acids in recombinant Lipomyces starkeyi through submerged fermentation

Bioconversion of alpha-linolenic acid into long chain polyunsaturated fatty acids by oleaginous fungi