BackgroundFatty alcohols are important oleochemicals widely used in detergents, surfactants and personal care products. Bio-synthesized fatty alcohol provides a promising alternative to traditional fatty alcohol industry. Harnessing oleaginous microorganisms for fatty alcohol production may offer a new strategy to achieve a commercially viable yield that currently still seems to be a remote target.ResultsIn this study, we introduced functional fatty acyl-CoA reductase (FAR), TaFAR1 to direct the conversion from fatty acyl-CoA to fatty alcohol in Yarrowia lipolytica (Y. lipolytica), an oleaginous non-conventional yeast showing great lipid-producing capability. Tri-module optimizations including eliminating fatty alcohol degradation pathway, enhancing TaFAR1 expression, and increasing fatty acyl-CoA supply were furtherly conducted, resulting in 63-fold increase in intracellular fatty alcohol-producing capability compared to the starting strain. Thus, this work demonstrated successful construction of first generation of Y. lipolytica fatty alcohol-producing cell factory. Through the study of effect of environmental nutrition on fatty alcohol production, up to 636.89 mg/L intracellular hexadecanol (high fatty alcohol-retaining capability) and 53.32 mg/L extracellular hexadecanol were produced by this cell factory through batch fermentation, which was comparable to the highest production of Saccharomyces cerevisiae under the similar condition.ConclusionThis work preliminarily explored fatty alcohol-producing capability through mobilization of FAR and fatty acid metabolism, maximizing the intracellular fatty alcohol-producing capability, suggesting that Y. lipolytica cell factory potentially offers a promising platform for fatty alcohol production.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0512-3) contains supplementary material, which is available to authorized users.
Abstract1-methylcyclopropene (1-MCP) in an ethylene receptor antagonist that blocks ethylene perception and downstream ripening responses in climacteric fruit imparting a longer shelf life. However, in European pear, the application of 1-MCP irreversibly obstructs the onset of system 2 ethylene production resulting in perpetually unripe fruit with undesirable quality. Application of exogenous ethylene, carbon dioxide and treatment to high temperatures is not able to reverse the blockage in ripening. We recently reported that during cold conditioning, activation of alternative oxidase (AOX) occurs pre-climacterically. In this study, we report that activation of AOX via exposure of 1-MCP treated ‘D’Anjou’ pear fruit to glyoxylic acid triggers an accelerated ripening response. Time course physiological analysis revealed that ripening is evident from decreased fruit firmness and increased internal ethylene. Transcriptomic and functional enrichment analyses revealed genes and ontologies implicated in glyoxylic acid-mediated ripening, including AOX, TCA cycle, fatty acid metabolism, amino acid metabolism, organic acid metabolism, and ethylene-responsive pathways. These observations implicate the glyoxylate cycle as a biochemical hub linking multiple metabolic pathways to stimulate ripening through an alternate mechanism. The results provide information regarding how blockage caused by 1-MCP may be circumvented at the metabolic level, thus opening avenues for consistent ripening in pear and possibly other fruit.
The oleaginous yeast Yarrowia lipolytica has attracted much attention due to its ability to utilize a wide range of substrates to accumulate high lipid content and its flexibility for genetic manipulation. In this study, intracellular lipid metabolism in Y. lipolytica was tailored to produce fatty acid, a renewable oleochemical and precursor for production of advanced biofuels. Two main strategies, including blocking activation and peroxisomal uptake of fatty acids and elimination of biosynthesis of lipids, were employed to reduce fatty acid consumption by the native pathways in Y. lipolytica. Both genetic modifications improved fatty acid production. However, disruption of the genes responsible for assembly of nonpolar lipid molecules including triacylglycerols (TAGs) and steryl esters resulted in the deleterious effects on the cell growth. The gene tesA encoding thioesterase from Escherichia coli was expressed in the strain with disrupted faa genes encoding fatty acyl-CoA synthetases and pxa1 encoding peroxisomal acyl-CoA transporter, and the titer of fatty acids resulted in 2.3 g/L in shake flask culture, representing 11-fold improvement compared with the parent strain. Expressing the native genes encoding acetyl-CoA carboxylase (ACC) and hexokinase also increased fatty acid production, although the improvement was not as significant as that with tesA expression. Saturated fatty acids including palmitic acid (C16:0) and stearic acid (C18:0) increased remarkably in the fatty acid composition of the recombinant bearing tesA compared with the parent strain. The recombinant expressing tesA gene resulted in high lipid content, indicating the great fatty acid producing potential of Y. lipolytica. The results highlight the achievement of fatty acid overproduction without adverse effect on growth of the strain. Results of this study provided insight into the relationship between fatty acid and lipid metabolism in Y. lipolytica, confirming the avenue to reprogram lipid metabolism of this host for overproduction of renewable fatty acids.
The advent of genome editing has opened new avenues for targeted trait enhancement in fruit, ornamental, industrial, and all specialty crops. In particular, CRISPR-based editing systems, derived from bacterial immune systems, have quickly become routinely used tools for research groups across the world seeking to edit plant genomes with a greater level of precision, higher efficiency, reduced off-target effects, and overall ease-of-use compared to ZFNs and TALENs. CRISPR systems have been applied successfully to a number of horticultural and industrial crops to enhance fruit ripening, increase stress tolerance, modify plant architecture, control the timing of flower development, and enhance the accumulation of desired metabolites, among other commercially-important traits. As editing technologies continue to
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