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The sesquiterpene α-humulene is an important plant natural product, which has been used in the pharmaceutical industry due to its anti-inflammatory and anticancer activities. Although phytoextraction and chemical synthesis have previously been applied in α-humulene production, the low efficiency and high costs limit the development. In this study, Yarrowia lipolytica was engineered as the robust cell factory for sustainable α-humulene production. First, a chassis with high α-humulene output in the cytoplasm was constructed by integrating α-humulene synthases with high catalytic activity, optimizing the flux of mevalonate and acetyl-CoA pathways. Subsequently, the strategy of dual cytoplasmic-peroxisomal engineering was adopted in Y. lipolytica; the best strain GQ3006 generated by introducing 31 copies of 12 different genes could produce 2280.3± 38.2 mg/l (98.7 ± 4.2 mg/g dry cell weight) α-humulene, a 100-fold improvement relative to the baseline strain. To further improve the titer, a novel strategy for downregulation of squalene biosynthesis based on Cu

Terpenes are a large class of secondary metabolites with diverse structures and functions that are commonly used as valuable raw materials in food, cosmetics, and medicine. With the development of metabolic engineering and emerging synthetic biology tools, these important terpene compounds can be sustainably produced using different microbial chassis. Currently, yeasts such as

Additional file 1: Fig. S1. Influence of WCO from different canteens on α-humulene production. Fig. S2. Influence of differential genes regulation on α-humulene production (DCW). (A) Overexpression of genes with significantly up-regulated transcript levels. (B) down-regulation of gene with significantly down-regulated transcript levels. The data represent the means ± standard deviations (n = 3). Fig. S3. Rational metabolic engineering to improve α-humulene production (DCW). YALI0_B21780g and YALI0_B21142g were down-regulated. YALI0_E32835g was overexpressed. The data represent the means ± standard deviations (n = 3).

The sesquiterpene α-humulene has been shown to have anti-inflammatory and anticancer activities, which has led to its vast application potential in medicine. However, α-humulene production methods including phytoextraction and chemical synthesis currently were limited to low yield, high costs, and expensive catalysts, which cannot meet the increasing market demand. In this study, Yarrowia lipolytica was developed as a robust cell factory for α-humulene production. The peroxisome in Y. lipolytica was first engineered to boost the synthesis of the sesquiterpene α-humulene. By compartmentalization of the α-humulene biosynthesis pathway, improving ATP and acetyl-CoA supply, and optimizing the gene copy numbers of rate-limiting enzymes, the engineered strain GQ2012 could produce 3.2 g/L α-humulene in a 5 L bioreactor, the highest α-humulene titer reported so far. Our study provides a valuable reference for highly sustainable production of terpenoids by peroxisome engineering in Y. lipolytica.

Arachidonic acid is an essential ω-6 polyunsaturated fatty acid, which plays a significant role in cardiovascular health and neurological development, leading to its wide use in the food and pharmaceutical industries. Traditionally, ARA is obtained from deep-sea fish oil. However, this source is limited by season and is depleting the already threatened global fish stocks. With the rapid development of synthetic biology in recent years, oleaginous fungi have gradually attracted increasing attention as promising microbial sources for large-scale ARA production. Numerous advanced technologies including metabolic engineering, dynamic regulation of fermentation conditions, and multiomics analysis were successfully adapted to increase ARA synthesis. This review summarizes recent advances in the bioengineering of oleaginous fungi for ARA production. Finally, perspectives for future engineering approaches are proposed to further improve the titer yield and productivity of ARA.

Microbial production of value-added chemicals derived from fatty acids is a sustainable alternative to petroleum-derived chemicals and unsustainable lipids from animals and plants. Fatty acids with different carbon chain lengths including short- (C6), medium- (C6-C12), long- (C14-C20), and very-long- (C20), with either even or odd number of carbons, have significantly different characteristics and wide applications in energy, material, medicine, and nutrition. Tailoring chain-length specificity of these compounds using metabolic engineering would be of high interest.

Microbial production of value-added chemicals derived from fatty acids is a sustainable alternative to petroleum-derived chemicals and unsustainable lipids from animals and plants. Fatty acids with different carbon chain lengths including short- (C20), with either even or odd number of carbons, have significantly different characteristics and wide applications in energy, material, medicine, and nutrition. Tailoring chain-length specificity of these compounds using metabolic engineering would be of high interest. Yarrowia lipolytica, as an oleaginous yeast, is a superior industrial chassis for the production of tailored chain-length fatty acids and their derivatives due to its hyper-oil-producing capability. In this Review, we cover metabolic engineering approaches that can lead to fatty acid chain length control in this microorganism. These approaches involve the manipulation of the fatty acid synthase, the thioesterase, the β-oxidation pathway, the elongation and desaturation pathway, the polyketide synthase-like polyunsaturated fatty acid synthase pathway, and the odd-chain fatty acids synthesis pathway. Finally, we also discuss alternative strategies that can be used in the future to tailored chain-length control.