62 Search Results

Abstract Background Sugar alcohols are widely used as low-calorie sweeteners in the food and pharmaceutical industries. They can also be transformed into platform chemicals. Yarrowia lipolytica , an oleaginous yeast, is a promising host for producing many sugar alcohols. In this work, we tested whether heterologous expression of a recently identified sugar alcohol phosphatase (PYP) from Saccharomyces cerevisiae would increase sugar alcohol production in Y. lipolytica . Results Y. lipolytica was found natively to produce erythritol, mannitol, and arabitol during growth on glucose, fructose, mannose, and glycerol. Osmotic stress is known to increase sugar alcohol production, and was found tomore » significantly increase erythritol production during growth on glycerol. To better understand erythritol production from glycerol, since it was the most promising sugar alcohol, we measured the expression of key genes and intracellular metabolites. Osmotic stress increased the expression of several key genes in the glycerol catabolic pathway and the pentose phosphate pathway. Analysis of intracellular metabolites revealed that amino acids, sugar alcohols, and polyamines are produced at higher levels in response to osmotic stress. Heterologous overexpression of the sugar alcohol phosphatase increased erythritol production and glycerol utilization in Y. lipolytica . We further increased erythritol production by increasing the expression of native glycerol kinase (GK), and transketolase (TKL). This strain was able to produce 27.5 ± 0.7 g/L erythritol from glycerol during batch growth and 58.8 ± 1.68 g/L erythritol during fed-batch growth in shake-flasks experiments. In addition, the glycerol utilization was increased by 2.5-fold. We were also able to demonstrate that this strain efficiently produces erythritol from crude glycerol, a major byproduct of the biodiesel production. Conclusions We demonstrated the application of a promising enzyme for increasing erythritol production in Y. lipolytica . We were further able to boost production by combining the expression of this enzyme with other approaches known to increase erythritol production in Y. lipolytica . This suggest that this new enzyme provides an orthogonal route for boosting production and can be stacked with existing designs known to increase sugar alcohol production in yeast such as Y. lipolytica . Collectively, this work establishes a new route for increasing sugar alcohol production and further develops Y. lipolytica as a promising host for erythritol production from cheap substrates such as glycerol.« less

Sugar metabolism by Saccharomyces cerevisiae produces ample amounts of CO₂ under both aerobic and anaerobic conditions. High solubility of CO₂ in fermentation media, contributing to enjoyable sensory properties of sparkling wine and beers by S. cerevisiae, might affect yeast metabolism. To elucidate the overlooked effects of CO₂ on yeast metabolism, we examined glucose fermentation by S. cerevisiae under CO₂ as compared to N₂ and O₂ limited conditions. While both CO₂ and N₂ conditions are considered anaerobic, less glycerol and acetate but more ethanol were produced under CO₂ condition. Transcriptomic analysis revealed that significantly decreased mRNA levels of GPP1 coding formore » glycerol-3-phosphate phosphatase in glycerol synthesis explained the reduced glycerol production under CO₂ condition. Besides, transcriptional regulations in signal transduction, carbohydrate synthesis, heme synthesis, membrane and cell wall metabolism, and respiration were detected in response to CO₂. Interestingly, signal transduction was uniquely regulated under CO₂ condition, where upregulated genes (STE3, MSB2, WSC3, STE12, and TEC1) in the signal sensors and transcriptional factors suggested that MAPK signaling pathway plays a critical role in CO₂ sensing and CO₂-induced metabolisms in yeast. Furthermore, our study identifies CO₂ as an external stimulus for modulating metabolic activities in yeast and a transcriptional effector for diverse applications.« less

Abstract Plant cell wall hydrolysates contain not only sugars but also substantial amounts of acetate, a fermentation inhibitor that hinders bioconversion of lignocellulose. Despite the toxic and non-consumable nature of acetate during glucose metabolism, we demonstrate that acetate can be rapidly co-consumed with xylose by engineered Saccharomyces cerevisiae . The co-consumption leads to a metabolic re-configuration that boosts the synthesis of acetyl-CoA derived bioproducts, including triacetic acid lactone (TAL) and vitamin A, in engineered strains. Notably, by co-feeding xylose and acetate, an enginered strain produces 23.91 g/L TAL with a productivity of 0.29 g/L/h in bioreactor fermentation. This strain also completely convertsmore » a hemicellulose hydrolysate of switchgrass into 3.55 g/L TAL. These findings establish a versatile strategy that not only transforms an inhibitor into a valuable substrate but also expands the capacity of acetyl-CoA supply in S. cerevisiae for efficient bioconversion of cellulosic biomass.« less

L-malic acid is widely used in the food, chemical, and pharmaceutical industries. Here, we report on production of malic acid from xylose, the second most abundant sugar in lignocellulosic hydrolysates, by engineered Saccharomyces cerevisiae. To enable malic acid production in a xylose-assimilating S. cerevisiae, we overexpressed PYC1 and PYC2, coding for pyruvate carboxylases, a truncated MDH3 coding for malate dehydrogenase, and SpMAE1, coding for a Schizosaccharomyces pombe malate transporter. Additionally, both the ethanol and glycerol-producing pathways were blocked to enhance malic acid production. The resulting strain produced malic acid from both glucose and xylose, but it produced much higher titersmore » of malic acid from xylose than glucose. Interestingly, the engineered strain had higher malic acid yield from lower concentrations (10 g L^–1) of xylose, with no ethanol production, than from higher xylose concentrations (20 and 40 g L^–1). As such, a fed-batch culture maintaining xylose concentrations at low levels was conducted and 61.2 g L^–1 of malic acid was produced, with a productivity of 0.32 g L^–1 h. Furthermore, these results represent successful engineering of S. cerevisiae for the production of malic acid from xylose, confirming that that xylose offers the efficient production of various biofuels and chemicals by engineered S. cerevisiae.« less

Although microalgae produce value-added products, such as lipids, pigments, and polysaccharides using light and carbon dioxide, these intracellular products require costly downstream processes such as extraction and purification. Thus, extracellular products are desirable for economic production. While reported before, the secretion of glycolic acid by microalgal photorespiration has not received attention for industrial applications. Here we developed a two-stage continuous cultivation system to increase glycolic acid production using a glycolate dehydrogenase (GYD1) deficient mutant of Chlamydomonas reinhardtii which produces high concentrations of glycolic acid. Specifically, 3% CO₂ was supplied in the first-stage culture for the production of biomass and ambientmore » air (0.03% CO₂) was supplied to the second stage for the production of glycolic acid. As a result, overall glycolic acid productivity reached 82.0 mg L^-1 d^-1 at a dilution rate of 0.34 d^-1. However, as the pH of the second stage decreased to 4.7 due to the increased glycolic acid production, we controlled the pH of the second stage at pH 6.0, resulting in 122.6 mg L^-1 d^-1 of glycolic acid productivity. Flux balance analysis revealed that the experimental glycolic acid production rate was 69% of the theoretical glycolic acid production rate. The deviation might be due to the toxicity of glycolic acid. When a techno-economic analysis was conducted based on the experimental results, the minimum glycolic acid production cost was estimated to be $31 kg^-1, indicating a potential for industrial production. Our findings suggest that microalgae can be utilized for the cost-effective industrial production of glycolic acid.« less

2,3-Butanediol (2,3-BDO) is a promising commodity chemical with various industrial applications. While petroleum-based chemical processes currently dominate the industrial production of 2,3-BDO, fermentation-based production of 2,3-BDO pro- vides an attractive alternative to chemical-based processes with regards to economic and environmental sustainability. The achievement of high 2,3-BDO titer, yield, and productivity in microbial fermentation is a prerequisite for the production of 2,3-BDO at large scales. Also, enantiopure production of 2,3-BDO production is desirable because 2,3-BDO stereoisomers have unique physicochemical properties. Pursuant to these goals, many metabolic engineering strategies to improve 2,3-BDO pro- duction from inexpensive sugars by Klebsiella oxytoca, Bacillus species,more » and Saccharomyces cerevisiae have been developed. Furthermore, this review summarizes the recent advances in metabolic engineering of non-pathogenic microorganisms to enable efficient and enantiopure production of 2,3-BDO.« less

Aldehydes are ubiquitous electrophilic compounds that ferment microorganisms including Saccharomyces cerevisiae encounter during the fermentation processes to produce food, fuels, chemicals, and pharmaceuticals. Aldehydes pose severe toxicity to the growth and metabolism of the S. cerevisiae through a variety of toxic molecular mechanisms, predominantly via damaging macromolecules and hampering the production of targeted compounds. Compounds with aldehyde functional groups are far more toxic to S. cerevisiae than all other functional classes, and toxic potency depends on physicochemical characteristics of aldehydes. The yeast synthetic biology community established a design–build–test–learn framework to develop S. cerevisiae cell factories to valorize the sustainable andmore » renewable biomass, including the lignin-derived substrates. However, thermochemically pretreated biomass-derived substrate streams contain diverse aldehydes (e.g., glycolaldehyde and furfural), and biological conversions routes of lignocellulosic compounds consist of toxic aldehyde intermediates (e.g., formaldehyde and methylglyoxal), and some of the high-value targeted products have aldehyde functional group (e.g., vanillin and benzaldehyde). Numerous studies comprehensively characterized both single and additive effects of aldehyde toxicity via systems biology investigations, and novel molecular approaches have been discovered to overcome the aldehyde toxicity. Based on those novel approaches, researchers successfully developed synthetic yeast cell factories to convert lignocellulosic substrates to valuable products, including aldehyde compounds. Here, we highlight the salient relationship of physicochemical charac- teristics and molecular toxicity of aldehydes, the molecular detoxification and macromolecules protection mechanisms of aldehydes, and the advances of engineering robust S. cerevisiae against complex mixtures of aldehyde inhibitors.« less

The halotolerant and osmotolerant yeast Zygosaccharomyces rouxii can produce multiple volatile compounds and has the ability to grow on lignocellulosic hydrolysates. We report the annotated genome sequence of Z. rouxii NRRL Y-64007 to support its development as a platform organism for biofuel and bioproduct production.