Optimizing hexanoic acid biosynthesis in Saccharomyces cerevisiae for the de novo production of olivetolic acid.

Medium chain fatty acids (MCFAs) are valuable platform compounds for the production of biotechnologically relevant chemicals such as biofuels and biochemicals. Two distinct pathways have been implemented in the yeast Saccharomyces cerevisiae for the biosynthetic production of MCFAs: (i) the mutant fatty acid biosynthesis (FAB) pathway in which the fatty acid synthase (FAS) complex is mutated and (ii) a heterologous multispecies-derived reverse β-oxidation (rBOX) pathway. Hexanoic acid has become of great interest as its acyl-CoA ester, hexanoyl-CoA, is required for the biosynthesis of olivetolic acid (OA), a cannabinoid precursor.

ME2 Deficiency Is Associated With Recessive Neurodevelopmental Disorder.

Malate is an important dicarboxylic acid produced from fumarate in the tricarboxylic acid cycle. Deficiencies of fumarate hydrolase (FH) and malate dehydrogenase (MDH), responsible for malate formation and metabolism, respectively, are known to cause recessive forms of neurodevelopmental disorders (NDDs). The malic enzyme isoforms, malic enzyme 1 (ME1) and 2 (ME2), are required for the conversion of malate to pyruvate.

A comparative analysis of NADPH supply strategies in Production of d-xylitol from d-xylose as a case study.

Enhancing the supply of the redox cofactor NADPH in metabolically engineered cells is a critical target for optimizing the synthesis of many product classes, such as fatty acids or terpenoids. In , several successful approaches have been developed in different experimental contexts. However, their systematic comparison has not been reported.

An optimized reverse β-oxidation pathway to produce selected medium-chain fatty acids in Saccharomyces cerevisiae.

Background: Medium-chain fatty acids are molecules with applications in different industries and with growing demand. However, the current methods for their extraction are not environmentally sustainable. The reverse β-oxidation pathway is an energy-efficient pathway that produces medium-chain fatty acids in microorganisms, and its use in Saccharomyces cerevisiae, a broadly used industrial microorganism, is desired.

Development of vitamin B12 dependency in Saccharomyces cerevisiae.

For decades, the industrial vitamin B12 (cobalamin) production has been based on bacterial producer strains. Due to limited methods for strain optimization and difficult strain handling, the desire for new vitamin B12-producing hosts has risen. As a vitamin B12-independent organism with a big toolbox for genomic engineering and easy-to-handle cultivation conditions, Saccharomyces cerevisiae has high potential for heterologous vitamin B12 production.

A yeast-based in vivo assay system for analyzing efflux of sugars mediated by glucose and xylose transporters.

Sugar transporter research focuses on the sugar uptake into cells. Under certain physiological conditions, however, the intracellular accumulation and secretion of carbohydrates (efflux) are relevant processes in many cell types. Currently, no cell-based system is available for specifically investigating glucose efflux.

EFR3 and phosphatidylinositol 4-kinase IIIα regulate insulin-stimulated glucose transport and GLUT4 dispersal in 3T3-L1 adipocytes.

Insulin stimulates glucose transport in muscle and adipocytes. This is achieved by regulated delivery of intracellular glucose transporter (GLUT4)-containing vesicles to the plasma membrane where they dock and fuse, resulting in increased cell surface GLUT4 levels. Recent work identified a potential further regulatory step, in which insulin increases the dispersal of GLUT4 in the plasma membrane away from the sites of vesicle fusion.

Identification of a glucose-insensitive variant of Gal2 from Saccharomyces cerevisiae exhibiting a high pentose transport capacity.

As abundant carbohydrates in renewable feedstocks, such as pectin-rich and lignocellulosic hydrolysates, the pentoses arabinose and xylose are regarded as important substrates for production of biofuels and chemicals by engineered microbial hosts. Their efficient transport across the cellular membrane is a prerequisite for economically viable fermentation processes. Thus, there is a need for transporter variants exhibiting a high transport rate of pentoses, especially in the presence of glucose, another major constituent of biomass-based feedstocks.

Subcellular Localization of Fad1p in : A Choice at Post-Transcriptional Level?

FAD synthase is the last enzyme in the pathway that converts riboflavin into FAD. In , the gene encoding for FAD synthase is , from which a sole protein product (Fad1p) is expected to be generated. In this work, we showed that a natural Fad1p exists in yeast mitochondria and that, in its recombinant form, the protein is able, per se, to both enter mitochondria and to be destined to cytosol.

Production of octanoic acid in Saccharomyces cerevisiae: Investigation of new precursor supply engineering strategies and intrinsic limitations.

The eight-carbon fatty acid octanoic acid (OA) is an important platform chemical and precursor of many industrially relevant products. Its microbial biosynthesis is regarded as a promising alternative to current unsustainable production methods. In Saccharomyces cerevisiae, the production of OA had been previously achieved by rational engineering of the fatty acid synthase.

High-Throughput Screening of an Octanoic Acid Producer Strain Library Enables Detection of New Targets for Increasing Titers in .

Octanoic acid is an industrially relevant compound with applications in antimicrobials or as a precursor for biofuels. Microbial biosynthesis through yeast is a promising alternative to current unsustainable production methods. To increase octanoic acid titers in , we use a previously developed biosensor that is based on the octanoic acid responsive promotor coupled to GFP.

Transcriptomic response of Saccharomyces cerevisiae to octanoic acid production.

The medium-chain fatty acid octanoic acid is an important platform compound widely used in industry. The microbial production from sugars in Saccharomyces cerevisiae is a promising alternative to current non-sustainable production methods, however, titers need to be further increased. To achieve this, it is essential to have in-depth knowledge about the cell physiology during octanoic acid production.

Improving 3-methylphenol (m-cresol) production in yeast via in vivo glycosylation or methylation.

Heterologous expression of 6-methylsalicylic acid synthase (MSAS) together with 6-MSA decarboxylase enables de novo production of the platform chemical and antiseptic additive 3-methylphenol (3-MP) in the yeast Saccharomyces cerevisiae. However, toxicity of 3-MP prevents higher production levels. In this study, we evaluated in vivo detoxification strategies to overcome limitations of 3-MP production.

Artificial ER-Derived Vesicles as Synthetic Organelles for Compartmentalization of Biochemical Pathways.

Compartmentalization in membrane-surrounded organelles has the potential to overcome obstacles associated with the engineering of metabolic pathways, such as unwanted side reactions, accumulation of toxic intermediates, drain of intermediates out of the cell, and long diffusion distances. Strategies utilizing natural organelles suffer from the presence of endogenous pathways. In our approach, we make use of endoplasmic reticulum-derived vesicles loaded with enzymes of a metabolic pathway ("metabolic vesicles").

On the natural spatio-temporal heterogeneity of South Pacific nitrous oxide.

Nitrous oxide (NO) is a powerful greenhouse gas and ozone depleting substance, but its natural sources, especially marine emissions, are poorly constrained. Localized high concentrations have been observed in the oxygen minimum zones (OMZs) of the tropical Pacific but the impacts of El Niño cycles on this key source region are unknown. Here we show atmospheric monitoring station measurements in Samoa combined with atmospheric back-trajectories provide novel information on NO variability across the South Pacific.

Fusing α and β subunits of the fungal fatty acid synthase leads to improved production of fatty acids.

Most fungal fatty acid synthases assemble from two multidomain subunits, α and β, into a heterododecameric FAS complex. It has been recently shown that the complex assembly occurs in a cotranslational manner and is initiated by an interaction between the termini of α and β subunits. This initial engagement of subunits may be the rate-limiting phase of the assembly and subject to cellular regulation.

biosynthesis of 8-hydroxyoctanoic acid via a medium-chain length specific fatty acid synthase and cytochrome P450 in .

Terminally hydroxylated fatty acids or dicarboxylic acids are industrially relevant compounds with broad applications. Here, we present the proof of principle for the biosynthesis of 8-hydroxyoctanoic acid from glucose and ethanol in the yeast . Toxicity tests with medium-chain length ω-hydroxy fatty acids and dicarboxylic acids revealed little or no growth impairments on yeast cultures even at higher concentrations.

Identification and characterisation of two high-affinity glucose transporters from the spoilage yeast Brettanomyces bruxellensis.

The yeast Brettanomyces bruxellensis (syn. Dekkera bruxellensis) is an emerging and undesirable contaminant in industrial low-sugar ethanol fermentations that employ the yeast Saccharomyces cerevisiae. High-affinity glucose import in B.

Improving isobutanol production with the yeast by successively blocking competing metabolic pathways as well as ethanol and glycerol formation.

Background: Isobutanol is a promising candidate as second-generation biofuel and has several advantages compared to bioethanol. Another benefit of isobutanol is that it is already formed as a by-product in fermentations with the yeast , although only in very small amounts. Isobutanol formation results from valine degradation in the cytosol via the Ehrlich pathway.

A superfolder variant of pH-sensitive pHluorin for in vivo pH measurements in the endoplasmic reticulum.

Many cellular processes are regulated via pH, and maintaining the pH of different organelles is crucial for cell survival. A pH-sensitive GFP variant, the so-called pHluorin, has proven to be a valuable tool to study the pH of the cytosol, mitochondria and other organelles in vivo. We found that the fluorescence intensity of Endoplasmic Reticulum (ER)-targeted pHluorin in the yeast Saccharomyces cerevisiae was very low and barely showed pH sensitivity, probably due to misfolding in the oxidative environment of the ER.