Structural characterization of the citrate transporter CexA uncovers the role of key residues S75, R192 and Q196.

The CexA transporter belongs to the DHA1 (Drug-H antiporter) family. CexA homologs are exclusively found in eukaryotic genomes, and CexA is the sole citrate exporter to have been functionally characterized in this family so far. In the present work, we expressed CexA in , demonstrating its ability to bind isocitric acid, and import citrate at pH 5.

Process engineering towards an oxidative cellular state improves 3-hydroxypropionic acid production with Lentilactobacillus diolivorans.

3-hydroxypropionic acid (3-HP) is among the top platform chemicals proposed for bio based production by microbial fermentation from renewable resources. A promising renewable substrate for 3-HP production is crude glycerol. Only a few microorganisms can efficiently convert glycerol to 3-HP.

Secretory expression of recombinant small laccase genes in Gram-positive bacteria.

Background: Laccases are multicopper enzymes that oxidize a wide range of aromatic and non-aromatic compounds in the presence of oxygen. The majority of industrially relevant laccases are derived from fungi and are produced in eukaryotic expression systems such as Pichia pastoris and Saccharomyces cerevisiae. Bacterial laccases for research purposes are mostly produced intracellularly in Escherichia coli, but secretory expression systems are needed for future applications.

Synthetic two-species allodiploid and three-species allotetraploid Saccharomyces hybrids with euploid (complete) parental subgenomes.

Combination of the genomes of Saccharomyces species has great potential for the construction of new industrial strains as well as for the study of the process of speciation. However, these species are reproductively isolated by a double sterility barrier. The first barrier is mainly due to the failure of the chromosomes to pair in allodiploid meiosis.

Synthetic activation of yeast stress response improves secretion of recombinant proteins.

Yeasts, such as Pichia pastoris (syn Komagataella spp.), are particularly suitable expression systems for emerging classes of recombinant proteins. Among them, recombinant antibody fragments, such as single-chain variable fragments (scFv) and single-domain antibodies (VHH), are credible alternatives to monoclonal antibodies.

Conversion of CO into organic acids by engineered autotrophic yeast.

The increase of CO emissions due to human activity is one of the preeminent reasons for the present climate crisis. In addition, considering the increasing demand for renewable resources, the upcycling of CO as a feedstock gains an extensive importance to establish CO-neutral or CO-negative industrial processes independent of agricultural resources. Here we assess whether synthetic autotrophic () can be used as a platform for value-added chemicals using CO as a feedstock by integrating the heterologous genes for lactic and itaconic acid synthesis.

Impact of ergosterol content on acetic and lactic acids toxicity to Saccharomyces cerevisiae.

Organic acid stress often represents a major hurdle in industrial bio-based microbial processes. Organic acids can be released from lignocellulosic feedstocks pretreatment and can also be desirable products obtained by microbial fermentation with applications in different industrial sectors. Yeasts are prominent cell factories.

Pushing and pulling proteins into the yeast secretory pathway enhances recombinant protein secretion.

Yeasts and especially Pichia pastoris (syn Komagataella spp.) are popular microbial expression systems for the production of recombinant proteins. One of the key advantages of yeast host systems is their ability to secrete the recombinant protein into the culture media.

From strain engineering to process development: monoclonal antibody production with an unnatural amino acid in Pichia pastoris.

Background: Expansion of the genetic code is a frequently employed approach for the modification of recombinant protein properties. It involves reassignment of a codon to another, e.g.

Artificial neural network modeling and statistical optimization of medium components to enhance production of exopolysaccharide by sp. EPS003.

Microbial Exopolysaccharides (EPS) have a wide range of applications in food, cosmetics, agriculture, pharmaceutical industries, and environmental bioremediation. The present study aims at enhancing the production of EPS from a soil-isolate sp. EPS003.

Genotypic and phenotypic diversity among Komagataella species reveals a hidden pathway for xylose utilization.

Background: The yeast genus Komagataella currently consists of seven methylotrophic species isolated from tree environments. Well-characterized strains of K. phaffii and K.

An artificial coculture fermentation system for industrial propanol production.

Converting plant biomass into biofuels and biochemicals microbial fermentation has received considerable attention in the quest for finding renewable energies and materials. Most approaches have so far relied on cultivating a single microbial strain, tailored for a specific purpose. However, this contrasts to how nature works, where microbial communities rather than single species perform all tasks.

Insights into the glycerol transport of Yarrowia lipolytica.

Cellular membranes separate cells from the environment and hence, from molecules essential for their survival. To overcome this hurdle, cells developed specialized transport proteins for the transfer of metabolites across these membranes. Crucial metabolites that need to cross the membrane of each living organism, are the carbon sources.

Yeasts from temperate forests.

Yeasts are ubiquitous in temperate forests. While this broad habitat is well-defined, the yeasts inhabiting it and their life cycles, niches, and contributions to ecosystem functioning are less understood. Yeasts are present on nearly all sampled substrates in temperate forests worldwide.

Going beyond the limit: Increasing global translation activity leads to increased productivity of recombinant secreted proteins in Pichia pastoris.

Yeasts are widely used cell factories for commercial heterologous protein production, however, specific productivities are usually tightly coupled to biomass formation. This greatly impacts production processes, which are commonly not run at the maximum growth rate, thereby resulting in suboptimal productivities. To tackle this issue, we evaluated transcriptomics datasets of the yeast Pichia pastoris (syn.

Improvement of 3-hydroxypropionic acid tolerance in Klebsiella pneumoniae by novel transporter YohJK.

3-Hydroxypropionic acid (3-HP) is a platform chemical which has potential applications in cosmetic and polymer industries. Microbial production of 3-HP is hampered by its toxic effect when its concentration is high (>300 mM). In this study, the effect of yohJK overexpression (via yieP deletion or episomal overexpression) on 3-HP tolerance was investigated in Klebsiella pneumoniae, Pseudomonas denitrificans and P.

What makes Komagataella phaffii non-conventional?

The important industrial protein production host Komagataella phaffii (syn Pichia pastoris) is classified as a non-conventional yeast. But what exactly makes K. phaffii non-conventional? In this review, we set out to address the main differences to the 'conventional' yeast Saccharomyces cerevisiae, but also pinpoint differences to other non-conventional yeasts used in biotechnology.

Adaptive laboratory evolution and reverse engineering enhances autotrophic growth in Pichia pastoris.

Synthetic biology offers several routes for CO conversion into biomass or bio-chemicals, helping to avoid unsustainable use of organic feedstocks, which negatively contribute to climate change. The use of well-known industrial organisms, such as the methylotrophic yeast Pichia pastoris (Komagataella phaffii), for the establishment of novel C1-based bioproduction platforms could wean biotechnology from feedstocks with alternative use in food production. Recently, the central carbon metabolism of P.

Utilizing yeasts for the conversion of renewable feedstocks to sugar alcohols - a review.

Sugar alcohols are widely marketed compounds. They are useful building block chemicals and of particular value as low- or non-calorigenic sweeteners, serving as sugar substitutes in the food industry. To date most sugar alcohols are produced by chemical routes using pure sugars, but a transition towards the use of renewable, non-edible feedstocks is anticipated.

The Plasma Membrane at the Cornerstone Between Flexibility and Adaptability: Implications for as a Cell Factory.

In the last decade, microbial-based biotechnological processes are paving the way toward sustainability as they implemented the use of renewable feedstocks. Nonetheless, the viability and competitiveness of these processes are often limited due to harsh conditions such as: the presence of feedstock-derived inhibitors including weak acids, non-uniform nature of the substrates, osmotic pressure, high temperature, extreme pH. These factors are detrimental for microbial cell factories as a whole, but more specifically the impact on the cell's membrane is often overlooked.

Fermenting Futures: an artistic view on yeast biotechnology.

BioArt is a new discipline where artists employ materials and techniques of modern life sciences and create novel meanings of biology, often involving living organisms such as tissue culture, bacteria and yeasts, which may also be genetically engineered. The authors have engaged in a collaboration to develop 'Fermenting Futures', a project designed to explore the significance of yeast for early human history by enabling baking and brewing, all the way to industrial biotechnology and synthetic biology with their potential contributions to fight the climate change. Research in two of the authors' lab provides the materials and thematic lines for the artists to develop their installations.

Sclerotia formed by citric acid producing strains of Aspergillus niger: Induction and morphological analysis.

Some strains of Aspergillus niger have been previously reported to produce sclerotia under certain conditions. Sclerotia are aggregations of hyphae which can act either as survival or as sexual structures in species related to A. niger.

Efficient conversion of hemicellulose sugars from spent sulfite liquor into optically pure L-lactic acid by Enterococcus mundtii.

Spent sulfite liquor (SSL), a waste stream from wood pulp production, has great potential as carbon source for future industrial fermentations. In the present study, SSL was separated into a hemicellulose derived sugar syrup (HDSS) and a lignosulfonic fraction by simulated moving bed chromatography. The recovery of SSL sugars in the HDSS was 89% and the fermentation inhibitors furfural, 5-hydroxymethylfurfural and acetic acid were removed by 98.

Established tools and emerging trends for the production of recombinant proteins and metabolites in Pichia pastoris.

Besides bakers' yeast, the methylotrophic yeast Komagataella phaffii (also known as Pichia pastoris) has been developed into the most popular yeast cell factory for the production of heterologous proteins. Strong promoters, stable genetic constructs and a growing collection of freely available strains, tools and protocols have boosted this development equally as thorough genetic and cell biological characterization. This review provides an overview of state-of-the-art tools and techniques for working with P.

Treatment with surfactants enables quantification of translational activity by O-propargyl-puromycin labelling in yeast.

Background: Translation is an important point of regulation in protein synthesis. However, there is a limited number of methods available to measure global translation activity in yeast. Recently, O-propargyl-puromycin (OPP) labelling has been established for mammalian cells, but unmodified yeasts are unsusceptible to puromycin.