Automated yeast cultivation control using a biosensor and flow cytometry.

Unlabelled: Effective microbial bioprocessing relies on maintaining ideal cultivation conditions, highlighting the necessity for tools that monitor and regulate cellular performance and robustness. This study evaluates a fed-batch cultivation control system based on at-line flow cytometry monitoring of intact yeast cells having a fluorescent transcription factor-based redox biosensor. Specifically, the biosensor assesses the response of an industrial xylose-fermenting Saccharomyces cerevisiae strain carrying the TRX2p-yEGFP biosensor for NADPH/NADP+ ratio imbalance when exposed to furfural.

Evaluation of Pyrophosphate-Driven Proton Pumps in under Stress Conditions.

In , pH homeostasis is reliant on ATP due to the use of proton-translocating ATPase (H-ATPase) which constitutes a major drain within cellular ATP supply. Here, an exogenous proton-translocating pyrophosphatase (H-PPase) from which uses inorganic pyrophosphate (PP) rather than ATP, was evaluated for its effect on reducing the ATP burden. The H-Ppase was localized to the vacuolar membrane or to the cell membrane, and their impact was studied under acetate stress at a low pH.

Re-evaluation of the impact of deletion on xylose utilization by .

Various rational metabolic engineering and random approaches have been applied to introduce and improve xylose utilization and ethanol productivity by . Among them, the gene was identified as an interesting candidate for enhancing xylose consumption as its deletion appeared to be sufficient to improve growth, substrate utilization and ethanol productivity on xylose, even in a laboratory strain lacking a heterologous xylose pathway. The present study aimed at studying the influence of deletion in recombinant strains carrying heterologous oxido-reductive xylose utilization pathway.

Assessment of the TRX2p-yEGFP Biosensor to Monitor the Redox Response of an Industrial Xylose-Fermenting Strain during Propagation and Fermentation.

The commercial production of bioethanol from lignocellulosic biomass such as wheat straw requires utilizing a microorganism that can withstand all the stressors encountered in the process while fermenting all the sugars in the biomass. Therefore, it is essential to develop tools for monitoring and controlling the cellular fitness during both cell propagation and sugar fermentation to ethanol. In the present study, on-line flow cytometry was adopted to assess the response of the biosensor TRX2p-yEGFP for redox imbalance in an industrial xylose-fermenting strain of during cell propagation and the following fermentation of wheat-straw hydrolysate.

Draft Genome Assembly of sp. Strain S1 and Achromobacter spanius Strain S4, Two Syringol-Metabolizing Bacteria Isolated from Compost Soil.

Two bacterial strains able to use syringol as a sole carbon source were isolated from compost. The isolates, named S1 and S4, were sequenced using the Illumina platform. The final assemblies contained 4.

Assessment of fluorescent protein candidates for multi-color flow cytometry analysis of .

Transcription factor-based biosensors represent promising tools in the construction and evaluation of efficient cell factories for the sustainable production of fuels, chemicals and pharmaceuticals. They can notably be designed to follow the production of a target compound or to monitor key cellular properties, such as stress or starvation. In most cases, the biosensors are built with fluorescent protein (FP) genes as reporter genes because of the direct correlation between promoter activity and fluorescence level that can be measured using, for instance, flow cytometry or fluorometry.

D-Xylose Sensing in : Insights from D-Glucose Signaling and Native D-Xylose Utilizers.

Extension of the substrate range is among one of the metabolic engineering goals for microorganisms used in biotechnological processes because it enables the use of a wide range of raw materials as substrates. One of the most prominent examples is the engineering of baker's yeast for the utilization of d-xylose, a five-carbon sugar found in high abundance in lignocellulosic biomass and a key substrate to achieve good process economy in chemical production from renewable and non-edible plant feedstocks. Despite many excellent engineering strategies that have allowed recombinant to ferment d-xylose to ethanol at high yields, the consumption rate of d-xylose is still significantly lower than that of its preferred sugar d-glucose.

Vanillin Production in : Whole-Genome Sequencing of sp. Strain 9.1 and Reannotation of Pseudomonas putida CalA as a Vanillin Reductase.

Microbial degradation of lignin and its related aromatic compounds has great potential for the sustainable production of chemicals and bioremediation of contaminated soils. We previously isolated sp. strain 9.

Identification of modifications procuring growth on xylose in recombinant Saccharomyces cerevisiae strains carrying the Weimberg pathway.

The most prevalent xylose-assimilating pathways in recombinant Saccharomyces cerevisiae, i.e. the xylose isomerase (XI) and the xylose reductase/xylitol dehydrogenase (XR/XDH) pathways, channel the carbon flux through the pentose phosphate pathway and further into glycolysis.

Exploring the xylose paradox in Saccharomyces cerevisiae through in vivo sugar signalomics of targeted deletants.

Background: There have been many successful strategies to implement xylose metabolism in Saccharomyces cerevisiae, but no effort has so far enabled xylose utilization at rates comparable to that of glucose (the preferred sugar of this yeast). Many studies have pointed towards the engineered yeast not sensing that xylose is a fermentable carbon source despite growing and fermenting on it, which is paradoxical. We have previously used fluorescent biosensor strains to in vivo monitor the sugar signalome in yeast engineered with xylose reductase and xylitol dehydrogenase (XR/XDH) and have established that S.

Physiological characterization and sequence analysis of a syringate-consuming Actinobacterium.

Hardwood lignin is made of up to 75% syringyl-units and the bioconversion of syringate and syringaldehyde is therefore of considerable interest for biological valorization of lignin. In the current study, we have isolated a syringate-consuming bacterium identified as Microbacterium sp. RG1 and characterized its growth on several lignin model compounds.

Mapping the diversity of microbial lignin catabolism: experiences from the eLignin database.

Lignin is a heterogeneous aromatic biopolymer and a major constituent of lignocellulosic biomass, such as wood and agricultural residues. Despite the high amount of aromatic carbon present, the severe recalcitrance of the lignin macromolecule makes it difficult to convert into value-added products. In nature, lignin and lignin-derived aromatic compounds are catabolized by a consortia of microbes specialized at breaking down the natural lignin and its constituents.

Bacterial conversion of depolymerized Kraft lignin.

Background: Lignin is a potential feedstock for microbial conversion into various chemicals. However, the microbial degradation rate of native or technical lignin is low, and chemical depolymerization is needed to obtain reasonable conversion rates. In the current study, nine bacterial strains belonging to the and genera were evaluated for their ability to grow on alkaline-treated softwood lignin as a sole carbon source.

Retraction Note to: Bacterial conversion of depolymerized Kraft lignin.

[This retracts the article DOI: 10.1186/s13068-018-1240-7.].

Bacterial conversion of depolymerized Kraft lignin.

Background: Lignin is a potential feedstock for microbial conversion into various chemicals. However, the degradation rate of native or technical lignin is low, and depolymerization is needed to obtain reasonable conversion rates. In the current study, base-catalyzed depolymerization-using NaOH (5 wt%)-of softwood Kraft lignin was conducted in a continuous-flow reactor system at temperatures in the range 190-240 °C and residence times of 1 or 2 min.

Exploring D-xylose oxidation in Saccharomyces cerevisiae through the Weimberg pathway.

Engineering of the yeast Saccharomyces cerevisiae towards efficient D-xylose assimilation has been a major focus over the last decades since D-xylose is the second most abundant sugar in nature, and its conversion into products could significantly improve process economy in biomass-based processes. Up to now, two different metabolic routes have been introduced via genetic engineering, consisting of either the isomerization or the oxido-reduction of D-xylose to D-xylulose that is further connected to the pentose phosphate pathway and glycolysis. In the present study, cytosolic D-xylose oxidation was investigated instead, through the introduction of the Weimberg pathway from Caulobacter crescentus in S.

Assessing the effect of d-xylose on the sugar signaling pathways of Saccharomyces cerevisiae in strains engineered for xylose transport and assimilation.

One of the challenges of establishing an industrially competitive process to ferment lignocellulose to value-added products using Saccharomyces cerevisiae is to get efficient mixed sugar fermentations. Despite successful metabolic engineering strategies, the xylose assimilation rates of recombinant S. cerevisiae remain significantly lower than for the preferred carbon source, glucose.

Conversion of lignin model compounds by Pseudomonas putida KT2440 and isolates from compost.

Starting from mature vegetable compost, four bacterial strains were selected using a lignin-rich medium. 16S ribosomal RNA identification of the isolates showed high score similarity with Pseudomonas spp. for three out of four isolates.

Effect of nitrogen availability on the poly-3-D-hydroxybutyrate accumulation by engineered Saccharomyces cerevisiae.

Poly-3-D-hydroxybutyrate (or PHB) is a polyester which can be used in the production of biodegradable plastics from renewable resources. It is naturally produced by several bacteria as a response to nutrient starvation in the excess of a carbon source. The yeast Saccharomyces cerevisiae could be an alternative production host as it offers good inhibitor tolerance towards weak acids and phenolic compounds and does not depolymerize the produced PHB.

Anaerobic poly-3-D-hydroxybutyrate production from xylose in recombinant Saccharomyces cerevisiae using a NADH-dependent acetoacetyl-CoA reductase.

Background: Poly-3-D-hydroxybutyrate (PHB) that is a promising precursor for bioplastic with similar physical properties as polypropylene, is naturally produced by several bacterial species. The bacterial pathway is comprised of the three enzymes β-ketothiolase, acetoacetyl-CoA reductase (AAR) and PHB synthase, which all together convert acetyl-CoA into PHB. Heterologous expression of the pathway genes from Cupriavidus necator has enabled PHB production in the yeast Saccharomyces cerevisiae from glucose as well as from xylose, after introduction of the fungal xylose utilization pathway from Scheffersomyces stipitis including xylose reductase (XR) and xylitol dehydrogenase (XDH).

Real-time monitoring of the sugar sensing in Saccharomyces cerevisiae indicates endogenous mechanisms for xylose signaling.

Background: The sugar sensing and carbon catabolite repression in Baker's yeast Saccharomyces cerevisiae is governed by three major signaling pathways that connect carbon source recognition with transcriptional regulation. Here we present a screening method based on a non-invasive in vivo reporter system for real-time, single-cell screening of the sugar signaling state in S. cerevisiae in response to changing carbon conditions, with a main focus on the response to glucose and xylose.

Biological valorization of low molecular weight lignin.

Lignin is a major component of lignocellulosic biomass and as such, it is processed in enormous amounts in the pulp and paper industry worldwide. In such industry it mainly serves the purpose of a fuel to provide process steam and electricity, and to a minor extent to provide low grade heat for external purposes. Also from other biorefinery concepts, including 2nd generation ethanol, increasing amounts of lignin will be generated.

Adaptation to low pH and lignocellulosic inhibitors resulting in ethanolic fermentation and growth of Saccharomyces cerevisiae.

Lignocellulosic bioethanol from renewable feedstocks using Saccharomyces cerevisiae is a promising alternative to fossil fuels owing to environmental challenges. S. cerevisiae is frequently challenged by bacterial contamination and a combination of lignocellulosic inhibitors formed during the pre-treatment, in terms of growth, ethanol yield and productivity.