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Computational strain optimisation methods (CSOMs) have been successfully used to exploit genome-scale metabolic models, yielding strategies useful for allowing compound overproduction in metabolic cell factories. Minimal cut sets are particularly interesting since their definition allows searching for intervention strategies that impose strong growth-coupling phenotypes, and are not subject to optimality bias when compared with simulation-based CSOMs. However, since both types of methods have different underlying principles, they also imply different ways to formulate metabolic engineering problems, posing an obstacle when comparing their outputs. ; "DeYeastLibrary – Designer yeast strain library optimized for metabolic engineering applications", Ref.ERA-IB-2/0003/2013, funded by national funds through FCT/MCTES, DD-DeCaf and SHIKIFACTORY100, both funded by the European Union through the Horizon 2020 research and innovation programme (grant agreements no. 686070 and 814408). This study was also supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2019 unit and BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte. The authors acknowledge the use of computing facilities within the scope of the Search-ON2: Revitalization of HPC infrastructure of UMinho" project (NORTE-07-0162-FEDER-000086), co-funded by the North Portugal Regional Operational Programme (ON.2 – O Novo Norte), under the National Strategic Reference Framework (NSRF), through the European Regional Development Fund (ERDF). VV also thanks funding from FCT/MCTES for the PhD studentship with reference SFRH/BD/118657/2016. ...

The Supplementary Material for this article can be found online at: https://www.frontiersin.org/articles/10.3389/fmicb. 2019.01283/full#supplementary-material ; A high-quality GSM model for Streptococcus pneumoniae R6 model strain (iDS372), comprising 372 genes and 529 reactions, was developed. The construction of this model involved performing a genome-wide reannotation to identify the metabolic capacity of the bacterium. A reaction representing the abstraction of the biomass composition was reconciled from several studies reported in the literature and previous models, and included in the model. The final model comprises two compartments and manifold automatically generated gene rules. The validation was performed with experimental data from recent studies, regarding the usability of carbon sources, the effect of the presence of oxygen and the requirement of amino acids for growth. This model can be used to better understand the metabolism of this major pathogen, provide clues regarding new drug targets and eventually design strategies for fighting infections by these bacteria. ; This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469 unit and COMPETE 2020 (POCI-01-0145-FEDER-006684) and BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 – Programa Operacional Regional doNorte. This project has received funding from the European Union's Horizon 2020 Research and Innovation Programme under grant agreementno.686070. ...

The polyphenol resveratrol (3,5,4-trihydroxystilbene) is a well-known plant secondary metabolite, commonly used as a medical ingredient and a nutritional supplement. Due to its health-promoting properties, the demand for resveratrol is expected to continue growing. This stilbene can be found in different plants, including grapes, berries (blackberries, blueberries and raspberries), peanuts and their derived food products, such as wine and juice. The commercially available resveratrol is usually extracted from plants, however this procedure has several drawbacks such as low concentration of the product of interest, seasonal variation, risk of plant diseases and product stability. Alternative production processes are being developed to enable the biotechnological production of resveratrol by genetically engineering several microbial hosts, such as Escherichia coli, Corynebacterium glutamicum, Lactococcus lactis, among others. However, these bacterial species are not able to naturally synthetize resveratrol and therefore genetic modifications have been performed. The application of emerging metabolic engineering offers new possibilities for strain and process optimization. This mini-review will discuss the recent progress on resveratrol biosynthesis in engineered bacteria, with a special focus on the metabolic engineering modifications, as well as the optimization of the production process. These strategies offer new tools to overcome the limitations and challenges for microbial production of resveratrol in industry. ; We would like to thank the European Union Framework Program 7 "BacHBerry" (http://www.bachb erry.eu), Project No. FP7-613793 for financial support, the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469 unit, COMPETE 2020 (POCI-01-0145FEDER-006684) and BiotecNorte operation (NORTE-01-0145FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte. ...

BACKGROUND The growing interest in polyphenols has led to the design of industrial-scale processes able to produce them by fermentation and recover them in a more sustainable way. The goal of this work is to present two integrated approaches for the recovery of resveratrol, obtained through fermentation. The production of resveratrol using Corynebacterium glutamicum and its continuous removal using a hydrophobic resin is described. Batch production is compared with in situ product removal, where Amberlite XAD-7HP is either directly added to the medium (direct adsorption) or is present in an external column (external adsorption). RESULTS For both adsorption strategies tested, the amount of extracellular resveratrol increased from 75% to at least 90% of the total amount produced. However, lower total resveratrol concentrations were attained 3.6 and 2.2mg L-1, for the external and direct contact strategies, respectively, versus 5.3mg L-1 for batch experiments. CONCLUSIONS The proposed in situ removal strategies demonstrated the potential of increasing the excretion of resveratrol produced intracellularly. These process configurations may not only lead to a simpler downstream process design, but also to the avoidance of potential problems with the toxicity of polyphenols to the cells, especially when larger titers are obtained. © 2017 Society of Chemical Industry ; We would like to thank the European Union Framework Program 7 'BacHBerry' (www.bachberry.eu), Project No. FP7- 613793 for financial support, the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2013 unit, COMPETE 2020 (POCI-01- 0145-FEDER-006684) and BioTecNorte operation (NORTE-01-0145- FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte. ...

The synthesis of galacto-oligosaccharides (GOS) by the action of Aspergillus oryzae β-galactosidase free and immobilized on magnetic polysiloxane-polyvinyl alcohol (mPOS-PVA) was studied. A maximum GOS concentration of 26% (w/v) of total sugars was achieved at near 55% lactose conversion from 50%, w/v lactose solution at pH 4.5 and 40 °C. Trisaccharides accounted for more than 81% of the total GOS produced. GOS formation was not considerably affected by pH and temperature. The concentrations of glucose and galactose encountered near maximum GOS concentration greatly inhibited the reactions and reduced GOS yield. GOS formation was not affected by enzyme immobilization in the mPOS-PVA matrix, indicating the absence of diffusional limitations in the enzyme carrier. Furthermore, this water insoluble magnetic derivative was reutilized 10-times and retained about 84% of the initial activity. In addition, the kinetic parameters for various initial lactose concentrations were determined and compared for the free and immobilized enzyme. ; David F. M. Neri gratefully acknowledges support by the Programme Alpan, the European Union Programme of High Level Scholarships for Latin America (Scholarship No. E05D057787BR). Luiz B. Carvalho Jr. is recipient of a scholarship of the Brazilian National Research Council ...

In recent years microorganisms have been engineered towards synthesizing interesting plant polyphenols such as flavonoids and stilbenes from glucose. Currently, low endogenous supply of malonylCoA, indispensable for plant polyphenol synthesis, impedes high product titers. Usually, limited malonylCoA availability during plant polyphenol production is avoided by supplementing fatty acid synthesisinhibiting antibiotics such as cerulenin, which are known to increase the intracellular malonylCoA pool as a side effect. Motivated by the goal of microbial polyphenol synthesis being independent of such expensive additives, we used rational metabolic engineering approaches to modulate regulation of fatty acid synthesis and flux into the citric acid cycle in Corynebacterium glutamicum strains capable of flavonoid and stilbene synthesis. Initial experiments showed that sole overexpression of genes coding for the native malonylCoAforming acetylCoA carboxylase is not sufficient for increasing polyphenol production in C. glutamicum. Hence, the intracellular acetylCoA availability was also increased by reducing the flux into the citric acid cycle through reduction of citrate synthase activity. In defined cultivation medium, the constructed C. glutamicum strains accumulated 24 mg/L (0.088 mM) naringenin or 112 mg/L (0.49 mM) resveratrol from glucose without supplementation of phenylpropanoid precursor molecules or any inhibitors of fatty acid synthesis. ; We would like to thank the European Union Framework Program 7 ''BacHBerry'' (www.bachberry.eu), Project No. FP7‐ 613793 for financial support and also the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/ BIO/04469 unit, COMPETE 2020 (POCI‐01–0145‐FEDER‐006684) and BiotecNorte operation (NORTE‐01–0145‐FEDER‐000004) funded by the European Regional Development Fund under the scope of Norte2020 ‐ Programa Operacional Regional do Norte. Furthermore, we would like to thank Dr. Tino Polen for providing the plasmids to replace the ...