iBB

Responding to the recent interest of the yeast research community in non-Saccharomyces cerevisiae species of biotechnological relevance, the N.C.Yeastract was associated to YEASTRACT+, a curated repository of known regulatory associations between transcription factors (TFs) and target genes in yeasts. A recent Minireview published in FEMS Yeast Research aims to advertise the update of the existing information since the release of N.C.Yeastract in 2019, and to raise awareness in the community about its potential to help the day-to-day work on non-Saccharomyces species, exploring all the information and bioinformatics tools available in YEASTRACT +. Using simple and widely used examples, a guided exploitation is offered. The usage potentialities of the new CommunityYeastract platform by the yeast community are also discussed. The Minireview is coauthored by a BSRG-iBB team coordinated by Isabel Sá-Correia and including Cláudia P. Godinho, Margarida Palma, Miguel C. Teixeira and the PhD students Miguel Antunes and Marta N. Mota, in collaboration with INESC-ID colleagues.

The nutritional composition and bioactive properties of roots and rhizomes of Asparagus stipularis were evaluated to demonstrate its potential in the food and pharmaceutical industries. HPLC-DAD-ESI/MS characterization of infusions allowed the identification and quantitation of 7 hydroxycinnamoyl derivatives, with caffeic acid as the most abundant. Roots infusion contained the highest amounts of these compounds. It also exhibited the highest antioxidant activity in all assays, with EC50 values of 0.44 ± 0.01, 0.98 ± 0.03 and 0.64 ± 0.01 mg/mL for DPPH, ABTS and FRAP assays, respectively, with no toxicity towards PLP2 primary cell cultures (GI50 > 400 μg/mL). The extract was encapsulated into PLGA nanoparticles obtaining a size of 260 nm and a polydispersity index around 0.1, with a zeta potential of -36 mV, as well as a good encapsulation efficiency of approximately 83%. The particles had a spherical morphology and smooth surface. FTIR and DSC assays confirmed the efficacy of the encapsulation methodology. This paper was published in Food Bioscience by Pedro Fonte from BERG-iBB in a transnational collaboration with researchers from Portugal, Spain, Italy and Tunisia. The developed systems will be used as delivery systems for bioactive compounds of A. stipularis and as an innovative dietary supplement.

Wound care is clinically demanding due to treatment inefficiency and represents an economic burden for healthcare systems. A promising therapeutic strategy is the use of exogenous growth factors that are decreased at the wound site and hence limit recovery of the skin. Insulin is one of the cheapest growth factors in the market able to accelerate the re-epithelialization and stimulate angiogenesis and cell migration. However, the effectiveness of topical insulin in wound healing is hampered by the proteases in the wound bed. The encapsulation into nanoparticles improves its stability in the wound, providing adhesion to the mucosal surface and allowing its sustained release. In a paper published in Materials, Pedro Fonte from BERG-iBB in a collaboration with researchers from CCMAR, University of Algarve and LAQV, University of Porto performed a standing point about a promising strategy to treat different types of wounds by the topical delivery of insulin-loaded nanocarriers.

Paraburkholderia sacchari has the capacity to produce xylonic acid and xylitol, compounds ranked in the top 30 high-value chemicals from biomass. In a recent paper in Biochemical Engineering Journal, Maryna Bondar, Manuela Fonseca and Teresa Cesário from BERG-iBB reveal the outstanding ability of this bacterium to metabolize D-xylose to xylonic acid. D-xylonic acid is a five-carbon sugar acid that can replace gluconic acid in several applications. The biotechnological production of D-xylonic acid is advantageous over gluconic acid because it uses xylose as carbon source. Xylose is a very abundant sugar in nature and only few native bacterial strains can metabolize it. Fed-batch cultivations in a single bioreactor attained xylonic acid titers of 390 g L-1 and a productivity of 7.7 g L-1 h-1. This simplified process can significantly affect process economics, potentiating its translation to an industrial scale.

Bacteria associated with coral hosts are diverse and abundant, with recent studies suggesting involvement of these symbionts in host resilience to anthropogenic stress. Combining published and unpublished data, a new article featuring iBB researchers Tina Keller-Costa and Rodrigo Costa among the authors provides a comprehensive overview of the diversity and function of culturable bacteria isolated from tropical, temperate, and cold-water corals. The study, published in the journal mSystems, compiles a total of 3,055 coral-associated isolates described in 52 reports from various laboratories around the world. The work presents a comparative genomic analysis of 74 strains and identifies signatures of potentially beneficial bacterium-coral symbioses among them. Such a resource is an important step in the selection of probiotic candidates, which are being investigated for promoting coral resilience and can potentially be applied in novel reef restoration and rehabilitation efforts. This genome and culture collection is available to the wider research community through the web site http://isolates.reefgenomics.org/ with the hope that many scientists across the globe will ask for access to these resources for future studies.

The determination of the recombination efficiency and the monitoring of parental plasmids (PP), minicircles (MC) and miniplasmid (MP) species during processing and in the final product are critical aspects of minicircle manufacturing. A paper published in Analytical Biochemistry by BERG-iBB researchers Cláudia Alves, Miguel Prazeres and Gabriel Monteiro describes a real-time PCR method for the specific identification of PP, MP or MC that uses sets of primers specific for each species. The results presented indicate that the method can be applied for the determination of recombination efficiency in pure DNA samples.

The mild operational conditions typically required by biobased production processes, together with the selective nature of biocatalysts, microbial diversity and tunability, and the century-long record of fermentation, clearly place bioprocesses as the primary contenders, by far, in achieving the generalized implementation of efficient, green manufacturing strategies. Moreover, by overlapping with the biorefinery approach, the foundations needed for bioprocesses to embrace the circular economy concept are set. In this Special Issue on “Bioprocess Engineering: Sustainable Manufacturing for a Green Society", guest edited by Pedro Fernandes and Carla de Carvalho (iBB-BERG) in Processes, the papers will discuss new bioproducts and concomitant bioengineering approaches to achieve a sustainable and environmentally friendly economy.

PI(4,5)P2 is a phospholipid found mostly in the plasma membrane of eukaryotic cells, where it plays a crucial role in processes like vesicle trafficking, cytoskeletal regulation, ion channel function, viral assembly and budding. While most phospholipids show considerable acyl-chain diversity, PI(4,5)P2 lipids are exceptionally enriched in specific acyl-chains, the most frequent composition in mammalian cells being 1-stearoyl-2-arachidonyl (18:0 20:4). The biological functions that call for this specific enrichment are still not fully clear. In a recent paper published in Communications Chemistry, a BSIRG-iBB team led by Fábio Fernandes together with the teams of Dr. Nuno Santos (IMM) and Dr. Manuel Melo (ITQB) identified a previously unreported increase in membrane order upon calcium-dependent PI(4,5)P2 clustering. Remarkably, the interaction of saturated PI(4,5)P2 with calcium culminated in the formation of gel nanodomains for fully saturated PI(4,5)P2, and the formation of these gel domains was abrogated in the presence of 18:0 20:4 polyunsaturated PI(4,5)P2. These results support a role of (18:0 20:4)PI(4,5)P2 in inhibiting the formation of highly ordered PI(4,5)P2 nanodomains in the plasma membrane.

Many studies that apply PnD in pre-clinical cutaneous wound healing models show large variations in the choice of the animal species (e.g., large animals, rodents), the choice of diabetic or non-diabetic animals, the type of injury (full-thickness wounds, burns, radiation-induced wounds, skin flaps), the source and type of PnD (placenta, umbilical cord, fetal membranes, cells, secretomes, tissue extracts), the method of administration (topical application, intradermal/subcutaneous injection, intravenous or intraperitoneal injection, subcutaneous implantation), and the type of delivery systems (e.g., hydrogels, synthetic or natural biomaterials as carriers for transplanted cells, extracts or secretomes). In a collaborative work coordinated by Prof. Pedro Fonte under the COST Action SPRINT (CA17116), the Postdoc researcher Ana Macedo and Master student Francisca Mendes from BERG-iBB, provided a comprehensive and integrative overview of the application of PnD in wound healing to assess its efficacy in preclinical animal models. The review was published in Frontiers in Bioengineering and Biotechnology.

Lipid hydroperoxides have recently been recognized as key mediators of diseases (such as neurodegenerative disorders or Type II diabetes) and cell death. In a recent work, structural and dynamic perturbations induced by the hydroperoxidized POPC lipid (POPC-OOH) in fluid POPC membranes were addressed using advanced small-angle X-ray scattering (SAXS) and fluorescence methodologies. Notably, this multidisciplinary approach revealed that the hydroperoxide group located at the membrane interface, promotes a higher membrane hydration and microviscosity, with a strikingly lower order and bending rigidity, an unusual trend in membrane biophysics, which ultimately compromises membrane structure organization. This international work co-led by Ana M. Melo (BSIRG-iBB) and Rosangela Itri (Institute of Physics, University of São Paulo) was recently published in Langmuir and involved other BSIRG-iBB researchers (Ana Coutinho, Alexander Fedorov and Manuel Prieto).

p28 is a 28 amino acids peptide derived from the bacterial protein azurin. It possesses cell-penetrating capabilities showing preferential enter in cancer cells. Moreover it has been subject in US to two phase I clinical trials as a anticancer agent. In a recent paper published in Journal of Controlled Release, a iBB team (Garizo AR, Dias TP, Fernandes F, Bernardes N, Fialho AM) together with a i3S/UP team (Castro F, Martins C, Almeida A, Barrias CC, Sarmento B) were able for the first time to fabricate p28-functionalized PLGA nanoparticles (NPs) loaded with the EGFR tyrosine kinase inhibitor gefitinib. The results obtained indicate that these NPs interact preferentially with lung cancer cells due to their decoration with p28 peptide. In vitro cytotoxicity assays demonstrate biological activity of the NPs against lung cancer cancer cells. Finally, in vivo studies demonstrated a great potential of the p28-NPs in enhancing the therapeutic effects of gefitinib.

Cassava (Manihot esculenta) cultivation is of great importance in many economies, particularly in Colombia. About 630,000 tons of C-rich cassava waste is produced annually and applications to high value products, applying the circular economy concept, must be developed. A recent publication in Journal of Polymers and the Environment assesses the potential use of cassava peel for the production of polyhydroxyalkanoates (PHAs) by Cupriavidus necator. A copolymer of P3HB-3HV was produced and processed into electrospun meshes of random and aligned microfibers, allowing the development of structures that can be applied in the context of tissue engineering. This work involved Manuela Fonseca, Frederico Ferreita and Teresa Cesário form BERG-IBB and has been done in collaboration with researchers from the University of Antioquia, Medellin-Colombia.

The production of recombinant proteins is gaining increasing importance as several applications request high quality proteins. However, several process parameters affect both the growth of cells and product yields. In a paper recently published in the journal Molecules, Ricardo FS Pereira and Carla CCR de Carvalho (iBB-BERG) used high throughput systems and statistical methods to assess the influence of fermentation conditions in lab-scale bioreactors. Using partial least squares, it was found that the height-to-diameter ratio of the bioreactor, aeration rate, and PID controller parameters contributed significantly to the final biomass and cytochrome concentrations. This information was used to fine-tune the process parameters, which increased cytochrome production and yield several-fold.

The role of the cell wall in yeast response and tolerance to stress is frequently neglected. A BSRG-iBB research paper just published in Scientific Reports, provides, for the first time, a comprehensive view of the alterations occurring at the cell wall in a yeast population adapting to sub-lethal stress induced by acetic acid. The results reveal changes to the cell wall polysaccharide composition and nanomechanical properties, as well as alterations in the transcript levels of key cell wall biosynthetic genes. This paper reinforces the notion that the adaptive yeast response to acetic acid involves coordinated alterations of the cell wall at the biophysical and molecular levels. The gathered knowledge is important for the design of superior industrial strains and for the efficient control of the deleterious activity of spoilage yeasts, particularly in the Food Industry. This research work is first-authored by the PhD student of the PhD programme in Biotechnology and Biosciences Ricardo Ribeiro (FCT_DP AEM fellowship), performed under the supervision of Isabel Sá-Correia. This collaborative study with Fábio Fernandes (BSIRG-iBB) and Mário S. Rodrigues and his team (BioISI, Faculty of Sciences, ULisboa), is also coauthored by Cláudia Godinho (posdoc researcher) and the PhD student Nuno Bourbon-Melo (FCT_DP BIOTECnico) from the BSRG-iBB team.

Materials with novel and enhanced functionalities can be obtained by modifying cellulose with a range of biomolecules. In a recent paper published in Materials, Mariana Barbosa, Hélvio Simões and Miguel Prazeres from BERG-iBB describe the modification of cellulose hydrogels using bi-functional biomolecular constructs based on carbohydrate binding modules (CBMs). The binding of CBM fusions, the capture of antibodies and the grafting of oligonucleotides onto hydrogels was successfully demonstrated. The CBM-based platform could contribute significantly to the development of advanced medical diagnostic sensors or specialized biomaterials, among others. The work was funded by project CBM-X.