Recombinant CBM-fusion technology : applications overview

Carbohydrate-binding modules (CBMs) are small components of several enzymes, which present an independent fold and function, and specific carbohydrate-binding activity. Their major function is to bind the enzyme to the substrate enhancing its catalytic activity, especially in the case of insoluble substrates. The immense diversity of CBMs, together with their unique properties, has long raised their attention for many biotechnological applications. Recombinant DNA technology has been used for cloning and characterizing new CBMs. In addition, it has been employed to improve the purity and availability of many CBMs, but mainly, to construct bi-functional CBM-fused proteins for specific applications. This review presents a comprehensive summary of the uses of CBMs recombinantly produced from heterologous organisms, or by the original host, along with the latest advances. Emphasis is given particularly to the applications of recombinant CBM-fusions in: (a) modification of fibers, (b) production, purification and immobilization of recombinant proteins, (c) functionalization of biomaterials and (d) development of microarrays and probes.Fundação para a Ciência e a Tecnologia (FCT), Portugal (grants SFRH/BDP/63831/ 2009 and SFRH/BPD/73850/2010, respectively). The authors thank the FCT GlycoCBMs Project REF. PTDC/AGR-FOR/3090/2012 — FCOMP-01- 0124-FEDER-027948, the FCT Strategic Project PEst-OE/EQB/LA0023/ 2013, and the Project “BioInd — Biotechnology and Bioengineering for improved Industrial and Agro-Food processes”, REF. NORTE-07-0124- FEDER-000028 Co-funded by the Programa Operacional Regional do Norte (ON.2 — O Novo Norte), QREN, FEDER

Modification of paper properties using carbohydrate-binding module 3 from the Clostridium thermocellum CipA scaffolding protein produced in Pichia pastoris: elucidation of the glycosylation effect

The carbohydrate-binding modules (CBMs) have emerged as an interesting alternative to enzymes for fibers modification, e.g. of pulp and paper. Glycosylation in CBMs is thought to have a key role in the improvement of cellulose fibers. Thus, in this work the non-glycosylated (CBM3mt) and glycosylated (CBM3wt) recombinant versions of CBM3 from Clostridium thermocellum CipAboth produced in Pichia pastoriswere studied. Binding assays showed that CBM3mt had a higher affinity for microcrystalline cellulose (Avicel) than CBM3wt. In addition, CBM3mt produced a much higher hydrophobization of Whatman paper than CBM3wt. However, the effects of the two CBM3s on pulp and paper were identical. The CBM3s did not affect the drainability of Eucalyptus globulus or a mixture of E. globulus and Pinus sylvestris pulps. On the other hand, both improved significantly strength-related properties of E. globulus papersheets, namely burst (up to 12 %) and tensile strength (up to 10 %) indexes. This is the first report showing the capacity of CBM3 from C. termocellum CipA to modify paper properties. The results showed that glycosylation did not influence the drainage of CBM3-treated pulps nor the properties of the produced papers. Thus, glycans in glycosylated CBM3 may not be related with fiber improvement, namely superior pulp drainage.C. Oliveira acknowledges support from Fundacao para a Ciencia e a Tecnologia (FCT), Portugal (Grant SFRH/BDP/63831/2009). The authors thank the FCT GlycoCBMs Project REF. PTDC/AGR-FOR/3090/2012-FCOMP-01-0124-FEDER-027948, the FCT Strategic Project PEst-OE/EQB/LA0023/2013, and the Project "BioInd-Biotechnology and Bioengineering for improved Industrial and Agro-Food processes", REF. NORTE-07-0124-FEDER-000028 Co-funded by the Programa Operacional Regional do Norte (ON.2-O Novo Norte), QREN, FEDER

Role of oxidative stress-induced systemic and cavernosal molecular alterations in the progression of diabetic erectile dysfunction

Background Erectile dysfunction (ED) is a prevalent complication of diabetes, and oxidative stress is an important feature of diabetic ED. Oxidative stress-induced damage plays a pivotal role in the development of tissue alterations. However, the deleterious effects of oxidative stress in the corpus cavernosum with the progression of diabetes remain unclear. The aim of this study was to evaluate systemic and penile oxidative stress status in the early and late stages of diabetes. Methods Male Wistar streptozotocin-diabetic rats (and age-matched controls) were examined 2 (early) and 8 weeks (late) after the induction of diabetes. Systemic oxidative stress was evaluated by urinary H2O2 and the ratio of circulating reduced/oxidized glutathione (GSH/GSSG). Penile oxidative status was assessed by H2O2 production and 3-nitrotyrosine (3-NT) formation. Cavernosal endothelial nitric oxide synthase (eNOS) was analyzed by quantitative immunohistochemistry. Dual immunofluorescence was also performed for 3-NT and α-smooth muscle actin (α-SMA) and eNOS–α-SMA. Results There was a significant increase in urinary H2O2 levels in both diabetic groups. The plasma GSH/GSSG ratio was significantly augmented in late diabetes. In cavernosal tissue, H2O2 production was significantly increased in late diabetes. Reactivity for 3-NT was located predominantly in cavernosal smooth muscle (SM) and was significantly reduced in late diabetes. Quantitative immunohistochemistry revealed a significant decrease in eNOS levels in cavernosal SM and endothelium in late diabetes. Conclusions The findings indicate that the noxious effects of oxidative stress are more prominent in late diabetes. Increased penile protein oxidative modifications and decreased eNOS expression may be responsible for structural and/or functional deregulation, contributing to the progression of diabetes-associated ED

Food losses and wastes in brazil: a systematic review

Globally, it is estimated that approximately a third of the food produced is lost or wasted. Reducing half of these losses would make it possible to increase the availability of food for another 1 billion people. Specifically, in Brazil, the amount of lost and wasted food, if used, would be enough to achieve food security, considering as a restriction for this security, the availability of food for purchase. In this context, the objective is to identify methodologies that measure losses and food waste in Brazil. For this, a systematic review of the subject was made. It can be observed that there are a few studies that quantify food losses and wastage in the country. The largest concentration of these studies is in the distribution and consumption stages of the food supply chain. Information is available from food lost and waste, mainly in the consumption and distribution phases. Otherwise, in the other phases of the supply chain, food loss and waste values in Brazil are still incipient or non-existent. Studies that identify these values can contribute to the generation of more accurate information on the subject, being able to raise the values referring to food loss and waste of the country.Keywords: supply chain; food secutity; sustainability; hungry.Perdas e desperdícios alimentares no Brasil: uma revisão sistemáticaRESUMOEstima-se que, globalmente, aproximadamente um terço dos alimentos produzidos sejam perdidos ou desperdiçados. No entanto, a redução de metade dessas perdas tornaria possível aumentar a disponibilidade de alimentos para mais 1 bilhão de pessoas. Especificamente, no Brasil, a quantidade de alimentos perdidos e desperdiçados, se utilizados, seria suficiente para alcançar a segurança alimentar, considerando como uma restrição para essa segurança, a disponibilidade de alimentos para compra. Nesse contexto, o objetivo é identificar metodologias que mensurem perdas e desperdício de alimentos no Brasil. Para isso, foi realizada uma revisão sistemática do assunto. Pode-se observar que existem poucos estudos que quantificam perdas e desperdícios de alimentos no país. A maior concentração desses estudos está nos estágios de distribuição e consumo da cadeia de suprimento de alimentos. As informações estão disponíveis a partir de alimentos perdidos e desperdiçados, principalmente nas fases de consumo e distribuição. Nas demais fases da cadeia de suprimentos, os valores de perda e desperdício de alimentos no Brasil ainda são incipientes ou inexistentes. Estudos que identifiquem esses valores podem contribuir para a geração de informações mais precisas sobre o assunto, podendo elevar os valores referentes à perda e desperdício de alimentos no país.Palavras-chave: cadeia de suprimentos; segurança alimentar; sustentabilidade; fome

Enhancing the enzymatic saccharification of whole slurry from autohydrolyzed Eucalyptus globulus wood by supplementation with a recombinant carbohydrate-binding module

Microbiotec'17 - Congress of Microbiology and Biotechnology 2017Background: Lignocellulosic biomass has a recognised potential as a sustainable platform for the production of biofuels and other biochemicals. However, lignin residues and other inhibitory compounds resulting from lignocellulosics pretreatment affect the digestibility of resulting whole slurries. The addition of synergistic proteins that can cooperate with cellulases is an emerging strategy for enhanced lignocellulosics hydrolysis. Carbohydrate-binding modules (CBMs) have been shown to improve the enzymatic hydrolysis of pure cellulose models but their effect on the enzymatic hydrolysis of lignocellulosic materials has not yet been evaluated. Thus, in this work, the potential synergistic effect of a family 3 CBM on the enzymatic saccharification of a pretreated lignocellulosic biomass was studied for the first time. [...]C. Oliveira and D. Gomes acknowledge support from Fundação para a Ciência e a Tecnologia (FCT), Portugal (grants SFRH/BPD/110640/2015 and SFRH/BD/88623/2012, respectively). This study was supported by the FCT under the scope of the GlycoCBMs Project PTDC/AGR-FOR/3090/2012–FCOMP-01-0124-FEDER-027948 and the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI-01-0145-FEDER-006684) and BioTecNorte operation (NORTE-010145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte.info:eu-repo/semantics/publishedVersio

Lipidomic approach of the biological activity of tacrine and two tacrine-analogues on non-synaptic rat brain mitochondria

Foundation for Science and Technology (FCT), FEDER and COMPET

Food losses and wastes in brazil: a systematic review

Globally, it is estimated that approximately a third of the food produced is lost or wasted. Reducing half of these losses would make it possible to increase the availability of food for another 1 billion people. Specifically, in Brazil, the amount of lost and wasted food, if used, would be enough to achieve food security, considering as a restriction for this security, the availability of food for purchase. In this context, the objective is to identify methodologies that measure losses and food waste in Brazil. For this, a systematic review of the subject was made. It can be observed that there are a few studies that quantify food losses and wastage in the country. The largest concentration of these studies is in the distribution and consumption stages of the food supply chain. Information is available from food lost and waste, mainly in the consumption and distribution phases. Otherwise, in the other phases of the supply chain, food loss and waste values in Brazil are still incipient or non-existent. Studies that identify these values can contribute to the generation of more accurate information on the subject, being able to raise the values referring to food loss and waste of the country.Keywords: supply chain; food secutity; sustainability; hungry.Perdas e desperdícios alimentares no Brasil: uma revisão sistemáticaRESUMOEstima-se que, globalmente, aproximadamente um terço dos alimentos produzidos sejam perdidos ou desperdiçados. No entanto, a redução de metade dessas perdas tornaria possível aumentar a disponibilidade de alimentos para mais 1 bilhão de pessoas. Especificamente, no Brasil, a quantidade de alimentos perdidos e desperdiçados, se utilizados, seria suficiente para alcançar a segurança alimentar, considerando como uma restrição para essa segurança, a disponibilidade de alimentos para compra. Nesse contexto, o objetivo é identificar metodologias que mensurem perdas e desperdício de alimentos no Brasil. Para isso, foi realizada uma revisão sistemática do assunto. Pode-se observar que existem poucos estudos que quantificam perdas e desperdícios de alimentos no país. A maior concentração desses estudos está nos estágios de distribuição e consumo da cadeia de suprimento de alimentos. As informações estão disponíveis a partir de alimentos perdidos e desperdiçados, principalmente nas fases de consumo e distribuição. Nas demais fases da cadeia de suprimentos, os valores de perda e desperdício de alimentos no Brasil ainda são incipientes ou inexistentes. Estudos que identifiquem esses valores podem contribuir para a geração de informações mais precisas sobre o assunto, podendo elevar os valores referentes à perda e desperdício de alimentos no país.Palavras-chave: cadeia de suprimentos; segurança alimentar; sustentabilidade; fome

Persistent organic pollutant levels in human visceral and subcutaneous adipose tissue in obese individuals - Depot differences and dysmetabolism implications

Background: The role of persistent organic pollutants (POPs) with endocrine disrupting activity in the aetiology of obesity and other metabolic dysfunctions has been recently highlighted. Adipose tissue (AT) is a common site of POPs accumulation where they can induce adverse effects on human health. Objectives: To evaluate the presence of POPs in human visceral (vAT) and subcutaneous (scAT) adipose tissue in a sample of Portuguese obese patients that underwent bariatric surgery, and assess their putative association with metabolic disruption preoperatively, as well as with subsequent body mass index (BMI) reduction. Methods: AT samples (n=189) from obese patients (BMI ≥35) were collected and the levels of 13 POPs were determined by gas chromatography with electron-capture detection (GC-ECD). Anthropometric and biochemical data were collected at the time of surgery. BMI variation was evaluated after 12 months and adipocyte size was measured in AT samples. Results: Our data confirm that POPs are pervasive in this obese population (96.3% of detection on both tissues), their abundance increasing with age (RS=0.310, p<0.01) and duration of obesity (RS=0.170, p<0.05). We observed a difference in AT depot POPs storage capability, with higher levels of ΣPOPs in vAT (213.9±204.2 compared to 155.1±147.4 ng/g of fat, p<0.001), extremely relevant when evaluating their metabolic impact. Furthermore, there was a positive correlation between POP levels and the presence of metabolic syndrome components, namely dysglycaemia and hypertension, and more importantly with cardiovascular risk (RS=0.277, p<0.01), with relevance for vAT (RS=0.315, p<0.01). Finally, we observed an interesting relation of higher POP levels with lower weight loss in older patients. Conclusion: Our sample of obese subjects allowed us to highlight the importance of POPs stored in AT on the development of metabolic dysfunction in a context of obesity, shifting the focus to their metabolic effects and not only for their recognition as environmental obesogens

Association of 3-Phenoxybenzoic Acid Exposure during Pregnancy with Maternal Outcomes and Newborn Anthropometric Measures: Results from the IoMum Cohort Study

The aims of this study were to characterize the exposure of pregnant women living in Portugal to 3-phenoxybenzoic acid (3-PBA) and to evaluate the association of this exposure with maternal outcomes and newborn anthropometric measures. We also aimed to compare exposure in summer with exposure in winter. Pregnant women attending ultrasound scans from April 2018 to April 2019 at a central hospital in Porto, Portugal, were invited to participate. Inclusion criteria were: gestational week between 10 and 13, confirmed fetal vitality, and a signature of informed consent. 3-PBA was measured in spot urine samples by gas chromatography with mass spectrometry (GC-MS). The median 3-PBA concentration was 0.263 (0.167; 0.458) μg/g creatinine (n = 145). 3-PBA excretion was negatively associated with maternal pre-pregnancy body mass index (BMI) (p = 0.049), and it was higher during the summer when compared to winter (p < 0.001). The frequency of fish or yogurt consumption was associated positively with 3-PBA excretion, particularly during the winter (p = 0.002 and p = 0.015, respectively), when environmental exposure is low. Moreover, 3-PBA was associated with levothyroxine use (p = 0.01), a proxy for hypothyroidism, which could be due to a putative 3-PBA—thyroid hormone antagonistic effect. 3-PBA levels were not associated with the anthropometric measures of the newborn. In conclusion, pregnant women living in Portugal are exposed to 3-PBA, particularly during summer, and this exposure may be associated with maternal clinical features.This article was supported by national funds through the FCT Foundation for Science and Technology, I.P., within the scope of the projects RISE - LA/P/0053/2020; CINTESIS, R&D UNIT (reference UIDB/4255/2020) and LAQV (references: UIDB/50006/2020 AND UIDP/50006/2020). Virgínia Cruz Fernandes was funded by FCT/MCTES (Foundation for Science and Technology and Ministry of Science, Technology and Higher Education) and the ESF (European Social Fund) through NORTE 2020 (North Region Operational Program) through a grant of Post-Doc (reference SFRH/BPD/109153/2015). Juliana Guimarães was funded by FCT/MCTES (Foundation for Science and Technology and Ministry of Science, Technology and Higher Education) under CINTESIS by a PhD scholarship (reference UI/BD/152087/2021). The authors would like to thank the kind participation of all pregnant women and the support of the recruitment activities by the health professionals of the Department of Obstetrics, Centro Hospitalar Universitário S. João, Porto, Portugal and also to the professionals of REQUIMTE/LAQV, Instituto Superior de Engenharia, Politécnico do Porto, for their collaboration in carrying out the analyzes of the 3-PBA metabolite. This work received support from PT national funds (FCT/MCTES, Fundação para a Ciência e Tecnologia and Ministério da Ciência, Tecnologia e Ensino Superior) through the projects LA/P/0053/2020; UIDB/4255/2020; UIDB/5006/2020 and UIDP/50006/2020. Virgínia Cruz Fernandes thanks FCT/MCTES (Fundação para a Ciência e Tecnologia and Ministério da Ciência, Tecnologia e Ensino Superior) and ESF (European Social Fund) through NORTE 2020 (Programa Operacional Região Norte) for his/her Post-Doc grant ref. SFRH/BPD/109153/2015). Juliana Guimarães was funded by FCT/MCTES (Foundation for Science and Technology and Ministry of Science, Technology and Higher Education) under CINTESIS by a PhD scholarship (reference UI/BD/152087/2021).info:eu-repo/semantics/publishedVersio