Engineering a biosynthetic pathway for high-value glycosaminoglycans production

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Saccharomyces cerevisiae as a host for chondroitin production

Chondroitin is a glycosaminoglycan that has gained widespread use in nutraceuticals and pharmaceuticals, mainly for treating osteoarthritis. Traditionally, it has been extracted from animal cartilage but recently, biotechnological processes have emerged as a commercial alternative to avoid the risk of viral or prion contamination and offer a vegan-friendly source. Typically, these methods involve producing the chondroitin backbone using pathogenic bacteria and then modifying it enzymatically through the action of sulfotransferases. Despite the challenges of expressing active sulfotransferases in bacteria, the use of eukaryotic microorganisms is still limited to a few works using Pichia pastoris. To create a safer and efficient biotechnological platform, we constructed a biosynthetic pathway for chondroitin production in S. cerevisiae as a proof-of-concept. Up to 125 mg/L and 200 mg/L of intracellular and extracellular chondroitin were produced, respectively. Furthermore, as genome-scale models are valuable tools for identifying novel targets for metabolic engineering, a stoichiometric model of chondroitin-producing S. cerevisiae was developed and used in optimization algorithms. Our research yielded several novel targets, such as uridine diphosphate (UDP)-Nacetylglucosamine pyrophosphorylase (QRI1), glucosamine-6-phosphate acetyltransferase (GNA1), or N-acetylglucosamine-phosphate mutase (PCM1) overexpression, that might enhance chondroitin production and guide future experimental research to develop more efficient host organisms for the biotechnological production process.This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UIDB/04469/2020 unit with DOI 10.54499/UIDB/04469/2020. The authors acknowledge FCT for funding MRC doctoral grant SFRH/BD/132998/2017 and further extension COVID/BD/152454/2022.info:eu-repo/semantics/publishedVersio

Development of a low-cost culture medium for biopolymer production by Rhizobium viscosum CECT 908 and its potential application in Microbial Enhanced Oil Recovery

Polymers are a versatile class of compounds that play an essential role in our society, being their production estimated in more than 180 million tons per year. Nowadays, the world market is dominated by synthetic and plant-derived polymers. Biopolymers of microbial origin are characterized by their better environmental compatibility and biodegradability when compared with the synthetic ones, and their production is faster than those obtained from plants. Microbial biopolymers usually exhibit excellent rheological properties, stability at a wide range of temperatures, salinities and pH values, as well as a broad variety of chemical structures, which results in different physicochemical and rheological properties. However, despite their outstanding properties, their application is still limited by their high production costs. In this work, an alternative low-cost culture medium was developed for biopolymer production by Rhizobium viscosum CECT 908, containing sugarcane molasses (60 g/L) and corn steep liquor (1%, v/v) as carbon and nitrogen sources, respectively. Using this low-cost medium, higher biopolymer production and apparent viscosity values (5.2 g/L and 6700 mPa s, respectively) were obtained comparing with the synthetic medium (2.3 g/L and 1100 mPa s), which contained glucose and yeast extract. As a result, the cost of the culture medium necessary to produce 1 Kg of biopolymer was reduced more than 20 times. The biopolymer produced in the alternative low-cost medium exhibited better rheological properties as compared to xanthan gum, including higher viscosity at the same concentration. Furthermore, it was found to be stable at temperatures up to 80ºC, NaCl concentrations as high as 200 g/L, and high shear rates (300 s-1). Polymers are widely used by the oil industry to increase the oil reservoirs productivity during the tertiary oil recovery processes. In sand-pack column assays performed using a heavy crude oil (40ºC= 170 mPa s), this biopolymer produced using the low-cost medium demonstrated a better performance than xanthan gum, recovering almost 50% of the entrapped oil. Results herein obtained highlight that the R. viscosum biopolymer is a promising candidate for application in MEOR as an alternative to the conventional microbial and synthetic polymers currently used.info:eu-repo/semantics/publishedVersio

Synthesis of novel psoralen analogues and their anti-proliferative effect on human cancer cell lines

We describe the synthesis of 3H-benzofuro[3,2-f]chromen-3-ones. The anti-proliferative effect on human cancer cell lines (MDA-MB231 and HeLa) was evaluated.FCT and FEDER, for National NMR Networ

A proposed methodology for understanding urban growth pattern : a case study in Siem Reap, Cambodia

In this paper, the main goal is to understand the relationship between urban growth and physical factors in order to determine the potential area for future urban expansion. A methodology is suggested for understanding urban growth pattern in Siem Reap which could effectively sustain archaeological sites and to balance the land use between urban and non-urban areas in Siem Reap, Cambodia. Remote sensing technique is used to analyze land use maps of Siem Reap from 1993 to 2011. Results show that urban-built up area increased significantly which causes the forest land to reduce steadily from 1993 to 2003 in the Siem Reap archaeological sites. In addition, Geographic Information System (GIS) is applied to analyze urban growth pattern. Geo-processing and logical functions are applied to detect and quantify the land use changes, especially urban changes. Two main factors are used to analyze the urban driving growth in Siem Reap, which are distance to road networks and population density. Pearson correlation statistics is applied to justify the relationship between the factors and urban area growth

Electric vehicle and conventional vehicle. Urban air pollution assessment

Road traffic is the main source of air pollutant emissions in urban areas. Pollutant emissions can be evaluated and calculated using mathematical models based on the characteristics of traffic flowing in roads. This article aims to assess the air pollution produced by a fleet of vehicles with conventional (diesel and petrol) and electrical (hybrid and electric) motorization, in the city centre of Braga. The scientific toolbox adopted to develop the studies includes emission models to estimate emission data and a GIS platform

Modeling urban growth pattern for sustainable archaeological sites : a case study in Siem Reap, Cambodia

In this paper, the main goal is to understand the relationship between urban growth and physical factors in order to determine the potential area for future urban expansion. A policy is suggested that could effectively sustain the archaeological sites and to balance the land use between urban and non-urban areas in Siem Reap, Cambodia. Remote sensing is used to analyze land use maps of Siem Reap from 1993 to 2011. Results show that urban-built up area increased significantly which causes the forest land to reduce in the Siem Reap archaeological sites. In addition, Geographic Information System (GIS) is used to analyze urban growth in potential suitable sites. Geo-processing and logical functions are applied to detect and quantify the land use changes, especially urban changes. The percentage of urban area in each year is compared with the population density and road buffers by using Pearson correlation. It is shown that the increasing in urban area is related with population density and road network factors

Development of an electrochemical aptasensor for the detection of human osteopontin

Electrochemical aptasensors, an emerging technology, enables the detection of protein biomarkers, which may be indicative of tumour activity. Osteopontin is a protein present in body fluids, being a possible biomarker since its overexpression has been related with breast cancer progression. An \RNA\ aptamer, described in the literature, with affinity for human osteopontin, was synthetized, immobilized in a microelectrode gold surface and used for development electrochemical aptasensor for human \OPN\ detection in standard solutions. Cyclic voltammetry results showed that this aptasensor allowed detecting human osteopontin with a detection limit of 8 nM, showing a satisfactory selectivity towards the target in the presence of others proteins, except for thrombin

Avaliação de zonas de criticidade acústica: aplicação das potencialidades de um SIG na avaliação do ruído de uma cidade de média dimensão

O crescimento urbano, pelas dimensões que actualmente assume, vem exercendo pressões de forma continuada nos recursos, nas infra-estruturas e nos equipamentos, afectando negativamente o standard de vida dos cidadãos. Neste contexto, a avaliação e monitorização da qualidade do ambiente urbano tornou se um tema de primordial importância quando considerado como um instrumento de apoio à decisão para a construção de cidades mais sustentáveis e com melhor qualidade de vida. Viana do Castelo é uma cidade de média dimensão localizada no litoral norte de Portugal e que aceitou o desafio de desenvolver uma programa para a melhoria da qualidade do seu ambiente urbano, com o objectivo de aderir ao Projecto Cidades Saudáveis e integrar a Rede Europeia das Cidades Saudáveis. Neste programa, a caracterização do ruído urbano, nomeadamente a determinação dos níveis de intensidade sonora na zona urbana e da exposição da população ao ruído ambiente, foi considerada uma das acções prioritárias. A ferramenta adoptada para desenvolver estes estudos inclui modelos de previsão de ruído numa plataforma de SIG. Com base em dados de tráfego e nas características físicas do local foram desenvolvidos mapas de ruído e procedeu-se ao seu cruzamento com o zonamento acústico do território e com a população residente. Esta combinação foi a base para a identificação das zonas de criticidade acústica, em termos de níveis de ruído e dos índices de exposição da população a esses níveis de ruído. O presente artigo apresenta uma abordagem deste problema, passando pelos fundamentos teóricos e pelo estudo desenvolvido para a cidade de Viana do Castelo

Synthesis of novel benzofurocoumarin analogues and their anti-proliferative effect on human cancer cell lines

The synthesis of five new tetracyclic benzofurocoumarin (benzopsoralen) analogues is described. Their inhibitory effects on the growth of three human tumour cell lines (MDA MB 231 (breast adenocarcinoma), HeLa (cervix adenocarcinoma) and TCC-SUP (bladder transitional cell carcinoma)) were evaluated, and discussed in terms of structure–activity relationship.FCT and FEDER, for National NMR Network (Bruker Avance II 400), REEQ/630/QUI/2005 (LC/MS instrument) and the PhD grant (SFRH/BD/48636/2008)