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- Eriksson, Dennis, et al.
(författare)
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Improving Risk Assessment in the European Food Safety Authority: Lessons From the European Medicines Agency
- 2020
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Ingår i: Frontiers in Plant Science. - : Frontiers Media SA. - 1664-462X. ; 11
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Tidskriftsartikel (refereegranskat)abstract
- The recent Regulation (EU) 2019/1381, published on the 6th September 2019, aims to improve the transparency and sustainability of the EU risk assessment in the food chain by amending the General Food Law Regulation (EC 178/2002) and a number of other regulations related to the food sector. This Regulation is introduced as a response to the Fitness Check of the General Food Law Regulation as well as a response to public concerns expressed by a European Citizens' Initiative on glyphosate and pesticides. This article evaluates the amendments introduced by Regulation 2019/1381with respect to the institutional and regulatory environment in the food chain and more specifically concerning the risk assessment procedure. For this purpose, we perform a comparison of the institutional and organizational characteristics of the European Food Safety Authority (EFSA) and European Medicines Agency (EMA) in relation to the processes of risk assessment and risk evaluation, especially the processes surrounding genetically modified foods and pesticides, and how these characteristics affect the politicization of these processes. We conclude that the risk assessment process followed by EFSA would have benefitted and become more effective and less politicized, if the recent Regulation 2019/1381 had introduced some of EMA's institutional structures and methods on risk evaluation.
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| 4. |
- Bagan Navarro, Hector, et al.
(författare)
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Synthesis and characterization of epitope-imprinted polymers for purification of human hemoglobin.
- 2017
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Ingår i: RSC Advances. - 2046-2069. ; 7:66, s. 41705-41712
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Tidskriftsartikel (refereegranskat)abstract
- One promising method to prepare protein-selective polymers is the epitope-imprinting approach, where surface-accessible peptides from a target protein are used as templates to create surface-exposed binding sites on molecularly imprinted polymers (MIPs). However, selection of a suitable peptide target is not always straightforward, and synthesis of peptide on a large scale can be costly. In this work, we developed a new approach that can be used to select peptide epitopes on protein surface to be used as templates to prepare protein-selective MIPs. In this case study, human hemoglobin (Hb) was immobilized on silica nanoparticles and then fragmented by tryptic digestion. The particle-supported peptides were then used as templates to synthesize the Hb-selective MIPs, which were obtained after removal of the silica support and the peptides. The MIPs were tested in equilibrium binding experiments to evaluate their protein separation performance. The new surface imprinted MIPs displayed high selectivity for Hb, and was able to separate different variants of Hb from protein mixtures and crude cell extracts.
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| 5. |
- Christensen, Simon, et al.
(författare)
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Oxidative Implications of Substituting a Conserved Cysteine Residue in Sugar Beet Phytoglobin BvPgb 1.2
- 2022
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Ingår i: Antioxidants. - : MDPI AG. - 2076-3921. ; 11:8
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Tidskriftsartikel (refereegranskat)abstract
- Phytoglobins (Pgbs) are plant-originating heme proteins of the globin superfamily with varying degrees of hexacoordination. Pgbs have a conserved cysteine residue, the role of which is poorly understood. In this paper, we investigated the functional and structural role of cysteine in BvPgb1.2, a Class 1 Pgb from sugar beet (Beta vulgaris), by constructing an alanine-substituted mutant (Cys86Ala). The substitution had little impact on structure, dimerization, and heme loss as determined by X-ray crystallography, size-exclusion chromatography, and an apomyoglobin-based heme-loss assay, respectively. The substitution significantly affected other important biochemical properties. The autoxidation rate increased 16.7- and 14.4-fold for the mutant versus the native protein at 25 °C and 37 °C, respectively. Thermal stability similarly increased for the mutant by ~2.5 °C as measured by nano-differential scanning fluorimetry. Monitoring peroxidase activity over 7 days showed a 60% activity decrease in the native protein, from 33.7 to 20.2 U/mg protein. When comparing the two proteins, the mutant displayed a remarkable enzymatic stability as activity remained relatively constant throughout, albeit at a lower level, ~12 U/mg protein. This suggests that cysteine plays an important role in BvPgb1.2 function and stability, despite having seemingly little effect on its tertiary and quaternary structure.
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| 6. |
- Cooper, Chris E., et al.
(författare)
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Engineering tyrosine residues into hemoglobin enhances heme reduction, decreases oxidative stress and increases vascular retention of a hemoglobin based blood substitute
- 2019
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Ingår i: Free Radical Biology and Medicine. - : Elsevier BV. - 0891-5849. ; 134, s. 106-118
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Tidskriftsartikel (refereegranskat)abstract
- Hemoglobin (Hb)-based oxygen carriers (HBOC) are modified extracellular proteins, designed to replace or augment the oxygen-carrying capacity of erythrocytes. However, clinical results have generally been disappointing due to adverse side effects, in part linked to the intrinsic oxidative toxicity of Hb. Previously a redox-active tyrosine residue was engineered into the Hb β subunit (βF41Y) to facilitate electron transfer between endogenous antioxidants such as ascorbate and the oxidative ferryl heme species, converting the highly oxidizing ferryl species into the less reactive ferric (met) form. We inserted different single tyrosine mutations into the α and β subunits of Hb to determine if this effect of βF41Y was unique. Every mutation that was inserted within electron transfer range of the protein surface and the heme increased the rate of ferryl reduction. However, surprisingly, three of the mutations (βT84Y, αL91Y and βF85Y) also increased the rate of ascorbate reduction of ferric(met) Hb to ferrous(oxy) Hb. The rate enhancement was most evident at ascorbate concentrations equivalent to that found in plasma (< 100 μM), suggesting that it might be of benefit in decreasing oxidative stress in vivo. The most promising mutant (βT84Y) was stable with no increase in autoxidation or heme loss. A decrease in membrane damage following Hb addition to HEK cells correlated with the ability of βT84Y to maintain the protein in its oxygenated form. When PEGylated and injected into mice, βT84Y was shown to have an increased vascular half time compared to wild type PEGylated Hb. βT84Y represents a new class of mutations with the ability to enhance reduction of both ferryl and ferric Hb, and thus has potential to decrease adverse side effects as one component of a final HBOC product.
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| 7. |
- Eriksson, Nélida Leiva, et al.
(författare)
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Sugar beet hemoglobins : Reactions with nitric oxide and nitrite reveal differential roles for nitrogen metabolism
- 2019
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Ingår i: Biochemical Journal. - 0264-6021. ; 476:14, s. 2111-2125
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Tidskriftsartikel (refereegranskat)abstract
- In contrast with human hemoglobin (Hb) in red blood cells, plant Hbs do not transport oxygen, instead research points towards nitrogen metabolism. Using comprehensive and integrated biophysical methods we characterized three sugar beet Hbs: BvHb1.1, BvHb1.2 and BvHb2. Their affinities for oxygen, CO, and hexacoordination were determined. Their role in nitrogen metabolism was studied by assessing their ability to bind NO, to reduce nitrite (NiR, nitrite reductase), and to form nitrate (NOD, NO dioxygenase). Results show that BvHb1.2 has high NOD-like activity, in agreement with the high nitrate levels found in seeds where this protein is expressed. BvHb1.1, on the other side, is equally capable to bind NO as to form nitrate, its main role would be to protect chloroplasts from the deleterious effects of NO. Finally, the ubiquitous, reactive, and versatile BvHb2, able to adopt ‘open and closed forms’, would be part of metabolic pathways where the balance between oxygen and NO is essential. For all proteins, the NiR activity is relevant only when nitrite is present at high concentrations and both NO and oxygen are absent. The three proteins have distinct intrinsic capabilities to react with NO, oxygen and nitrite; however, it is their concentration which will determine the BvHbs’ activity.
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| 8. |
- Gutiérrez Román, Ana I.F., et al.
(författare)
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Production of Sustainable Proteins Through the Conversion of Insects to Proteins Using Beauveria bassiana Cultures
- 2022
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Ingår i: Frontiers in Sustainable Food Systems. - : Frontiers Media SA. - 2571-581X. ; 5
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Tidskriftsartikel (refereegranskat)abstract
- Various strategies are being suggested to solve the challenges in the food system, such as changing the source of nutrients, including the use of non-traditional food sources such as insects. Although insects are promoted as a cheap and sustainable source of protein, consumers are reluctant to eat them. The mycoproteins produced by fungi, on the other hand, are very well received and appreciated by consumers. Thus, in this work we have studied the use of the entomopathogenic fungi Beauveria bassiana (Ascomycota: Hypocreales) for the production of protein using insects as feed. B. bassiana was cultivated in culture medium containing entire insects from the species Eurysacca and Hypothenemus or single carbon sources such as glucose or laminarin from Laminaria digitata. The results showed that B. bassiana can produce up to 16-fold more biomass and 8-fold more protein when grown in insect-based medium than when grown in glucose. The results also indicated that the production of proteins continuously increased when B. bassiana was grown in medium containing insects, reaching its maximum at 9 days (up to 3 mg/mL). On the other hand, when cultivated in glucose-supplemented medium, the production of proteins was constantly low (~0.5 mg/mL). In conclusion, B. bassiana was a large biomass producer and exuded a large amount of protein when grown in medium containing insect powder, making it an ideal intermediate link between insects and protein. Furthermore, the proteins produced by fungi such as B. bassiana can be used in the food, health, and cosmetic industries.
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| 10. |
- Ishchuk, Olena, 1980, et al.
(författare)
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Improved production of human hemoglobin in yeast by engineering hemoglobin degradation
- 2021
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Ingår i: Metabolic Engineering. - : Elsevier BV. - 1096-7176 .- 1096-7184. ; 66, s. 259-267
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Tidskriftsartikel (refereegranskat)abstract
- With the increasing demand for blood transfusions, the production of human hemoglobin (Hb) from sustainable sources is increasingly studied. Microbial production is an attractive option, as it may provide a cheap, safe, and reliable source of this protein. To increase the production of human hemoglobin by the yeast Saccharomyces cerevisiae, the degradation of Hb was reduced through several approaches. The deletion of the genes HMX1 (encoding heme oxygenase), VPS10 (encoding receptor for vacuolar proteases), PEP4 (encoding vacuolar proteinase A), ROX1 (encoding heme-dependent repressor of hypoxic genes) and the overexpression of the HEM3 (encoding porphobilinogen deaminase) and the AHSP (encoding human alpha-hemoglobin-stabilizing protein) genes — these changes reduced heme and Hb degradation and improved heme and Hb production. The reduced hemoglobin degradation was validated by a bilirubin biosensor. During glucose fermentation, the engineered strains produced 18% of intracellular Hb relative to the total yeast protein, which is the highest production of human hemoglobin reported in yeast. This increased hemoglobin production was accompanied with an increased oxygen consumption rate and an increased glycerol yield, which (we speculate) is the yeast's response to rebalance its NADH levels under conditions of oxygen limitation and increased protein-production.
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