| 1. |
- Agnantiari, G., et al.
(författare)
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A Purified α-galactosidase from aspergillus niger with enhanced kinetic characteristics
- 1991
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Ingår i: Acta Biotechnologica. - : Wiley. - 0138-4988 .- 1521-3846. ; 11:5, s. 479-484
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Tidskriftsartikel (refereegranskat)abstract
- Extracellular α-galactosidase from Aspergillus niger was purified 128-fold over the crude extract by gel filtration, ion exchange chromatography and chromatofocusing. Certain substrates and end products affected enzyme activity. Among the former p-nitrophenyl-α-galactopyranoside (PNPG) inhibited the enzyme at 1.4 mM while melibiose did not inhibit α-galactosidase at concentrations up to 50 mM. Enzymic end products such as glucose did not inhibit the enzyme at concentrations up to 100 mM while galactose exhibited a competitive inhibition with a Ki = 1.29 mM. The kinetic characteristics of the enzyme compared favourably to other microbial α-galactosidases and make it suitable for food process applications.
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| 2. |
- Anasontzis, George E, 1980, et al.
(författare)
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Challenges in ethanol production with fusarium oxysporum through consolidated bioprocessing
- 2014
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Ingår i: Bioengineered Bugs. - : Informa UK Limited. - 1949-1018 .- 1949-1026 .- 2165-5979 .- 2165-5987. ; 5:6, s. 393-395
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Tidskriftsartikel (refereegranskat)abstract
- Fusarium oxysporum has been reported as being able to both produce the enzymes necessary to degrade lignocellulosic biomass to sugars and also ferment the monosaccharides to ethanol under anaerobic or microaerobic conditions. However, in order to become an economically feasible alternative to other ethanol-producing microorganisms, a better understanding of its physiology, metabolic pathways, and bottlenecks is required, together with an improvement in its efficiency and robustness. In this report, we describe the challenges for the future and give additional justification for our recent publication.
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| 3. |
- Anasontzis, George E, 1980, et al.
(författare)
-
Constitutive homologous expression of phosphoglucomutase and transaldolase increases the metabolic flux of Fusarium oxysporum
- 2014
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Ingår i: Microbial Cell Factories. - : Springer Science and Business Media LLC. - 1475-2859. ; 13:1
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Tidskriftsartikel (refereegranskat)abstract
- Background: Fusarium oxysporum is among the few filamentous fungi that have been reported of being able to directly ferment biomass to ethanol in a consolidated bioprocess. Understanding its metabolic pathways and their limitations can provide some insights on the genetic modifications required to enhance its growth and subsequent fermentation capability. In this study, we investigated the hypothesis reported previously that phosphoglucomutase and transaldolase are metabolic bottlenecks in the glycolysis and pentose phosphate pathway of the F. oxysporum metabolism.Results: Both enzymes were homologously overexpressed in F. oxysporum F3 using the gpdA promoter of Aspergillus nidulans for constitutive expression. Transformants were screened for their phosphoglucomutase and transaldolase genes expression levels with northern blot. The selected transformant exhibited high mRNA levels for both genes, as well as higher specific activities of the corresponding enzymes, compared to the wild type. It also displayed more than 20 and 15% higher specific growth rate upon aerobic growth on glucose and xylose, respectively, as carbon sources and 30% higher biomass to xylose yield. The determination of the relative intracellular amino and non-amino organic acid concentrations at the end of growth on glucose revealed higher abundance of most determined metabolites between 1.5- and 3-times in the recombinant strain compared to the wild type. Lower abundance of the determined metabolites of the Krebs cycle and an 68-fold more glutamate were observed at the end of the cultivation, when xylose was used as carbon source.Conclusions: Homologous overexpression of phosphoglucomutase and transaldolase in F. oxysporum was shown to enhance the growth characteristics of the strain in both xylose and glucose in aerobic conditions. The intracellular metabolites profile indicated how the changes in the metabolome could have resulted in the observed growth characteristics. © 2014 Anasontzis et al.; licensee BioMed Central Ltd.
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| 4. |
- Anasontzis, George E., et al.
(författare)
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Metabolic Engineering of Fusarium oxysporum to Improve Its Ethanol-Producing Capability
- 2016
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Ingår i: Frontiers in Microbiology. - : Frontiers Media SA. - 1664-302X. ; 7
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Tidskriftsartikel (refereegranskat)abstract
- Fusarium oxysporum is one of the few filamentous fungi capable of fermenting ethanol directly from plant cell wall biomass. It has the enzymatic toolbox necessary to break down biomass to its monosaccharides and, under anaerobic and microaerobic conditions, ferments them to ethanol. Although these traits could enable its use in consolidated processes and thus bypass some of the bottlenecks encountered in ethanol production from lignocellulosic material when Saccharomyces cerevisiae is used-namely its inability to degrade lignocellulose and to consume pentoses-two major disadvantages of F. oxysporum compared to the yeast-its low growth rate and low ethanol productivity-hinder the further development of this process. We had previously identified phosphoglucomutase and transaldolase, two major enzymes of glucose catabolism and the pentose phosphate pathway, as possible bottlenecks in the metabolism of the fungus and we had reported the effect of their constitutive production on the growth characteristics of the fungus. In this study, we investigated the effect of their constitutive production on ethanol productivity under anaerobic conditions. We report an increase in ethanol yield and a concomitant decrease in acetic acid production. Metabolomics analysis revealed that the genetic modifications applied did not simply accelerate the metabolic rate of the microorganism; they also affected the relative concentrations of the various metabolites suggesting an increased channeling toward the chorismate pathway, an activation of the γ-aminobutyric acid shunt, and an excess in NADPH regeneration
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| 5. |
- Antonopoulou, Io, 1989-, et al.
(författare)
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CO2 to Methanol: A Highly Efficient Enzyme Cascade
- 2022. - 1
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Ingår i: Multienzymatic Assemblies. - New York, NY : Springer Nature. ; , s. 317-344
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Bokkapitel (övrigt vetenskapligt/konstnärligt)abstract
- Carbon dioxide (CO2) has been increasingly regarded not only as a greenhouse gas but also as a valuable feedstock for carbon-based chemicals. In particular, biological approaches have drawn attention as models for the production of value-added products, as CO2 conversion serves many natural processes. Enzymatic CO2 reduction in vitro is a very promising route to produce fossil free and bio-based fuel alternatives, such as methanol. In this chapter, the advances in constructing competitive multi-enzymatic systems for the reduction of CO2 to methanol are discussed. Different integrated methods are presented, aiming to address technological challenges, such as the cost effectiveness, need for material regeneration and reuse and improving product yields of the process.
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| 6. |
- Antonopoulou, Io, 1989-
(författare)
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Development of biocatalytic processes for selective antioxidant production
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Feruloyl esterases (FAEs, EC 3.1.1.73) represent a subclass of carboxylic acid esterases that under normal conditions catalyze the hydrolysis of the ester bond between hydroxycinnamic acids (ferulic acid, sinapic acid, caffeic acid, p-coumaric acid) and sugar residues in plant cell walls. Based on their specificity towards monoferulates and diferulates, substitutions on the phenolic ring and on their amino acid sequence identity, they have been classified into four types (A-D) while phylogenetic analysis has resulted in classification into thirteen subfamilies (SF1-13). Under low water content, these enzymes are able to catalyze the esterification of hydroxycinnamic acids or the transesterification of their esters (donor) with alcohols or sugars (acceptor) resulting in compounds with modified lipophilicity, having a great potential for use in the tailor-made modification of natural antioxidants for cosmetic, cosmeceutical and pharmaceutical industries. The work described in this thesis focused on the selection,characterization and application of FAEs for the synthesis of bioactive esters with antioxidant activity in non-conventional media. The basis of the current classification systems was investigated in relation with the enzymes’ synthetic and hydrolytic abilities while the developed processes were evaluated for their efficiency and sustainability.Paper I was dedicated to the screening and evaluation of the synthetic abilities of 28 fungal FAEs using acceptors of different lipophilicity at fixed conditions in detergentless microemulsions. It was revealed that FAEs classified in phylogenetic subfamilies related to acetyl xylan esterases (SF5 and 6) showed increased transesterification rates and selectivity. In general, FAEs showed preference on more hydrophilic alcohol acceptors and in descending order to glycerol > 1-butanol > prenol. Homology modeling and small molecule docking simulations were employed as tools for the identification of a potential relationship between the predicted surface and active site properties of selected FAEs and the transesterification selectivity.Papers II- IV focused on the characterization of eight promising FAEs and the optimization of reaction conditions for the synthesis of two bioactive esters (prenyl ferulate and L-arabinose ferulate) in detergentless microemulsions. The effect of the medium composition, the donor and acceptor concentration, the enzyme load, the pH, the temperature and the agitation on the transesterification yield and selectivity were investigated. It was observed that the acceptor concentration and enzyme load were crucial parameters for selectivity. Fae125 (Type A, SF5) iiexhibited highest prenyl ferulate yield (81.1%) and selectivity (4.685) converting 98.5% of VFA to products after optimization at 60 mM VFA, 1.5 M prenol, 0.04 mg FAE mL-1, 40oC, 24 h, 53.4:43.4:3.2 v/v/v n-hexane: t-butanol: 100 mM MOPS-NaOH pH 8.0. On the other hand, FaeA1 (Type A, SF5) showed highest L-arabinose ferulate yield (52.2 %) and selectivity (1.120) at 80 mM VFA, 55 mM L-arabinose, 0.02 mg FAE mL-1, 50oC, 8 h, 19.8: 74.7: 5.5 v/v/v n-hexane: t-butanol: 100 mM MOPS-NaOH pH 8.0.In paper V, the effect of reaction media on the enzyme stability and transesterification yield and selectivity was studied in different solvents for the synthesis of two bioactive esters: prenyl ferulate and L-arabinose ferulate. The best performing enzyme (Fae125) was used in the optimization of reaction conditions in the best solvent (n-hexane) via response surface methodology. Both bioconversions were best described by a two-factor interaction model while optimal conditions were determined as the ones resulting in highest yield and selectivity.Highest prenyl ferulate yield (87.5%) and selectivity (7.616) were observed at 18.56 mM prenol mM-1VFA, 0.04 mg FAE mL-1, 24.5 oC, 24.5 h, 91.8: 8.2 v/v n-hexane: 100 mM sodium acetate pH 4.7. Highest L-arabinose ferulate yield (56.2%) and selectivity (1.284) were observed at 2.96 mM L-arabinose mM-1VFA, 0.02 mg FAE mL-1, 38.9 oC, 12 h, 90.5: 5.0: 4.5 v/v/v n-hexane: dimethyl sulfoxide: 100 mM sodium acetate pH 4.7. The enzyme could be reused for six consecutive reaction cycles maintaining 66.6% of its initial synthetic activity. The developed bioconversions showed exceptional biocatalyst productivities (> 300 g product g-1FAE) and the waste production was within the range of pharmaceutical processes.Paper VI focused on the investigation of the basis of the type A classification of a well-studied FAE from Aspergillus niger(AnFaeA) by comparing its activity towards methyl and arabinose hydroxycinnamic acid esters. For this purpose, L-arabinose ferulateand caffeate were synthesized enzymatically. kcat/Kmratios revealed that AnFaeA hydrolyzed arabinose ferulate 1600 times and arabinose caffeate 6.5 times more efficiently than methyl esters. This study demonstrated that short alkyl chain hydroxycinnamate esters which are used nowadays for FAE classification can lead to activity misclassification, while L-arabinose esters could potentially substitute synthetic esters in classification describing more adequately the enzyme specificitiesin the natural environment.
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| 7. |
- Antonopoulou, Io, et al.
(författare)
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Enzymatic synthesis of bioactive compounds with high potential for cosmeceutical application
- 2016
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Ingår i: Applied Microbiology and Biotechnology. - : Springer Science and Business Media LLC. - 0175-7598 .- 1432-0614. ; 100:15, s. 6519-6543
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Tidskriftsartikel (refereegranskat)abstract
- Cosmeceuticals are cosmetic products containing biologically active ingredients purporting to offer a pharmaceutical therapeutic benefit. The active ingredients can be extracted and purified from natural sources (botanicals, herbal extracts, or animals) but can also be obtained biotechnologically by fermentation and cell cultures or by enzymatic synthesis and modification of natural compounds. A cosmeceutical ingredient should possess an attractive property such as anti-oxidant, anti-inflammatory, skin whitening, anti-aging, anti-wrinkling, or photoprotective activity, among others. During the past years, there has been an increased interest on the enzymatic synthesis of bioactive esters and glycosides based on (trans)esterification, (trans)glycosylation, or oxidation reactions. Natural bioactive compounds with exceptional theurapeutic properties and low toxicity may offer a new insight into the design and development of potent and beneficial cosmetics. This review gives an overview of the enzymatic modifications which are performed currently for the synthesis of products with attractive properties for the cosmeceutical industry
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| 8. |
- Antonopoulou, Io, 1989-, et al.
(författare)
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Ferulic Acid From Plant Biomass: A Phytochemical With Promising Antiviral Properties
- 2022
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Ingår i: Frontiers in Nutrition. - : Frontiers Media S.A.. - 2296-861X. ; 8
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Forskningsöversikt (refereegranskat)abstract
- Plant biomass is a magnificent renewable resource for phytochemicals that carry bioactive properties. Ferulic acid (FA) is a hydroxycinnamic acid that is found widespread in plant cell walls, mainly esterified to polysaccharides. It is well known of its strong antioxidant activity, together with numerous properties, such as antimicrobial, anti-inflammatory and neuroprotective effects. This review article provides insights into the potential for valorization of FA as a potent antiviral agent. Its pharmacokinetic properties (absorption, metabolism, distribution and excretion) and the proposed mechanisms that are purported to provide antiviral activity are presented. Novel strategies on extraction and derivatization routes, for enhancing even further the antiviral activity of FA and potentially favor its metabolism, distribution and residence time in the human body, are discussed. These routes may lead to novel high-added value biorefinery pathways to utilize plant biomass toward the production of nutraceuticals as functional foods with attractive bioactive properties, such as enhancing immunity toward viral infections.
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| 9. |
- Antonopoulou, Io, 1989-, et al.
(författare)
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Inhibition of the main protease of SARS-CoV-2 (Mpro) by repurposing/designing drug-like substances and utilizing nature’s toolbox of bioactive compounds
- 2022
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Ingår i: Computational and Structural Biotechnology Journal. - : Elsevier. - 2001-0370. ; 20, s. 1306-1344
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Forskningsöversikt (refereegranskat)abstract
- The emergence of the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) has resulted in a long pandemic, with numerous cases and victims worldwide and enormous consequences on social and economic life. Although vaccinations have proceeded and provide a valuable shield against the virus, the approved drugs are limited and it is crucial that further ways to combat infection are developed, that can also act against potential mutations. The main protease (Mpro) of the virus is an appealing target for the development of inhibitors, due to its importance in the viral life cycle and its high conservation among different coronaviruses. Several compounds have shown inhibitory potential against Mpro, both in silico and in vitro, with few of them also having entered clinical trials. These candidates include: known drugs that have been repurposed, molecules specifically designed based on the natural substrate of the protease or on structural moieties that have shown high binding affinity to the protease active site, as well as naturally derived compounds, either isolated or in plant extracts. The aim of this work is to collectively present the results of research regarding Mpro inhibitors to date, focusing on the function of the compounds founded by in silico simulations and further explored by in vitro and in vivo assays. Creating an extended portfolio of promising compounds that may block viral replication by inhibiting Mpro and by understanding involved structure–activity relationships, could provide a basis for the development of effective solutions against SARS-CoV-2 and future related outbreaks.
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| 10. |
- Antonopoulou, Io, et al.
(författare)
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Optimization of enzymatic synthesis of l-arabinose ferulate catalyzed by feruloyl esterases from Myceliophthora thermophila in detergentless microemulsions and assessment of its antioxidant and cytotoxicity activities
- 2018
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Ingår i: Process Biochemistry. - : Elsevier. - 1359-5113 .- 1873-3298. ; 65, s. 100-108
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Tidskriftsartikel (refereegranskat)abstract
- The feruloyl esterases FaeA1, FaeA2, FaeB1, FaeB2 from Myceliophthora thermophila C1 and MtFae1a from M. thermophila ATCC 42464 were used as biocatalysts for the transesterification of vinyl ferulate (VFA) with l-arabinose in detergentless microemulsions. The effect of parameters such as the microemulsion composition, the substrate concentration, the enzyme load, the pH, the temperature and the agitation was investigated. FaeA1 offered the highest transesterification yield (52.2 ± 4.3%) after 8 h of incubation at 50 °C using 80 mM VFA, 55 mM l-arabinose and 0.02 mg FAE mL−1 in a mixture comprising of 19.8: 74.7: 5.5 v/v/v n-hexane: t-butanol: 100 mM MOPS-NaOH pH 8.0. The ability of l-arabinose ferulate (AFA) to scavenge 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals was significant (IC50 386.5 μM). AFA was not cytotoxic even at high concentrations (1 mM) however was found to be pro-oxidant at concentrations higher than 20 μM when the antioxidant activity was determined with the dichloro-dihydro-fluorescein diacetate (DCFH-DA) assay in human skin fibroblasts.
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