| 1. |
- 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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| 2. |
- Antonopoulou, Io, 1989-, et al.
(författare)
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Optimized Enzymatic Synthesis of Feruloyl Derivatives Catalyzed by Three Novel Feruloyl Esterases from Talaromyces wortmannii in Detergentless Microemulsions
- 2018
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Ingår i: Computational and Structural Biotechnology Journal. - : Elsevier. - 2001-0370. ; , s. 361-369
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Tidskriftsartikel (refereegranskat)abstract
- Three novel feruloyl esterases (Fae125, Fae7262 and Fae68) from Talaromyces wortmanniioverexpressed in the C1 platform were evaluated for the transesterification of vinyl ferulatewith two acceptors of different size and lipophilicity (prenol and L-arabinose) in detergentless microemulsions. The effect of reaction conditions such as the microemulsion composition, the substrate concentration, the enzyme load, the pH, the temperature and the agitation were investigated. The type A Fae125 belonging to the subfamily 5 (SF5) of phylogenetic classification showed highest yields for the synthesis of both products after optimization of reaction conditions: 81.8% for prenyl ferulate and 33.0% for L-arabinose ferulate. After optimization, an 8-fold increase in the yield and a 12-fold increase in selectivity were achieved for the synthesis of prenyl ferulate.
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| 3. |
- Antonopoulou, Io, 1989-, et al.
(författare)
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Screening of novel feruloyl esterases from Talaromyces wortmannii for the development of efficient and sustainable syntheses of feruloyl derivatives
- 2019
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Ingår i: Enzyme and microbial technology. - : Elsevier. - 0141-0229 .- 1879-0909. ; 120, s. 124-135
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Tidskriftsartikel (refereegranskat)abstract
- The feruloyl esterases Fae125, Fae7262 and Fae68 from Talaromyces wortmannii were screened in 10 different solvent: buffer systems in terms of residual hydrolytic activity and of the ability for the transesterification of vinyl ferulate with prenol or L-arabinose. Among the tested enzymes, the acetyl xylan-related Fae125 belonging to the phylogenetic subfamily 5 showed highest yield and selectivity for both products in alkane: buffer systems (n-hexane or n-octane). Response surface methodology, based on a 5-level and 6-factor central composite design, revealed that the substrate molar ratio and the water content were the most significant variables for the bioconversion yield and selectivity. The effect of agitation, the possibility of DMSO addition and the increase of donor concentration were investigated. After optimization, competitive transesterification yields were obtained for prenyl ferulate (87.5-92.6%) and L-arabinose ferulate (56.2-61.7%) at reduced reaction times (≤ 24 h) resulting in good productivities (> 1 g/L/h, >300 kg product/kg FAE). The enzyme could be recycled for six consecutive cycles retaining 66.6% of the synthetic activity and 100% of the selectivity.
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| 4. |
- Bonzom, Cyrielle, 1987, et al.
(författare)
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Feruloyl esterases, effects of glycosylation on activity, stability and immobilization
- 2017
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Feruloyl esterases (FAE; EC 3.1.1.73) are a subclass of carboxylic ester hydrolases. They catalyze the hydrolysis of ester linkages in plant cell walls materials releasing ferulic acid and other hydroxycinnamic acids and therefore have been classified in the CAZy database [1] in the carbohydrate esterase (CE) family. Currently, all FAEs in the CAZY database belong to the CE1 family. Immobilization is a powerful tool to allow enzyme reuse and therefore decrease their cost in the overall process. Immobilization on solid carriers can be achieved by different approaches, one of them is by physical adsorption. This technique relies mainly on the surface charges of the support and enzyme. Mesoporous silica particles (MPS) are porous supports made of silica which pore size can be tuned from 2 to 50 nm depending on the synthesis conditions [2]. Because they are negatively charged, silica particles allow for rapid enzyme immobilization by physical adsorption. Furthermore, the diameter of their pore size can be tuned to match the one of the enzyme used and greatly reduces leaching after immobilization.Depending on the production host of the enzyme, the glycosylation extent and/or pattern can vary from none in Escherichia coli to hyper-glycosylation in Pichia pastoris. Glycosylation has been shown to alter activity and stability of enzymes [3]. Our hypothesis was that glycosylation would influence the immobilization behavior of the studied FAE in MPS as well as its activity.In this study, the same enzyme MtFae1a from the fungus Myceliophtora thermophila has been expressed in three different host organisms: E. coli, M. thermophila and P. pastoris to achieve different degrees of glycosylation. The proteins were then purified and biochemically characterized. Different behaviors were observed depending on the production host of the enzymes, for instance in terms of specific activity and temperature optima. Furthermore, glycosylation affected the immobilization capacity of the enzymes on MPS, as well as the activity of immobilized FAEs.[1]B.L. Cantarel, P.M. Coutinho, C. Rancurel, T. Bernard, V. Lombard, B. Henrissat, The Carbohydrate-Active EnZymes database (CAZy): an expert resource for Glycogenomics, Nucleic Acids Res. 37 (2009) D233–238. doi:10.1093/nar/gkn663.[2]D. Zhao, J. Feng, Q. Huo, N. Melosh, G.H. Fredrickson, B.F. Chmelka, G.D. Stucky, Triblock Copolymer Syntheses of Mesoporous Silica with Periodic 50 to 300 Angstrom Pores, Science. 279 (1998) 548–552. doi:10.1126/science.279.5350.548.[3]A. Basso, P. Braiuca, S. Cantone, C. Ebert, P. Linda, P. Spizzo, P. Caimi, U. Hanefeld, G. Degrassi, L. Gardossi, In Silico Analysis of Enzyme Surface and Glycosylation Effect as a Tool for Efficient Covalent Immobilisation of CalB and PGA on Sepabeads®, Adv. Synth. Catal. 349 (2007) 877–886. doi:10.1002/adsc.200600337.
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| 5. |
- Cerullo, Gabriella, et al.
(författare)
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The Synthetic Potential of Fungal Feruloyl Esterases: A Correlation with Current Classification Systems and Predicted Structural Properties
- 2018
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Ingår i: Catalysts. - : MDPI AG. - 2073-4344. ; 8:242
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Tidskriftsartikel (refereegranskat)abstract
- Twenty-eight fungal feruloyl esterases (FAEs) were evaluated for their synthetic abilities in a ternary system of n-hexane: t-butanol: 100 mM MOPS-NaOH pH 6.0 forming detergentless microemulsions. Five main derivatives were synthesized, namely prenyl ferulate, prenyl caffeate, butyl ferulate, glyceryl ferulate, and l-arabinose ferulate, offering, in general, higher yields when more hydrophilic alcohol substitutions were used. Acetyl xylan esterase-related FAEs belonging to phylogenetic subfamilies (SF) 5 and 6 showed increased synthetic yields among tested enzymes. In particular, it was shown that FAEs belonging to SF6 generally transesterified aliphatic alcohols more efficiently while SF5 members preferred bulkier l-arabinose. Predicted surface properties and structural characteristics were correlated with the synthetic potential of selected tannase-related, acetyl-xylan-related, and lipase-related FAEs (SF1-2, -6, -7 members) based on homology modeling and small molecular docking simulations.
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| 6. |
- Hüttner, Silvia, 1984, et al.
(författare)
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Immobilisation on mesoporous silica and solvent rinsing improve the transesterification abilities of feruloyl esterases from Myceliophthora thermophila
- 2017
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Ingår i: Bioresource Technology. - : Elsevier BV. - 0960-8524 .- 1873-2976. ; 239, s. 57-65
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Tidskriftsartikel (refereegranskat)abstract
- The immobilisation of four feruloyl esterases (FAEs) (FaeA1, FaeA2, FaeB1, FaeB2) from the thermophilic fungus Myceliophthora thermophila C1 was studied and optimised via physical adsorption onto various mesoporous silica particles with pore diameters varying from 6.6 nm to 10.9 nm. Using crude enzyme preparations, enrichment of immobilised FAEs was observed, depending on pore diameter and protein size. The immobilised enzymes were successfully used for the synthesis of butyl ferulate through transesterification of methyl ferulate with 1-butanol. Although the highest butyl ferulate yields were obtained with free enzyme, the synthesis-to-hydrolysis ratio was higher when using immobilised enzymes. Over 90% of the initial activity was observed in a reusability experiment after nine reaction cycles, each lasting 24 h. Rinsing with solvent to remove water from the immobilised enzymes further improved their activity. This study demonstrates the suitability of immobilised crude enzyme preparations in the development of biocatalysts for esterification reactions.
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| 7. |
- Zerva, Anastasia, et al.
(författare)
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Cross-Linked Enzyme Aggregates of Feruloyl Esterase Preparations from Thermothelomyces thermophila and Talaromyces wortmannii
- 2018
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Ingår i: Catalysts. - : MDPI. - 2073-4344. ; 8:5
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Tidskriftsartikel (refereegranskat)abstract
- Cross-linked enzyme aggregates (CLEA®) technology is a well-established method in the current literature for the low-cost and effective immobilization of several enzymes. The main advantage of this particular method is the simplicity of the process, since it consists of only two steps. However, CLEA immobilization must be carefully designed for each desired enzyme, since the optimum conditions for enzymes can vary significantly, according to their physicochemical properties. In the present study, an investigation of the optimum CLEA immobilization conditions was carried out for eight feruloyl esterase preparations. Feruloyl esterases are a very important enzyme group in the valorization of lignocellulosic biomass, since they act in a synergistic way with other enzymes for the breakdown of plant biomass. Specifically, we investigated the type and concentration of precipitant and the crosslinker concentration, for retaining optimal activity. FAE68 was found to be the most promising enzyme for CLEA immobilization, since in this case, the maximum retained activity, over 98%, was observed. Subsequently, we examined the operational stability and the stability in organic solvents for the obtained CLEA preparations, as well as their structure. Overall, our results support that the maximum activity retaining and the stability properties of the final CLEAs can vary greatly in different FAE preparations. Nevertheless, some of the examined FAEs show a significant potential for further applications in harsh industrial conditions.
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| 8. |
- Zerva, Anastasia, et al.
(författare)
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Optimization of Transesterification Reactions with CLEA-Immobilized Feruloyl Esterases from Thermothelomyces thermophila and Talaromyces wortmannii
- 2018
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Ingår i: Molecules. - : MDPI. - 1431-5157 .- 1420-3049. ; 23:9
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Tidskriftsartikel (refereegranskat)abstract
- Feruloyl esterases (FAEs, E.C. 3.1.1.73) are biotechnologically important enzymes with several applications in ferulic acid production from biomass, but also in synthesis of hydroxycinnamic acid derivatives. The use of such biocatalysts in commercial processes can become feasible by their immobilization, providing the advantages of isolation and recycling. In this work, eight feruloyl esterases, immobilized in cross-linked enzyme aggregates (CLEAs) were tested in regard to their transesterification performance, towards the production of prenyl ferulate (PFA) and arabinose ferulate (AFA). After solvent screening, comparison with the activity of respective soluble enzymes, and operational stability tests, FAE125 was selected as the most promising biocatalyst. A central composite design revealed the optimum conditions for each transesterification product, in terms of water content, time, and substrate ratio for both products, and temperature and enzyme load additionally for prenyl ferulate. The optimum product yields obtained were 83.7% for PFA and 58.1% for AFA. FAE125 CLEAs are stable in the optimum conditions of transesterification reactions, maintaining 70% residual activity after five consecutive reactions. Overall, FAE125 CLEAs seem to be able to perform as a robust biocatalyst, offering satisfactory yields and stability, and thus showing significant potential for industrial applications.
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