| 1. |
- Adlercreutz, Patrick, et al.
(författare)
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Enzymatic conversions of polar lipids. Principles, problems and solutions
- 2001
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Ingår i: Journal of Molecular Catalysis - B Enzymatic. - 1381-1177. ; 11:4-6, s. 173-178
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Forskningsöversikt (refereegranskat)abstract
- This text provides a brief overview of the principles of enzymatic lipid conversion and some recent advances in the enzymatic conversion of glycerophospholipids and galactolipids. Lipases and phospholipases are used to exchange fatty acids or the polar group in the lipids. The reactions can be carried out either as hydrolysis-esterification sequences or as one-step transferase reactions. The scope and limitations of the different methods are discussed.
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| 2. |
- Adlercreutz, Patrick, et al.
(författare)
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Enzymatic fatty acid exchange in glycerophospholipids
- 2003
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Ingår i: European Journal of Lipid Science and Technology. - : Wiley. - 1438-7697 .- 1438-9312. ; 105:10, s. 638-645
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Forskningsöversikt (refereegranskat)abstract
- Lipases can be used to exchange fatty acids in the sn-1 position of glycerophospholipids and phospholipase A2 is useful for the corresponding exchange reaction in the sn-2 position. In both cases, the exchange can be done in a one-step acidolysis process or in a two-step process. In the latter case, the original fatty acid in the desired position is removed by enzymatic hydrolysis or alcoholysis and after isolation of the resulting lysophospholipid, the new fatty acid is introduced, using the same enzyme, in an esterification reaction. Several synthesis examples from the literature are reviewed. Incorporation of a new fatty acid into the sn-1 position is more favourable than incorporation into the sn-2 position because of the magnitudes of the equilibrium constants of the reactions and because lipases can be used at much lower water activity than phospholipase A2. With the consecutive use of both enzymes highly pure products with defined fatty acids in both positions can be obtained.
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| 3. |
- Andersson, Mats, et al.
(författare)
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A kinetic study of almond-β-glucosidase catalysed synthesis of hexyl-glycosides in low aqueous media : Influence of glycosyl donor and water activity
- 2001
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Ingår i: Journal of Molecular Catalysis - B Enzymatic. - 1381-1177. ; 14:4-6, s. 69-76
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Tidskriftsartikel (refereegranskat)abstract
- A variety of alkyl and aryl glycosides were investigated as substrates for almond β-glucosidase catalysed synthesis of hexyl-β-D-glycosides in low aqueous hexanol media. The rate-limiting step in the organic media was determined to be the glycosylation of the enzyme. The kinetic constants V max , K m (glycosyl donor) and V max /K m were all influenced by the water activity and they all increased in value with increasing water activity. The increase in V max /K m was mainly determined by the increase in V max and a plot of log(V max /K m ) versus water activity resulted in a straight line with similar slopes for all glycosides but with different absolute values and thus the most reactive substrate p-nitrophenyl glucoside was the best one in the entire water activity range studied (0.53-0.96). The preference for the two competing acceptors, hexanol and water, was not affected by the aglycon part of the glucoside. Surprisingly, the ratio between trans glycosylation and hydrolysis increased with increasing water activity. A decrease in water activity caused an increase in equilibrium yield of hexyl glycoside, as expected, but was not beneficial for the kinetically controlled yield.
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| 4. |
- Andersson, Mats, et al.
(författare)
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Stabilisation of chloroperoxidase towards peroxide dependent inactivation
- 2000
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Ingår i: Biocatalysis and Biotransformation. - : Informa UK Limited. - 1024-2422 .- 1029-2446. ; 18:6, s. 457-469
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Tidskriftsartikel (refereegranskat)abstract
- The addition of polyethyleneimine with a molecular weight of 2000 to chloroperoxidase from Caldariomyces fumago dramatically improved the stability of the enzyme towards peroxide dependent inactivation. The rate constant for the H 2 O 2 -dependent inactivation of chloroperoxidase decreased from 0.0016s -1 to 1.1 * 10 -5 s -1 in the presence of 1% polyethyleneimine. The stabilising effect towards tert-butyl hydroperoxide was even more impressive. The half-life of the chloroperoxidase when exposed to a solution of 40 mM tert-butyl hydroperoxide increased from 3.2 minutes to ≥ 70 hours in presence of 0.1% polyethyleneimine.
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| 5. |
- Andersson, Mats, et al.
(författare)
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Toward an enzyme-based oxygen scavenging laminate. Influence of industrial lamination conditions on the performance of glucose oxidase
- 2002
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Ingår i: Biotechnology and Bioengineering. - : Wiley. - 1097-0290 .- 0006-3592. ; 79:1, s. 37-42
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Tidskriftsartikel (refereegranskat)abstract
- The laminate consisted of several polymer layers, aluminium, and one cellulose-based layer containing the active enzymatic system (e.g., glucose oxidase, catalase, glucose, and CaCO3). During the industrial lamination process, the enzyme layer was exposed to three temperature spikes up to 325degreesC without significant enzyme inactivation. Ninety-seven percent of the glucose oxidase activity still remained after the lamination process. The best laminate had an oxygen absorbing capacity of 7.6 +/- 1.0 L/m(2). A reference that was not laminated expressed a corresponding oxygen absorbing capacity of 7.1 +/- 0.8 L/m(2).
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| 6. |
- Barros, Raúl J., et al.
(författare)
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Effect of mass-transfer limitations on the selectivity of immobilized α-chymotrypsin biocatalysts prepared for use in organic medium
- 2000
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Ingår i: Biotechnology and Bioengineering. - 0006-3592. ; 67:3, s. 319-326
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Tidskriftsartikel (refereegranskat)abstract
- The selectivity of preparations of α-chymotrypsin immobilized on Celite or polyamide and carrying out syntheses of di- and tripeptides in acetonitrile medium were studied. The study concerns the effect of mass- transfer limitations on three different kinds of selectivity: acyl donor, stereo- and nucleophile selectivities, defined respectively as the ratio of initial rates with different acyl donors; the enantioselectivity factor (E); and the ratio of initial rates of peptide synthesis and hydrolysis of the acyl donor. Strong mass-transfer limitations caused by increased enzyme loading had a very strong effect on acyl donor selectivity, with reductions of up to 79%, and on stereoselectivity, with reductions of up to 77% in relation to optimum values, both on Celite. Nucleophile selectivity was not affected as strongly by mass-transfer limitations. Using a small molecule (AlaNH2) as nucleophile, the onset of these limitations caused only minor reductions in selectivity, while when using a larger nucleophilic species (AlaPheNH2) it was reduced by up to 60% when increasing enzyme loading on Celite from 2 to 100 mg/g. The different way these kinds of selectivity are affected by the onset of mass-transfer limitations can be explained by a combination of different aspects: the kinetic behavior of the enzyme toward nucleophile and acyl donor concentrations, the relative concentrations of reagents used in the reaction media, and their relative diffusion coefficients. In short, higher concentrations of nucleophile than acyl donor are generally used, and the nucleophile most often used in the experiments hereby described (AlaNH2) diffuses faster than the acyl donors employed. These factors combined are expected to give rise to concentration gradients inside porous biocatalyst particles higher for acyl donor than for nucleophile under conditions of mass-transfer limitations. This explains why acyl donor selectivity and stereoselectivity are much more influenced by mass transfer limitations than nucleophile selectivity.
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| 7. |
- Barros, Raúl J., et al.
(författare)
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Modeling the performance of immobilized α-chymotrypsin catalyzed peptide synthesis in acetonitrile medium
- 2001
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Ingår i: Journal of Molecular Catalysis - B Enzymatic. - 1381-1177. ; 11:4-6, s. 841-850
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Tidskriftsartikel (refereegranskat)abstract
- A model was developed which describes simultaneous reaction and internal diffusion for kinetically controlled, immobilized α-chymotrypsin-catalyzed, oligopeptide synthesis in acetonitrile medium. The model combines the equations that describe the intrinsic kinetics of four different reactions and the physical characteristics of three different support materials, as determined experimentally, to predict the apparent initial activity and nucleophile selectivity of the immobilized biocatalyst. The model is able to predict reasonably well the experimentally observed initial rate and nucleophile selectivity vs. enzyme loading profiles. The reduction in observed initial rate with enzyme loading when fast reactions are carried out with α-chymotrypsin immobilized on celite, and the larger influence of mass transfer limitations on the initial reaction rates than on nucleophile selectivities are correctly predicted by the numerical calculations. The model is general in terms of its application to other systems - enzymes, reactions, support materials and/or kinetic schemes - as long as the intrinsic kinetics and the characteristics of the enzyme and support material are known.
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| 8. |
- Brocca, Stefania, et al.
(författare)
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Mutants provide evidence of the importance of glycosydic chains in the activation of lipase 1 from Candida rugosa
- 2000
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Ingår i: Protein Science. - : Wiley. - 0961-8368 .- 1469-896X. ; 9:5, s. 985-990
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Tidskriftsartikel (refereegranskat)abstract
- Sequence analysis of Candida rugosa lipase 1 (LIP1) predicts the presence of three N-linked glycosylation sites at asparagine 291, 314, 351. To investigate the relevance of sugar chains in the activation and stabilization of LIP1, we directed site mutagenesis to replace the above mentioned asparagine with glutamine residues. Comparison of the activity of mutants with that of the wild-type (wt) lipase indicates that both 314 and 351 Asn to Gln substitutions influence, although at a different extent, the enzyme activity both in hydrolysis and esterification reactions, but they do not alter the enzyme water activity profiles in organic solvents or temperature stability. Introduction of Gln to replace Asn35 is likely to disrupt a stabilizing interaction between the sugar chain and residues of the inner side of the lid in the enzyme active conformation. The effect of deglycosylation at position 314 is more difficult to explain and might suggest a more general role of the sugar moiety for the structural stability of lipase 1. Conversely, Asn291Gln substitution does not affect' the lipolytic or the esterase activity of the mutant that behaves essentially as the wt enzyme. This observation supports the hypothesis that changes in activity of Asn314Gln and Asn351Gln mutants are specifically due to deglycosylation.
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| 9. |
- Costes, David, et al.
(författare)
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Cross-linked crystals of hydroxynitrile lyase as catalyst for the synthesis of optically active cyanohydrins
- 2001
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Ingår i: Journal of Molecular Catalysis - B Enzymatic. - 1381-1177. ; 11:4-6, s. 607-612
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Tidskriftsartikel (refereegranskat)abstract
- Purified hydroxynitrile lyase (HNL) from Manihot esculenta was crystallized by the sitting-drop vapour-diffusion method. The bipyramidal crystals formed (10-20 μm) were cross-linked with different amounts of glutaraldehyde and used as biocatalyst for the synthesis of optically active cyanohydrins. The cross-linked crystals were more stable than Celite-immobilized enzymes when incubated in organic solvents, especially in polar solvents. After six consecutive batch reactions in dibutylether, the remaining activity of the cross-linked crystals was more than 70 times higher than for the immobilized enzymes. Nevertheless, the specific activity of the cross-linked crystals (per milligram protein) was reduced compared to the activity of immobilized enzymes. The product enantiopurity was independent of the type of enzyme preparation used.
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| 10. |
- Costes, David, et al.
(författare)
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Stability and stabilization of hydroxynitrile lyase in organic solvents
- 2001
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Ingår i: Biocatalysis and Biotransformation. - : Informa UK Limited. - 1024-2422 .- 1029-2446. ; 19:2, s. 119-130
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Tidskriftsartikel (refereegranskat)abstract
- The stability of hydroxynitrile lyase from Hevea brasiliensis has been studied in organic solvents. In dry solvents, the enzyme had half-lives in the range 1400-2500 hours. The enzyme half-life was one order of magnitude lower if the medium was water saturated. The substrates, aldehyde and hydrogen cyanide, were found to promote enzyme deactivation. The deactivation increased with substrate concentration, but was reduced in hydrophilic solvent. At high substrate concentration (2 M) in tert-butyl methyl ether, the enzyme half-life was 1.7 h when incubated with hydrogen cyanide while it was 1.0 h with 3-phenylpropionaldehyde. The addition of polyethylenimine, 125 mg per g of enzyme preparation, increased the enzyme half-life to 110 h when incubated with hydrogen cyanide and to 3.2 h with 3-phenylpropanaldehyde in tert-butyl methyl ether. Albumin and poly(ethylene glycol) gave similar stabilization effect.
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