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- Madsen, Mikkel, et al.
(författare)
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Unaided efficient transglutaminase cross-linking of whey proteins strongly impacts the formation and structure of protein alginate particles
- 2022
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Ingår i: Food Chemistry: Molecular Sciences. - : Elsevier BV. - 2666-5662. ; 5
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Tidskriftsartikel (refereegranskat)abstract
- There is a dogma within whey protein modification, which dictates the necessity of pretreatment to enzymatic cross-linking of β-lactoglobulin (β-Lg). Here microbial transglutaminase (MTG) cross-linked whey proteins and β-Lg effectively in 50 mM NaHCO3, pH 8.5, without pretreatment. Cross-linked β-Lg spanned 18 to >240 kDa, where 6 of 9 glutamines reacted with 8 of 15 lysines. The initial isopeptide bond formation caused loss of β-Lg native structure with t1/2 = 3 h, while the polymerization occurred with t1/2 = 10 h. Further, cross-linking effects on protein carbohydrate interaction have been overlooked, leaving a gap in understanding of these complex food matrices. Complexation with alginate showed that β-Lg cross-linking decreased onset of particle formation, hydrodynamic diameter, stoichiometry (β-Lg/alginate) and dissociation constant. The complexation was favored at higher temperatures (40 °C), suggesting that hydrophobic interactions were important. Thus, β-Lg was cross-linked without pretreatment and the resulting polymers gave rise to altered complexation with alginate.
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| 2. |
- Teze, David, et al.
(författare)
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Rational Enzyme Design without Structural Knowledge : A Sequence-Based Approach for Efficient Generation of Transglycosylases
- 2021
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Ingår i: Chemistry: A European Journal. - : Wiley. - 1521-3765 .- 0947-6539. ; 27:40, s. 10323-10334
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Tidskriftsartikel (refereegranskat)abstract
- Glycobiology is dogged by the relative scarcity of synthetic, defined oligosaccharides. Enzyme-catalysed glycosylation using glycoside hydrolases is feasible but is hampered by the innate hydrolytic activity of these enzymes. Protein engineering is useful to remedy this, but it usually requires prior structural knowledge of the target enzyme, and/or relies on extensive, time-consuming screening and analysis. Here we describe a straightforward strategy that involves rational rapid in silico analysis of protein sequences. The method pinpoints 6-12 single mutant candidates to improve transglycosylation yields. Requiring very little prior knowledge of the target enzyme other than its sequence, the method is generic and procures catalysts for the formation of glycosidic bonds involving various d / l -, α/β-pyranosides or furanosides, and exo - and endo -action. Moreover, mutations validated in one enzyme can be transposed to others, even distantly related enzymes.
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