β-Galactosidase was isolated from Ranunculus arvensis seeds using DEAE-cellulose,Sephadex G-100,and Con A sepharose-4B chromatography.The enzyme was purified to electrophoretic homogeneity with a specific activity of...β-Galactosidase was isolated from Ranunculus arvensis seeds using DEAE-cellulose,Sephadex G-100,and Con A sepharose-4B chromatography.The enzyme was purified to electrophoretic homogeneity with a specific activity of 50 U/mg of protein and a yield of 7.1%.The molecular mass of the isolatedβ-galactosidase,as estimated by SDS-PAGE,was 18 kDa,indicating that it was a monomeric form The purifiedβ-galactosidase has a glycoproteinic nature when applied to Con-A-Sepharose-4B chromatography.An activation energy(Ea)of 11 kcal/mol of lactose,pH 5.0,and 50℃ were found to be the optimum parameters to purify β-galactosidase from R.arvensis seeds.The residual activity test was carried at 55-75℃,allowing calculating the half-lives of 533-48 min,enthalpy(ΔH°=110.38-110.21 kJ/mol),free energy(ΔG°=109.88-109.77 kJ/mol),and entropy(ΔS°=1.52-1.26 J/mol·K).The β-galactosidase produced from this species is better than the previously described enzyme due to its kinetic and thermodynamic properties,and it could be used in various industrial applications.Purifiedβ-galactosidase,when incubated with high lactose concentration,showed transgalactosylation activity,leading to trisaccharides as a major product of total galactooligosaccharide(GOS).Therefore,the purifiedβ-galactosidase could be used as a potential alternative in the food industry and would be further explained for trisaccharide synthesis.展开更多
基金funding this research work though the project number KKU-IFP2-P-1.
文摘β-Galactosidase was isolated from Ranunculus arvensis seeds using DEAE-cellulose,Sephadex G-100,and Con A sepharose-4B chromatography.The enzyme was purified to electrophoretic homogeneity with a specific activity of 50 U/mg of protein and a yield of 7.1%.The molecular mass of the isolatedβ-galactosidase,as estimated by SDS-PAGE,was 18 kDa,indicating that it was a monomeric form The purifiedβ-galactosidase has a glycoproteinic nature when applied to Con-A-Sepharose-4B chromatography.An activation energy(Ea)of 11 kcal/mol of lactose,pH 5.0,and 50℃ were found to be the optimum parameters to purify β-galactosidase from R.arvensis seeds.The residual activity test was carried at 55-75℃,allowing calculating the half-lives of 533-48 min,enthalpy(ΔH°=110.38-110.21 kJ/mol),free energy(ΔG°=109.88-109.77 kJ/mol),and entropy(ΔS°=1.52-1.26 J/mol·K).The β-galactosidase produced from this species is better than the previously described enzyme due to its kinetic and thermodynamic properties,and it could be used in various industrial applications.Purifiedβ-galactosidase,when incubated with high lactose concentration,showed transgalactosylation activity,leading to trisaccharides as a major product of total galactooligosaccharide(GOS).Therefore,the purifiedβ-galactosidase could be used as a potential alternative in the food industry and would be further explained for trisaccharide synthesis.
