Activity modulation and reusability of beta-D-galactosidase confined in sol-gel derived porous silicate glass

作者：Crescimbeni Marina C; Nolan Veronica; Clop Pedro D; Marin Graciela N; Perillo Maria A^*

来源：Colloids and Surfaces B: Biointerfaces , 2010, 76(2): 387-396.

DOI：10.1016/j.colsurfb.2009.11.005

• beta-D-Galactosidase
• Silicate matrix
• Sol-gel
• Encapsulation
• ONPG hydrolysis
• Reusability

In the present work we applied thesol-gel method to obtain glass lentils entrapping beta-D-galactosidase (beta-Gal) (E(beta-Gal)) within a silicate matrix. The effect of pH, temperature, polarity and salt concentration on the activity of E(beta-Gal) was Studied. Apparent kinetic parameters for ortlio-nitro-phenyl-beta-D-galactopyranoside hydrolysis catalyzed by E beta-Gal (V(max)', K(M)') were lower compared to the soluble enzyme (S(beta-Gal)), reflecting the solute diffusion restriction imposed by the matrix observed in the time Curves, a partial protein inactivation upon encapsulation, and an improvement in the affinity of for the substrate as compared with S(beta-Gal). At pH < 4, E(beta-Gal) stability was higher than that of S(beta-Gal), E(beta-Gal) Could be reused after storage at 4 degrees C for up to 90 days, and retained its activity profile within the range of pH = 2-10 and saline concentration 0-400 mM. Pre-incubation at 75 degrees C for 30 min fully inactivated S(beta-Gal) while E(beta-Gal) retained approximately 90% of its activity, even in the reused samples. Encapsulation did not introduce additional impairments to the reaction rate measured in heterogeneous dispersions, beyond those derived from their own particle-crowded environment. This reusable E(beta-Gal) was resistant to typical technological conditions applied in milk processing that Would lead to the Unfolding and inactivation of S(beta-Gal). The results are discussed from the biophysical viewpoint.

• 出版日期2010-4-1