CONICET | Buscador de Institutos y Recursos Humanos

It is widely accepted that water molecules inside the cell compartments exhibit physical properties that differ of dilute solutions. Then, the microenvironment in which metabolic activity actually occurs in cells is not equivalent to the conditions used to determine the properties of enzymes in vitro. In this work the kinetic activity of β-Galactosidase confined in a nanopore of a silicate gel was studied and it was related to confined water dynamics. Encapsulation of proteins in silicate gels obtained by the sol-gel method has been of interest for the synthesis of biosensors [1-3] , delivery nanosystems [4, 5], and for the possibility of studying protein properties in conditions far from dilute solutions. The silicate gel obtained showed highly porous structure, quite narrow pore diameter distribution, around 40 nm, and high surface area. The results observed for the encapsulated β-Galactosidase (Eβ-Gal), which catalyses a hydrolysis reaction against two different substrates, ONPG and PNPG, let us take into consideration the differences between bulk and confined water molecules inside the nanopores and the influence of these differences on the catalytic activity. The Vmax value calculated for Eβ-Gal was higher than the Vmax obtained for the free soluble enzyme (Sβ-Gal) when the substrate was ONPG, and was even higher in aged gels. The kinetic results were correlated with a study of water dynamics performed with H1-NMR transversal relaxation time experiments [6]. In view of these results, it is quite plausible to propose that some characteristic of confined water inside the nanopores influence the hydrolytic activity of Eβ-Gal, shedding light on the biophysical processes inside nano environments present in the nanobiotechnology.