Immobilization of Kluyveromyces lactis β‐Galactosidase on Meso‐macroporous Silica: Use of Infrared Spectroscopy to Rationalize the Catalytic Efficiency

Abstract

Enzyme immobilization on adequate carriers is a challenging strategy. Understanding the enzyme-carrier interactions and their effects on enzyme conformation and bioactivity is critical. In this study, a meso-macropores silica (MMS) was used to immobilize β-galactosidase from the yeast Kluyveromyces lactis (β-gal-KL) by physical adsorption. The bioactivity of the immobilized β-gal-KL was altered, evidenced by the increased K m , decreased V max and k cat , and increased activity at alkaline values. By performing infrared spectroscopy analysis and subsequent secondary structure assessment from the amide I band, the immobilized β-gal-KL suffered a β-sheet (~31-35 %) to α-helix (~15-19 %) transition with increased turns (~21-22 %) with respect to the free β-gal-KL having ~12 % α-helix, ~42 % β-sheet, and ~17 % turns. These findings led us to correlate the observed bioactivity performance to structural alterations to a non-native conformation. The presented line of thought can lead to a better understanding of the reasons causing bioactivity alterations upon enzyme immobilization.