Preparation and studies on immobilized α-glucosidase from baker’s yeast Saccharomyces cerevisiae

Abstract

α-Glucosidase from S. cerevisiae was covalently immobilized onto Sepabeads EC-EA by the glutaraldehyde method. An analysis of the variables controlling the immobilization process is first presented and it is shown that the highest coupling of α-glucosidase occurred within 24 h. Also, a loading of 30 mg/g support proved to be effective, resulting in a rather high activity of around 45 U g-1 with a satisfactory degree of enzyme fixed. Both free and immobilized enzymes were then characterized by determining the activity profile as a function of pH, temperature and thermal stability. The obtained immobilized preparation showed the same optimum pH, but a higher optimum temperature compared with the soluble one. In addition, the immobilized enzyme treated at 45 ºC for 1 h still retained an activity of around 20 %, whereas the free enzyme completely lost its original activity under this condition. In conclusion, the developed immobilization procedure is quite simple, easily reproducible and provides a promising solution for the application of immobilized α-glucosidase.

Maltaza iz S. cerevisiae je kovalentno imobilizovana na Sepabeads EC-EA nakon aktivacije nosača rastvorom glutaraldehida. Ispitivanjem kinetike imobilizacije utvrđeno je da se 25 % enzima imobilizuje nakon 24 časa. Imobilizovana α-glukozidaza ima isti pH optimum kao i rastvorni enzim, dok je optimalna temperatura za aktivnost imobilizovanog enzima uvećana za 10 °C u poređenju sa rastvornim enzimom. Kada se uporede zaostale aktivnosti rastvorne i imobilizovane forme α-glukozidaze, nakon inkubacije od 1 h na 45 °C rastvorni enzim ne pokazuje aktivnost dok imobilizovana forma zadržava oko 20 % početne aktivnosti. Imobilizovana forma enzima zadržava 20 % početne aktivnosti čak i posle 3 h inkubacije na 45 °C.