Study of Immobilized and Extracelullar Lactase in Mouse-ear Cress
a Botanic Gardens, b Department of Molecular and Subcelullar Biology of Drugs, Faculty of Pharmacy, Comenius University, Bratislava, Slovakia, c Institute of Biochemistry and Biotechnology, Martin Luther University, Halle/Saale, Germany, d Department of Molecular Biology, Institute of Pharmaceutical Biology and Pharmacology, Martin Luther University, Halle/Saale, Germany, e Department of Botany, Faculty of Natural Sciences, Comenius University, Bratislava, Slovakia
Cells in suspension culture of mouse-ear cress (Arabidopsis thaliana (L.), Heynh. cultivar, Columbia were permeabilized with Tween 20, Tween 80, ethanol, and hexadecyltrimethylammonium bromide or hexadecylpyridinium chloride, and immobilized using glutaraldehyde. Lactase showed a pH optimum at 4.7 and the optimal temperatures for immobilized cells and cell cultures was 60 °C and 56 °C, respectively. Four-hour enzyme hydrolysis of the tested substrate proceeded with a conversion of 60??????70 %. The immobilized cells showed a high lactase activity, fair stability in long-term storage and convenient physicomechanical properties. The culture medium after removing cells was used for identification and determination of enzyme activity. The extracellular activity of lactase estimated in cell suspension accounts for 61 % of the total activity, the rest being due to the intracellular activity. The specific extratracellular activity is 4.15 times higher than the intracellular one. Using a histochemical method, secretion of lactase by root tips, root hairs, callus and suspension cells was detected. The described method permits a rapid, simple and specific identification of lactase.
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