Ingénierie enzymatique
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Reuse of Cyclodextrin Glycosyltransferase through Immobilization on Magnetic Carriers

Kate Cristina Blanco, Francisco José dos Santos, Natalia Sozza Bernardi, Miguel Jafelicci Júnior, Rubens Monti and Jonas Contiero

CGTase from Bacillus lehensis isolated from wastewater of a cassava flour mill in the state of São Paulo, Brazil was immobilized after being partially purified using precipitation with ammonium sulfate. The partially purified CGTase was immobilized by covalent binding on a magnetite support, which was silanized with 3-aminopropyltrimethoxysilane and activated with glutaraldehyde, resulting in a yield of 16.27% and final activity of 17.54% of the initial activity. The physicochemical properties and kinetics of cyclodextrin glycosyltransferase from Bacillus lehensis were determined. The optimum temperature of the immobilized CGTase was higher than that of the soluble enzyme (70°C and 55°C, respectively). The optimum pH of the immobilized enzyme was the same as that of the soluble enzyme (pH 8.0). In the pH and thermo stability assays, 50% of enzyme activity was maintained after 24 h and 26.22% of the initial activity was retained after 160 min, respectively. The Michaelis-Menten constant was 0.82 mg.ml-1 and maximum velocity was 45.45 mol.ml−1.min−1. In the reuse study of the immobilized enzyme after four cycles, 16.66% of the initial catalytic activity was maintained, demonstrating the ability to recover and reuse the enzyme immobilized on magnetite.