Properties of alpha-chymotrypsin covalently immobilized on poly(acrylic acid)-grafted magnetite particles

Enzymatic properties of alpha-chymotrypsin covalently immobilized on magnet ic particles rin graft polymerization of acrylic acid were investigated. Gr aft polymerization was carried out in a redox system consisting of mercapto groups introduced onto the surfaces of magnetite particles and eerie ions, alpha-Chymotrypsin was immobilized on magnetite particles by condensation with the carboxyl groups of the grafted poly(acrylic acid). The amount of a lpha-chymotrypsin immobilized on 1 g of magnetic was 13-17mg and the activi ty of the immobilized alpha-chymotrypsin (at 37 degrees C, pH 8.0) was 70% the maximum activity of the native enzyme. Due to immobilization, optimum p H for alpha-chymotrypsin shifted to a slightly higher value, whereas optimu m temperature did not change. A kinetic study of the enzymatic reaction wit h immobilized alpha-chymotrypsin showed that the immobilization limited acc essibility of substrate molecules to the active sites of the enzyme bur cau sed little decrease of the maximum reaction rate. In water at 37 degrees C, immobilized alpha-chymotrypsin kept 93% of its original activity over a pe riod of 25 days, though the native enzyme was completely deactivated within 5 days by autolytic denaturalization.