A novel immobilization method for nuclease P ₁ on macroporous absorbent resin with glutaraldehyde cross-linking and determination of its properties

Microbial nuclease P ₁ from Penicllium citrinum was immobilized on macroporous absorbent resins: strong polar poly (styrene-co-DVB) resin (SPPSD), polymethacrylic ester resin and poly (styrene-co-DVB)-Br resin. The results showed that SPPSD was the best carrier. Three methods of glutaraldehyde cross-linking were used and simultaneous immobilization and cross-linking (CIS) was demonstrated to be the best method. The functional properties of immobilized nuclease P ₁ were studied and compared to those of the free enzyme. The highest enzyme activities of free and immobilized nuclease P ₁ were obtained in 0.2 M acetate buffer at pH 4.5 and a temperature of 70 °C. An increase in K _m (from 3.165 to 18.125 mg mL ^-1) and a decrease in V _max (from 1667.18 to 443.95 U min ^-1 mL ^-1) were recorded after immobilization. SPPSD-glutaraldehyde-nuclease P ₁ exhibited better thermal stability than the free enzyme. The apparent activation energy (E _a) of the free and immobilized nuclease P ₁ was 137.04 kJ mol ^-1 and 98.43 kJ mol ^-1, respectively, implying that the catalytic efficiency of the immobilized enzyme was restricted by mass-transfer rather than kinetic limit.