Structures and properties of α-amylase and glucoamylase immobilized by ZIF-8 via one-pot preparation

The immobilization of α-amylase and glucoamylase using a metal-organic framework (enzyme@ZIF-8) was prepared in situ through a one-pot method. The morphology, crystal structure, and molecular characteristics of the free enzyme and enzyme@ZIF-8 were characterized. The enzyme@ZIF-8 exhibited the rhombic dodecahedron morphology, with a decrease in particle size. Successful immobilization of α-amylase and glucoamylase within ZIF-8 was confirmed, with 30-40 % loading rate. The immobilization process did not significantly alter the crystal structure of ZIF-8. The changes in secondary structure of enzyme after immobilization resulted in modification of catalytic activity of enzyme. The melting enthalpy of enzyme @ZIF-8 increased with the increase of enzyme content. The melting peak temperature of the enzyme immobilized by ZIF-8 increased. The activity of free and immobilized enzymes was influenced by the different time, pH and temperature. At pH 5-8 and temperature 60-80 °C, the activity of the immobilized enzyme was significantly greater than that of the free enzyme. The repeatability of enzyme@ZIF-8 was 61.52 % after three cycles. The kinetic parameters of Michaelis-Menten model for enzymatic reaction were determined by fitting the initial rate of reactions and initial substrate concentration data. The Michaelis-Menten constant (K_M) values of immobilized enzyme were lower than that of free enzyme, indicating the greater affinity between the enzyme and the substrate.