Magnetic Ni0.5Zn0.5Fe2O4@SiO2 nanocomposite was prepared via the rapid combustion process, and its surface was modified to obtain amino-functionalized magnetic Ni0.5Zn0.5Fe2O4@SiO2-NH2 nanocomposite. The modified nanocomposite was loaded on graphene oxide (GO), on which penicillin G acylase (PGA) was covalently immobilized. The structure for docking was visualized between PGA and penicillin G using the PyMol program, which revealed the configuration of the active site. Selections of immobilization conditions including immobilization concentration and time of fixation, were explored. The catalytic performance of the immobilized PGA was characterized. The immobilized and free PGA had the highest activity at pH 8.0 and 45 °C. Compared with the activity of the free PGA, the activity of the immobilized PGA was affected less by pH and temperature. The immobilized PGA exhibited the high-effective activity and good stability. Vmax and Km of immobilized PGA were 0.8123 mol·min-1 and 0.0399 mol·L-1, respectively. Free PGA's Vmax and Km were 0.6854 mol·min-1 and 0.0328 mol·L-1. Immobilized PGA remained >70% in relative activity after 9 successive cycles.