This work describes the performance of a tyrosinase/nickel oxide nanoparticles/glassy carbon (Tyr/NiO NPs/GC) electrode. This electrode was prepared by first applying a NiO NPs electrochemical deposition onto the GC electrode surface and then tyrosinase immobilization was applied to the surface of electrodeposited NiO NPs. Scanning electron microscopy (SEM) and atomic force microscopy (AFM) procedures demonstrated the existence of different NiO NP geometrical structures. These geometrical structures could lead to better immobilization of proteins on their surfaces. The copper containing enzyme tyrosinase successfully achieved electrical contact with the electrode because of the unique structural alignment of tyrosinase enzyme on the nanometer-scale nickel oxide surfaces. This method could be suitable for application to nanofabricated devices facilitating better performance. It was concluded that tyrosinase can be effectively applied to nanometer-scale nickel oxide surfaces.