A novel method of immobilizing horse radish peroxidase enzyme (HRP) onto the surface of gold nanoparticles (GNPs) was developed. As a result, a high-activity bionanoconjugates was obtained through utilizing the biotin-streptavidin (SA) system. The HRP-SA-GNP bionanoconjugate with high activity was conveniently prepared through the biotin- avidin system. Compared with the HRP-GNP bioconjugate prepared through the traditional electrostatic absorption method, the enzyme activity per GNPs of this new bionanoconjugate was enhanced by 10 times. Moreover, the enzyme activity of this bionanoconjugate was controllable. The above method of bionanoconjugation preparation has promising applications in the fields including preparing highly active bio-nanoprobe and immobilized enzyme.