We synthesized the nanocomposite of chitosan-graft-polyaniline/montmorillonite/zinc oxide (CTS-g-PANi/MMT/ZnO). ZnO nanoparticles (NPs) were successfully synthesized from ZnO powder in the Cts/MMT matrix, using traditional method. CTS-g-PANi co-polymer was synthesized in the presence of ammonium peroxydisulfate (APS). The CTS-g-PANi and MMT were used as the natural polymeric stabilizer and the solid support, respectively. The morphology and the particle size of ZnO NPs and synthesized nanocomposite were investigated by Fourier-transform infrared (FT-IR) spectroscopy, X-ray diffraction (XRD) analysis, and scanning electron microscopy (SEM). According to the XRD results, the nanoparticle size was found to be in the range of 22-35 nm. SEM images showed the hexagonal shape of ZnO NPs, which is suitable for the evaluation of antibacterial activity. Finally, the bactericidal property of prepared nanocomposites was also evaluated against the bacteria of Gram positive Staphylococcus aureus (Staph.) and Gram negative Escherichia coli (E. coli), using the paper disc diffusion method.