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The rise in antimicrobial resistance requires the development of new antibacterial agents. Herein, we develop nanocomposite by growing zinc oxide nanoparticles on the surface of chitosan (CS) modified graphene oxide (GO), obtaining GO@CS/ZnO as a novel antibacterial material. The crystal structures, surface functional groups and morphology were analyzed by using X-ray diffraction (XRD) pattern, Fourier transform infrared spectroscopy (FTIR) scanning electron microscopy (SEM) images, respectively. The Minimum Inhibitory concentration (MIC) and the Minimum Bactericidal Concentration (MBC) of GO@CS/ZnO show more antibacterial potency toward both Gram-negative bacteria Escherichia coli (E. coli) and Gram-positive bacteria Staphylococcus aureus (S. aureus). In this study the GO@CS/ZnO expresses its more potency than reported graphene based nanocomposites. Investigation of intercellular reactive oxygen species (ROS) generation, NO generation and catalase activity in E. coli and S. aureus reveal that GO@CS/ZnO treatment also augments the intracellular bacterial killing by inducing reactive oxygen species production that causes oxidative damage. The minimal inhibition concentrations (MIC) of GO@CS/ZnO against E.coli and S. aureus are only 2.5 µg/mL and 5 µg/mL respectively. Compared with graphene based nanocomposite, which have been widely used as antibacterial agents, our GO@CS/ZnO shows better antibacterial effect. We envision that this study offers novel insights into antimicrobial actions and also demonstrates GO@CS/ZnO is a novel class of topical antibacterial agent in the areas of healthcare and environmental engineering.