Traditional methods to incorporate metals into graphene oxide (GO) usually require toxic reagents or high temperatures. This study proposes an innovative and sustainable method to incorporate silver (Ag) into graphene oxide using electron beam and evaluate its antibacterial activities. The method is based on green synthesis, without toxic reagents or hazardous wastes, and can be carried out at room temperature, in short reaction times. To synthesize the Ag/rGO nanocomposite, a water/isopropanol solution with dispersed graphene oxide and silver nitrate was submitted to a dose range from 150 to 400 kGy. The product was characterized by thermogravimetry analysis, X-ray diffraction and transmission electron microscopy. The antibacterial activity of Ag/rGO was observed against Gram-negative Escherichia coli by plate count method and atomic force microscopy. The results showed that concentrations as low as 100 μg/mL of produced Ag/rGO were enough to inactivate the cells.