Modification of paint with nanoparticles (NPs) provides self-cleaning, water/dirt-repellent, and other properties. Therefore, the aim of the present study was to biosynthesize silver (Ag) and copper oxide (CuO) NPs and to prepare NP-modified paint. To this end, AgNPs and CuONPs were biosynthesized usingBacillus atrophaeusspores and commercial and crude dipicolinic acid (DPA) extracted from the spore of this bacterium. The synthesized NPs were characterized using electron microscopy, Fourier-transform infrared (FTIR), X-ray diffraction analysis (XRD), and energy-dispersive X-ray spectroscopy (EDS) methods. A minimum inhibitory concentration (MIC) assay of NPs againstEscherichia coliATCC8739 andStaphylococcus aureusATCC6538 was carried out. The antibacterial effects of prepared NP–paint complexes were assessed using an optical density (OD) comparison before and after adding metal sheets coated with NP–paint complexes into the nutrient broth medium. Four different types of NPs were synthesized in this research: AgNPs synthesized by spore (A), AgNPs synthesized by commercial DPA (B), AgNPs synthesized by crude DPA (C), and CuONPs synthesized by spore (D). SEM analysis confirmed the spherical shape of NPs. According to the results, NPs A, B, and D showed higher antibacterial activity againstS. aureuscompared toE. coli. Furthermore, the analysis of the antibacterial effects of NP–paint complexes suggested that paint–NPs A, B, and C displayed higher activity onE. colicompared toS. aureus. Moreover, the antibacterial effect of paint–NP D was significantly lower than other NPs. According to this robust antibacterial effect on pathogenic bacteria, it seems that these NP–paint complexes could be useful in public places such as hospitals, airports, dormitories, schools, and office buildings, where the rate of transmission of infection is high.