Copper nanoparticles (Cu-NPs) have a wide range of applications as heterogeneous catalysts. In this study, a novel green biosynthesis route for producing Cu-NPs using the metal-reducing bacterium, Shewanella oneidensis is demonstrated. Thin section transmission electron microscopy shows that the Cu-NPs are predominantly intracellular and present in a typical size range of 20-40 nm. Serial block-face scanning electron microscopy demonstrates the Cu-NPs are well-dispersed across the 3D structure of the cells. X-ray absorption near-edge spectroscopy and extended X-ray absorption fine-structure spectroscopy analysis show the nanoparticles are Cu(0), however, atomic resolution images and electron energy loss spectroscopy suggest partial oxidation of the surface layer to Cu2 O upon exposure to air. The catalytic activity of the Cu-NPs is demonstrated in an archetypal "click chemistry" reaction, generating good yields during azide-alkyne cycloadditions, most likely catalyzed by the Cu(I) surface layer of the nanoparticles. Furthermore, cytochrome deletion mutants suggest a novel metal reduction system is involved in enzymatic Cu(II) reduction and Cu-NP synthesis, which is not dependent on the Mtr pathway commonly used to reduce other high oxidation state metals in this bacterium. This work demonstrates a novel, simple, green biosynthesis method for producing efficient copper nanoparticle catalysts.