The biosynthesis of metallic nanomaterials, especially noble metal nanomaterials, has attracted extensive attention due to the multifarious potential applications of these materials. In this study we have explored the effects of Cu(II) on Pd(II) removal, and of Cu(II) as well as phosphate on the morphologies and composition of biogenic nanoparticles (NPs) from aqueous Pd(II) systems by Shewanella oneidensis MR‐1 (MR‐1). Cu(II) causes a slight efficiency decrease of Pd(II) removal by MR‐1, but a high removal efficiency of 89.8 % is still achieved, which contrasts sharply with the significant inhibition to other microbes. Without phosphate buffer in the medium, 5‐nm PdNPs densely envelop MR‐1 and accumulate significantly in the periplasm in the absence of Cu(II), while 4‐nm PdNPs are evenly distributed in the periplasm, and the cellular surface is smooth in the presence of Cu(II). The following effect of Cu(II) on PdNP synthesis by MR‐1 is therefore proposed: Cu(II) significantly affects PdNPs morphologies by inhibiting Pd(0) transfer across the outer membrane and PdNP crystal growth in the periplasm, but not the sorption of Pd(II). Phosphate addition only slightly increases Pd harvest but also precipitates Cu(II) and poses difficulties for the biosynthesis. The study highlights the effect of co‐existing metals during the biosynthesis of metallic nanomaterials and offers new insight into the mechanism of PdNP formation and the electron transfer during Pd(II) reduction by MR‐1.