Silver has long been used as an antimicrobial agent in general and medicinal use. Here, we observe that exposure of the Gram-positive, endospore-forming bacterium Bacillus subtilis to Ag(I) effects growth in a biphasic manner. In the first phase at Ag(I) concentrations below 50 µM B. subtilis growth is not affected, but activity of the respiratory enzyme cytochrome c oxidase is disrupted completely. Between 50 to 100 µM Ag(I) B. subtilis growth is drastically diminished and completely absent above 100 µM Ag(I). Synthesis of cytochrome c oxidase, or SCO proteins, have been shown to play a role in assembly of the CuA center of cytochrome c oxidase and we suppose that the effects observed here of silver on Bacillus subtilis in culture may be explained at least in part by the interaction of Bacillus SCO (BsSCO) with Ag(I). We find that Ag(I) forms a high affinity complex with BsSCO in vitro that blocks SCO’s interaction with copper indicating competition between the metals for binding BsSCO. The interaction of BsSCO with Ag(I) exhibits multiple phases and is more complex than that observed for the high-affinity, 1:1 copper complex with BsSCO. We propose that the initial response of B. subtilis cultures is due to high affinity binding of Ag(I) to BsSCO that blocks the functionality of BsSCO required for assembly of cytochrome c oxidase. Our results provide evidence of a specific effect of silver on Bacillus subtilis cells and implies that SCO proteins play a role in sensitivity to Ag(I).