Abstract
The diffuse and renewed use of silver as antimicrobial agent has caused the development of resistance to silver ions in some bacterial strains, posing a serious threat for health systems. In order to cast light on the mechanistic features of resistance, here, we aimed to understand how silver interacts with the periplasmic metal-binding protein SilE which is engaged in bacterial silver detoxification. This aim was addressed by studying two peptide portions of SilE sequence (SP2 and SP3) that contain the putative motifs involved in Ag+ binding. We demonstrate that SP2 model peptide is involved in silver binding through its histidine and methionine residues in the two HXXM binding sites. In particular, the first binding site is supposed to bind the Ag+ ion in a linear fashion, while the second binding site complexes the silver ion in a distorted trigonal planar fashion. We propose a model where the SP2 peptide binds two silver ions when the concentration ratio Ag+/SP2 is ≥10.0. We also suggest that the two binding sites of SP2 have different affinity for silver. This evidence comes from the change in the path direction of the Nuclear Magnetic Resonance (NMR) cross-peaks upon the addition of Ag+. Here, we report the conformational changes of SilE model peptides occurring upon silver binding, monitored at a deep level of molecular details. This was addressed by a multifaceted approach, combining NMR, circular dichroism, and mass spectrometry experiments.