Insight into the Cation−π Interaction at the Metal Binding Site of the Copper Metallochaperone CusF

The periplasmic Cu⁺/Ag⁺ chaperone CusF features a novel cation−π interaction between a Cu⁺/Ag⁺ ion and Trp44 at the metal binding site. The nature and strength of the Cu⁺/Ag⁺–Trp44 interactions were investigated using computational methodologies. Quantum-mechanical (QM) calculations showed that the Cu⁺ and Ag⁺ interactions with Trp44 are of similar strength (∼14 kcal/mol) and bond order. Quantum-mechanical/molecular-mechanical (QM/MM) calculations showed that Cu⁺ binds in a distorted tetrahedral coordination environment in the Trp44Met mutant, which lacks the cation−π interaction. Molecular dynamics (MD) simulations of CusF in the apo and Cu⁺-bound states emphasized the importance of the Cu⁺–Trp44 interaction in protecting Cu⁺ from water oxidation. The protein structure does not change over the time scale of hundreds of nanoseconds in the metal-bound state. The metal recognition site exhibits small motions in the apo state but remains largely preorganized toward metal binding. Trp44 remains oriented to form the cation−π interaction in the apo state and faces an energetic penalty to move away from the metal ion. Cu⁺ binding quenches the protein’s internal motions in regions linked to binding CusB, suggesting that protein motions play an essential role in Cu⁺ transfer to CusB