Reversible Stereoisomer-Specific Cotton Effect of the Ligand Field Transitions at a Copper(II) Binding Site of the Prion Protein

The prion protein (PrP) is a metal binding protein that plays multifunctional roles in the brain and it is associated with spongiform encephalopathy disorders, a group of neurodegenerative infectious diseases. PrP can bind six copper ions at its N-terminal region, one of the binding sites involving His96. Copper(II) ions bind to His96 forming two coordination modes at physiological pH: a 3N1O species involving the His imidazole group, two deprotonated amides (from His96 and Thr95) and one backbone carbonyl oxygen; and a 4N coordination mode where the oxygen-based ligand is replaced by a third deprotonated amide (from Gly94). In this study, synthetic PrP fragments 92-99 were used to evaluate the impact of different substitutions at positions 94 and 95 of the sequence in the Cotton effect of the ligand field transitions for the 4N species, as studied by circular dichroism (CD) spectroscopy. Specifically, the T95A substitution had no impact, while the G94A substitution yields a change in the sign of the ligand field transitions, which can be reverted when substituting Gly94 by the stereoisomer D-Ala. The electronic structure of the Cu(II)-PrP(92-96) complex and its variants was studied, while time-dependent density functional theory (TD-DFT) calculations provided a prediction of the CD spectra in the ligand field region that is consistent with observed experimental results. Furthermore, the calculated CD spectra for other Cu(II)-PrP(92-96) complexes with substitutions (alkyls and halides) at position 94 indicate that the complexes with L-stereoisomers displays the opposite Cotton effect as compared to their D-stereoisomer counterparts. Our analysis indicates this impact in the Cotton effect is a consequence of the geometric distortion that the nearly square planar 4N Cu(II)-PrP(92-99) complex suffers when an L-substituent is placed in position 94; the distortion from planarity is reversed by the D-stereoisomer substitution. Overall, this study provides insights into the origin of the Cotton effect of the ligand field transitions of nearly square planar Cu(II)-peptide complexes.