Direct evidence that all three histidine residues coordinate to Cu(II) in amyloid-β1-16

We provide direct evidence that all three histidine residues in amyloid-beta(1-16) (A beta(1-16)) coordinate to Cu(II). In our approach, we generate A beta(1-16) analogues, in each of which a selected histidine residue is isotopically enriched with (15)N. Pulsed electron spin resonance (ESR) experiments such as electron spin echo envelope modulation (ESEEM) and hyperfine sublevel correlation (HYSCORE) spectroscopy clearly show that all three histidine imidazole rings at positions 6, 13 and 14 in A beta(1-16) bind to Cu(II). The method employed here does not require either chemical side chain modification or amino acid residue. replacement, each of which is traditionally used to determine whether an amino acid residue in a protein binds to a metal ion. We find that the histidine coordination in the A beta(1-16) peptide is independent of the Cu(II)-to-peptide ratio, which is in contrast to the A beta(1-40) peptide. The ESR results also suggest tight binding between the histidine residues and the Cu(II) ion, which is likely the reason for the high binding affinity of the A beta peptide for Cu(II).