Metal-binding properties of the peptide APP170-188:: A model of the ZnII-binding site of amyloid precursor protein (APP)

Amyloid precursor protein (APP) plays a key role in Alzheimer's disease (AD), although the function of this membrane protein is still unclear. Metal ions are implicated in AD and they also interact with APP. APP possesses a strong Zn-II binding site, which is evolutionary conserved. In this paper a synthetic peptide, APP(170-188), with a sequence corresponding to the conserved Zn-II-binding domain of APP, was synthesised and its metal-binding properties analysed. Titration experiments pointed to the binding of a stoichiometric amount of divalent ions. Further studies indicated that the binding of divalent metals like Zn-II, Cd-II and Co-II induces the dimerisation of the peptide. This dimer contains a dinuclear cluster in which the two divalent metals are bridged by two thiolate ligands from cysteine residues. The other two ligands of the tetrahedral coordination sites of each metal ion are terminal thiolate ligands. This structure was supported by the following arguments. The complex formed with Co-II presents the characteristic features for tetrahedral tetrathiolate coordination in its UV-visible spectrum. The sequence of APP(170-188). contains only three cysteine residues, which is incompatible with a monomeric Co-II-APP(170-188) complex. EPR measurements of the complex with one equivalent of Co-II show almost no signal at 4 K, which is compatible with an antiferromagnetic spin-coupling of the metal ions in a cluster structure. Size-exclusion chromatography indicated that the elution time for the complexes with Zn-II and Cd-II corresponds to the expected molecular weight of a dimer. The circular dichroism (CD) spectrum of the complex with one equivalent of Cd-II shows a band at 265 nm(+), and an ellipticity similar to those observed for similar Cd-II-thiolate clusters. Possible biological implications of the Zn-II binding site and the metal-induced dimerisation are discussed.