Mutants of Metal Binding Site M1 in APP E2 Show Metal Specific Differences in Binding of Heparin but Not of sorLA

The amyloid precursor protein (APP) and its neurotoxic cleavage product A beta are key players in the development of Alzheimers disease (AD) and appear to be essential for neuronal development and cell homeostasis. Proteolytic processing of APP and its physiological function depend on its interaction with heparin and are influenced by the binding of metal ions and sorLA. We created various mutations of metal binding site M1 residing within the extracellular E2 domain of APP. Using isothermal titration calorimetry and circular dichroism spectroscopy, we analyzed the binding of Cu2+ and Zn2+ to APP E2 and identified two mutations that are most suited for functional studies to dissect ion specific effects of metal binding. The H313A mutation abrogates only copper-based effects, whereas the H382A mutation weakens any metal binding at M1 of APP E2. Subsequently, we tested the effect of Cu2+ and Zn2+ on the binding of heparin and sorLA to APP E2 using a chromatographic technique and surface plasmon resonance. We show that Zn2+ and to a larger degree also Cu2+ enhance the binding of heparin to APP E2, consistent with an extracellular regulation of the function of APP by both metal ions. In contrast, neither ion seemed to affect the interaction between APP E2 and sorLA. This supports an intracellular interaction between the latter two partners that would not sense extracellular variations of metal ions upon synaptic activity.