Exploration of the Potential Role for Aβ in Delivery of Extracellular Copper to Ctr1

Amyloid beta (A beta) peptides are notorious for their involvement in Alzheimer's disease (AD), by virtue of their propensity to aggregate to form oligomers, fibrils, and eventually plaques in the brain. Nevertheless, they appear to be essential for correct neurophysiology on the synaptic level and may have additional functions including antimicrobial activity, sealing the blood-brain barrier, promotion of recovery from brain injury, and even tumor suppression. A beta peptides are also avid copper chelators, and coincidentally copper is significantly dysregulated in the AD brain. Copper (Cu) is released in significant amounts during calcium signaling at the synaptic membrane. A beta peptides may have a role in maintaining synaptic Cu homeostasis, including as a scavenger for redox-active Cu and as a chaperone for clearing Cu from the synaptic cleft. Here, we employed the A beta(1-16) and A beta(4-16) peptides as well-established non-aggregating models of major A beta species in healthy and AD brains, and the Ctr(1-14) peptide as a model for the extracellular domain of the human cellular copper transporter protein (Ctr1). With these model peptides and a number of spectroscopic techniques, we investigated whether the Cu complexes of A beta peptides could provide Ctr1 with either Cu(II) or Cu(I). We found that A beta(1-16) fully and rapidly delivered Cu(II) to Ctr(1-14) along the affinity gradient. Such delivery was only partial for the A beta(4-16)/Ctr(1-14) pair, in agreement with the higher complex stability for the former peptide. Moreover, the reaction was very slow and took ca. 40 h to reach equilibrium under the given experimental conditions. In either case of Cu(II) exchange, no intermediate (ternary) species were present in detectable amounts. In contrast, both A beta species released Cu(I) to Ctr(1-14) rapidly and in a quantitative fashion, but ternary intermediate species were detected in the analysis of XAS data. The results presented here are the first direct evidence of a Cu(I) and Cu(II) transfer between the human Ctr1 and A beta model peptides. These results are discussed in terms of the fundamental difference between the peptides' Cu(II) complexes (pleiotropic ensemble of open structures of A beta(1-16) vs the rigid closed-ring system of amino-terminal Cu/Ni binding A beta(4-16)) and the similarity of their Cu(I) complexes (both anchored at the tandem His13/His14, bis-His motif). These results indicate that Cu(I) may be more feasible than Cu(II) as the cargo for copper clearance from the synaptic cleft by A beta peptides and its delivery to Ctr1. The arguments in favor of Cu(I) include the fact that cellular Cu export and uptake proteins (ATPase7A/B and Ctr1, respectively) specifically transport Cu(I), the abundance of extracellular ascorbate reducing agent in the brain, and evidence of a potential associative (hand-off) mechanism of Cu(I) transfer that may mirror the mechanisms of intracellular Cu chaperone proteins.