Redox reactivities of membrane-bound amyloid-β-Cu complexes and their targeting by metallothionein-3

Abstract

Alzheimers disease (AD) is characterized by the accumulation of amyloid-{beta} peptide (A{beta}1-40/42) in the central nervous system (CNS) and its aggregation in senile amyloid plaques. Copper coordination to A{beta} triggers A{beta}1-40/42 aggregation and, in the presence of biological reducing agents, it promotes the catalytic generation of reactive oxygen species (ROS) via Fenton-type and Haber-Weiss reactions. Due to its amphiphilic nature, A{beta}1-40/42 can interact with cell membranes and compromise their integrity by thinning the lipid bilayer and forming channel-like structures potentially leading to cell death. In this work, by applying biophysical and biochemical approaches, we characterized the insertion of A{beta}1-42 into an artificial lipid bilayer system mimicking cell membranes and demonstrate that the A{beta}1-42-lipid interaction does not prevent the Cu2+ coordination to A{beta}1-42. We performed a comparative analysis of the redox reactivities of membrane-bound A{beta}1-42 (memA{beta}1-42-Cu2+) species with soluble A{beta}1-42-Cu2+ establishing that membrane insertion leads to memA{beta}1-42-Cu2+ complexes featuring an enhanced detrimental catechol oxidase activity towards the neurotransmitter dopamine. Moreover, memA{beta}1-42-Cu2+ efficiently catalyzes A{beta} di-tyrosine crosslinking and hydroxyl radical production in the presence of ascorbate. In addition, we establish that memA{beta}1-42-Cu2+ redox reactivity catalyze lipid peroxidation in membranes containing polyunsaturated fatty acids (PUFAs), such as arachidonic acid (AA), leading to the generation of malondialdehyde (MDA) toxic end-products. This reactivity compromises the structural integrity of the lipid bilayers resulting in membrane leakage, further substantiating how important is to control aberrant A{beta}1-40/42-Cu2+ interactions in AD. Metallothioneins (MTs) are key metalloproteins central to neuronal and astrocytic transition metal homeostasis and buffering. These cysteine-rich proteins bind with high affinity d10 metals (Cu+ and Zn2+) forming two metal thiolate clusters in their N-terminal {beta}-domain and C-terminal -domain. The metallothionein-3 (MT-3) isoform is central to metal homeostasis in the CNS, but it is downregulated in AD patients, possessing a neuroprotective role in AD. MT-3 can control aberrant protein-Cu2+ interactions and the Cu-centered redox reactivities of amyloidogenic protein-Cu2+ complexes such as -synuclein (Parkinsons disease), PrP (prion disease), and soluble and aggregated A{beta}1-40 (AD). In this work, we unravel that the detrimental memA{beta}1-42-Cu2+ catechol oxidase and redox reactivities can be efficiently silenced by MT-3 via metal swap reactions, effectively scavenging and reducing Cu2+ to Cu+ in its {beta}-domain using thiolates as electron source, forming the redox-inert species Cu+4Zn2+4MT-3. Consequently, MT-3 can efficiently prevent lipid peroxidation and protect membrane structural integrity. New strategies targeting membrane-bound A{beta}1-42-Cu2+ complexes as key players of the AD etiology could be envisioned.