Redox Silencing of Copper in Metal-linked Neurodegenerative Disorders

Abstract

Dysregulation of copper and zinc homeostasis in the brain plays a critical role in Alzheimer disease (AD). Copper binding to amyloid-␤ peptide (A␤) is linked with the neurotoxicity of A␤and free radical damage. Metallothionein-3 (MT-3) is a small cysteine-and metal-rich protein expressed in the brain and found down-regulated in AD. This protein occurs intra-and extracellularly, and it plays an important role in the metabolism of zinc and copper. In cell cultures Zn 7 MT-3, by an unknown mechanism, protects neurons from the toxicity of A␤. We have, therefore, used a range of complementary spectroscopic and biochemical methods to characterize the interaction of Zn 7 MT-3 with free Cu 2؉ ions. We show that Zn 7 MT-3 scavenges free Cu Essential transition metals like copper and zinc play a critical role in neurobiology. The homeostasis of both metals is tightly regulated and essential for brain physiology (1). In vitro evidence suggests that in physiological conditions micromolar concentrations of zinc and copper are actively released from neurons during neurotransmission processes into pre-and postsynaptic clefts. However, at present it is not known whether the released copper is present in a chemically exchangeable form (2). Dysregulated metal metabolism (zinc and copper) occurs in neurodegenerative disorders such as Alzheimer (3, 4), Parkinson (5), and prion (6) diseases. In these diseases increased extracellular copper concentrations and free-radical production have been found. There is now direct evidence that copper is bound to amyloid-␤ peptide (A␤) 2 in senile plaque of Alzheimer disease (AD) (7). Copper is also linked with the neurotoxicity of A␤ and free radical damage (8).Because of its redox-active nature (Cu ϩ /Cu 2ϩ ), its reactivity with molecular oxygen (O 2 ) generates the reactive oxygen species (ROS) such as superoxide, hydrogen peroxide, and hydroxyl radicals (9, 10). The redox cycling of copper requires its reduction by biological components, including ascorbate, which actively accumulates in the brain at concentrations between 0.5 and 10 mM (11). Hence, copper chelation and its redox-silencing may represent critical events in preventing the progression of neurodegenerative diseases. In recent years a metal chelation therapy has emerged as a promising tool to attenuate abnormal metal-protein interactions that lead to increased free-radical toxicity (10).The natural metal chelator metallothionein-3 (MT-3), also known as the growth inhibitory factor, is a small non-inducible cysteine-and metal-rich protein mainly expressed in the brain. In the brain its expression was found in zinc-enriched neurons. Like other mammalian metallothioneins (MTs), MT-3 binds with a high affinity essential monovalent and divalent d 10 metal ions Cu(I) and Zn(II). Recently, the interaction of Zn 7 MT-3 with the small-GTPase Rab3a, which is strictly linked to the exo-endocytotic cycle of synaptic vesicles, has been demonstrated. It has been suggested that Zn 7 MT-3 actively participates in synaptic cycle of zinc vesicles...

Metal swap between Zn7-metallothionein-3 and amyloid-β–Cu protects against amyloid-β toxicity

“…1b). The newly developed CysS-Cu(I) charge transfer CD bands at about (+)255 and (-)285 nm were identical in the size and the position to those already observed in the reaction of Zn 7 MT-3 with free Cu(II) 16 ( Supplementary Fig. 2 online).…”

“…This study builds on our recent work in which, by complementary spectroscopic techniques, we provided evidence that Zn 7 MT-3 through the cooperative formation of an air stable Cu(I) 4 Zn 4 MT-3 species efficiently 4 scavenges and redox silences free extracellularly occurring Cu(II) 16 . In Aβ 1-40 -Cu(II) copper is tightly bound in a square-planar geometry through three histidine ligands (His 6 , His 13 , and His 14 ) and an ill defined fourth ligand [17][18][19] .…”

“…Nevertheless, the stability of the Cu(I) 4 -thiolate cluster in air oxygen is striking and not understood. We reasoned that the structural constraints and the short Cu(I)···Cu(I) distances (< 2.8Å) in the Cu(I) 4 -thiolate cluster, leading to peculiar metal-metal interactions, may be important for its stability 16 .…”

“…These results and the mass of the isolated protein of 7,431.0 Da, obtained by electrospray ionization time-of-flight mass spectrometry (ESI-TOF MS) (Supplementary Table 1 online), suggested that also in this case Cu(I) 4 Zn 4 MT-3 containing two disulfide bonds was formed. Evidence for the presence of a Cu(I) 4 -thiolate cluster in Cu(I) 4 Zn 4 MT-3 was provided by the appearance of its characteristic low temperature Cu(I) emission spectrum 16 (Fig. 1c).…”

“…1c). This Accordingly, the high energy emissive band in Cu(I) 4 Zn 4 MT-3 has been assigned to 3 CC (cluster centered) origin and that at low energy to triplet charge transfer 6,16 . Consequently, peculiar metal-metal interactions in this cluster exist.…”