Abstract
Aβ4–42 is the major subspecies of Aβ peptides characterized by avid Cu(II) binding via the ATCUN/NTS motif. It is thought to be produced in vivo proteolytically by neprilysin, but in vitro experiments in the presence of Cu(II) ions indicated preferable formation of C-terminally truncated ATCUN/NTS species including CuIIAβ4–16, CuIIAβ4–9, and also CuIIAβ12–16, all with nearly femtomolar affinities at neutral pH. Such small complexes may serve as shuttles for copper clearance from extracellular brain spaces, on condition they could survive intracellular conditions upon crossing biological barriers. In order to ascertain such possibility, we studied the reactions of CuIIAβ4–16, CuIIAβ4–9, CuIIAβ12–16, and CuIIAβ1–16 with reduced glutathione (GSH) under aerobic and anaerobic conditions using absorption spectroscopy and mass spectrometry. We found CuIIAβ4–16 and CuIIAβ4–9 to be strongly resistant to reduction and concomitant formation of Cu(I)–GSH complexes, with reaction times ∼10 h, while CuIIAβ12–16 was reduced within minutes and CuIIAβ1–16 within seconds of incubation. Upon GSH exhaustion by molecular oxygen, the CuIIAβ complexes were reformed with no concomitant oxidative damage to peptides. These finding reinforce the concept of Aβ4–x peptides as physiological trafficking partners of brain copper.
Highlights
Aβ4−42 is the major subspecies of Aβ peptides characterized by avid Cu(II) binding via the ATCUN/NTS motif
A β peptides are products of extracellular hydrolysis of amyloid precursor protein (APP) present in neuronal synaptic membranes.[1−3] Overproduction or excessive aggregation of Aβ has been long implicated as an upstream cause of neuronal death in Alzheimer’s disease (AD).[4,5]
A number of brain proteases with other known functions have been implicated in this process, including neprilysin (NEP), angiotensinogen converting enzyme (ACE), and insulin degrading enzyme (IDE).[21]
Summary
ADetermined from the initial decay of the d−d band at 525 nm. Velocities are given in nM/s; all data are shown ± SD. They could be assigned to the Cu(I) complex of GSH, reported previously by others.[33,34] This CuII reduction phase lasted for about 9 h and reproducibly reached about 65% CuII conversion at 25 °C, as calculated from the intensity of the CuII(Aβ4−16) d−d band at 525 nm (Table 1). The comparison of kinetic traces indicated the similarity of the early phase of the reduction process between the aerobic and anaerobic conditions (Figure S10). These traces exhibited the mathematical form of first order kinetics for all conditions, only differing by the degree of CuII reduction: ca. ■ AUTHOR INFORMATION (9) Alies, B.; Renaglia, E.; Roź ga, M.; Bal, W.; Faller, P.; Hureau, C
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