The Pathogenic A2V Mutant Exhibits Distinct Aggregation Kinetics, Metal Site Structure, and Metal Exchange of the Cu²⁺–Aβ Complex

Abstract

A prominent current hypothesis is that impaired metal ion homeostasis may contribute to Alzheimer's disease (AD). We elucidate the interaction of Cu with wild-type (WT) Aβ and the genetic variants A2T and A2V which display increasing pathogenicity as A2T<WT<A2V. Cu significantly extends the lag phase in aggregation kinetics, in particular for the pathogenic A2V variant. Additionally, a rapid, initial, low intensity ThT response is observed, possibly reflecting formation of Cu induced amorphous aggregates, as supported by atomic force microscopy (AFM) and circular dichroism (CD) spectroscopy, again most notably for the A2V variant. Electron paramagnetic resonance (EPR) spectroscopy gives pK values for transition between two Cu coordination geometries (component I and II) of 7.4 (A2T), 7.9 (WT), and 8.4 (A2V), that is, component I is stabilized at physiological pH in the order A2T<WT<A2V. H NMR relaxation exhibits the same trend for the non-coordinating aromatic residues (A2T<WT<A2V), and implies markedly faster inter-peptide Cu exchange for the A2V variant than for WT and A2T. We therefore hypothesize that component I of the Cu-Aβ complex is related to pathogenicity, accounting for both the pathogenic nature of the A2V variant and the protective nature of the A2T variant.

The Aggregation Pattern of Aβ1–40 is Altered by the Presence of N‐Truncated Aβ4–40 and/or CuII in a Similar Way through Ionic Interactions

“…The experiments performed on the apopeptides yielded results very similar to those presented in the literature for Aβ 1–40 and Aβ 4–40 [33, 35, 62, 84] . The ThT fluorescence intensity spectra of the Aβ 1–40 aggregation follows a steep and asymmetrical S‐shaped curve with a growth phase that slows with time.…”

Cu and Zn coordination to amyloid peptides: From fascinating chemistry to debated pathological relevance

“…Furthermore, in the study of Somavarapu et al [53], reviewed in section 2.1, Cu(II) shows a more pronounced propensity to extend the lag phase, especially for the pathogenic A2V. A fast, low intensity ThT response has been observed at the first hours of the in vitro aggregation experiment.…”

“…The mutation of Ala2 is an interesting one: A2V is highly pathogenic [50], while A2T has shown a protective effect [51]. EPR studies show a similar Cu(II) coordination for the WT and the A2 mutants, but changes on the pK a values of the transition between component I and component II, which could be key for a pathological role of Cu-Aβ binding [52,53]. In fact, with a pK a for A2V of 8.4 (7.4 for A2T, 7.8 for WT), component I is stabilized at physiological pH.…”

“…In fact, with a pK a for A2V of 8.4 (7.4 for A2T, 7.8 for WT), component I is stabilized at physiological pH. Furthermore, this mutant shows a faster inter-peptide Cu(II) exchange linked to a higher pathogenic character of component I [53]. Similar features were observed for the Tottori mutation D7N, involved in early onset AD, although the shift of pK a value between components I and II is less accused [54,55].…”

Metal ion coordination delays amyloid-β peptide self-assembly by forming an aggregation–inert complex

“…Super-stoichiometric concentrations of Cu(II) and Zn(II) easily induce rapid formation of amorphous aggregates without amyloid structures, where the metal ion might bind to a second binding site and possibly bridge between Aβ peptides (9,31,32). In contrast, substoichiometric Cu(II) (33,34) or Zn(II) concentrations retard the overall fibrillization process, and our previous study on Zn(II) revealed the specific reduction of the fibril-end elongation (14). Besides Cu(II) and Zn(II), several other transition metal ions bind to the Aβ peptide, competing for similar ligands with slightly different coordination modes (13,(19)(20)(21).…”