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. 2021 May 14;22(10):5190.
doi: 10.3390/ijms22105190.

Oxidase Reactivity of CuII Bound to N-Truncated Aβ Peptides Promoted by Dopamine

Chiara Bacchella  1 , Simone Dell'Acqua  1 , Stefania Nicolis  1 , Enrico Monzani  1 , Luigi Casella  1
Affiliations

Oxidase Reactivity of CuII Bound to N-Truncated Aβ Peptides Promoted by Dopamine

Chiara Bacchella et al. Int J Mol Sci. .

Abstract

The redox chemistry of copper(II) is strongly modulated by the coordination to amyloid-β peptides and by the stability of the resulting complexes. Amino-terminal copper and nickel binding motifs (ATCUN) identified in truncated Aβ sequences starting with Phe4 show very high affinity for copper(II) ions. Herein, we study the oxidase activity of [Cu-Aβ4-x] and [Cu-Aβ1-x] complexes toward dopamine and other catechols. The results show that the CuII-ATCUN site is not redox-inert; the reduction of the metal is induced by coordination of catechol to the metal and occurs through an inner sphere reaction. The generation of a ternary [CuII-Aβ-catechol] species determines the efficiency of the oxidation, although the reaction rate is ruled by reoxidation of the CuI complex. In addition to the N-terminal coordination site, the two vicinal histidines, His13 and His14, provide a second Cu-binding motif. Catechol oxidation studies together with structural insight from the mixed dinuclear complexes Ni/Cu-Aβ4-x reveal that the His-tandem is able to bind CuII ions independently of the ATCUN site, but the N-terminal metal complexation reduces the conformational mobility of the peptide chain, preventing the binding and oxidative reactivity toward catechol of CuII bound to the secondary site.

Keywords: Alzheimer’s disease; amyloid-β peptides; copper; dopamine; neurodegeneration; oxidative stress.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Kinetic profiles of DA (3 mM, panel (a)) and MC (3 mM, panel (b)) oxidation with time in 50 mM HEPES buffer at pH 7.4 and 20 °C in the presence of CuII alone (25 µM, green trace) and with 1 equiv. Aβ1–16 (25 µM, pink), 2 equiv. Aβ1–16 (50 µM, grey), 1 equiv. Aβ4–16 (25 µM, light blue), and 2 equiv. Aβ4–16 (50 µM, orange).
Figure 2
Figure 2
Kinetic profiles of DA (0.3 mM, panel (a)) and MC (0.3 mM, panel (b)) oxidation with time in 50 mM HEPES buffer at pH 7.4 and 20 °C in the presence of CuII alone (25 µM, green trace) and with 1 equiv. Aβ1–16 (25 µM, pink), 2 equiv. Aβ1–16 (50 µM, grey), 1 equiv. Aβ4–16 (25 µM, light blue), and 2 equiv. Aβ4–16 (50 µM, orange).
Figure 3
Figure 3
Kinetic profiles of DA (3 mM, panel (a)) and MC (3 mM, panel (b)) oxidation with time in 50 mM HEPES buffer at pH 7.4 and 20 °C in the presence of CuII alone (25 µM, green trace) and with 1 equiv. Aβ1–28 (25 µM, pink), 2 equiv. Aβ1–28 (50 µM, grey), 1 equiv. Aβ4–28 (25 µM, light blue), and 2 equiv. Aβ4–28 (50 µM, orange).
Figure 4
Figure 4
Kinetic profiles of DA (0.3 mM, panel (a)) and MC (0.3 mM, panel (b)) oxidation with time in 50 mM HEPES buffer at pH 7.4 and 20 °C in the presence of CuII alone (25 µM, green trace) and with 1 equiv. Aβ1–28 (25 µM, pink), 2 equiv. Aβ1–28 (50 µM, grey), 1 equiv. Aβ4–28 (25 µM, light blue), and 2 equiv. Aβ4–28 (50 µM, orange).
Figure 5
Figure 5
Kinetic profiles of MC (3 mM) oxidation with time in 50 mM HEPES buffer at pH 7.4 and 20 °C in the presence of CuII alone (25 μM, green trace). The same experiments were also performed upon the addition of [Cu–Aβ4–16] at 1:1 molar ratio (25 µM, light blue) and [Ni–Cu–Aβ4–16] at 1:1:1 molar ratio (25 µM, red).
Figure 6
Figure 6
Kinetic profiles of MC (0.3 mM) oxidation with time in 50 mM HEPES buffer at pH 7.4 and 20 °C in the presence of CuII alone (25 μM, green trace). The same experiments were also performed upon the addition of Aβ4–16 complexes: [Cu–Aβ4–16] at 1:1 molar ratio (25 µM, light blue), [Ni–Cu–Aβ4–16] at 1:1:1 molar ratio (25 µM, red) and at 2:1:1 molar ratio (25 µM, violet).
Figure 7
Figure 7
Titration of nickel(II)–Aβ4–16 complex (blue spectrum, at 1:1 ratio, 0.5 mM) with 0.0–0.55 mM copper(II) (final point as violet spectrum) in 5 mM phosphate buffer solution at pH 7.4. The pink spectrum corresponds to the absorption of free nickel(II).
Figure 8
Figure 8
Far-UV CD spectra of 1.1 equiv. Aβ4–16 peptide (10 µM, panel a) and 1.1 equiv. Aβ4–28 peptide (5.5 µM, panel b) in 5 mM phosphate buffer solution at pH 7.4 (blue traces) and upon the addition of copper(II) (1 equiv., light blue) and DA (1 equiv., pink).
Figure 9
Figure 9
Far-UV CD spectra of Aβ4–16 peptide (10 µM) in 5 mM phosphate buffer solution at pH 7.4 (blue trace) and upon the addition of NiII (9.5 µM), after few seconds of incubation (light blue) and 15 min (green). Then, 1 (pink) and 2 equiv. of copper(II) (grey) were added to the Ni–Aβ4–16 complex.
Figure 10
Figure 10
(a) Vis-CD spectra of of copper(II) (2 mM) alone in 5 mM phosphate buffer solution at pH 7.4 (grey trace) and upon the addition of 1 equiv. Aβ4–16 peptide (2 mM, pink) and further addition of 1 equiv. of NiII to the initial complex (blue). (b) Vis-CD spectra of nickel(II) (2 mM) alone (grey trace) and upon the addition of 1 equiv. Aβ4–16 peptide (2 mM, pink) and further addition of 1 equiv. of CuII (blue).
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