The amino‐terminal copper and nickel/N‐terminal site (ATCUN/NTS) present in proteins and bioactive peptides exhibits high affinity towards Cu II ions and have been implicated in human copper physiology. Little is known, however, about the rate and exact mechanism of formation of such complexes. We used the stopped‐flow and microsecond freeze‐hyperquenching (MHQ) techniques supported by steady‐state spectroscopic and electrochemical data to demonstrate the formation of partially coordinated intermediate Cu II complexes formed by glycyl–glycyl–histidine (GGH) peptide, the simplest ATCUN/NTS model. One of these novel intermediates, characterized by two‐nitrogen coordination, t 1/2 ≈100 ms at pH 6.0 and the ability to maintain the Cu II /Cu I redox pair is the best candidate for the long‐sought reactive species in extracellular copper transport.

Copper complexes of metal binding domains of synthesized amyloid-β peptides -Aβ(1-16) and N-truncated Aβ(4-16) containing a novel N-terminal FRH sequence, as well as its shorter mutants were characterized by cyclic voltammetry. The influence of the peptide sequence and peptide to copper molar ratio on the electrochemical properties of the obtained structures were studied and discussed. The reversibility of the studied redox processes in copper complexes with Aβ(4-x) derivatives was also investigated. The results indicate the crucial role of Tyr10 in the redox process of the Aβ(4-x) complex, including the removal of reversibility of the Cu(II)/Cu(III) redox couple.

Copper(II) complexes of peptides containing a histidine (His) residue at the third position (known as ATCUN or NTS motif) gain interest for their biological roles and biotechnological applications. We characterized three such peptides, Ala-Ala-His-NH 2 (AAHam), Ala-βAla-His-NH 2 (ABH-am) and βAla-Ala-His-NH 2 (BAH-am) by cyclic voltammetry in order to assess the influence of substitution of the α-alanine residue (Ala) by β-alanine (βAla) on their redox properties. The obtained results, complementary to the previously reported potentiometric and spectroscopic studies confirmed strong effects of size and position of component chelate rings, 5-membered for Ala and 6-membered for βAla, on the Cu(II) binding affinity and redox properties. The detailed voltammetric study revealed the effects of the composition of the supporting electrolyte and the selection of starting potential on the observed redox processes. The conclusions are relevant for designing artificial nucleases and proteases and for proper practice of studies of redox properties of complexes of Cu(II) carried out in buffer solutions.

Gly-His-Lys (GHK) is a tripeptide present in the human bloodstream that exhibits a number of biological functions. Its activity is attributed to the copper-complexed form, Cu(II)GHK. Little is known, however, about the molecular aspects of the mechanism of its action. Here, we examined the reaction of Cu(II)GHK with reduced glutathione (GSH), which is the strongest reductant naturally occurring in human plasma. Spectroscopic techniques (UV–vis, CD, EPR, and NMR) and cyclic voltammetry helped unravel the reaction mechanism. The impact of temperature, GSH concentration, oxygen access, and the presence of ternary ligands on the reaction were explored. The transient GSH-Cu(II)GHK complex was found to be an important reaction intermediate. The kinetic and redox properties of this complex, including tuning of the reduction rate by ternary ligands, suggest that it may provide a missing link in copper trafficking as a precursor of Cu(I) ions, for example, for their acquisition by the CTR1 cellular copper transporter.

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