The angiotensin metabolite His-Leu is a strong copper chelator forming highly redox active species

His-Leu is a hydrolytic byproduct of angiotensin metabolism, whose concentration in the bloodstream could be at least micromolar. This encouraged us to investigate its Cu(II) binding properties and the concomitant redox reactivity. The Cu(II) binding constants were derived from isothermal titration calorimetry and potentiometry, while identities and structures of complexes were obtained from ultraviolet–visible, circular dichroism, and room-temperature electronic paramagnetic resonance spectroscopies. Four types of Cu(II)/His-Leu complexes were detected. The histamine-like complexes prevail at low pH. At neutral and mildly alkaline pH and low Cu(II):His-Leu ratios, they are superseded by diglycine-like complexes involving the deprotonated peptide nitrogen. At His-Leu:Cu(II) ratios of ≥2, bis-complexes are formed instead. Above pH 10.5, a diglycine-like complex containing the equatorially coordinated hydroxyl group predominates at all ratios tested. Cu(II)/His-Leu complexes are also strongly redox active, as demonstrated by voltammetric studies and the ascorbate oxidation assay. Finally, numeric competition simulations with human serum albumin, glycyl-histydyl-lysine, and histidine revealed that His-Leu might be a part of the low-molecular weight Cu(II) pool in blood if its abundance is >10 μM. These results yield further questions, such as the biological relevance of ternary complexes containing His-Leu

Aβ5−xPeptides: N‑Terminal Truncation Yields Tunable Cu(II)Complexes

The Aβ5−x peptides (x = 38, 40, 42) are minor Aβ species in normal brains but elevated upon the application of inhibitors of Aβ processing enzymes. They are interesting from the point of view of coordination chemistry for the presence of an Arg-His metal binding sequence at their N-terminus capable of forming a 3-nitrogen (3N) three-coordinate chelate system. Similar sequences in other bioactive peptides were shown to bind Cu(II) ions in biological systems. Therefore, we investigated Cu(II) complex formation and reactivity of a series of truncated Aβ5−x peptide models comprising the metal binding site: Aβ5−9, Aβ5−12, Aβ5−12Y10F, and Aβ5−16. Using CD and UV−vis spectroscopies and potentiometry, we found that all peptides coordinated the Cu(II) ion with substantial affinities higher than 3 × 1012 M−1 at pH 7.4 for Aβ5−9 and Aβ5−12. This affinity was elevated 3-fold in Aβ5−16 by the formation of the internal macrochelate with the fourth coordination site occupied by the imidazole nitrogen of the His13 or His14 residue. A much higher boost of affinity could be achieved in Aβ5−9 and Aβ5−12 by adding appropriate amounts of the external imidazole ligand. The 3N Cu-Aβ5−x complexes could be irreversibly reduced to Cu(I) at about −0.6 V vs Ag/AgCl and oxidized to Cu(III) at about 1.2 V vs Ag/AgCl. The internal or external imidazole coordination to the 3N core resulted in a slight destabilization of the Cu(I) state and stabilization of the Cu(III) state. Taken together these results indicate that Aβ5−x peptides, which bind Cu(II) ions much more strongly than Aβ1−x peptides and only slightly weaker than Aβ4−x peptides could interfere with Cu(II) handling by these peptides, adding to copper dyshomeostasis in Alzheimer brains

Gly-Gly-His immobilized on monolayer modified back-side contact miniaturized sensors for complexation of copper ions

Miniaturized planar back-side contact transducers (BSC) with chemically modified gold surface have been utilized as electrochemical sensors. The electrodes have been functionalized by sequential immobilization of aryl diazonium salts or alkanethiols and short peptide Gly-Gly-His. The applicability of gold substrates modified with aryl diazonium salts in voltammetric detection of copper(II) ions in aqueous solutions has been studied. The combination of two fundamental elements of the solid-state electrode, i.e. back-side contact (BSC) gold sensor and self-assembled monolayers, allowed one to obtain reliable miniaturized copper(II) ion sensors. It can have important future applications in environmental sensing or in implantable biodevices.11 page(s

Key Intermediate Species Reveal the Copper(II)‐Exchange Pathway in Biorelevant ATCUN/NTS Complexes

The amino-terminal copper and nickel/N-terminal site (ATCUN/NTS) present in proteins and bioactive peptides exhibits high affinity towards CuII 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 CuII complexes formed by glycyl-glycyl-histidine (GGH) peptide, the simplest ATCUN/NTS model. One of these novel intermediates, characterized by two-nitrogen coordination, t1/2 ≈100 ms at pH 6.0 and the ability to maintain the CuII /CuI redox pair is the best candidate for the long-sought reactive species in extracellular copper transport

Copper Exchange and Redox Activity of a Prototypical 8‑Hydroxyquinoline: Implications for Therapeutic Chelation

The N-truncated β-amyloid (Aβ) isoform Aβ_4–<i>x</i> is known to bind Cu²⁺ via a redox-silent ATCUN motif with a conditional <i>K</i>_d = 30 fM at pH 7.4. This study characterizes the Cu²⁺ interactions and redox activity of Aβ_{<i>x</i>–16} (<i>x</i> = 1, 4) and 2-[(dimethylamino)-methyl-8-hydroxyquinoline, a terdentate 8-hydroxyquinoline (8HQ) with a conditional <i>K</i>_d(CuL) = 35 pM at pH 7.4. Metal transfer between Cu­(Aβ_1–16), CuL, CuL₂, and ternary CuL­(N_Im^Aβ) was rapid, while the corresponding equilibrium between L and Aβ_4–16 occurred slowly via a metastable CuL­(N_Im^Aβ) intermediate. Both CuL and CuL₂ were redox-silent in the presence of ascorbate, but a CuL­(N_Im) complex can generate reactive oxygen species. Because the N_Im^Aβ ligand will be readily exchangeable with N_Im ligands of ubiquitous protein His side chains in vivo, this class of 8HQ ligand could transfer Cu²⁺ from inert Cu­(Aβ_4–<i>x</i>) to redox-active CuL­(N_Im). These findings have implications for the use of terdentate 8HQs as therapeutic chelators to treat neurodegenerative disease