Liviu M. Mirica

Bifunctional compounds for controlling metal-mediated aggregation of the aβ42 peptide.

Abnormal interactions of Cu and Zn ions with the amyloid β (Aβ) peptide are proposed to play an important role in the pathogenesis of Alzheimers disease (AD). Disruption of these metal-peptide interactions using chemical agents holds considerable promise as a therapeutic strategy to combat this incurable disease. Reported herein are two bifunctional compounds (BFCs) L1 and L2 that contain both amyloid-binding and metal-chelating molecular motifs. Both L1 and L2 exhibit high stability constants for Cu(2+) and Zn(2+) and thus are good chelators for these metal ions. In addition, L1 and L2 show strong affinity toward Aβ species. Both compounds are efficient inhibitors of the metal-mediated aggregation of the Aβ(42) peptide and promote disaggregation of amyloid fibrils, as observed by ThT fluorescence, native gel electrophoresis/Western blotting, and transmission electron microscopy (TEM). Interestingly, the formation of soluble Aβ(42) oligomers in the presence of metal ions and BFCs leads to an increased cellular toxicity. These results suggest that for the Aβ(42) peptide-in contrast to the Aβ(40) peptide-the previously employed strategy of inhibiting Aβ aggregation and promoting amyloid fibril dissagregation may not be optimal for the development of potential AD therapeutics, due to formation of neurotoxic soluble Aβ(42) oligomers.

Organometallic nickel(III) complexes relevant to cross-coupling and carbon-heteroatom bond formation reactions.

Nickel complexes have been widely employed as catalysts in C-C and C-heteroatom bond formation reactions. In addition to Ni(0) and Ni(II) intermediates, several Ni-catalyzed reactions are proposed to also involve odd-electron Ni(I) and Ni(III) oxidation states. We report herein the isolation, structural and spectroscopic characterization, and organometallic reactivity of Ni(III) complexes containing aryl and alkyl ligands. These Ni(III) species undergo transmetalation and/or reductive elimination reactions to form new C-C or C-heteroatom bonds and are also competent catalysts for Kumada and Negishi cross-coupling reactions. Overall, these results provide strong evidence for the direct involvement of organometallic Ni(III) species in cross-coupling reactions and oxidatively induced C-heteroatom bond formation reactions.

The effect of Cu2+ and Zn2+ on the Aβ42 peptide aggregation and cellular toxicity

The coordination chemistry of Cu and Zn metal ions with the amyloid β (Aβ) peptides has attracted a lot of attention in recent years due to its implications in Alzheimers disease. A number of reports indicate that Cu and Zn have profound effects on Aβ aggregation. However, the impact of these metal ions on Aβ oligomerization and fibrillization is still not well understood, especially for the more rapidly aggregating and more neurotoxic Aβ42 peptide. Here we report the effect of Cu(2+) and Zn(2+) on Aβ42 oligomerization and aggregation using a series of methods such as Thioflavin T (ThT) fluorescence, native gel and Western blotting, transmission electron microscopy (TEM), and cellular toxicity studies. Our studies suggest that both Cu(2+) and Zn(2+) ions inhibit Aβ42 fibrillization. While presence of Cu(2+) stabilizes Aβ42 oligomers, Zn(2+) leads to formation of amorphous, non-fibrillar aggregates. The effects of temperature, buffer, and metal ion concentration and stoichiometry were also studied. Interestingly, while Cu(2+) increases the Aβ42-induced cell toxicity, Zn(2+) causes a significant decrease in Aβ42 neurotoxicity. While previous reports have indicated that Cu(2+) can disrupt β-sheets and lead to non-fibrillar Aβ aggregates, the neurotoxic consequences were not investigated in detail. The data presented herein including cellular toxicity studies strongly suggest that Cu(2+) increases the neurotoxicity of Aβ42 due to stabilization of soluble Aβ42 oligomers.

Tale of a twist: magnetic and optical switching in copper(II) semiquinone complexes.

An intermediate (C) that is observed in both phenol hydroxylation and catechol oxidation with the side-on peroxide species [Cu(2)O(2)(DBED)(2)](2+) (DBED = N(1),N(2)-di-tert-butylethane-1,2-diamine) is identified as a copper(II) semiquinone species ([1](+)) through independent synthesis and characterization. The reaction of the redox-active 3,5-di-tert-butylquinone ligand with [(DBED)Cu(I)(MeCN)](+) yields a copper(II) semiquinone [1](+) complex with a singlet ground state and an intense purple chromophore (ε(580) ~ 3500 M(-1) cm(-1)). All other copper(II) semiquinone complexes characterized to date are paramagnetic and weakly colored (ε(800) ~ 500 M(-1) cm(-1)). Antiferromagnetic coupling between the Cu(II) center and the semiquinone radical in [1](+) is characterized by paramagnetic (1)H NMR and SQUID magnetometry. Comparative X-ray crystal structures along with density functional theory calculations correlate the geometric structures of copper(II) semiquinone complexes with their magnetic and optical properties. The unique observable properties of [1](+) originate from an increase in the overlap of the Cu 3d and semiquinone π orbitals resulting from a large rhombic distortion in the structure with a twist of 51°, attributable to the large isotropic demands of the tert-butyl substituents of the DBED ligand. Independent characterization of [1](+) allows the spectroscopic yields of intermediate C to be quantified in this intriguing hydroxylation reaction.

Pulsed hydrogen–deuterium exchange mass spectrometry probes conformational changes in amyloid beta (Aβ) peptide aggregation

Probing the conformational changes of amyloid beta (Aβ) peptide aggregation is challenging owing to the vast heterogeneity of the resulting soluble aggregates. To investigate the formation of these aggregates in solution, we designed an MS-based biophysical approach and applied it to the formation of soluble aggregates of the Aβ42 peptide, the proposed causative agent in Alzheimer’s disease. The approach incorporates pulsed hydrogen–deuterium exchange coupled with MS analysis. The combined approach provides evidence for a self-catalyzed aggregation with a lag phase, as observed previously by fluorescence methods. Unlike those approaches, pulsed hydrogen–deuterium exchange does not require modified Aβ42 (e.g., labeling with a fluorophore). Furthermore, the approach reveals that the center region of Aβ42 is first to aggregate, followed by the C and N termini. We also found that the lag phase in the aggregation of soluble species is affected by temperature and Cu2+ ions. This MS approach has sufficient structural resolution to allow interrogation of Aβ aggregation in physiologically relevant environments. This platform should be generally useful for investigating the aggregation of other amyloid-forming proteins and neurotoxic soluble peptide aggregates.

Galactose Oxidase as a Model for Reactivity at a Copper Superoxide Center

The mononuclear copper enzyme, galactose oxidase, has been investigated under steady-state conditions via O(2)-consumption assays using 1-O-methyl-alpha-D-galactopyranoside as the sugar substrate to produce an aldehyde at the C-6 position. The rate-determining step of the oxidative half-reaction was probed through the measurement of substrate and solvent deuterium and O-18 isotope effects on k(cat)/K(m)(O(2)). The reaction conforms to a ping-pong mechanism with the kinetic parameters for the reductive half, k(cat)/K(m)(S) = 8.3 x 10(3) M(-1) s(-1) at 10 degrees C and pH 7.0, comparing favorably to literature values. The oxidative half-reaction yielded a value of k(cat)/K(m)(O(2)) = 2.5 x 10(6) M(-1) s(-1). A substrate deuterium isotope effect of 32 was measured for the k(cat)/K(m)(S), while a smaller, but significant value of 1.6-1.9 was observed on k(cat)/K(m)(O(2)). O-18 isotope effects of 1.0185 with either protiated or deuterated sugar, together with the absence of any solvent isotope effect, lead to the conclusion that hydrogen atom transfer from reduced cofactor to a Cu(II)-superoxo intermediate is fully rate-determining for k(cat)/K(m)(O(2)). The measured O-18 isotope effects provide corroborative evidence for the reactive superoxo species in the dopamine beta-monooxygenase/peptidylglycine alpha-hydroxylating monooxygenase family, as well as providing a frame of reference for copper-superoxo reactivity. The combination of solvent and substrate deuterium isotope effects rules out solvent deuterium exchange into reduced enzyme as the origin of the relatively small substrate deuterium isotope effect on k(cat)/K(m)(O(2)). These data indicate fundamental differences in the hydrogen transfer step from the carbon of substrate vs the oxygen of reduced cofactor during the reductive and oxidative half-reactions of galactose oxidase.

Dinuclear Palladium(III) Complexes with a Single Unsupported Bridging Halide Ligand: Reversible Formation from Mononuclear Palladium(II) or Palladium(IV) Precursors

Along with the well-known involvement of Pd and Pd oxidation states in a large number of palladium-catalyzed reactions, recent reports have proposed the intermediacy of less-common Pd and Pd oxidation states in several chemical transformations. Among these systems, dinuclear and mononuclear Pd complexes have been shown to act as active intermediates in both twoand one-electron oxidative C H functionalization and C C bond formation reactions. For example, dinuclear organometallic Pd complexes stabilized by a Pd Pd bond have been recently reported and shown to be a catalytically competent alternative to mononuclear Pd species in carbon–heteroatom bond-formation reactions. In this context, we have recently shown that mononuclear Pd complexes stabilized by a tetradentate diazapyridinophane ligand can be isolated and have shown that they exhibit C C bond-formation reactivity. Continuing our work in the study of high-valent Pd complexes, we report herein novel cationic dinuclear Pd and mononuclear Pd complexes supported by a common tridentate nitrogendonor ligand, N,N’,N’’-trimethyl-1,4,7-triazacyclononane (Me3tacn). Moreover, we provide evidence for the involvement of a Pd species in the Kharasch addition reaction and confirm the ability of Pd to catalyze one-electron radical reactions. In addition, the reported Pd systems are the first group 10 d–d dinuclear complexes bridged by a single unsupported halide ligand and represent a model of the delocalized Pd X Pd electronic structure that has been proposed to exist in some Pd X Pd X one-dimensional (1D) chains. While triazacyclonane (tacn) 10] and other tacn derivatives have been employed in the synthesis of Pd complexes, only one complex, [(Me3tacn)Pd (MeCN)2](PF6)2, has been reported for Me3tacn. [10] We have synthesized the Pd complexes [(Me3tacn)Pd X2] (X = Cl 1a, X = Br 1b) through the reaction of Me3tacn with [(MeCN)2Pd X2]. [12] The X-ray structure analysis of 1a reveals a square-planar arrangement of two Cl ions and two N atoms around Pd, while the third N atom of Me3tacn is not in close proximity to the metal center (Figure 1a), in contrast to the five-coordinate geometry

Dinuclear Co(II)Co(III) mixed-valence and Co(III)Co(III) complexes with N- and O-donor ligands: characterization and water oxidation studies.

The tridentate ligand N-methyl-N,N-bis(2-pyridylmethyl)amine (L) has been employed to synthesize a dinuclear Co(II)Co(III) mixed-valence complex containing μ-methoxo and μ-carboxylato bridging ligands, [LCo(II)(μ-carboxylato)bis(μ-methoxo)Co(III)L](ClO(4))(2). In this complex, the two pseudo-octahedral Co centers have an identical ligand environment, yet the average Co-N and Co-O bond distances at the two Co ions differ significantly. Electrochemical, spectroscopic, and magnetic susceptibility measurements confirm that it belongs to a localized Class II mixed-valence system, despite the presence of a short Co···Co distance of 3.021 Å. Oxidation of this Co(II)Co(III) complex leads to formation of the corresponding Co(III)Co(III) complex that was characterized structurally and spectroscopically. In addition, dinuclear and trinuclear μ-hydroxo Co(III) complexes have been obtained in the presence of phosphate anions and absence of methanol, respectively, suggesting that an additional bridging ligand is needed to stabilize the Co(III)bis(μ-hydroxo)Co(III) fragment. Moreover, the ability of the mixed-valence Co(II)Co(III) complex and the three related Co(III) complexes to electrocatalytically oxidize water was also investigated. The observed limited water oxidation catalytic ability for these systems suggests that a multinuclear Co cluster and/or presence of O-rich ligands may be needed for the generation of efficient molecular Co-based water oxidation catalysts.

Kinetic Analysis of Iron-Dependent Histone Demethylases: α‑Ketoglutarate Substrate Inhibition and Potential Relevance to the Regulation of Histone Demethylation in Cancer Cells

The Jumonji C domain-containing histone demethylases (JmjC-HDMs) are α-ketoglutarate (αKG)-dependent, O(2)-activating, non-heme iron enzymes that play an important role in epigenetics. Reported herein is a detailed kinetic analysis of three JmjC-HDMs, including the cancer-relevant JMJD2C, that was achieved by employing three enzyme activity assays. A continuous O(2) consumption assay reveals that HDMs have low affinities for O(2), suggesting that these enzymes can act as oxygen sensors in vivo. An interesting case of αKG substrate inhibition was found, and the kinetic data suggest that αKG inhibits JMJD2C competitively with respect to O(2). JMJD2C displays an optimal activity in vitro at αKG concentrations similar to those found in cancer cells, with implications for the regulation of histone demethylation activity in cancer versus normal cells.

Aromatic Methoxylation and Hydroxylation by Organometallic High-Valent Nickel Complexes.

Herein we report the synthesis and reactivity of several organometallic Ni(III) complexes stabilized by a modified tetradentate pyridinophane ligand containing one phenyl group. A room temperature stable dicationic Ni(III)-disolvento complex was also isolated, and the presence of two available cis coordination sites in this complex offers an opportunity to probe the C-heteroatom bond formation reactivity of high-valent Ni centers. Interestingly, the Ni(III)-dihydroxide and Ni(III)-dimethoxide species can be synthesized, and they undergo aryl methoxylation and hydroxylation that is favored by addition of oxidant, which also limits the β-hydride elimination side reaction. Overall, these results provide strong evidence for the involvement of high-valent organometallic Ni species, possibly both Ni(III) and Ni(IV) species, in oxidatively induced C-heteroatom bond formation reactions.