A mixed-metal derivative of laccase containing mercury(II) in the type 1 binding site

Morie-Bebel, M. Michelle; Morris, Margaret C.; Menzie, Janelle L.; McMillin, David R.

doi:10.1021/ja00324a048

Cited by 67 publications

(65 citation statements)

References 4 publications

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“…The explanations offered in the following are speculative, because only bulk T cells and rel atively crude preparations of proteins were used in the pre sent study, so that the precise antigenic determinants in duced by Hg2 could not be identified. Hg2 ions preferen tially bind to protein thiol (-SH) groups [36] and, albeit with lower affinity, amino (-NH2) groups [37,38], Protein-Hg2 complexes consist of two, three, or four ligands, including -N2 -S2, -Sj, -N2S2, and Sj coordination sites [37][38][39][40][41], Hence, Hg2 ions can interact with a great variety o f differ ent proteins and, by undergoing multipoint high-avidity binding with up to four different ligands, some Hg2 ions may generate very stable intramolecular coordination com plexes. Taking into account the large number of different Hg protein complexes possible and the obvious stability of some o f them, three different possibilities of how Hg may render self proteins immunogenic may be envisaged.…”

Section: Discussionmentioning

confidence: 99%

Murine Systemic Autoimmune Disease Induced by Mercuric Chloride: T Helper Cells Reacting to Self Proteins

Kubicka-Muranyi

Kremer²,

Rottmann

et al. 1996

Int Arch Allergy Immunol

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HgCl₂ induces a CD4+ T-cell-dependent systemic autoimmune disease in susceptible strains of rats and mice. In rats, autoreactive T cells were shown to be involved, whereas in mice, attention has focussed on the demonstration of ‘Hg-specific’ T cells. To clarify these seemingly different T cell involvements, T cells from B10.S mice treated with HgCl₂ for 1 or 8 weeks were analyzed for their capacity to mount anamnestic responses against various self antigens (Ags) which either contained Hg or did not. T cells from donors short-term treated with HgCl₂ failed to mount memory responses to Hg-free Ags, but mounted a significant response to HgCl₂ and also reacted with Hg-containing self Ags. Interestingly, T cells from donors long-term treated with HgCl₂ showed a different pattern of reactivity. They hardly reacted to HgCl₂ and reacted poorly to Hg-containing splenic proteins, but responded vigorously to nuclei and fibrillarin irrespective of whether these self constituents had been treated with HgCl₂ or not. Conceivably, the initial activation of T cells that recognize Hg in combination with nuclear self proteins, such as fibrillarin, eventually results in activation of T cells specific for the unaltered self proteins.

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Section: Discussionmentioning

confidence: 99%

Murine Systemic Autoimmune Disease Induced by Mercuric Chloride: T Helper Cells Reacting to Self Proteins

Kubicka-Muranyi

Kremer²,

Rottmann

et al. 1996

Int Arch Allergy Immunol

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“…However, in these altered enzymes, the T2 and T3 sites can still be reduced with smaller, inner-sphere reductants, and the reduced TNC, even in the absence of the T1, will react with and reduce O 2 . Elimination of T1 reactivity was accomplished either through site-selective mutation which inhibits Cu loading at the T1, resulting in a T1 depleted enzyme (T1D) [81], or through a derivative of tree laccase in which the T1 Cu has been replaced by the redox innocent Hg 2+ ion (T1Hg) [82, 83]. These derivatives have been crucial to the elucidation of the mechanism of O 2 reduction at the TNC, as described below.…”

Section: O2 Reduction Mechanismmentioning

confidence: 99%

Electron transfer and reaction mechanism of laccases

Jones

Solomon

2015

Cell. Mol. Life Sci.

429

358

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Laccases are part of the family of multicopper oxidases (MCOs), which couple the oxidation of substrates to the four electron reduction of O2 to H2O. MCOs contain a minimum of four Cu's divided into Type 1 (T1), Type 2 (T2), and binuclear Type 3 (T3) Cu sites that are distinguished based on unique spectroscopic features. Substrate oxidation occurs near the T1, and electrons are transferred approximately 13 Å through the protein via the Cys-His pathway to the T2/T3 trinuclear copper cluster (TNC) where dioxygen reduction occurs. This review outlines the electron transfer (ET) process in laccases, and the mechanism of O2 reduction as elucidated through spectroscopic, kinetic, and computational data. Marcus theory is used to describe the relevant factors which impact ET rates including the driving force (ΔG°), reorganization energy (λ), and electronic coupling matrix element (HDA). Then the mechanism of O2 reaction is detailed with particular focus on the intermediates formed during the two 2e− reduction steps. The first 2e− step forms the peroxide intermediate (PI), followed by the second 2e− step to form the native intermediate (NI), which has been shown to be the catalytically relevant fully oxidized form of the enzyme.

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“…27,28 (Scheme 1, top) Derivatives of the native enzyme have been prepared where the T1 is either eliminated 29,30 (replacement of the Cys ligand of the T1 with a Ser to generate a type 1 depleted or T1D form) or replaced by a redox innocent mercuric ion (the T1Hg derivative). 31,32 These have valid TNCs which, when reduced, react with O 2 with essentially the same rate constant as the native enzyme. (k Nat = 1.7 × 10 6 M −1 s −1 , k T1Hg = 2.2 × 10 6 M −1 s −1 ) 33 This reaction generates a species with at least one less electron transferred to O 2 which we have, in fact, determined to be a peroxide intermediate (PI).…”

Section: O 2 Reactivity: the Trinuclear Cu Cluster (Tnc)mentioning

confidence: 99%

O2 Reduction to H2O by the multicopper oxidases

2008

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In nature the four electron reduction of O 2 to H 2 O is carried out by Cytochrome c Oxidase (CcO) and the multicopper oxidases (MCOs). In the former, Cytochrome c provides electrons for pumping protons to produce a gradient for ATP synthesis, while in the MCOs the function is the oxidation of substrates, either organic or metal ions. In the MCOs the reduction of O 2 is carried out at a trinuclear Cu cluster (TNC). Oxygen intermediates have been trapped which exhibit unique spectroscopic features that reflect novel geometric and electronic structures. These intermediates have both intact and cleaved O-O bonds, allowing the reductive cleavage of the O-O bond to be studied in detail both experimentally and computationally. These studies show that the topology of the TNC provides a unique geometric and electronic structure particularly suited to carry out this key reaction in Nature.The multicopper oxidases (MCOs) couple four 1-electron oxidations of substrate to the four electron reductive cleavage of the O-O bond of dioxygen using a minimum of four Cu atoms (table 1). 1,2 Among these four Cu's is a type 1 (T1) or blue Cu site, characterized by an intense S Cys → Cu(II) charge transfer (CT) transition at around 600 nm in the absorption spectrum and a uniquely small A || in its electron paramagnetic resonance (EPR) spectrum. This is the site of substrate oxidation, and from table 1, the MCOs can be divided into two classes depending upon the identity of the substrate. For enzymes such as laccase 3 and ascorbate oxidase, 4 redox active organic molecules which can interact weakly with the enzyme provide the electrons. For MCOs like Fet3p 5 and Ceruloplasmin, 6 the substrate is a metal ion (ferrous in these cases) which binds tightly to a substrate binding site. As shown in figure 1, these substrate binding sites are located near the His ligands of the T1 Cu center. The electron from substrate is first transferred to the T1 and then over >13Å through a Cys-His pathway to a trinuclear Cu cluster (TNC) where O 2 is reduced to water (vide infra). 7 We first consider the electron transfer (ET) pathways to the TNC. ET PathwaysHere we focus on the Fe(II) binding site of the enzyme Fet3p, which is involved in the uptake of iron by yeast. 8 (Studies on this enzyme were performed in collaboration with Prof. Dan Kosman and coworkers.) A variable-temperature, variable-field magnetic circular dichroism (VTVH MCD) methodology we developed in other studies was applied to probe this ferrous site. [9][10][11] From figure 2A dark blue, there is a characteristic feature at 8900 cm −1 in the MCD spectrum corresponding to Fe(II) binding with a high affinity (K B > 10 5 M −1 , from MCD titration studies) to a 6 coordinate site in the protein. 12 In the light blue spectrum this feature is eliminated and a peak at 9700 cm −1 , corresponding to aqueous Fe(II) (green) is observed when Zn(II) is first bound to the substrate site, inhibiting ferroxidase activity.From mutagenesis studies we have found that three carboxylates are involved...

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A mixed-metal derivative of laccase containing mercury(II) in the type 1 binding site

Cited by 67 publications

References 4 publications

Murine Systemic Autoimmune Disease Induced by Mercuric Chloride: T Helper Cells Reacting to Self Proteins

Murine Systemic Autoimmune Disease Induced by Mercuric Chloride: T Helper Cells Reacting to Self Proteins

Electron transfer and reaction mechanism of laccases

O2 Reduction to H2O by the multicopper oxidases

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