Electron galactose oxidase

Copper(II) complexes with phenoxo ligands have attracted great interest, in order to develop basic coordination chemistry for their possible use as models for tyrosinase activity (dimeric complexes) and fungal enzyme galactose oxidase (GO) (monomeric complexes). The latter enzyme catalyzes the two-electron oxidation of primary alcohols with dioxygen to yield aldehyde and... 

Galactose oxidase, 39 28-30 Gallaborane, 41 211-216 electron diffraction, 41 214 infrared spectra, 41 212-213 synthesis, 41 215... 

So-called blue multinuclear copper oxidase enzymes, such as laccase and ascorbate oxidase, catalyze the stepwise oxidation of organic substrates (most likely in successive one-electron steps) in tandem with the four-electron reduction of O2 to water, i.e. no oxygen atom(s) from O2 are incorporated into the substrate (Eq. 4) [15]. Catechol oxidase, containing a type 3 center, mediates a two-electron substrate oxidation (o-diphenols to o-chinones), and turnover of two substrate molecules is coupled to the reduction of O2 to water [34,35]. The non-blue copper oxidases, e.g. galactose oxidase and amine oxidases [27,56-59], perform similar oxidation catalysis at a mononuclear type 2 Cu site, but H2O2 is produced from O2 instead of H2O, in a two-electron reduction. 

In contrast to the active site of galactose oxidase, to pre-catalyst 13, and to the system reported by Stack et al., the proposed catalytic species 15 does not imdergo reduction to Cu intermediates, as the oxidation equivalents needed for the catalysis are provided for solely by the phenoxyl radical Hgands. Since the conversion of alcohols into aldehydes is a two-electron oxidation process, only a dinuclear Cu species with two phenoxyl ligands is thought to be active. Furthermore, concentrated H2O2 is formed as byproduct in the reaction instead of H2O, as in the system described by Marko et al. [159]. 

Nonblue. These include galactose oxidase (GO) and amine oxidases (e.g., plasma amine oxidase, diamine oxidase, lysyl oxidase), which produce dihydrogen peroxide by the two-electron reduction of 02 [33], For GO (stereospecific primary alcohol oxidation), spectroscopic studies by Whittaker [70,71] suggest that the two-electron oxidation carried out by a mononuclear copper center is aided by a stabilized ligand-protein radical (i.e., (L)Cu(I) + 02 —> (L +)Cu(lI) + H202), obviating the need to get to Cu(III) in the catalytic cycle. Protein x-ray structures [33,72] reveal a novel copper protein cofactor, which would seem... 

Galactose oxidase is an extracellular enzyme secreted by the fungus Dactylium den-droides. It is monomeric (M = 68000), contains a single copper site and catalyses the oxidation of a wide range of primary alcohols to the corresponding aldehydes. The two-electron transfer reaction RCH2OH - RCHO + 2H+ + 2e does not utilise a Cu(III)/Cu(I) couple, but a second redox site, involving a tyrosine radical which mediates the transfer of the second electron. 

The first spectral study of galactose oxidase was the report of the electron spin resonance spectrum by Blumberg et al. (19). More recently, Cleveland et al. (20) reported a further ESR study which was based on a computer fit to the spectrum. They concluded that four nitrogens were bound to the Cu(II) atom. 

Given some notion of the nature of the endogenous ligands, we can next ask how exogenous ligands perturb the copper atom and what this can tell us about the electronic transitions exhibited by the Cu(II) atom in galactose oxidase. 

Table II. Electronic Transitions Exhibited by Galactose Oxidase"...

Figure 5. Electron spin resonance spectrum of galactose oxidase (A) and galactose oxidase and Fe(CN)63 at a 1 1 (B) and 1 6 (C) molar ratio spectra were recorded at 100°K, with a power of 20 mw (9.115 GHz) and a modulation amplitude of 2G [galactose oxidase] = 0.5 raM.

Transition metal ions with organic radicals exist in the active sites of metalloproteins. The best understood example is galactose oxidase, which features a single Cu(II) ion coordinated to a modified tyrosyl radical. Many combined experimental and theoretical studies have focused on electronic properties of metal complexes with redox active ligands, yet reactivity beyond characterization has been limited. We will demonstrate the influence of the metal complex redox state on H2 activation by anilino-phenolate noninnocent ligands. 

Galactose oxidase exhibits a mononuclear copper-active site, which is flanked by a tyrosinyl radical. In a single step, a two-electron oxidation of alcohols can be performed by this enzyme, where one electron is extracted by copper and the other by the tyrosine residue [19]. 

Fig. 10. Interconversion of redox states for galactose oxidase. Three distinct states (AGO, lAGO, and RGO) may be prepared and interconverted by either one-electron or two-electron redox steps.

Fig. 21. Proposed catalytic mechanism for substrate oxidation by galactose oxidase. (A) Substrate binding displaces Tyr-495 phenolate which serves as a general base for abstracting the hydroxylic proton. (B) Stererospecihc pro- hydrogen abstraction by the Tyr-Cys phenoxyl radical. (C) Inner sphere electron transfer reducing Cu(II) to Cu(I). (D) Dissociation of the aldehyde product.

Babcock, G. T., El-Deeb, M. K., Sandusky, P. O., Whittaker, M. M., and Whittaker, J. W., 1992, Electron paramagnetic resonance and electron nuclear double resonance spectroscopies of the radical site in galactose oxidase and of thioether-substituted phenol model compounds, J. Am. Chem. Soc. 114 372793734. 

Itoh, S., Takayama, S., Arakawa, R., Furuta, A., Komatsu, M., Ishida, A., Takamuku, S., and Fukuzumi, S., 1997, Active Site Models for Galactose Oxidase. Electronic Effects of the Thioether Group in the Novel Organic Cofactor, Inorg. Chem. 36 1407nl416. 

Most copper-containing oxygenases and oxidases use multiple metal centers to conduct their biotransformations. Copper amine oxidases (CAO) and galactose oxidase (GalO), instead, employ posttranslationahy derived amino acid side chains as cofactors to supply additional electrons (24). 

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