Enzyme multicopper oxidase

S / V CONTENTS Preface, Robert W. Hay. Structure and Function of Manganese-Containing Biomolecules, David C. Weather-bum. Repertories of Metal Ions as Lewis Acid Catalysts in Organic Reactions, Junghan Suh. The Multicopper-Enzyme Ascorbate Oxidase, Albrecht Messerschmidt. The Bioinorganic Chemistry of Aluminum, Tomas Kiss and Etelka Farkas. The Role of Nitric Oxide in Animal Physiology, Anthony R. Butler, Frederick Flitney and Peter Rhodes. Index. 

Other Copper Oxidases. A number of additional multicopper oxidases have been detected [22], including phenoxazinone synthase (Table 1). This enzyme catalyzes the overall six-electron oxidative coupling of 2-aminophenols to form 1-aminophenoxazinone, the final step in the bacterial (Streptomyces) biosynthesis of the antineoplastic agent actinomycin. 

The multicopper oxidases (laccase, ascorbate oxidase, and ceruloplasmin) catalyze a four-electron reduction of dioxygen to water (285-287). Consistent with the four-electron stoichiometry, the enzymes contain four copper ions. One of the copper ions is type I, causing an intensely blue color of the proteins, thus the enzymes of this family are referred to as blue oxidases. They also contain a monomeric copper site that exhibits normal spectroscopic features, whereas the other two copper... 

What distinguishes multicopper oxidases from other copper proteins is that they contain one each of these three types of copper site (Solomon and Lowery, 1993 Solomon et al., 1996). Not only does this make them excellent models for all copper proteins, but because they have four redox-active metal ions, they also serve as paradigms for other enzymes that couple a one-electron reductant to a four-electron oxidant, most notably cytochrome c oxidase. Indeed, the three copper sites (and four copper atoms) in the multicopper oxidases play essentially equivalent roles in comparison to the two heme groups and two copper atoms in cytochrome c oxidase. 

Based on present sequence data, known or likely ferroxidase enzymes can be identihed in several eukaryotes. These enzymes are listed in Table 11. All are multicopper oxidases, by sequence homology at least. In mammals, they include ceruloplasmin and, most likely, hephaestin (Hp), although only mouse Hp (mHp) has been characterized at this time (Vulpe et al., 1999). The alignments in Fig. 5A show that mHp is essentially... 

In vitro experiments with Cp and Fet3p have shown dehnitively that both enzymes have a strong substrate specihcity toward Fe(II) that is not shared by the other well-characterized multicopper oxidases. Furthermore, several independent analyses have demonstrated that the formation of Tf Fe(III) in a mixture of apo iand Fe(II) is strongly catalyzed by Cp. At the other end of the experimental spectrum are the well-established correlations between organismal abnormalities in iron handling in... 

In the area of copper metabolism, four topics are covered bacterial copper transport reviewed by Huat Lu and Sohoz copper P-type ATPases reviewed by Voskoboinik, Camakaris, and Mercer copper chaperones reviewed by Stine Elam et al. and copper metaUoregulation of gene expression reviewed by Winge. An important related topic is the link between copper and iron metabolism. In this area, Kosman has reviewed the multicopper oxidase enzymes, such as FetSp and ceruloplasmin, which catalyze the conversion of iron(II) to iron(III) in preparation for its specific transport by partner transporter proteins. 

The three copper sites which represent a total of four one-electron acceptors place the multicopper oxidases in the small family of enzymes that can couple the oxidation of four equivalents of a one-electron reducing... 

The type I copper sites function as electron transfer centers in the blue copper proteins and in multicopper enzymes, particularly oxidases (33). They are characterized by their intense blue color, their unusually small A values, and their very positive redox potentials (Table II). X-ray crystal structures of several blue copper proteins have been determined, notably plastocyanin (34), azurin (35), cucumber basic blue protein (36), and pseudoazurin (37). The active site structures show marked similarities but also distinct differences (Fig. 8). 

Figure 5 Illustration of possible partial reaction cycles of some copper- and flavin-dependent oxidase enzymes, (a) Copper amine oxidase 30, 31 (b) galactose oxidase (32) (c) catechol oxidase (10) (d) multicopper oxidases (10) (e) flavin oxidases (30) (f) cytochrome c oxidase (38).

The high-affinity pathway involves oxidation of Fe to Fe by the ferroxidase FET3 and subsequent transport of Fe " " across the plasma membrane by the permease FTRl. FET3p is a member of the family of multicopper oxidases, which include ascorbate oxidase, laccase, and ceruloplasmin (see Chapter 14), and does not become functional until it is loaded with copper intracellularly through the activities of the copper chaperone ATX Ip and the copper transporter CCC2p. It appears that Fe " " produced by FET3 is transferred directly to FTRl, and does not equilibrate with the bulk phase, as is illustrated in Fig. 7.13. This is almost certainly achieved by the classic metabolite-channeling mechanism, a common feature of multifunctional enzymes. 

Intramolecular ET between distinct copper centers is part of the catalytic cycles of many copper-containing redox enzymes, such as the multicopper oxidases, ascorbate oxidase, and ceruloplasmin, as well as the copper-containing nitrite reductases. Examination of internal LRET in these proteins is of considerable interest as it may also provide insights into the evolution of selected ET pathways in particular, whether and how the enzymes have evolved in order to optimize catalytic functions. With the increase in the number of known high-resolution 3D structures of transition metal containing redox enzymes, studies of structure-reactivity relationships have become feasible and indeed many have been carried out during the last two decades. 

Laccase, ascorbate oxidase, and ceruloplasmin are the classical members of the multicopper oxidase family also known as blue oxidases. Recently, a small number of bacterial members of this family have been characterized, including CueO from E. coli a spore-coat laccase (CotA) from Bacillus suhtilis and phenoxazinone synthase from Streptomyces antibioticus The catalyzed reaction of these enzymes except for phenoxazinone synthase is given in Equation (11). A comprehensive overview of the broad and active research on blue copper oxidases is presented in Messerschmidt. Recent results have been included in a review on the reduction of dioxygen by copper-containing enzymes. The nature and number of the different copper sites in blue oxidases has been described in the sections about the type-1 copper site and the trinuclear copper cluster. 

The accumulation of iron is dependent on its transport into the cell. Askwith and Kaplan (Chapter 4) discuss iron transport mechanisms in eukaryotic cells, developing models based on studies carried out in the yeast, Saccharomyces cerevisiae. These cells possess both siderophore-dependent and elemental iron transport systems. The latter system relies on cell surface ferrireductases to convert extracellular ferric chelates to ferrous iron, which can be transported through either a high or low affinity iron transport system. Studies on a high affinity ferrous iron transporter (FET3) revealed that the multicopper oxidase will oxidize ferrous to ferric iron, which is then mobilized across the membrane by a ferric transmembrane permease (Ftrlp). This is a highly specific transport system in yeast it only transports iron. In humans, the copper enzyme, ceruloplasmin, is responsible for the radical-free oxidase activity. This plasma protein oxidizes the ferrous iron that is excreted from cells into the transferrin-usable ferric form. 

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