Introduction

Copper is an essential trace element in living systems, where it serves as a cofactor in many enzymatic redox reactions and oxygen transport (Fig. 13.1). The physiological oxidation states of copper are Cu1+ and Cu2+, whereas Cu3+ is not a biologically relevant species because of the high redox potential of the Cu3+/Cu2+ couple (Shleev et al. 2005). The copper at the active sites of redox proteins has been divided into three main classes (Table 13.1): type 1 (T1), blue copper; type 2 (T2), normal copper, and; type 3 (T3), a binuclear copper center (Malkin and Malmström 1970; Reinhammar 1984; Solomon et al. 1996, 2004; Kaim and Rall 1996).

T1 copper confers a typical blue color on the protein, which results from an intense electronic absorption band (around 600 nm) due to the covalent copper-cysteine bond. These sites are found in mononuclear copper proteins involved in intermolecular electron transfer pathways (azurin, plastocyanin, amicyanin, stella-cyanin, rusticyanin), multicopper proteins (ascorbate oxidase, bilirubin oxidase, laccase, ceruloplasmin), and in a subclass of nitrite reductases, where they function in intramolecular electron transfer.

T2 copper in proteins yields positive EPR signals and only weak absorption in the visible spectrum. Type 2 sites are present in all blue multicopper oxidases, as well as in galactose oxidase, prokaryotic and eukaryotic copper amine oxidases, copper-containing superoxide dismutase, and cytochrome c oxidase.

The T3 binuclear copper center contains two ligand-bridged spin-coupled copper ions (CuA and CuB). T3 sites are diamagnetic and display a distinctive absorption band near 330 nm as well as a characteristic luminescence spectrum (Wynn et al. 1983; Solomon et al. 1996; Shin and Lee 2000; Shleev et al. 2005). This site is present in tyrosinase and in hemocyanin, the oxygen carrier found in molluscs and arthropods. In blue multicopper oxidases, the T2 and T3 sites form a trinuclear

H. Claus

Institut für Mikrobiologie und Weinforschung, Johannes Gutenberg-Universität Mainz, Becherweg 15, 55099, Mainz, Germany e-mail: [email protected]

I. Sherameti and A. Varma (eds.), Soil Heavy Metals, Soil Biology, Vol 19, DOI 10.1007/978-3-642-02436-8_13, © Springer-Verlag Berlin Heidelberg 2010

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Fig. 13.1 Copper enzymes and their reactions (adapted from Shleev et al. 2005)

copper cluster (the T2/T3 cluster) (Allendorf et al. 1985; Messerschmidt and Huber 1990; Messerschmidt et al. 1992).

Tyrosinases and laccases are ubiquitously distributed in nature, and their corresponding activities can be observed intra- and/or extracellularly in soil microorganisms. A common feature is the existence of a T3 copper center, and both enzyme classes use molecular oxygen for substrate oxidation with the formation of water (Fig. 13.1).

Tyrosinases are involved in the initial steps of melanin synthesis. They catalyze the ortho-hydroxylation of monophenols to ortho-diphenols, and the latter into reactive ortho-quinones, which are then polymerized into dark pigments. Laccases oxidize various aromatic and nonaromatic compounds through a radical mechanism. They contribute to host defense mechanisms and the metabolic turnover of complex organic substances such as lignin and humic matter.

Both of the copper oxidases have been proposed for various biotechnological applications, such as the treatment of wastewaters or polluted soils, the removal of

Table 13.1 Some features of copper in proteins (modified from Lewis and Tolman 2004; Shleev et al. 2005)

Features

Type 1 copper

Type 2 copper

Type 3 copper

Cu atoms/

1 (mononuclear)

1 (mononuclear)

2 (binuclear,

protein

spin-coupled

CuA/CuB pair)

EPR signal

Paramagnetic

Paramagnetic

Diamagnetic

Light adsorption

High at 610 nm in ox.

Low

High at 330 nm in ox.

state: blue color

state

Coordination

Cys, 2 His, Met or Leu

3 His (2 His and 1 H20

6 His

or Phe in multicopper

in the T2/T3 cluster

proteins

of multicopper

proteins)

Function

Electron transfer,

Electron transfer,

Binding of O2 for

catalysis

catalysis

transport and/or

catalysis

Examples

• Multicopper proteins

• Multicopper proteins

• Multicopper

• Nitrite reductase

• Nitrite reductase

proteins:Ascorbate

• Small blue

• Amine oxidase

oxidase

Cu proteins:

• Cytochrome c oxidase

Billirubin oxidase

Azurin

(+Fe)

Ceruloplasmina

Pseudoazurin

• Galactose oxidase

Fet3 protein

Amicyanin

• Glyoxal oxidase

(Saccharomyces)

Plastocyanin

• Quercetin

Laccase

Stellacyanin

2,3-dioxygenase

• Laccase-like proteins

Rusticyanin

• Superoxide dismutase

(bacteria):

Metallo oxidases

(Mn, Cu, Fe)

Phenoxazinone

synthase

• Tyrosinase

• Hemocyaninb

• Dopamine

b-monooxygenasec

• Peptidylglycine

a-amidating

monooxygenasec

a Also exhibits cuprous oxidase activity (Stoj and Kosman 2003) b Displays tryosinase activity after specific activation (Decker et al. 2007) c Contains two uncoupled Cu ions; it is not known if one or both activate oxygen polyphenols from breweries, the synthesis of pharmaceutical drugs and new biopolymers, or for use as additives in food and cosmetic products (Couto and Herrera 2006; Halaouli et al. 2006).

This contribution provides an overview of the general biochemical and structural properties of tyrosinases and laccases, focusing on their occurrence and relevance in soil microorganisms and giving some examples of biotechno-logical applications of them.

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