Laccase biological activity

As one of the blue oxidases, laccase contains four copper ions with strong intermetal-lic interactions between one pair. The involvement of a pair of copper ions in the biologically active role of reduction of dioxygen has added to the interest shown in this enzyme for more than ten years. Reviews in 197087, 197388 and 197589,90 contain general discussions of the copper oxidases including laccase. 

SSF containing wheat bran and soybean as a substrate was chosen for the production of ligninolytic enzymes for Funalia trogii ATCC200800 [18] as it mimics the natural environment of the WRF and permits the concentration of dyes by absorption process prior to biological treatment [4, 38, 39]. It is possible to stimulate the yield of laccase activity of Trametes versicolor by using several agricultural wastes [40]. 

The multi-copper oxidases include laccase, ceruloplasmin, and ascorbate oxidase. Laccase can be found in tree sap and in fungi ascorbate oxidase, in cucumber and related plants and ceruloplasmin, in vertebrate blood serum. Laccases catalyze oxidation of phenolic compounds to radicals with a concomitant 4e reduction of O2 to water, and it is thought that this process may be important in the breakdown of lignin. Ceruloplasmin, whose real biological function is either quite varied or unknown, also catalyzes oxidation of a variety of substrates, again via a 4e reduction of O2 to water. Ferroxidase activity has been demonstrated for it, as has SOD activity. Ascorbate oxidase catalyzes the oxidation of ascorbate, again via a 4e reduction of O2 to water. Excellent reviews of these three systems can be found in Volume 111 of Copper Proteins and Copper Enzymes (Lontie, 1984). 

Fueled by the biological relevance, the initial steps of Cu-based O2 activation have attracted much interest in the last decades. Various, very different Cux/02 species that result from the reaction of dioxygen with Cu complexes have meanwhile been identified (Fig. 2) [70-90], where mononuclear species A and B as well as dinuclear type E and F species (and some tricopper systems as in, for example, laccase) are considered the most relevant in nature. 

Copper proteins are involved in a variety of biological functions, including electron transport, copper storage and many oxidase activities. A variety of reviews on this topic are available (Sykes, 1985 Chapman, 1991). Several copper proteins are easily identified by their beautiful blue colour and have been labelled blue copper proteins. The blue copper proteins can be divided into two classes, the oxidases (laccase, ascorbate oxidase, ceruloplasmin) and the electron carriers (plastocyanin, stellacyanin, umecyanin, etc.). 

The low-temperature MCD and absorption titration studies (Figure 10) have determined that azide binds to both the type 2 and type 3 centers with similar binding constants. A series of chemical perturbations and stoichiometry studies have shown that these effects are associated with the same azide. This demonstrates that one N3 bridges between the type 2 and type 3 centers in laccase. These and other results from MCD spectroscopy first defined the presence of a trinuclear copper cluster active site in biology (89). At higher azide concentration, a second azide binds to the trinuclear site in laccase. Messerschmidt et al. have determined from X-ray crystallography that a trinuclear copper cluster site is also present in ascorbate oxidase (87, 92) and have obtained a crystal structure for a two-azide-bound derivative (87). It appears that some differences exist between the two-azide-bound laccase and ascorbate oxidase derivatives, and it will be important to spectroscopically correlate between these sites. 

In order to mimic biological cell membranes, biomolecules have been used either to decorate the membrane surface or be inserted directly inside the membrane. Various enzymes, such as laccase, GOx, or HRP, have been reported for decorating planar membranes. These enzymes were either chemically linked to the membrane by reactions such as esterification and amidation, or just physically adsorbed. For example, physically adsorbed laccase on copolymer membranes displayed a higher activity than free... 

---