Laccases, role

M. Smith C. E. Thurston D. A. Wood, Eungal Laccases Role in Delignification and Possible Industrial Applications. In Muiti-Copper Oxidases] A. Messerschmidt, Ed. World Scientific Publishing Singapore, 1997 pp 201-224. 

Fungal laccases (benzenediokoxygen oxidoreductase, EC 1.10.3.2) belong to the multicopper blue phenoloxidases. They comprise glycosylated proteins expressed in multiple forms and variable molecular weight, ranging from 59 to 110 kDa. Laccase is expressed as multiple constitutive and induced isoenzymes [30, 64]. The enzyme contains four copper atoms (Cu), in different states of oxidation (I, II, III) [65], which play an important role in the catalytic mechanism. Laccase oxidizes different compounds while reducing O2 to H20, a total reduction of four electrons. 

Reported redox potentials of laccases are lower than those of non-phenolic compounds, and therefore these enzymes cannot oxidize such substances [7]. However, it has been shown that in the presence of small molecules capable to act as electron transfer mediators, laccases are also able to oxidize non-phenolic structures [68, 69]. As part of their metabolism, WRF can produce several metabolites that play this role of laccase mediators. They include compounds such as /V-hvdi oxvacetan i I ide (NHA), /V-(4-cyanophenyl)acetohydroxamic acid (NCPA), 3-hydroxyanthranilate, syringaldehyde, 2,2 -azino-bis(3-ethylben-zothiazoline-6-sulfonic acid) (ABTS), 2,6-dimethoxyphenol (DMP), violuric acid, 1-hydroxybenzotriazole (HBT), 2,2,6,6-tetramethylpipperidin-iV-oxide radical and acetovanillone, and by expanding the range of compounds that can be oxidized, their presence enhances the degradation of pollutants [3]. 

Figure 4.5. Role of mediators in laccase-catalyzed reaction.

In Phanerochaete flavido-alba, an induction of ligninolytic activities that was ascribed to phenolic compounds was evidenced [69]. Phenols have also been shown to have an important role as redox mediators for dye degradation with laccases from Pycnoporus cinnabarinus and Trametes villosa, and they resulted to be necessary to degrade a strongly recalcitrant azo dye, the Reactive Black 5 [70]. 

Polyphenol oxidase occurs within certain mammalian tissues as well as both lower (46,47) and higher (48-55) plants. In mammalian systems, the enzyme as tyrosinase (56) plays a significant role in melanin synthesis. The PPO complex of higher plants consists of a cresolase, a cate-cholase and a laccase. These copper metalloproteins catalyze the one and two electron oxidations of phenols to quinones at the expense of 02. Polyphenol oxidase also occurs in certain fungi where it is involved in the metabolism of certain tree-synthesized phenolic compounds that have been implicated in disease resistance, wound healing, and anti-nutrative modification of plant proteins to discourage herbivory (53,55). This protocol presents the Triton X-114-mediated solubilization of Vida faba chloroplast polyphenol oxidase as performed by Hutcheson and Buchanan (57). 

Copper oxidases are widely distributed in nature, and enzymes from plants, microbes, and mammals have been characterized (104,105). The blue copper oxidases, which include laccases, ascorbate oxidases, and ceruloplasmin, are of particular interest in alkaloid transformations. The principle differences in specificity of these copper oxidases are due to the protein structures as well as to the distribution and environment of copper(II) ions within the enzymes (106). While an in vivo role in metabolism of alkaloids has not been established for these enzymes, copper oxidases have been used in vitro for various alkaloid transformations. 

Laccases are oxidoreductases, primarily secreted by fungi, available in industrial quantities for use in the fabrics industry. Their natural role is in the breakdown of... 

The redox potential of blue copper oxidases varies from species to species. The high redox potential of around 700 mV in fungal laccase is primarily attributed to nonaxial methionine ligand, a geometry that stabilizes the reduced state. Other factors such as solvent accessibility, dipole orientation, and hydrogen bonding also play an important role. ... 

No differences were found between the native and deglycosylated enzymes for the characteristics of pH range of stability, optimum pH, and adsorption to cellulose and milled wood. It can be concluded that the most important role of the carbohydrate chains in laccase III is the resistance to proteolysis in wood decay (15),... 

Scheme 3.1 Illustration of the role of a mediator in a redox reaction cycle, using laccase as the example.

A possible reductive role for veratryl alcohol oxidase is proposed in Figure 5. Laccases from C. versicolor can produce both polymerization and depolymerization of lignin (29). In phenolic lignin model dimers, laccase can perform the same electron abstraction and subsequent bond cleavage as found for lignin peroxidase (30). The phenolic radical is however likely to polymerize unless the quinoid-type intermediates can be removed, for example by reduction back to the phenol. Veratryl alcohol oxidase, in... 

Figure 5. Proposed role of laccase and VA0 in lignin depolymerisation.

---