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Laccases are members of a large family of multicopper oxidases that catalyze the oxidation of a wide range of organic and inorganic substrates accompanied by the reduction of dioxygen to water. A new laccase was isolated from the basidiomycete Coriolopsis caperata strain 0677 and its amino-acid sequence was determined. According to its physicochemical properties and spectroscopic features, the laccase from C. caperata is a high redox-potential blue laccase. Attempts to crystallize the native enzyme were unsuccessful. The copper type 2-depleted (T2D) laccase was prepared and crystallized. The structure of T2D laccase from C. caperata was solved at 1.6 Å resolution, and attempts to reconstruct the T2 copper centre were performed using Cu+ and Cu2+ ions. The structure of T2D+Cu+ laccase was solved at 1.89 Å resolution. It was shown that the T2D+Cu+ laccase structure contained four copper ions in the active site. Reconstruction could not be achieved when the T2D laccase crystals were treated with CuSO4. [ABSTRACT FROM AUTHOR]

Laccases are oxidases that contain several copper atoms, and catalyse single-electron oxidations of phenolic compounds with concomitant reduction of oxygen to water. The enzymes are particularly widespread in ligninolytic basidiomycetes, but also occur in certain prokaryotes, insects and plants. Depending on the species, laccases are involved in various biosynthetic processes contributing to carbon recycling in land ecosystems and the morphogenesis of biomatrices, wherein low-molecular-weight naturally occurring phenols serve as key enzyme substrates. Studies of these in vivo synthetic pathways have afforded new insights into fungal laccase applicability in green synthetic chemistry. Thus, we here review fungal laccase-catalysed oxidations of naturally occurring phenols that are particularly relevant to the synthesis of fine organic chemicals, and we discuss how the discovered synthetic strategies mimic laccase-involved in vivo pathways, thus enhancing the green nature of such reactions. Laccase-catalysed in vivo processes yield several types of biopolymers, including those of cuticles, lignin, polyflavonoids, humus and the melanin pigments, using natural mono- or poly-phenols as building blocks. The in vivo synthetic pathways involve either phenoxyl radical-mediated coupling or cross-linking reactions, and can be adapted to the design of in vitro oxidative processes involving fungal laccases in organic synthesis; the laccase substrates and the synthetic mechanisms reflect in vivo processes. Notably, such in vitro synthetic pathways can also reproduce physicochemical properties (e.g. those of chromophores, and radical-scavenging, hydration and antimicrobial activities) found in natural biomaterials. Careful study of laccase-associated in vivo metabolic pathways has been rewarded by the discovery of novel green applications for fungal laccases. This review comprehensively summarizes the available data on laccase-catalysed biosynthetic pathways and associated applications in fine chemical syntheses. [ABSTRACT FROM AUTHOR]

• In forest soils, ectomycorrhizal and saprotrophic Agaricales differ in their strategies for carbon acquisition, but share common gene families encoding multi-copper oxidases (MCOs). These enzymes are involved in the oxidation of a variety of soil organic compounds. • The MCO gene family of the ectomycorrhizal fungus Laccaria bicolor is composed of 11 genes divided into two distinct subfamilies corresponding to laccases ( lcc) sensu stricto ( lcc1 to lcc9), sharing a high sequence homology with the coprophilic Coprinopsis cinerea laccase genes, and to ferroxidases ( lcc10 and lcc11) that are not present in C. cinerea. The fet3-like ferroxidase genes lcc10 and lcc11 in L. bicolor are each arranged in a mirrored tandem orientation with an ftr gene coding for an iron permease. Unlike C. cinerea, L. bicolor has no sid1/ sidA gene for siderophore biosynthesis. • Transcript profiling using whole-genome expression arrays and quantitative reverse transcriptase–polymerase chain reaction (qRT-PCR) revealed that some transcripts were very abundant in ectomycorrhizas ( lcc3 and lcc8), in fruiting bodies ( lcc7) or in the free-living mycelium grown on agar medium ( lcc9 and lcc10), suggesting a specific function of these MCOs. • The amino acid composition of the MCO substrate binding sites suggests that L. bicolor MCOs interact with substrates different from those of saprotrophic fungi. [ABSTRACT FROM AUTHOR]

A new exocellular laccase was purified from the basidiomycete C30. LAC2 is an acidic protein (pI = 3.2) preferentially produced upon a combined induction by copper and p -hydroxybenzoate. The spectroscopic signature (UV/visible and EPR) of this isoform is typical of multicopper oxidases, but its enzymatic and physico-chemical properties proved to be markedly different from those of LAC1, the constitutive laccase previously purified from the same organism. In particular, the LAC2 k cat values observed for the oxidation of the substrates syringaldazine (k cat = 65 600 min-1 ), ABTS (2,2-azino-bis-[3-ethylthiazoline-6-sulfonate] (k cat = 41 000 min-1 ) and guaiacol (k cat = 75 680 min-1 ) are 10–40 times those obtained with LAC1 and the redox potential of its T1 copper is 0.17 V lower than that of LAC1 (E ° = 0.73 V). This is the first report on a single organism producing simultaneously both a high and a low redox potential laccase. The cDNA, clac2 , was cloned and sequenced. It encodes a protein of 528 amino acids that shares 69% identity (79% similarity) with LAC1 and 81% identity (95% similarity) with Lcc3-2 from Polyporus ciliatus (AF176321-1), its nearest neighbor in database. Possible reasons for why this basidiomycete produces, in vivo , enzyme forms with such different behaviors are discussed. [ABSTRACT FROM AUTHOR]

The effects of cadmium Cd (II) ions on the physiology and biological activity of Trametes versicolor, a strain belonging to white-rotting Basidiomycetes, were examined. Cd (II) ions were added to 10-day-old cultures grown on a liquid medium, or at the time of inoculation. Our experiments showed that T. versicolor is a good cadmium biosorbent from aqueous solution, this strain removing almost all the Cd (ll) ions over the first 2 h of incubation by what appears to be a rapid, energy-independent surface binding phenomenon, at the rate of ∼2 mg Cd per g mycelial dry weight. An additional slower and energy-dependent transport mechanism was also present, taking in ∼0.3 mg Cd (II) per g dry weight. It is also shown that these Cd (II) ions significantly stimulate the activity of extracellular laccase when added to 10-day-old cultures. [Copyright &y& Elsevier]

The blue laccase from the white-rot basidiomycete fungus Panus tigrinus, an enzyme involved in lignin biodegradation, has been crystallized. P. tigrinus laccase crystals grew within one week at 296 K using the sitting-drop vapour- diffusion method in 22%(w/v) PEG 4000, 0.2 M CaCl2, 100 mM Tris-HCl pH 7.5. The crystals belong to the monoclinic space group P21, with unit-cell parameters a = 54.2, b = 111.6, c = 97.1, β = 97.7°, and contain 46% solvent. A complete native data set was collected to 1.4 Å resolution at the copper edge. Molecular replacement using the Coprinus cinereus laccase structure (PDB code 1 hfu) as a starting model was performed and initial electron-density maps revealed the presence of a full complement of copper ions. Model refinement is in progress. The P. tigrinus laccase structural model exhibits the highest resolution available to date and will assist in further elucidation of the catalytic mechanism and electron-transfer processes for this class of enzymes. [ABSTRACT FROM AUTHOR]