Purification and characterization of a fungal laccase from the ascomycete Thielavia sp. and its role in the decolorization of a recalcitrant dye.

A laccase-producing ascomycete was isolated from arid soil in Tunisia. This fungus was identified as Thielavia sp. using the phylogenetic analysis of rDNA internal transcribed spacers. The extracellular laccase produced by the fungus was purified to electrophoretic homogeneity, showing a molecular mass around 70 kDa. The enzyme had an optimum pH of 5.0 and 6.0 for ABTS and 2,6­DMP, respectively and it showed remarkable high thermal stability, showing its optimal temperature at 70 °C (against 2,6­DMP). It presented slight inhibiting effect by EDTA, SDS and l­cyst although this effect was more marked by sodium azide (0.1 mM). On the other hand, it showed tolerance to up to 300 mM NaCl, retaining around 50% of its activity at 900 mM. Among the metal ions tested on TaLac1, Mn2+ showed an activating effect. Their kinetic parameters Km and kcat were 23.7 µM and 4.14 s-1 for ABTS, and 24.3 µM and 3.46 s-1 towards 2,6­DMP. The purified enzyme displayed greater efficiency in Remazol Brilliant Blue R decolorization (90%) in absence of redox mediator, an important property for biotechnological applications.

A halotolerant laccase from Chaetomium strain isolated from desert soil and its ability for dye decolourization.

A novel fungal laccase produced by the ascomycete Chaetomium sp. isolated from arid soil was purified and characterized and its ability to remove dyes was determined. Extracellular laccase was purified 15-fold from the crude culture to homogeneity with an overall yield of 50% using ultrafiltration and anion-exchange chromatography. The purified enzyme was found to be a monomeric protein with a molecular mass of 68 kDa, estimated by SDS-PAGE, and with an isoelectric point of 5.5. The optimal temperature and pH value for laccase activity toward 2,6-DMP were 60 °C and 3.0, respectively. It was stable at temperatures below 50 °C and at alkaline conditions. Kinetic study showed that this laccase showed higher affinity on ABTS than on 2,6-DMP. Its activity was enhanced by the presence of several metal ions such as Mg2+, Ca2+ and Zn2+, while it was strongly inhibited by Fe2+, Ag+ and Hg2+. The novel laccase also showed high, remarkable sodium chloride tolerance. Its ability to decolorize different dyes, with or without HBT (1-hydroxy-benzotriazole), as redox mediator, suggests that this protein may be useful for different industrial applications and/or bioremediation processes.