Regulation of expression of CRL1 laccase gene from Coriolopsis rigida during detoxification of ‘alpeorujo’

SAPARRAT M. C. N.; JURADO, M.; DÍAZ, R.; MARTÍNEZ, M. J.

Barcelona

Congreso; 14th European Congress on Biotechnology; 2009

European Federation of Biotechnology

Coriolopsis rigida is a white rot fungus with an outstanding ability to degrade lignin and other aromatic recalcitrant compounds from contaminated soils. Laccase secreted from C. rigida is involved in the polymerization of free phenols present in olive mill wastewater and detoxification of this industrial effluent. In Spain, the olive extraction process have been modifying to eliminate liquid effluents being carried out in two-phase centrifugation system, which produces a single semisolid byproduct called ‘alpeorujo’. This residue has a high potential as organic substrate for agronomic purposes, but it contains high levels of free phenols which makes its application difficult. However, some reports suggest that this problem could be solved by treating the residues with C. rigida. Recently, it has been shown that a C. rigida laccase isoenzyme, encoded by the CRL1 gene, is involved in the decrease of the levels of free phenols in the aqueous fraction of ‘alpeorujo’ (AFA) and therefore reducing its toxicity. To optimize and better understand the role of laccase in the process, we analyzed the effect of AFA (2.5, 10 and 20%) and/or Cu2+ (300 uM) on fungal growth, laccase activity and CRL1 gene expression analyzed by real time RT-PCR. C. rigida cultures grown on the basal glucose-peptone medium revealed low levels of extracellular laccase activity, although CRL1 transcripts increased during the incubation time. An increase in fungal biomass was observed in cultures supplemented with Cu2+ and/or AFA. In addition, both Cu2+ and AFA induced similarly CRL1 gene expression, although the effect of Cu2+ on extracellular laccase activity was significantly higher than the effect of AFA compounds at all concentrations assayed. The highest effect of Cu2+ on laccase activity could be related with its role as cofactor required for the catalytic activity of this enzyme. This study suggests that additional doses of Cu2+ during ‘alpeorujo’ treatment with C. rigida could improve the efficiency of the detoxification of this agro-industrial residue.