A Thermo-Active Laccase Isoenzyme From Trametes trogii and Its Potential for Dye Decolorization at High Temperature

Abstract

A thermo-activation and thermostable laccase isoenzyme (Lac 37 II) produced by <i>Trametes trogii</i> S0301 at 37°C was purified to apparent homogeneity by anionic exchange chromatography and sephadex G-75 chromatography, with 12.3% of yeiled and a specific activity of 343.1 U mg^-1. The molecular weight of the purified Lac 37 II was estimated to be approximately 56 kDa in 12% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The optimal pH and temperature for the protein was 2.7 and 60°C, respectively. The purified Lac 37 II showed higher resistance to all tested metal ions and organic solvents except for Fe²⁺ and Cd²⁺ at 37°C and the activity of the purified Lac 37 was significantly enhanced by Cu²⁺ at 50 mM. The <i>K</i> _<i>cat</i> , <i>K</i> _<i>m</i> , and <i>K</i> _<i>cat</i> /<i>K</i> _<i>m</i> of Lac 37 II were 2.977 s^-1, 16.1 μM, and 184.9 s^-1 μM^-1, respecively, in the condition of pH 2.7 and 60°C using ABTS as a substrate. Peptide-mass fingerprinting analysis showed that the Lac 37 II matched to the gene-deduced sequences of <i>lcc3</i> in <i>T. trogii</i> BAFC 463, other than <i>Lcc1</i>, <i>Lcc 2</i>, and <i>Lcc 4</i>. Compared with laccase prepared at 28°C, the onset of thermo-activation of Lac 37 II activity occurred at 30°C with an increase of 10%, and reached its maximum at the temperatures range of 40-60°C with an increase of about 40% of their original activity. Furthermore, Lac 37 II showed the efficient decolorization ability toward triphenylmethane dyes at 60°C, with decolorization rates of 100 and 99.1% for 25 mg L^-1 malachite and crystal violet in 5 h, respectively, when hydroxybenzotriazole (HBT) was used as a mediator. In conclusion, it is the first time to report a thermo-activation laccase from a thermophilic <i>T. trogii</i> strain, which has a better enzyme property and higher decolorization ability among fungal laccases, and it also has a further application prospective in the field of biotechnology.