Kinetic characterization of purified laccase from Trametes hirsuta: a study on laccase catalyzed biotransformation of 1,4-dioxane.

Abstract

Laccase is one of the best known biocatalysts which degrade wide varieties of complex molecules that are both non-cyclic and cyclic in structure. The study focused on enzyme kinetics of a purified laccase from Trametes hirsuta L. fungus and its application on biotransformation of a carcinogenic molecule 1,4-dioxane. Laccase was purified from white-rot fungus T. hirsuta L. which showed specific activity of 978.34 U/mg after the purification fold of 54.08. The stable laccase activity (up to 16h) is shown at 4-6 pH and 20-40°C temperature range. The purified enzyme exhibited significant stability for 10 metal ions up to 10mM concentration, except for Fe2+ and Hg2+. The Cu2+ ion induced laccase activity up to 142% higher than the control at 10mM concentration. The laccase enzyme kinetic parameters Km was 20 ± 5µM and 400 ± 60µM, whereas Kcat was 198.29 ± 0.18/s and 80.20 ± 1.59/s for 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and guaiacol respectively. The cyclic ether 1,4-dioxane (100ppm) was completely degraded in presence of purified laccase within 2h of incubation and it was confirmed by HPLC and GC analysis. The oxidation reaction was accelerated by 25, 22, 6 and 19% in presence of 1mM syringaldehyde, vanillin, ABTS and guaiacol mediators respectively. In this study, fungal laccase (a natural biocatalyst) based degradation of synthetic chemical 1,4-dioxane was reported for the first time. This method has added advantages over the multiple methods reported earlier being a natural remedy.