Induction and Purification of Diphenol Oxidase from Lenzite elegans

Abstract

Laccase (EC.1.10.3.2. Diphenol oxidase) are multicopper oxidases that catalyze oxidation of many substituted phenolic compounds, aromatic amines including some inorganic substances by using molecular oxygen as electron acceptor. Nevertheless, most laccases that have been studied so far are not well-suited for industrial applications due to their low stability at high temperatures or pH values. This research focused on identifying and characterizing novel fungal laccases that have potential for industrial applications as well as developing efficient production methods for laccases. Lenzite elegans, a white rot fungi was screened by plate tests using indicator compound 2,2’ Azinobis(3-ethylbenzthiazoline-6-sulfonic ) acid (ABTS). The ability of Lenzite elegans to produce laccase was studied in liquid media. This fungal species produced significant amounts of laccase in titers (958 U), and the enzyme was preliminarily characterized. The novel laccase was found to be rather typical basidiomycete laccase. The novel laccase from Lenzite elegans was purified and biochemically characterized and has a relatively high specific activity (30.9 U). This enzyme also showed remarkable stability and active at high temperatures (60oC). Medium I and II supported the elaboration of LiP, MnP and Laccase enzymes than 2 % ME and CCWM. However, the highest Laccase titers were recorded in CCWM. The enzyme was inhibited by NaN3, EDTA and NaI, and the inhibition was dose dependent. Divalent cations Ca2+, Zn2+, Mg2+, Fe2+, Co2+ and Cu2+ were tested against the enzyme, Fe2+, Cu2+ exhibited similar characteristics with Fe2+ effect higher than all the cations used. Laccase from Lachnocladium sp. had a KM of 0.067 μM and Vmax of 0.505 U. with low protein concentrations of 0.118-0.196 mg recorded in this study could be traced to fungal growth and purification procedures.