Production and characterization of novel thermostable CotA-laccase from Bacillus altitudinis SL7 and its application for lignin degradation.

Laccases are multi-copper oxidases and found in ligninolytic bacteria catalyzing the oxidation of both phenolic and non-phenolic compounds, however its application in lignin degradation suffers due to low oxidation rate, which have intensified the search for new laccases. In the present study, spore coat A protein (CotA) encoding gene having laccase like activity from Bacillus altitudinis SL7 (CotA-SL7) was cloned and expressed in Escherichia coli. The purified CotA-SL7 was active at wide range of temperature and pH with optimum activity at 55 °C and pH 5.0. The kinetic parameters of CotA-SL7 was determined with K, V and k values 0.4 mM, 2777 μmol/min/mg, and 5194 s, respectively. Molecular docking revealed the presence of Pro, Phe, Asp, Asn, His, and Ile residues at the active site taking part in the oxidation of ABTS. The purified CotA-SL7 reduced lignin contents by 31 % and changes in lignin structure were analyzed through fourier transformed infrared spectroscopy (FTIR), scanning electron microsscopy (SEM) and gas chromatography mass-spectrometry (GC-MS). The appearance of low molecular size compounds clearly indicates the cleavage of lignin polymer and opening of the benzene ring by purified CotA-SL7. Thus, high catalytic efficiency of CotA-SL7 makes it a suitable bio-catalyst for remediation of lignin contaminated wastewater from pulp and paper industries with clear insights into lignin degradation at molecular level.