Optimization, purification and characterization of laccase from Ganoderma leucocontextum along with its phylogenetic relationship

Abstract

The aim of this work to study an efficient laccase producing fungus Ganoderma leucocontextum, which was identified by ITS regions of DNA and phylogenetic tree was constructed. This study showed the laccase first-time from G. leucocontextum by using medium containing guaiacol. The growth cultural (pH, temperature, incubation days, rpm) and nutritional (carbon and nitrogen sources) conditions were optimized, which enhanced the enzyme production up to 4.5-folds. Laccase production increased 855 U/L at 40 °C. The pH 5.0 was suitable for laccase secretion (2517 U/L) on the 7th day of incubation at 100 rpm (698.3 U/L). Glucose and sucrose were good carbon source to enhance the laccase synthesis. The 10 g/L beef (4671 U/L) and yeast extract (5776 U/L) were the best nitrogen source for laccase secretion from G. leucocontextum. The laccase was purified from the 80% ammonium sulphate precipitations of protein identified by nucleotides sequence. The molecular weight (65.0 kDa) of purified laccase was identified through SDS and native PAGE entitled as Glacc110. The Glacc110 was characterized under different parameters. It retained > 90% of its activity for 16 min incubation at 60 °C in acidic medium (pH 4.0). This enzyme exerted its optimal activity at pH 3.0 and temperature 70 °C with guaiacol substrate. The catalytic parameters Km and Vmax was 1.658 (mM) and 2.452 (mM/min), respectively. The thermo stability of the laccase produced by submerged fermentation of G. leucocontextum has potential for industrial and biotechnology applications. The results remarked the G. leucocontextum is a good source for laccase production.