Production of extracellular laccase from the newly isolated <i>Bacillus</i> sp. PK4

Abstract

Laccase belongs to the family of multicopper oxidase and have wide range of biotechnological applications starting from the food industry to bioremediation. However, the use of laccase at commercial scale is hindered by factors like high enzyme cost, low activity and /or stability under given conditions. This study was carried out with aim of screening for extracellular thermostable laccase producing bacteria. Twenty-two (22) laccase positive strains were isolated from the selected environmental samples while extracellular laccase activity was detected only in six strains namely TM1, TMT1, PK4, PS1, TMS1 and ASP3. The laccase enzyme produced from PK4 was found to be more thermostable with a half-life of 60 min at 80°C. The strain PK4 was identified and designated as Bacillus sp. PK4. The laccase production was optimized using one- factor-at-a-time method and maximum enzyme production were observed at the temperature of 37°C, pH 7.5, 10% inoculum size with yeast extract and glucose as the nitrogen and carbon sources, respectively at an agitation rate of 150 rpm. Copper sulphate at 0.1 mM concentration was found to maximize the laccase production among the tested inducers. Among the trace elements, FeSO₄ and ZnSO₄ gave the maximum laccase production for the isolated strain in comparison with the control. The effect of copper-induced time showed that the addition of copper before inoculation effectively increased the laccase production compared to the addition of copper after 2, 4, 6 and 8 h of inoculation. The optimization of the media resulted in 11.8 fold increase in laccase production.

Keywords: Laccase, thermostability, Bacillus sp., optimization