Expression, purification, and characterization of a novel laccase from Setosphaeria turcica in Eschericha coli.

Laccases are multicopper oxidases (E.C. 1.10.3.2) that catalyze the oxidation of many phenolic compounds. In this study, a novel laccase, Stlac4, from Setosphaeria turcica was cloned and expressed in Escherichia coli by insertion into the pET-30a expression plasmid. The recombinant laccase was purified and visualized on SDS-PAGE as a single band with an apparent molecular weight of 71.5 KDa, and confirmed by Western blot. The maximum activity of the purified laccase was 127.78 U · mg , the optimum temperature and pH value were 60 °C and 4.0 respectively, measured by oxidation of 2,2'-Azinobis-(3-ethylbenzthiazoline-6-sulphonate) (ABTS). Purified laccase activity under different metal ions and an inhibitor were tested, revealing that laccase activity increased by approximately 434.8% with Fe , and 217.4% with Cu at 10 mmol · L concentrations, Mn increased the laccase activity only at 5 mmol · L , while Na increased activity at 1 mmol · L but inhibited activity at 5 and 10 mmol · L . SDS increased laccase activity at 1 mmol · L , and inhibited activity at 5 and 10 mmol · L .