Prokaryotic Expression and Enzymatic Properties of Lipase from Schizochytrium Pombe

Based on the codon preference of Escherichia coli, the gene of lipase from Schizochytrium pombe was optimized and the codon adaptation index was 0.8 and the gene was expressed in Escherichia coli as a His-tag fusion protein using pET-30a as the expression vector. The recombinant strain with OD600 about 0.6 was induced with 1 mmol/L isopropyl-β-D-thiogalactoside at 30 °C for 12 h. The purified recombinant lipase STGL3 had a molecular mass of 32 kDa, and the purified lipase had a specific enzyme activity of 2.29 μmol/(min•mg), which was 2.3 times higher than before, and a protein content of 650.00 mg/L. The recombinant STGL3 showed the highest hydrolysis activity at 40 °C and pH 7.5, respectively. The effect of different metal ion concentrations on enzyme activity varied, lipase STGL3 was stable in the presence of Ca2+, while Mg2+, Mn2+, Zn2+, Fe2+ and EDTA had an inhibitory effects on lipase activity. Different types of surfactants and organic solvents inhibited the enzymatic activity of recombinant lipase to different extents. The recombinant lipases showed a preference towards pNP-esters with medium and longer acyl-chains. (C12, C14 and C16) and the recombinant lipases showed relatively weak lipase activity towards the shorter carbon chains. The kinetic constants of lipase were 0.29 mmol/L for Km, 2.28 mmol/(L•min-1 ) for Vmax, and 6.19 S-1 for Kcat. The present study not only achieved the expression and characterization of enzymatic properties of lipase from Leptosphaeria japonica, but also provided some basis for the study of enhancing the quality and quality of algal oil by means of genetic engineering, and further exploring the potential industrial applications of Leptosphaeria japonica lipase.