Heterologous expression and characterization of a novel thermostable and alkali stable recombinant lipase enzyme from Bacillus thuringensis into E. coli BL21(DE3) for detergent formulation

Abstract

Present study concerned with expression and biochemical characterization of lipase enzyme for potential use in detergent formulation. Lipase gene (1242 bp) of Bacillus thuringensis was cloned and expressed in Escherichia coli BL21 strain using pET-21a(+) expression vector. Maximum expression of cloned lipase gene was obtained at 37°C with an induction of 0.4 mM IPTG (Isopropyl ß-D-1-thiogalactopyranoside) after 4 h of induction. Recombinant lipase was purified to homogeneity using immobilized metal ion affinity chromatography carrying 109.80 U/mg specific activity with 38.79 purification folds. Molecular mass of purified lipase was determined as 45 kDa using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Purified recombinant lipase showed stability up to 90°C and retained significant activity (52%) after 4 h at 90°C. It was also found to be stable at a wide range of pH and in the presence of higher concentrations of several inhibitors (sodium dodecyl sulfate, dimethylsulfoxide, sodium azide, β-mercaptoethanol, polysorbate 80, dithiothreitol) as well as organic solvents (acetone, methanol, ethanol, isopropanol, n-butanol). The activity of recombinant lipase was enhanced in the presence of various metal ions and activated up to 200% by Ca²⁺. The compatibility of recombinant lipase with commercial detergents and other additives as well as its broad substrate specificity endorse the potential application of this enzyme in detergent formulations.