Cloning, expression, purification, and characterization of lipase 3646 from thermophilic indigenous Cohnella sp. A01

• Lipase from Cohnella sp. A01is a thermostable enzyme.
• We purified lipase 3646 using two steps of DE52 ion exchange chromatography only.
• Lipase 3646 showed high stability in alkaline pH.
• Lipase 3646 could be exploited in certain industries such as detergent.
• Lipase 3646 is highly stable in the presence of organic solvents and detergents.

Lipases form one of the most important groups of biocatalysts used in biotechnology. We studied the lipase from the bacterium Cohnella sp. A01 due to the versatility of thermophilic lipases in industry. In this study lipase 3646 gene from the thermophilic bacterium Cohnella sp. A01 was expressed in Escherichia coli and the enzyme was purified by a two-steps anion exchange chromatography. The purified lipase appeared to have a molecular weight of approximately 29.5 kDa on SDS–PAGE. The values of Km and Vmax, calculated by the Michaelis–Menten equation, were 1077 μM and 61.94 U/mg, respectively. The kinetic characterization of the purified enzyme exhibited maximum activity at 70 °C and pH 8.5. Activities at 50, 55 and 60 °C for 120 min were measured 58%, 47% and 41%, respectively. The enzyme was also highly stable at the pH range of 8.5–10.0 for 180 min. The effect of EDTA indicated that the enzyme is not a metalloenzyme. The stability of lipase 3646 in the presence of organic solvents, detergents, metal ions and inhibitors suggested that this lipase could be exploited in certain industries such as detergent and leather. Lipase 3646 was determined structurally to be 37.5% α-helix, 12.8% β-sheet, 22.7% β-turn and 27% random coil.