Purification, characterization and thermostability of lipase from a thermophilic Bacillus sp J33

A thermostable lipase produced by a thermophilic Bacillus sp. J33 was purified to 175-fold with 15.6% recovery by ammonium sulphate and Phenyl Sepharose column chromatography. The enzyme is a monomeric protein having molecular weight of 45 kDa. It hydrolyzes triolein at all positions. The fatty acid specificity of lipase is broad with little preference for C12 and C4. The K-m and V-max for lipase with pNP-laurate as substrate was calculated to be 2.5 mM and 0.3 mu M min(-1) ml(-1) respectively. The immobilized enzyme was stable for 12 h at 60 degrees C. Polyhydric alcohols such as ethylene glycol (2.5 M), sorbitol (2.5 M) and glycerol (2.5 M) were used as thermostabilizers. Lipase acquired a remarkable stability, since no deactivation occurred at 70 degrees C for 150 min in the presence of additives.