The effect of carbon sources on the expression level of thermostable L2 lipase in Pichia pastoris

Thermostable lipases are enzymes that are particularly pleasing for industrial purposes such as in the production of detergents, animal skin-based industry and food processing endeavours. Thermostable L2 lipase gene fished out from Bacillus sp. L2 was cloned into Pichia pastoris strain GS115 under constitutive expression system of pGAPZαA. Study performed on various carbon sources revealed that glucose and glycerol support the growth of Pichia pastoris and expression of L2 lipase. In addition, the by-product of sugar refinery processes (molasses) was found to be a potential carbon source for the growth of P. pastoris and L2 lipase expression. Since the thermostable L2 lipase will be extracellularly secreted into the medium, the use of P. pastoris expression system is seen as an alternative to the conventional expression system of Escherichia coli that applies intracellular expression of the L2 lipase gene encoded.Key words: Thermostable lipase, Pichia pastoris, constitutive expression, carbon source

Ability of T1 lipase to degrade amorphous P(3HB): structural and functional study

An enzyme with broad substrate specificity would be an asset for industrial application. T1 lipase apparently has the same active site residues as polyhydroxyalkanoates (PHA) depolymerase. Sequences of both enzymes were studied and compared, and a conserved lipase box pentapeptide region around the nucleophilic serine was detected. The alignment of 3-D structures for both enzymes showed their active site residues were well aligned with an RMSD value of 1.981 Å despite their sequence similarity of only 53.8%. Docking of T1 lipase with P(3HB) gave forth high binding energy of 5.4 kcal/mol, with the distance of 4.05 Å between serine hydroxyl (OH) group of TI lipase to the carbonyl carbon of the substrate, similar to the native PhaZ7 Pl . This suggests the possible ability of T1 lipase to bind P(3HB) in its active site. The ability of T1 lipase in degrading amorphous P(3HB) was investigated on 0.2% (w/v) P(3HB) plate. Halo zone was observed around the colony containing the enzyme which confirms that T1 lipase is indeed able to degrade amorphous P(3HB). Results obtained in this study highlight the fact that T1 lipase is a versatile hydrolase enzyme which does not only record triglyceride degradation activity but amorphous P(3HB) degradation activity as well