Abstract
A novel cold-active true lipase from Pseudomonas sp. KE38 was cloned, sequencing and expressed in E. coli by degenerate PCR and genome walking technique. The open reading frame of the cloned gene encoded a polypeptide chain of 617 amino acids with a confirmed molecular weight of 64 kD. Phylogenetic analysis of the deduced amino acid sequence of the lipase indicated that it had high similarity with lipases of subfamily Ι.3 of bacterial lipases. Recombinant lipase was purified in denatured form as inclusion bodies, which were then renatured by urea followed by dialysis. Lipase activity was determined titrimetrically using olive oil as substrate. The enzyme showed optimal activity at 25 °C, pH 8.5 and was highly stable in the presence of various metal ions and organic solvents. Low optimal temperature and high activity in the presence of methanol and ethanol make this lipase a potential candidate for transesterification reactions and biodiesel production.
Highlights
Lipolytic enzymes catalyze the hydrolysis of long-chain triacylglycerols to glycerol and free fatty acids in the presence of excess water
Special attention has been focused on Pseudomonas lipases due to their thermo-resistance and activity at alkaline pHs which are not common among the lipases produced by other microorganisms[10]
In a previous study performed in our lab, we described the purification and characterization of an extracellular lipase from a pcychro-tolerant bacterium, Pseudomonas fluorescens KE3821
Summary
Lipolytic enzymes catalyze the hydrolysis of long-chain triacylglycerols to glycerol and free fatty acids in the presence of excess water. True lipases on the other can catalyze the hydrolysis of triacylglycerols with an acyl chain length ≥ 10 carbon a toms[4,5,6] Because of their activities in both aqueous and nonaqueous environments, lipases have specific applications in industry and medicine[7]. KE38Lip_R and other materials[11] Another useful property of cold-active enzymes is their ability to catalyze reactions at low or moderate temperatures[11,12]. Due to these two important features, cold-active lipases have recently attracted attention and found various applications in the field of detergent industry (cold-washing), bioremediation of oil-contaminated cold environments and biotransformation reactions of heat-labile compounds[13].
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