Overexpression of Recombinant Lipase from Burkholderia Cepacia in Escherichia Coli

Abstract

This study attempts to clone and express the extracellular lipase from Burkholderia cepacia in Escherichia coli using pET system as well as to determine the enzyme activity of recombinant lipase. The extracted DNA from B. cepacia was used as a template for amplifying lipase gene, and then the lipase gene was subcloned into pET-32a and subsequently transformed into E. coli BL21. Media assay and SDS-PAGE were carried out to analyse the results. Nucleotide sequencing of the DNA insert from the clone revealed that the lipase activity corresponded to an open reading frame consisting of 1092 bp coding for a 37.5-kDa protein. The successful expression of lipase was confirmed by obtaining blue color colonies on Nile Blue Sulphate Agar and big band at 37.5-kD size on SDS-PAGE. The enzyme activity assay also showed the high lipase activity around 590 μg lipase ml−1 culture 30 min−1 of recombinant E. coli BL21. The specific lipolytic activity of the recombinant lipase was 185 U/mL which is around 35-fold higher than the native baseline. The findings suggest that the crude recombinant lipase has potential application in digestion of lipids and fatty acids. In conclusion, the results of the current study showed a lipase gene encoding an enzyme with non-specific hydrolysis activity, which could be applied as lipase biosensor for digestion of lipids in food and medicine as well as oil-contamination treatment.