Heat-induced overexpression of the thermophilic lipase from Bacillus thermocatenulatus in Escherichia coli by fermentation and its application in preparation biodiesel using rapeseed oil.

ABSTRACT

Due to its simple, less by-product and environment friendly properties, enzymatic transesterification of oil with short-chain alcohol to biodiesel, fatty acid methyl esters (FAMEs) is considered to be a promising way of green production and has attracted much attention. In this study, FAMEs were synthesized by an enzymatic method with recombinant lipase as catalysts. A thermophilic Bacillus thermocatenulatus lipase 2 (BTL2) was overexpressed in Escherichia coli BL21(DE3) through relative and quantitative analysis using real-time quantitative PCR. The results suggested that the BTL2 gene was overexpressed in E. coli at the mRNA level, and the recombinant strain harboring a high-copy number vectors was selected and applied to fermentation to produce BTL2 with enzyme activity of 35.54 U/mg cells. The recombinant BTL2 solution exhibited excellent resistance to neutral pH, high temperature, and organic solvents after a certain treatment. Finally, the effects of enzymatic transesterification for preparing biodiesel were studied, using rapeseed oil as raw material, as well as BTL2 solution as catalysts, which resulted in 86.04% yield of FAMEs under 50°C for 36 h. The liquid BTL2 was directly used to prepare FAMEs at a higher temperature efficiently, making the thermophilic BTL2 had the potential application value in biodiesel reproduction subsequently.