Improving of the Thermostability of Penicillium expansum Lipase by Creating Mutation P197E

Abstract:

In order to improve the thermostability of Penicillium expansum lipase (PEL), a mutation P197E was created by site-directed mutagenesis. The mutation is generated by overlap extension PCR using the cDNA of a single site mutant lipase ep8 as the template and two special primers that corresponding to mutation P197E. Recombinant vector pPIC3.5K-ep8-P197E which contain double-mutant genes was constructed and electroporated into Pichia pasteris GS115. The recombinant transformant was selected and grow in the methanol containing media for expressing double-mutant lipase, PEL-ep8-P197E. Thermostability analysis reveals that the residual activity of the double-mutant are 42.13% and 37.3% greater than that of the wild type PEL and PEL-ep8 after incubated at 40°C for 30 min. The Tm of the double-mutant lipase is 41.51°C, 2.81°C higher than PEL and 2.25°C higher than PEL-ep8.

Key words:Penicillum expansum lipase, Mutagenesis, Thermostability