ObjectiveTo obtain active lipases for biodiesel production by refolding Proteus sp. lipase inclusion bodies expressed in E. coli.ResultsA lipase gene lipPN1 was cloned from Proteus sp. NH 2-2 and expressed in E. coli BL21(DE3). Non-reducing SDS-PAGE revealed that recombinant LipPN1(rLipPN1) were prone to form inclusion bodies as disulfide-linked dimers in E. coli. Site-directed mutagenesis confirmed that Cys85 in LipPN1 was involved in the dimer formation. After optimizing the inclusion body refolding conditions, the maximum lipase activity reached 1662 U/L. The refolded rLipPN1 exhibited highest activity toward p-nitrophenyl butyrate at pH 9.0 and 40 degrees C. It could be activated by Ca2+ with moderate tolerance to organic solvents. It could also convert soybean oil into biodiesel at a conversion ratio of 91.5%.ConclusionPreventing the formation of disulfide bond could enhance the refolding efficiency of rLipPN1 inclusion bodies.