Development of a Pichia pastoris whole-cell biocatalyst with overexpression of mutant lipase I PCL G47I from Penicillium cyclopium for biodiesel production.

Penicillium cyclopium lipase I (PCL) is a thermolabile triacylglycerol lipase with very low activity against monoacylglycerols, and there have been no reports on the transesterification of oil to produce biodiesel. A mutant PCL G47I with an improved thermostability was previously obtained through replacing Gly47 with Ile in PCL. In this study, a novel Pichia pastoris whole-cell biocatalyst (WCB) with overexpression of PCL G47I was constructed and characterized for biodiesel production from soybean oil. The optimum conditions for biodiesel preparation were 1 g soybean oil, 1 : 2 initial oil/methanol molar ratio with 3 times methanol addition of 1 : 0.75 oil/methanol molar ratio at 4 h intervals, 7% water content, 400 U lipase, temperature of 25 °C, and reaction time of 20 h. Under the optimum conditions, the FAME yield reached 60.7% and remained 47.3% after 4 batch cycles, and no glycerol was generated as a byproduct. These findings indicated that this WCB is a promising biocatalyst for biodiesel production in a relatively cost-effective manner. Additionally, the resulting enzymatic process may provide a potential method for biodiesel production at an industrial scale.