Biodiesel Production via Genetically Altered Lipase from Proteus mirabilis Transesterification in Methyl Acetate

Lipase catalyzed biodiesel production presents a potential methodology that exceeds the traditionally utilized acid/base protocols. Advantages include economic favorability and convenience due to more efficient catalysis, which leads to less time, starting material, and energy required for production. A lipase has been previously genetically engineered for methanol tolerance and was shown to be an effective catalysis for transesterification reactions utilizing methanol to produce fatty acid methyl esters, or biodiesel, and glycerol. An alternate acyl acceptor to methanol, methyl acetate, produces a fuel stabilizer, triacetin, as opposed to glycerol when used in the transesterification. This methodology has not been previously utilized with the characterization of these genetically engineered lipases, which are also described as dieselzymes. Dieselzyme mutations have been expressed in E.coli using a pET28 bacterial expression vector and purified utilizing immobilized metal ion affinity chromatography. Kinetic optimization was performed for three mutants using p‐nitrophenyl palmitate as substrate at varying pHs and in methyl acetate versus methanol reactions. Further investigation of the dieselzymes includes use of synthetic triglycerides as substrate to continue kinetic optimization. Support or Funding Information University of Tampa Department of Biochemistry, Chemistry and Physics Summer Fellowship; University of Tampa Honors Program Fellowship; University of Tampa David Delo Research Professor Grant; University of Tampa Dana Foundation Grant