title: Enhancing the Acyltransferase Activity of Candida antarctica Lipase A by Rational Design.
abstract: A few lipases, such as Candida antarctica lipase A (CAL-A), are known to possess acyltransferase activity. This enables the enzyme to synthesize fatty acid esters from natural oils and alcohols even in the presence of bulk water. Unfortunately, fatty acids are still formed in these reactions as undesired side-products. To reduce the amount of fatty acids, several CAL-A variants were rationally designed based on its crystal structure. These variants were expressed in Escherichia coli and Pichia pastoris, purified, and their acyltransferase/hydrolase activities were investigated by various biocatalytic approaches. Among the investigated variants, mutant Asp122Leu showed a significant decrease in the hydrolytic activity, thus reducing the side-product yield during acylation. As desired, this variant retained wild-type process-relevant features like pH profile and thermostability.
authors.affiliation: Institute of Biochemistry, Department of Biotechnology and Enzyme Catalysis, Greifswald University, Felix-Hausdorff-Strasse 4, 17487, Greifswald (Germany).
authors.forename: Janett
authors.initials: J
authors.lastname: Müller
authors.affiliation: Institute of Biochemistry, Department of Biotechnology and Enzyme Catalysis, Greifswald University, Felix-Hausdorff-Strasse 4, 17487, Greifswald (Germany).
authors.forename: Miriam A
authors.initials: MA
authors.lastname: Sowa
authors.affiliation: Institute of Biochemistry, Department of Biotechnology and Enzyme Catalysis, Greifswald University, Felix-Hausdorff-Strasse 4, 17487, Greifswald (Germany).
authors.forename: Birte
authors.initials: B
authors.lastname: Fredrich
authors.affiliation: Enzymicals AG, Walther-Rathenau-Strasse 49a, 17489 Greifswald (Germany).
authors.forename: Henrike
authors.initials: H
authors.lastname: Brundiek
authors.affiliation: Institute of Biochemistry, Department of Biotechnology and Enzyme Catalysis, Greifswald University, Felix-Hausdorff-Strasse 4, 17487, Greifswald (Germany). uwe.bornscheuer@uni-greifswald.de.
authors.forename: Uwe T
authors.initials: UT
authors.lastname: Bornscheuer
mesh_terms: D000217:Acyltransferases / Q000737:chemistry / Q000378:metabolism*; D002175:Candida / Q000201:enzymology*; D020134:Catalytic Domain; D003198:Computer Simulation; D004591:Electrophoresis, Polyacrylamide Gel; D008049:Lipase / Q000378:metabolism*; D008958:Models, Molecular; D015394:Molecular Structure; D015202:Protein Engineering
chemical_list: D000217:Acyltransferases; D008049:Lipase
keywords: CAL-A; acyltransferases; biocatalysis; ester synthesis; immobilization