title: Rational design of a new one-step purification strategy for Candida antarctica lipase B by ion-exchange chromatography.
abstract: A fast and efficient one-step method for purification of lipase B from Candida antarctica by ion-exchange chromatography was developed by rational design. The electrostatic properties of the enzyme were calculated and validated by isoelectric focusing and measurement of the titration curve. C. antarctica lipase B shows an unusual pH profile with a broad isoelectric region from pH 4 to 8. At pH 3 C. antarctica lipase B can be bound to a cation-exchange chromatography column and was purified to homogeneity with a purification factor of 2.4. It was stable at pH 3, the residual activity was still 80% after 6 days incubation at 20 degrees C. The broad isoelectric region of C. antarctica lipase B is unique as compared to almost all other alpha/beta-hydrolases which have a well-defined isoelectric point. A search in the lipase engineering database resulted in only one further alpha/beta-hydrolase, the Fusarium solani cutinase, which also has a broad isoelectric region.
authors.affiliation: Institute of Technical Biochemistry, University of Stuttgart, Allmandring 31, D-70569 Stuttgart, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Trodler
authors.forename: Jens
authors.initials: J
authors.lastname: Nieveler
authors.forename: Monika
authors.initials: M
authors.lastname: Rusnak
authors.forename: Rolf D
authors.initials: RD
authors.lastname: Schmid
authors.forename: Jürgen
authors.initials: J
authors.lastname: Pleiss
mesh_terms: D002175:Candida / Q000201:enzymology*; D002852:Chromatography, Ion Exchange / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005656:Fungal Proteins; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D008049:Lipase / Q000302:isolation & purification*; D008958:Models, Molecular
chemical_list: D005656:Fungal Proteins; D008049:Lipase; C431349:lipase B, Candida antarctica