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Combinatorial Library Based Engineering of Candida antarctica Lipase A for Enantioselective Transacylation of sec-Alcohols in Organic Solvent
KTH, School of Biotechnology (BIO), Industrial Biotechnology. Stockholm University, Sweden . (Albanova University Center)ORCID iD: 0000-0003-2371-8755
2015 (English)In: Angewandte Chemie International Edition, ISSN 1433-7851, E-ISSN 1521-3773, Vol. 54, no 14, 4284-4288 p.Article in journal (Refereed) Published
Abstract [en]

A method for determining lipase enantioselectivity in the transacylation of sec-alcohols in organic solvent was developed. The method was applied to a model library of Candida antarctica lipase A (CalA) variants for improved enantioselectivity (E values) in the kinetic resolution of 1-phenylethanol in isooctane. A focused combinatorial gene library simultaneously targeting seven positions in the enzyme active site was designed. Enzyme variants were immobilized on nickel-coated 96-well microtiter plates through a histidine tag (His(6)-tag), screened for transacylation of 1-phenylethanol in isooctane, and analyzed by GC. The highest enantioselectivity was shown by the double mutant Y93L/L367I. This enzyme variant gave an E value of 100 (R), which is a dramatic improvement on the wild-type CalA (E=3). This variant also showed high to excellent enantioselectivity for other secondary alcohols tested.

Place, publisher, year, edition, pages
2015. Vol. 54, no 14, 4284-4288 p.
Keyword [en]
biocatalysis, kinetic resolution, lipase A, protein engineering, secondary alcohols
National Category
Chemical Sciences
URN: urn:nbn:se:kth:diva-165210DOI: 10.1002/anie.201410675ISI: 000351679600024PubMedID: 25676632ScopusID: 2-s2.0-84925582964OAI: diva2:810384

QC 20150507

Available from: 2015-05-07 Created: 2015-04-24 Last updated: 2015-05-07Bibliographically approved

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Humble, Maria Svedendahl
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