Directed Evolution of an Esterase from Pseudomonas fluorescens Yields a Mutant with Excellent Enantioselectivity and Activity for the Kinetic Resolution of a Chiral Building Block
2006; Wiley; Volume: 7; Issue: 5 Linguagem: Inglês
10.1002/cbic.200500546
ISSN1439-7633
AutoresMarlen Schmidt, Daniel Hasenpusch, Markus Kähler, U. Kirchner, Kerstin Wiggenhorn, Walter Langel, Uwe T. Bornscheuer,
Tópico(s)Pharmacogenetics and Drug Metabolism
ResumoAbstract A triple mutant of an esterase from Pseudomonas fluorescens (PFE) that was created by directed evolution exhibited high enantioselectivity ( E =89) in a kinetic resolution and yielded the building block ( S )‐but‐3‐yn‐2‐ol. Surprisingly, a mutation close to the active site caused the formation of inclusion bodies, but remote mutations were found to be responsible for the high selectivity. Back mutations gave a variant (double mutant PFE Ile76Val/Val175Ala) that showed excellent selectivity ( E =96) and activity (20 min for 50 % conversion, which corresponds to 1.25 U per mg of protein).
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