Back to Search
Start Over
Learning from Directed Evolution: Further Lessons from Theoretical Investigations into Cooperative Mutations in Lipase Enantioselectivity
- Source :
- ChemBioChem. 8:106-112
- Publication Year :
- 2007
- Publisher :
- Wiley, 2007.
-
Abstract
- An earlier experimental study, which involved the directed evolution of enantioselective lipase variants from Pseudomonas aeruginosa as catalysts in the hydrolytic kinetic resolution of 2-methyl-decanoic acid p-nitrophenyl ester, provided a mutant with six mutations. Consequently, the selectivity factor was found to increase from E = 1.1 for the wild-type to E = 51 for the best mutant. Only one of the amino acid exchanges in this mutant was found to occur next to the binding pocket, the other mutations being remote. Our previous theoretical analysis with molecular-dynamics simulations helped to unveil the source of enhanced enantioselectivity: a relay mechanism that involves two of the six mutations was shown to induce strong cooperativity. In this investigation, single, double, and triple mutants were constructed and tested as enantioselective catalysts. This study supports our original postulate regarding the relay mechanism, offers further mechanistic insight into the role of individual mutations, and provides mutants that display even higher enantioselectivity (E of up to 64).
- Subjects :
- Models, Molecular
Molecular model
Stereochemistry
Mutant
Molecular Conformation
Cooperativity
Biology
Crystallography, X-Ray
Biochemistry
Catalysis
Kinetic resolution
Lipase
Molecular Biology
DNA Primers
chemistry.chemical_classification
Organic Chemistry
Enantioselective synthesis
Esters
Stereoisomerism
Directed evolution
Amino acid
Kinetics
Genetic Techniques
Models, Chemical
chemistry
Mutation
Pseudomonas aeruginosa
biology.protein
Molecular Medicine
Directed Molecular Evolution
Software
Subjects
Details
- ISSN :
- 14397633 and 14394227
- Volume :
- 8
- Database :
- OpenAIRE
- Journal :
- ChemBioChem
- Accession number :
- edsair.doi.dedup.....0547e5cb841060e0b098621819e9f1e4